|
 
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| ABSTRACTS |
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| Year : 2022 | Volume
: 13
| Issue : 4 | Page : 146-231 |
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Abstracts of 5th Asian congress of radiation research (5th ACRR) and 3rd biennial meeting of the society for radiation research
| Date of Web Publication | 04-Nov-2022 |
Correspondence Address:
 Source of Support: None, Conflict of Interest: None  | Check |
DOI: 10.4103/2588-9273.360475
How to cite this article:
. Abstracts of 5th Asian congress of radiation research (5th ACRR) and 3rd biennial meeting of the society for radiation research. J Radiat Cancer Res 2022;13:146-231 |
How to cite this URL:
. Abstracts of 5th Asian congress of radiation research (5th ACRR) and 3rd biennial meeting of the society for radiation research. J Radiat Cancer Res [serial online] 2022 [cited 2023 Jun 7];13:146-231. Available from: https://www.journalrcr.org/text.asp?2022/13/4/146/360475 |
Organised by
Bio-Science Group, Bhabha Atomic Research Centre, Mumbai
in association with
Society for Radiation Research-India and Asian Association for Radiation Research (AARR)
DAE Convention Centre, Anushaktinagar, Mumbai
Nov. 17-20, 2022
Contents
Special Invited Lecture (SL)
Plenary Lecture (PL)
Invited Lecture (IL)
AARR Award Recipient Lecture (AARRL)
SRR Young Scientist Award Presentation (SRR-YSA)
Oral Presentation (OP)
O1: Radiation Biology for Risk assessment, Radioprotection and Cancer Radiotherapy
O2: Radiation Physics/Chemistry, Food Irradiation, Nuclear Agriculture and Environment
Poster Presentation (P)
| Special Invited Lecture (SL) | |  |
| SL-1 | | |
| Dual Immunomodulatory and Radiotherapeutic Nanoradiopharmaceuticals as a New Paradigm in Cancer Therapy – Interventions of Green Nanotechnology and Nano-Ayurvedic Medicine toward the Development of MGF-198AuNPs Nanomedicine Agent | | |
Kattesh V. Katti
Institute of Green Nanotechnology, University of Missouri, Columbia, Missouri, USA
E-mail: [email protected]
Introduction: Various radiative energies, including Gamma radiation-upon interaction with tumor cells-results in irreparable and indiscriminate damage to both the normal and cancerous cells-all resulting in cascade of immune system cell suppressing signaling pathways that obliterate preventing tumor cell growth, resulting in uncontrolled tumor cell division, and ultimately causing drug resistance and whole-body metastases.Radiotherapeutic agents with capabilities to deliver effective radiation dose with concomitant immunomodulatory characteristics-all built within a singular radiopharmaceutical matrix-will provide the best means to treat myriad of human cancers effectively. Materials and Methods: This lecture focuses on the development of Mangiferin functionalized Au-198 nanoparticles (MGF-198AuNPs) as an innovative dual action-immunomodulatory NF-κB suppressing and radiation therapy tool-with tumor microenvironment targeting characteristics. MGF-198AuNPs provides a desirable beta energy emission and half-life that destroys tumor cells/tumor tissue (βmax = 0.96 MeV; half-life of 2.7 days) while the presence of mangiferin suppresses pro tumor NF-κB signaling pathways. Results: Motivated by ancient Ayurvedic medicine, the nonradioactive surrogate MGF-AuNPs was developed to expand the scope of this nanomedicine agent as an immunomodulatory Nano Ayurvedic Medicine. Our invention of a new medical modality called as Nano Ayurvedic Medicine, approved by the US Patents and Trademarks Office, has been recently validated through successful human clinical trials of treating triple negative breast cancer patients. This lecture will provide: (a) full in vivo investigations on therapeutic properties of MGF-198AuNP agent in treating prostate tumors; (b) Immunomodulatory effects of MGF-198AuNPs through targeting the tumor microenvironments and thereby suppressing NF-κB transcription factors while promoting antitumor M-1 macrophages; and (c) Estimation of the dose distribution delivered by radioactive gold nanoparticles (MGF-198AuNPs ) to the tumor inside the human prostate using Monte-Carlo N-Particle code (MCNP-6.1.1 code). Conclusions: The overall implications of developing of dual radiotherapy and immunomodulatory MGF-198AuNPs nanoradiopharmaceuticals exhibiting immunomodulatory therapeutics for applications in oncology will be discussed.
| SL-2 | | |
| The Linear No-Threshold Dose Response Model: An Assessment of its Historical and Scientific Foundations | | |
E. J. Calabrese
Department of Environmental Health Sciences, School of Public Health and Health Sciences, University of Massachusetts, Amherst, Massachusetts, USA
E-mail: [email protected]
The linear no-threshold (LNT) single-hit (SH) dose response model for cancer risk assessment is assessed with respect to its historical foundations. This presentation also examines how mistakes, ideological biases, and scientific misconduct by key scientists affected the acceptance, validity, and applications of the LNT model for cancer risk assessment. In addition, the analysis demonstrates that the LNT single-hit model was inappropriately adopted for governmental risk assessment, regulatory policy, practices, and for risk communication.
| SL-3 | | |
| Mechanisms of DNA Double Strand Break Repair in Mammalian Cells | | |
George Iliakis1,2, Emil Mladenov1,2, Aashish Soni1,2, Fanghua Li2, Veronika Mladenova1,2, Lisa Krieger1,2, Gabriel E. Pantelias3, and Martin Stuschke1,4
1Department of Radiation Therapy, Division of Experimental Radiation Biology, University of Duisburg-Essen, University Hospital Essen, 2Institute of Medical Radiation Biology, University of Duisburg-Essen Medical School, 4German Cancer Research Center (DKFZ) and German Cancer Consortium (DKTK), Partner Site University Hospital Essen, Essen, Germany,3Institute of Nuclear Technology and Radiation Protection, National Centre for Scientific Research ”Demokritos,”Aghia Paraskevi Attikis, Athens, Greece
E-mail: [email protected]
In vertebrates, DNA double-strand breaks (DSBs) are removed by end-joining: classical non-homologous end-joining (c-NHEJ) and alternative end-joining (alt-EJ); or by homology-dependent processes: homologous recombination (HR) and single-strand annealing (SSA). These DSB processing options are not alternatives operating with equal efficiency, but show instead striking differences in speed, accuracy and cell-cycle-phase dependence and are associated with widely different risks to genome stability. Therefore, characterization of engagement-parameters is essential for a mechanistic understanding of the effects of DSB-inducing agents, such as ionizing radiation (IR), on cells. We address pathway selection in G2-phase of the cell cycle, where all above pathways are functional. Using detailed analysis of the formation and decay of -H2AX and RAD51 foci at different doses of IR, we discover regulatory confinements in HR with direct consequences for SSA- and c-NHEJ-engagement. Most notably, we find pronounced suppression of HR with increasing DSB-load that is not due to RAD51 availability and which is delimited but not defined by 53BP1 and RAD52. Strikingly, at low DSB loads, HR repairs ~50% of DSBs, whereas at high DSB-loads its contribution is undetectable. With increasing DSB-load and the associated suppression of HR, SSA gains ground, while alt-EJ is suppressed. These observations explain earlier, apparently contradictory results and advance our understanding of logic and mechanisms underpinning the wiring between DSB repair pathways. They show that cells of higher eukaryotes are programmed to use error-free DSB processing pathways, as HR, when low numbers of DSBs are present in their genome, but shift gears and involve c-NHEJ and ultimately also alt-EJ and SSA as the load of DSBs increases.
Acknowledgement: Grant support by German Federal Ministry of Education and Research (BMBF-02NUK037B, BMBF-02NUK043B and BMBF-02NUK054B), German Research Foundation (DFG-GRK1739, DFG-IL51/10, DFG-IL51/11), German Federal Ministry for Economic Affairs (BMWi-50WB1836 and BMWi-50WB2118), German Academic Exchange Service (DAAD-575138800).
| SL-4 | | |
| Low-Dose and Low-Dose-Rate Radiation Effects: What Have We learned from Long-Term Animal Studies? | | |
Gayle E. Woloschak
Northwestern University School of Medicine, Chicago, Illinois
E-mail: [email protected]
Introduction: The Department of Energy embarked on the study of long-term effects of radiation beginning from around 1940 until 1995 by examining life-shortening effects of ionizing radiation on burros, pigs, rats, mice, dogs, primates and other species. Materials and Methods: Of these many studies, data are available for most while tissues remain only from some rats, dogs, and mice. A total of over 120,000 animals were studied (and probably the number is much larger!). The experiments were done in a systematic way with tissues studied and archived, detailed pathologies and diseases recorded, and much of the information provided in database format. Many of the logistics of the experiments, data on each individual animal for a variety of endpoints, and available tissues from the animal can be found at the janus.northwestern.edu/wololab website. Results: Experiments were done to evaluate effects of radiation quality, dose and dose-rate, effects of in utero exposures, external beam and internal emitters, and many more. Among the results to be presented are findings that relate to low-dose and low-dose rate effects on life-shortening and cancer in animals as well as applications that relate to DDREF. Conclusions: This work demonstrates that studies done over 50 years ago can be analyzed differently to provide new insights about the field. In addition, experiments comparing multiple species can be important in uncovering common themes in radiation effects.
| SL-5 | | |
| Application of Machine Learning in Gamma and Neutron Spectroscopy | | |
Omololu Makinde
Mechanical Engineering, University of South Florida, Florida, USA
E-mail: [email protected]
Neutron spectra and effects of varying energies on absorbed dose, dose-equivalent measurement are critical in interpretation of instruments and dosimeter responses. However, they are more significant in bio-dosimetry operations for radiation effects. Most irradiation for research focuses on the ratio of neutrons to gamma in a mixed field as a characteristic of the radiation field being used. However, the effects of fast neutrons of varying energy bands on biological samples can be significant and could change the efficacy of any drug developed as a countermeasure. The biggest reason for not characterizing radiation fields for neutron spectra is related to the difficulty in measurement and determination of neutron spectra. Current methodology involves Neutron spectrum unfolding using activation foils of select material, the use of multi-sphere or Bonner Sphere and finally a helium-3 spectrometer. The cumbersome nature of these experiments forces radiation-providing facilities to only conduct them once in a long period for specific configurations, which then set the only known configurations the facility can use. The availability of data and current machine learning techniques can bridge this gap by utilizing historical data sets from facilities and stochastic monte carlo models to develop deep neural networks that can predict the neutron spectra accurately. The Armed Forces Radiobiology Research Facility's mission is to conduct radiobiological research in support of the United States Department of Defense and other health agencies that could possibly work in radiation fields. The institute provides sources for scientists to determine biological effects and develop possible countermeasures. The primary source for providing mixed field radiation is a GA TRIGA nuclear reactor. Exposure is done in a large room that allows for varying targets and shielding conditions to generate varying radiation fields. However, only a handful of configurations have been characterized for neutron spectra, thereby limiting the ability to use the source. This characterization was conducted and published by Verbinski, et al. In 1981 and validated later by the National Institute of Standards and Technology, hence providing a link to national standard for spectra and source for research. In order to allow the facility to use all the space and create varying radiation fields that can be traced to a national standard, we will use sparse data from the historical measured spectra and spectra developed from a US Department of Energy Code to create a regression model. This sparse model will then be used to predict results in multiple configurations in the exposure room. The error will then be minimized by collecting data at the predicted configurations. Finally, a recommendation system will be built using a deep neural network to provide reactor configurations to deliver any desired neutron and gamma spectra in the exposure room.
| Plenary Lecture (PL) | | |
| PL-1 | | |
| Radiation-Induced Damage to Cellular DNA: Product Formation and Mechanistic Insights | | |
Jean Cadet, J. Richard Wagner
Département De Médecine Nucléaire Et Radiobiologie, Faculté De Médecine Et Des Sciences De La Santé, Université De Sherbrooke, Sherbrooke, Canada
E-mail: [email protected]
Major advances have been made during the last two decades on the formation in cells of several classes of radiation-induced genomic damage that include 20 single oxidized pyrimidine and purine bases and a few tandem lesions. This has necessitated extensive mechanistic studies initially performed on simple model compounds that were progressively extended to oligonucleotides and isolated DNA in order to take into consideration several key structural and conformational features. Concerning the reactive oxygen species potentially generated by radiolysis of water molecules only hydroxyl radical (•OH), unlike superoxide anion radical and hydrogen peroxide, is able in aqueous solution to react with both the bases and the 2-deoxyribose moieties. Therefore exposure of aerated aqueous solutions of DNA samples gives rise to several dozen of modified base compounds that were unambiguously identified and their mechanism of formation established. Thus, evidence was provided that the 5,6-double bond of pyrimidine bases is highly susceptible to •OH addition whereas the methyl group of thymine and 5-methylcytosine is subject to •OH-mediated hydrogen atom abstraction. It was also found that •OH is able to add to the C8 of purine bases thus generating a transient radical that in the case of guanine is converted into 8-oxo-7,8-dihydroguanine (8-oxoGua) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyGua) through one-electron and one-reduction respectively. It may be pointed out that ionization processes upon exposure of aqueous solutions of DNA components concerns essentially water molecules. However DNA bases could be ionized in order to mimic qualitatively the direct effects of ionizing radiation using either type I photosensitizers or more suitably high intensity nanosecond UVC laser sources. Interestingly the oxidation reactions issued from the generation of base radical cations mimic partially the effects of •OH in terms of final products. However, evidence was shown that in native DNA efficient positive hole transfer occurs leading to preferential formation of 8-oxoGua and FapyGua at the expense of pyrimidine oxidation products. The development of sensitive and accurate methods (HPLC-ESI-MS/MS, modified alkaline comet assay) has allows the measurement of very low amounts of several radiation-induced modifications in cellular DNA. Thus as an illustration it was reported that 20 8-oxoGua lesions are formed per 10-9 nucleobases and per Gy. Several conclusions may be drawn from the comparison of the measurement of radiation-induced (•OH and ionization) and bi-photonic (ionization) UVC mediated formation of oxidized nucleobases in cells with model studies:
- Aerated aqueous systems are convenient to mimic conditions of oxidation reactions mediated by both •OH and ionization in cells
- Efficient charge transfer reactions occur in cellular DNA as in isolated DNA
- Damage mediated by •OH (indirect effect) is predominant other ionization (direct effect)
- Several putative lesion (purine 5', 8-cyclo-2'-deoxyribonucleosides, secondary 8-oxoGua oxidation products are not detectable even upon exposure to high doses of ionizing radiation.
| PL-2 | | |
| Molecular Radiation Biology on DNA Damage Response for Radiotherapy and Radioprotection in Next Generation | | |
Y. Matsumoto
Laboratory for Zero-Carbon Energy, Institute of Innovative Research, Tokyo Institute of Technology, Tokyo, Japan
E-mail: [email protected]
DNA damage is considered the key issue in the biological effects of ionizing radiation. The researches in the past three decades have revealed the molecular frameworks of DNA damage response (DDR), including DNA repair and cell cycle checkpoint. There are extensive studies to develop therapeutic strategies for cancer targeting DDR. There are also increasing attention to inherent variation in radiosensitivity and cancer susceptibility among individuals from the aspect of radiotherapy and radioprotection. Among various types of DNA damages, DNA double-strand break (DSB) is considered the most deleterious type. DNA-dependent protein kinase (DNA-PK) as well as Ataxia-telangiectasia mutated (ATM) is thought to act as the sensor for DSB and orchestrate various cellular responses to DSB. DSBs are repaired mainly through homologous recombination (HR) and non-homologous end joining (NHEJ), in which DNA-PK play a central role with DNA ligase IV and its associated proteins XRCC4 and XLF. We have sought to elucidate the role of protein phosphorylation by DNA-PK in NHEJ and also the relationship between the abundance or status of NHEJ factors and the outcome of radiotherapy among individuals.Mutations, which arise from the failure in DDR, lead to cancer and hereditary effects. While classical studies on mutation were conducted looking at specific loci, there are increasing number of studies employing the whole genome sequencing (WGS). In addition to superior sensitivity, WGS can detect the mutations in non-coding region and synonymous mutations and is bringing a revolution on the concept of mutation. We are applying this approach to the studies on the effects of low dose/low dose-rate radiation in terms of mutagenesis. I will overview the current understanding and unsettled questions in the molecular mechanisms of DDR, especially in DSB repair, and discuss the future directions toward the contribution in radiotherapy and radioprotections in the next generation.
| PL-3 | | |
| Are Many Patients Getting High Doses (Over 100 mSv) in a Few Years and What Can Be Done? | | |
Madan Rehani
Massachusetts General Hospital, Boston, Massachusetts, USA
E-mail: [email protected]; [email protected]
Recent publications from our group have shown that the number of patients who are receiving cumulative doses in three digits of mSv of effective dose or in three digits of mGy as organ doses in a short period of one day to a few years are much more than ever known or anticipated. There is a better understanding now of the age distribution of these patients, the disease types, and clinical reasons that lead to recurrent imaging. This has resulted in joint position statements led by International Atomic Energy Agency (IAEA) with several professional societies, and this is a hot topic currently. The purpose of this talk is to create awareness of this rare situation never witnessed before since the discovery of X-rays and radioactivity which has implications on patient radiation safety.The major contribution to the dose comes through computed tomography (CT), fluoroscopy guided interventional (FGI) procedures, hybrid imaging and imaging in radiotherapy with CT being the largest contributor. The most significant change in future can be envisaged from technological advances in imaging equipment. There has been development of photon counting detector and also of monochromatic source of X-ray that is similar to current day X-ray tube. With nearly a million patients getting added every year globally, there is great potential for radiation biologists/ epidemiologists and radiation scientists to use this patient cohort to study radiation effects.
| PL-4 | | |
| Current Status and Translational Challenges of Emerging Radiotherapy Techniques | | |
S. D. Sharma1,2
1Radiological Physics and Advisory Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, NTS, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Radiotherapy is one of the important techniques of cancer management. Varieties of radiotherapy techniques and equipments are used in treatment of a number of cancer cases. Newer techniques and technologies are being introduced at a rapid rate to achieve the goal of radiotherapy. A number of new innovations have taken place in the recent past which needs to be reviewed in terms of physics, technology, biology, dosimetry and translational challenges. Materials and Methods: The radiotherapy techniques included in this review includes Ultra-high dose rate radiotherapy (popularly known as FLASH-RT), nanoparticle assisted radiotherapy (NPART), microbeam radiotherapy (MRT) and intensity modulated brachytherapy (IMBT). FLASH-RT has gained significant momentum after publications of a number of reports of in-vivo studies. Metallic nanoparticles are thought to be a good enhancer of radiation dose to the tumour. Kilovolatge (kV) x-ray based MRT has emerged as a promising alternative to conventional radiotherapy because of unconventionally high dose delivery to tumour keeping normal tissue dose well below the tolerance level. IMBT uses anisotropic radiation profile generated through shielding designs incorporated onto sources and applicators to achieve highly conformal isodose coverage to tumour. Results: These emerging techniques of radiotherapy have been demonstrated to be effective in managing varieties of cancer cases. FLASH-RT is thought to be a potential techniqe to revolutionize the radiotherapy. However, there are some translational challenges associated with these radiotherapy techniques. Conclusions: Emerging radiotherapy techniques are highly promising and efforts are on to overcome the translational challenges.
| PL-5 | | |
| The Cancer Cell Adhesion Resistome: Mechanisms and Its Diagnostic and Therapeutic Potential | | |
N. Cordes1,2,3,4
1OncoRay – National Center for Radiation Research in Oncology, Faculty of Medicine Carl Gustav Carus, Technische Universität Dresden, 2Department of Radiotherapy and Radiation Oncology, University Hospital Carl Gustav Carus, Technische Universität Dresden, 3Helmholtz-Zentrum Dresden – Rossendorf, Institute of Radiooncology – OncoRay, Dresden, 4German Cancer Consortium (DKTK), Partner Site Dresden, and German Cancer Research Center (DKFZ), Heidelberg, Germany
E-mail: [email protected]
Introduction: A large variety of extracellular and intracellular factors contribute to inherent and acquired cancer resistance mechanisms. Cell-extracellular matrix (ECM) interactions as part of multiprotein complexes called focal adhesions (FAs) are one of them. The several hundred signaling and adaptor proteins coalesced in FAs allow fundamental influence on numerous signaling networks dysfunctional in cancer cells that promote survival. My review presents the current knowledge on the Cancer Cell Adhesion Resistome and outlines their diagnostic and therapeutic potential. Materials and Methods: Own studies and studies by others. Results and Conclusions: The Cancer Cell Adhesion Resistome offers numerous diagnostic and therapeutic opportunities for intervening in resistance, proliferation, migration/invasion, and metastasis.
| PL-6 | | |
| Rationale for Combining Hyperthermia and Radiotherapy: Current Status and Future Directions | | |
H. Crezee1,2
1Department of Radiation Oncology, Amsterdam University Medical Centers, Location University of Amsterdam, 2Cancer Center Amsterdam, Amsterdam, Netherlands
E-mail: [email protected]
Introduction: Hyperthermia (HT), raising the tumour temperature to 40-43°C, is a potent Radiosensitizer and is clinically applied in combination with radiotherapy (RT) for many different tumour sites. Multiple synergistic working mechanisms are responsible for the therapeutic effect of HT. Research is ongoing to gain better understanding of the contribution of different working mechanisms to hyperthermic radiosensitisation, this knowledge may help to further optimize its effectiveness during clinical application of HT. Goal: Providing an overview and understanding of the clinical and biological rationale of using HT as a radiosensitizer. Results: Mechanisms are known to include direct cell kill of hypoxic tumour cells, blockage of DNA damage repair, enhanced perfusion, re-oxygenation of tumour cells, and immunogenic effects. Each of these effects has a different dose-effect relationship and a different ideal timing of hyperthermia with respect to radiotherapy delivery. For instance, temperatures exceeding 41°C and a short time interval between RT and HT are required to block Homologous Recombination repair of DNA double Strand Breaks. HT is also the optimal clinical strategy to overcome radioresistance associated with tumor hypoxia. Current research is evaluating and optimizing combinations of radiotherapy, hyperthermia and immunotherapy, with promising results. Conclusions: The proven clinical and biological rationale warrants further research and wider clinical application of HT in combination with radiotherapy.
| PL-7 | | |
| Radiation Biology Research at IUAC | | |
A. C. Pandey
Inter-University Accelerator Centre, Aruna Asaf Ali Marg, New Delhi, India
The ongoing radiation biology research at IUAC investigates the effects of accelerated charged particles on the biological systems at the molecular level. There is a dedicated beamline for Heavy Ion Radiation Biology for experiments using the low flux of accelerated ions. The areas of research include modeling of radiation effects, exploration of pathways to radiation induced effects with potential applications in many important areas such as Heavy Ion Cancer therapy and Radiation protection in manned space flights. IUAC has a dedicated radiation biology beamline having ASPIRE [Automated Sample Positioning and Irradiation system for radiation biology Experiments] where cells can be irradiated in air at atmospheric pressure to the preset dose of accelerated heavy ions with a very good dose uniformity. Apart from that, there is a dedicated radiation biology laboratory having basic equipment to facilitate the sample preparation and post irradiation treatments like CO2 incubators, Image based cell counter, an upright Fluorescence microscope, Multimode Plate Reader, Field Inversion Gel electrophoresis two biosafety cabinets, one small laminar flow bench, refrigerated centrifuges, a microplate washer and deep refrigerators. The facilities allow for conducting analytical procedures involving gene expression studies using PCR, Western Blot, Fluorescence Immunostaining studies etc. by the university users. The talk will discuss some of the studies undertaken utilizing IUAC facilities. Signaling pathways of activation and secretion of Matrix Metalloproteinases from human lung carcinoma cells and Radiosensitization of human cancer cells using G-quadruplex ligands have been studied. Recently, the group has intensively studied the effects of 62 MeV carbon ion beam on mitochondrial fission-fusion homeostasis and electron transport chain (ETC) function in HEK 293T cells. In another study, modulation of autophagy and the related mechanism by 62 MeV C ion beam in human embryonic intestinal cells (INT407 cells) were investigated without and with treatment with autophagy inducers/blockers. There is an indication that LiCl treatment can be used as a radio-protective agent against autophagy. Conversely, pre-treatment with autophagy inducers increased the accumulation of monodansylcadaverine and altered the distribution and size of the autophagic vacuoles in ion irradiated cells.
| PL-8 | | |
| Hypo-Fractionated Radiotherapy a Paradigm Shift in the Concept and Practices of Radiobiology | | |
S. P. Mishra, Anoop Kr. Srivastava, Avinav Bharati, Farhana Khatoon, Madhup Rastogi
Department of Radiation Oncology, Dr. Ram Manohar Lohia Institute of Medical Sciences, Lucknow, Uttar Pradesh, India
E-mail: [email protected]
Introduction: Fractionation and fraction size has remained in debate from the beginning of the radiotherapy in cancer treatment. The early breakthrough in radiobiological understanding advocated a fraction size of 2Gy per fraction and it has remained the most sacrosanct regimen. Large or single fraction using x-rays were used very early which may have been as large as 20Gy. A paradigm shift has been introduced by SRS, SBRT (>20Gy per ffraction) and flash RT (>40Gy/sec) in hypo-fractionation. Hypo-fractionated RT is being practiced in treatment of cancer of breast, prostate, lung and many oligometastatic sites with various fractionation sizes. The fractionation used varies from 300cGy to 20-25 Gy per fraction which has various implications which may be futuristic. Materials and Methods: SRS, SBRT (one to five fractions) and FLASH radiotherapy (FLASH-RT) delivers a single high dose at a mean dose rate >40 Gy/s in milliseconds to achieve similar tumor control to CONV-RT while sparingnormal tissues from detrimental injury. It provides an intriguing perspective in improving clinical outcomes fortumor patients as well as a novel way to enhance the differential responses between normal and tumor tissues. An attempt has been made to utilized the evaluations based BED, TCP, NTCP to arrive at a common denominator for comparison. Results: The established radiobiological tools for obtaining equivalence between the conventional fraction and other hypo fractionated may be only providing quantitative assessment. Conclusions: The large fraction size and flash RT are poised to challenge the established concept and practices and would demand clinic-pathological analysis to model the equivalence in RT.
| PL-9 | | |
| Modeling the Ageing of Immune Responses using Radiation and the Modulatory Effects of an Herbal Composition Hemohim | | |
S. K. Jo1,2, H. R. Park1, U. Jung3
1Research Division for Radiation Science, Advanced Radiation Technology Research Institute, Korea Atomic Energy Research Institute, Jeongeup-Si, 2Atomy Corp., Advisory Board, Gongju, 3Environmental and Disaster Assessment Research Division, Korea Atomic Energy Research Institute, Daejeon, South Korea
E-mail: [email protected]
Introduction: Ionizing radiation (IR) is known to accelerate ageing. In the ageing-related changes in immunity, the imbalance of helper Th1- and Th2-related immune responses is most interested. Materials and Methods: We investigated chronic effects on immune responses after ionizing radiation exposure (Gamma-ray, 5Gy; single or fractionated) in C57BL/6 mice in comparison with those of old mice. Also we evaluated the modulatory effects of a new herbal composition, HemoHIM, on the IR-induced immunological ageing model. HemoHIM had developed to protect the immune system against acute irradiation. Results: At 6 months after irradiation, spleen lymphocyte proliferation declined similarly to that of 18-months-old mice. In irradiated mice, Th1-related responses (IFN-γ, IgG2a) were lower but Th2-related responses (IL-4, IL-5, IgG1, IgE) were higher compared to the normal control mice of same age, showing similarity to those of old normal mice. The levels of IL-12p70, IL-12 receptors, and p-STAT4 were lower in the irradiated mice. The decrease of NK cell activity was noticeable in the irradiated mice showing lower values than those of old mice. Next, a new herbal composition, HemoHIM, was evaluated for the modulation of IR-induced immunological ageing. HemoHIM ameliorated the persistent immunological imbalance in fractionated-IR exposed mice by regulating IL-12/pSTAT4/SOCS3 pathway. Conclusions: IR accelerated the immunological ageing with respect to immune cell activity and Th1/Th2 balance. A new herbal composition, HemoHIM, modulated IR-induced immunological ageing and could be a good recommendation for the alleviation of the long-term complications after radiotherapy and for the immunomodulation in the elderly and the weak.
| PL-10 | | |
| Biomarkers of Radiation Injury: Identification of Novel Biomarkers for Acute Radiation Injury using Multi-Omics Approach and Nonhuman Primate Model | | |
Vijay K. Singh1,2
1Department of Pharmacology and Molecular Therapeutics, Division of Radioprotectants, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, 2Uniformed Services University of the Health Sciences, Armed Forces Radiobiology Research Institute, Bethesda, Maryland, USA
E-mail: [email protected]
Introduction: The availability of validated biomarkers to assess radiation exposure and to assist in developing medical countermeasures remains an unmet need. Materials and Methods: We used a cobalt-60 gamma-irradiated nonhuman primate (NHP) model to delineate a multi-omics-based serum probability index of radiation exposure. Both male and female NHPs were irradiated with different doses ranging from 6.0 to 8.5 Gy, with 0.5 Gy increments between doses. We leveraged high-resolution mass spectrometry for analysis of metabolites, lipids, and proteins at 1, 2, and 6 days post-irradiation in NHP serum. Results: A logistic regression model was implemented to develop a 4-analyte panel to stratify irradiated NHPs from unirradiated with high accuracy that was agnostic for all doses of gamma-rays tested in the study, up to six days after exposure. This panel was comprised of Serpin Family A9, acetylcarnitine, PC (16:0/22:6), and suberylglycine, which showed 2 – 4 fold elevation in serum abundance upon irradiation in NHPs and can potentially be translated as a molecular diagnostic for human use following larger validation studies. Conclusions: Taken together, this study, for the first time, demonstrates the utility of a combinatorial molecular characterization approach using an NHP model for developing minimally invasive assays from small volumes of blood that can be effectively used for radiation exposure assessments.
| PL-11 | | |
| Stress Responses in Mice Exposed to Space Radiation: Early and Late Onset | | |
Edouard I. Azzam1,2
1Radiobiology and Health Branch, Isotopes, Radiobiology and Environment Directorate (IRED), Canadian Nuclear Laboratories (CNL), Chalk River, Canada,2Department of Radiology, Division of Radiation Research, RUTGERS New Jersey Medical School, Newark, New Jersey, USA
E-mail: [email protected], [email protected]
Introduction: As space exploration missions to the moon and Mars are being contemplated, the need to translate/interpret the dose received by crew, during these prolonged journeys, into health risks (cancer & non-cancer) at a later time is essential. In this talk, aspects of oxidative stress, inflammatory responses and degenerative conditions in tissues of mice exposed to space radiation will be discussed. The role of the biophysical characteristics of the radiation and its mode of delivery will be considered. Materials and Methods: Middle-aged male CBA/CaJ mice (9-10 month old) were exposed (whole or partial body) to isovelocity 1 GeV/u protons, 40Ca20+, 28Si14+, or16O8+particles with respective average Linear Energy Transfer (LET) values of ~0.24, 14, 44, and 88 keV/μm. A different set of mice was exposed to 137Cs γ rays as reference radiation (LET ~ 0.9 keV/μm) to examine oxidative stresses and inflammatory responses in various organs that differ in their radiosensitivity. Results: Exposure to high charge and energy (HZE) particles resulted in significant early/transient as well as delayed effects on abundance of circulating hematopoietic cells and bone marrow stem cells. At 15 months after irradiation prominent injury reflective of oxidative and degenerative effects were detected in various tissues of mice exposed to absorbed doses as low as 20 cGy of HZE particles. Conclusions: Our ongoing analyses in male CBA/CaJ mice revealed that exposure of 10 month-old mice to HZE radiation leads to both short- and long-term biological changes: (i) perturbs abundance of hematopoietic cells; (ii) induces persistent oxidative stress and inflammatory effects in various tissues; (iii) is associated with changes in protein levels and post-translational modifications; (iv) induces non-targeted effects; (v) fractionation of the HZE particle dose often results in greater level of tissue damage than induced by a single bolus.
| PL-12 | | |
| Epidemiological and Biological Studies in High Level Natural Radiation Areas of Kerala Coast, India: An Overview | | |
Birajalaxmi Das
Low Level Radiation Research Section, Radiation Biology and Health Sciences Division, Bio Science Group, Bhabha Atomic Research Centre, Trombay, Mumbai, Maharashtra, India
E-mail: [email protected]
High level natural radiation areas (HLNRA) provide unique opportunity to study the effect of low dose and low dose rate ionizing radiation (LDIR) on human population. There are few prominent areas in the world, such as Kerala (India), Yangjiang (China), Guarapari (Brazil) and Ramsar (Iran), where human population is exposed to chronic LDIR since generations. The HLNRAs provide data below 100mSv which gives important information about the recent debate on Linear no threshold (LNT) hypothesis. The coastal belt of Kerala contains highest Th-232 deposits in its beach sand and due to patchy distribution of monazite, the level of radiation varies from <1.0 mGy to 45 mGy/year. Epidemiological and biological studies are conducted in this area to find out the effects and mechanism of chronic LDIR in human population. Till date, data on incidence of cancer, hereditary anomalies and late onset diseases did not show any adverse health effects. Biological end points such as chromosomal aberrations, micronuclei, premature chromosome condensation, etc. showed similar frequency in control and exposed population. The frequency of DNA damage in terms strand breaks, base damages and telomere length etc. did not show any increase in HLNRA Somatic and germinal mutations were similar in both the areas. However, efficient DNA repair and upregulation of DNA damage response (DDR), DNA repair and telomere specific genes are observed in HLNRA. Global transcriptome profile indicated abundance of DDR, repair, immune response, histone and chromatin modification genes along with survival pathways to be active in HLNRA. HLNRA population showed radio-adaptive response at multiple endpoints. High throughput next generation sequencing data has shed new insights on biological mechanisms in response to LDIR. In fact, the shape of the dose response curve seems to be non-linear both at epidemiological and biological end points, which provides important clue on recent debate of LNT hypothesis.
| PL-13 | | |
| Techniques for Retrospective Dosimetry in Radiological Emergency | | |
H. Yasuda
Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima, Japan
E-mail: [email protected]
Introduction: While ionizing radiation (”radiation” hereafter) has contributed to the development of our civilized society, a number of accidents accompanied with loss of human lives have occurred. In such radiological emergency situations, it is crucially important to estimate immediately and accurately the individual doses of subjects who were/might be exposed to high dose radiations. Meaning of Retrospective Dosimetry (RD): The term “retrospective dosimetry (RD)” means the determination of dose after the occurrence of a radiological emergency where the level of radiation exposure of individual(s) needs to be assessed in a retrospective way for medical decisions. In an unexpectedly occurred event, conventional personal dosimeters are usually unavailable and thus some of biological samples or fortuitous materials should be used for RD. The approaches for RD can be classified to three categories: clinical, biological and physical dosimetry. Clinical and Biological Approach: Clinical approach for RD is based on observed clinical conditions such as nausea, vomiting and diarrhea; and can be applied only to high dose exposure which induces those prodromal symptoms. Biological dosimetry uses some of the human tissues such as blood, tooth, bone and nail; and can work for the dose range lower than that of clinical dosimetry. The cytogenetic technique using peripheral blood lymphocytes is the most popular for biological dosimetry in radiological emergency. Physical Approach: Physical approach for RD normally uses some fortuitous objects that can stably keep readable radiation-induced changes. The most popular phenomena employed for this approach are electron spin/paramagnetic resonance (ESR/EPR) and luminescence emissions (thermoluminescence/TL, optically stimulated luminescence/OSL and radiophotoluminescence/RPL). Nuclear track detection (NTD) can be useful for RD of heavy charged particles including neutron-recoiled protons. New RD techniques based on recent scientific findings and novel technologies are being developed for more suitable response to radiological emergency.
| PL-14 | | |
| Radiation Tolerance and Probable DNA Repair Pathways in Nitrogen-Fixing Cyanobacterium, Nostoc sp. Strain PCC7120 | | |
Anurag Kirti1, Arvind Kumar1, Mitali Pradhan1,2, Sarita Pandey1, Hema Rajaram1,2
1Molecular Biology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Cyanobacteria are one of the ancient organisms having inhabited the earth over 2.8 billion years ago. The environmental conditions being harsh resulted in cyanobacteria developing several stress amelioration pathways. One such ability is the high radioresistance exhibited by some cyanobacterial strains such as Nostoc PCC7120, with LD50 of 6 kGy. They exhibited efficient DNA repair ability as observed with re-stitching of γ-irradiation damaged genomic DNA within 2 days of recovery. Inability to detect DNA repair proteins in the proteome mapping of these cells during radiation or post-irradiation recovery (PIR) resulted in very little information on its DNA repair mechanisms. Bioinformatic analysis of the genome revealed absence of full length RecB, RecC and Ku proteins, thus negating the presence of RecBCD pathway of HR and the NHEJ pathways of DNA repair. In order to understand the DNA repair mechanism, we took up an approach of investigating individual genes. No DNA repair can take place without a functional SSB. The annotated SSB proteins of Nostoc PCC7120 were truncated, capable of binding ssDNA but not interacting with other proteins. All4779 was identified the full length SSB and contributed to tolerance of Nostoc. Unlike E. coli where in SbcCD complex is required for DNA repair, the genes could individually contributed towards tolerance to γ-irradiation in Nostoc. In the absence of RecBCD pathway, the RecF pathway is expected to be the major pathway. They exhibited differential expression in response to γ-irradiation during PIR. These genes are probably required in tandem in specific proportions for efficient DNA repair. Several DNA repair genes were found to be regulated by two global regulators LexA and NtcA. DNA repair is one of the crucial component of radioresistance, and in Nostoc the genes are under multi-level regulation leading to a well-controlled and efficient DNA repair.
| PL-15 | | |
| New Decorporation Strategies for Reducing Risk from Intakes of Lanthanides and Actinides | | |
R. J. Abergel1,2
1Department of Nuclear Engineering, University of California, 2Chemical Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA
E-mail: [email protected]
The threat of a major radiological event presents a danger of not only large-scale external radiation exposure of the population but also internal contamination with radionuclides, including actinides and lanthanide fission products. Numerous factors influence the potential health effects after contamination, including the contaminant physicochemical properties or the contamination route, but preventing or reversing tissue deposition remain the most efficacious ways to reduce the risk associated with intake. Current treatment options are limited and address only a few of the many radioisotopes that would pose a significant medical threat after a nuclear event. To seek regulatory approval for new decorporation agents or new formulations, a number of efficacy and safety studies must respond to the selective criteria of the Animal Rule from the U.S. Food and Drug Administration (FDA). The FDA approved the first clinical study for a new f-element decorporation agent 3, 4, 3-LI (1,2-HOPO) in August 2014. The Investigational New Drug (IND) application was the result of a large number of studies performed to optimize the drug candidate, demonstrate its safety, activity and efficacy, and establish a mechanism of action. These studies will be summarized, with an accent on the extensive regulatory process guiding such development work, and a brief description of on-going clinical safety evaluations. Finally, while initially targeted radionuclides included isotopes from the actinide series (Am, Cf, Cm, Pu, U), recent developments in the medical use of f-elements (Gd, Lu, Ac, Th) have uncovered a persisting need for such decorporation agents much beyond the national security threat. A perspective on applying 3, 4, 3-LI (1, 2-HOPO) to prevent internal Gd deposition and subsequent toxicity from MRI contrast agents will be given. This work has been supported over the years by grants from the National Institute of Allergy and Infectious Diseases and the Biomedical Advanced Research and Development Authority, through the U.S. Department of Energy under Contract #DE-AC02-05CH11231, as well as from the U.S. Department of Defense through the Alliance for Radiological Exposures and Mitigation Science.
| Invited Lecture (IL) | | |
| IL-1 | | |
| Nanoparticle Agents for Tumor Microenvironment Phenotyping and Cancer Therapy | | |
Ketan B. Ghaghada
Department of Radiology, Texas Children's Hospital/Baylor College of Medicine, Houston, Texas, USA
E-mail: [email protected]
High atomic weight (Z) element-based nanoparticle agents can serve as theranostic agents (therapeutic and diagnostic) in cancer medicine. When used as a contrast agent for X-ray/computed tomography (CT) imaging, high Z-based nanoparticles enable in vivo profiling of tumor microenvironment in mouse models of solid tumor. Specifically, data mining of nanoparticle contrast-enhanced CT images, a process we termed nano-radiomics, enabled differentiation of tumors based on immunosuppressive myeloid burden. In comparison, conventional CT imaging-based tumor metrics were unable to profile the tumor microenvironment in solid tumors. When combined with radiation therapy, high Z-based nanoparticle agents enhanced radiation effects, increased tumor vascular permeability and led to improved tumor outcomes. Overall, this work describes novel applications on the use of high Z nanoparticle agents for imaging-based profiling of tumor microenvironment in solid tumors and cancertherapy.
| IL-2 | | |
| Distinct Roles of HR and NHEJ in the Repair of Ionizing Radiation-Induced Chromosomal Double-Strand Breaks in Mammalian Cells | | |
Aashish Soni1,2, Tamara Murmann-Konda2, Maria Siemann-Loekes1, Emil Mladenov1,2, Gabriel E. Pantelias3, Martin Stuschke1,4, George Iliakis1,2
1Department of Radiation Therapy, Division of Experimental Radiation Biology, University Hospital Essen, University of Duisburg-Essen, 2Institute of Medical Radiation Biology, University of Duisburg-Essen Medical School, 4German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Partner Site University Hospital Essen, Essen, Germany, 3Institute of Nuclear Technology and Radiation Protection, National Centre for Scientific Research “Demokritos,”Aghia Paraskevi Attikis, Athens, Greece
E-mail: [email protected]
Introduction: DNA double-strand breaks (DSBs) are among the most deleterious lesions induced by ionizing radiation (IR). Unrepaired or misrepaired DSBs can cause genomic instability in the form of chromosome translocations (CTs), leading to cell death or cancer. Classical non-homologous end-joining (c-NHEJ) has been considered the prime DSB repair pathway in mammalian cells. On the other hand, Homologous recombination (HR) has been shown to be crucial for DSB repair in yeast. However, its significance in mammalian cells remained obscured for a long time and started emerging only recently. Alternative end-joining (alt-EJ) acts as a backup pathway to abrogated c-NHEJ or HR, operating with slower kinetics, and is highly error-prone. HR functions strictly in a cell cycle-dependent manner (late S and G2-phase), while c-NHEJ and alt-EJ function throughout the cell cycle. Materials and Methods: Here, we analyzed the relative contributions of c-NHEJ, HR, and alt-EJ in mammalian cells, which were explicitly irradiated in the G2-phase of the cell cycle and irradiated with low and high doses of IR. Such G2-specific analysis facilitated the simultaneous investigation of all DSB repair pathways at varying degrees of DSB loads. Results: Our results suggest that HR is indispensable for repairing chromosomal DSBs at physiologically relevant low IR doses (≤1Gy). In contrast, a shift towards c-NHEJ is evident when cells are exposed to high doses of IR. Both c-NHEJ and HR suppress CT formation, which strongly depends on PARP-1 mediated alt-EJ.Furthermore, cells opt to actively suppress c-NHEJ at physiologically relevant IR doses and HR with increasing DSB loads at high doses of IR. Conclusions: Our results highlight the importance of HR in repairing chromosomal DSBs in mammalian cells exposed to physiologically relevant IR doses. The underlying molecular mechanisms behind such DSB load-dependent repair pathway switch are currently under investigation.
Acknowledgements: DFG Graduate Training Program 1431 and 1739, BMBF and BMWi.
| IL-3 | | |
| Caenorhabditis elegans Ribosomal Protein S3 Cooperates with Uracil DNA Glycosylase to Process Oxidative DNA Lesions | | |
Dindial Ramotar1, Noha Elsakrmy1,2, Mustapha Aouida1, Balasubramanian Moovarkumudalvan1, Ashima Mohanty1,2, Reem Ali1
1Division of Biological and Biomedical Sciences, 2Division of Genomics and Precision Medicine, College of Health and Life Sciences, Hamad Bin Khalifa University, Education City, Ar-Rayyan, Qatar
E-mail: [email protected]
Introduction: Carcinogenicity and cytotoxicity are severe consequences of DNA damage. Base Excision Repair (BER) is a conserved DNA repair pathway that replaces many damaged bases caused by oxidation. Aberrations in BER are associated with carcinogenesis, neurodegeneration, and aging. The nematode C. elegans is an attractive model system for studying BER. However, in this organism, the complete pathway is not fully delineated. Materials and Methods: We searched for new components of the BER process in C. elegans by using affinity tag chromatography and mass spectrometry to identify the interactome of uracil DNA glycosylase-1 (CeUNG-1), an enzyme that acts during the first step of the BER pathway. Results: We discovered that CeUNG-1 is associated with the 40S ribosomal protein S3 (CeRPS-3), homologs of which have been shown to process 8-oxoguanine and abasic site lesions in other organisms. We report a strong in silico interaction between CeUNG-1 and CeRPS-3, which was confirmed using the yeast two-hybrid system. Downregulation of the Cerps-3 gene reduced the viability of wild-type worms upon exposure to the chemical oxidant hydrogen peroxide. Further analysis shows that Cerps-3 knockdown significantly sensitized the AP endonuclease APN-1-deficient strain, apn-1 to the lethal effects of hydrogen peroxide. A cross-species complementation experiment reveals that the expression of CeRPS-3 rescued the hydrogen peroxide sensitivity, and suppressed the high mutation frequency of the yeast AP endonuclease-deficient strain lacking Apn1 and Apn2. Conclusion: We propose that CeRPS-3 may function as an auxiliary DNA repair enzyme in C. elegans to process oxidative DNA lesions.
| IL-4 | | |
| Cell Cycle-Specific Epigenetic Alteration and Reprogramming for Sensitization of Radio-Resistant Cancer Cells | | |
Sanjay Gupta1,2
1Epigenetics and Chromatin Biology Group, Gupta Laboratory, Cancer Research Institute, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, Navi Mumbai, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
The dynamic nature of epigenome changes in a cell cycle-dependent manner, and radioresistance varies in different cell cycle phases, most resistant in G0/G1, most sensitive in S, and sensitive in the M phase. Moreover, a significant clinical problem in treating tumors is the poor response by radiotherapy. The presentation will discuss the identification of cell cycle-specific alterations and the potential of epigenetic modifiers as an adjunct to enhance the efficacy of radiotherapy. The present study has shown that a specific population of binucleated tetraploid cells, characterized by reduced H3S10/S28ph but increased H3K9ac, was generated after radiation and mitotic progression of M-phase cells only. These epigenetic features correlated with decreased survival potential of the mitotic population. However, the tetraploid cells are tumorigenic, and their survival potentially contributes to tumor relapse. The low levels of H3S10/28ph were attributed to decreased protein translation, chromatin recruitment of kinase Msk1, and persistent levels of protein phosphatase PP1α. Further, the nucleosomal organization studies demonstrated the presence of compact chromatin architecture coupled with decreased levels of site-specific histone post-translational modifications, H3K9ac, H3K27ac, and H3S10pK14ac in the G0/G1 and M-phases of the radio-resistant breast cancer cells. The radio-resistant cells possessed high HDAC and low HAT activity. In vitro and in vivo studies showed that treatment of radio-resistant cells with class 1 HDAC inhibitor led to the retention of γH2AX and decreased H3S10p after irradiation leading to cell death. Additionally, an analysis of human patient samples showed variable tumor HDAC activity, thus demonstrating inter-tumoral epigenetic heterogeneity in a patient population. The study highlights that sub-grouping patients for treatment with HDAC inhibitors in combination with radiotherapy will reduce the incidence of tumor recurrence and prove to be more effective in better clinical outcomes.
| IL-5 | | |
| Low Dose Gamma Rays Enhances Myogenic Differentiation in Long Term Myoblast Cultures | | |
Lindsey Bertrand1, Jais Kavumkal1, Mathew Flegal1, Priya Chandran1, Jeffrey F. Dilworth2,3, Dmitry Klokov1,4, Soji Sebastian1,3
1Radiobiology and Health, Canadian Nuclear Laboratories, Deep River, 2Sprott Centre for Stem Cell Research, Ottawa Hospital Research Institute, 3Department of Cellular and Molecular Medicine, University of Ottawa, 4Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, Canada
E-mail: [email protected]
Cellular exposure to high doses of ionizing radiation causes DNA damage, genome instability, immunosuppression, and lethality. In contrast, there is increasing evidence showing that radiation at low doses (below 100 mGy) may be harmless and even beneficial. For example, we and others have shown that exposure to low dose radiation (LDR) significantly enhances the anti-tumor effects of immune checkpoint therapy (ICT), reduces tumor volume, and shortens the response time of PD-L1 ICT. Here, we extend our work to muscle stem cells and show that exposure to an acute dose of LDR (cobalt-60) is sufficient to induce changes in skeletal muscle gene expression leading to favourable effects. Although much research has described radiation-responsive signaling pathways underlying transcriptional regulation, the focus here is on histone variant H3.3 mediated epigenetic mechanisms responsible for muscle stem cell memory in the context of long term aged myoblast cultures. We investigated how exposure of muscle stem cells to a single acute dose of 10 or 100 mGy of gamma rays modulates myogenic gene expression and differentiation into mature muscle fibers. Whereas a substantial decrease in differentiation capacity was observed in control unirradiated myoblasts with age/time in culture, the loss of differentiation potential was partially restored in cultures exposed to LDR at early passages. High throughput RNA-seq and mass spectrometry analyses of LDR-exposed stem cells showed a unique signature of gene expression and histone modifications, providing a mechanistic insight into the enhanced myogenesis regulated by LDR. While exposure to LDR did not affect the proliferation status for the time points examined, LDR preserved H3.3 acetylation and enhanced the tissue-specific transcriptional memory. Together, the results show that LDR results in myogenic stimulation of exposed myoblasts, an effect that persists in long term cultures. These results demonstrate the potential beneficial effects of LDR for use in regenerative medicine.
| IL-6 | | |
| Responses of Human Cells to Low-Dose Ionizing Radiation: Implications for the Linear No-Threshold Paradigm | | |
S. Nishad, Neha R. Paraswani, Anu Ghosh
Animal House Facility and Radiation Signaling Section, Radiation Biology and Health Sciences Division, Bio-Science Group, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
There is a keen interest among the medical and scientific community to understand the health effects of low-dose ionizing radiation (IR) on humans as this can influence public perception and broader societal applications of radiation. To assess responses of human cells to low-dose IR, a quantitative discovery proteomics approach was used. For this purpose, human peripheral blood cells were collected from individuals residing in a narrow 55-km long coastal strip in the state of Kerala on the south-west coast of Indian subcontinent which is known for high levels of natural radiation. This belt has abundance of localized thorium-rich monazite sand deposits due to which it shows continous high background radiation levels ranging from ≤1 to ≥45 mGy per year. An integrated gel-based (two-dimensional gel electrophoresis-mass spectrometry; 2DE-MS) and gel-free (isobaric Tags for Relative and Absolute Quantification; iTRAQ) analysis was performed to obtain an exhaustive index of proteins for individuals residing in these areas. The data showed for the first time that in human cells, low doses of chronic radiation can induce several proteins in vivo, which can detect the damage, initiate a DNA damage response signaling and repair and mediate a radioadaptive response. Protein-protein interaction networks highlighted the role of YWHAZ (14-3-3ζ) as a stress-adaptive signaling hub in low-dose radiation response. Additionally, in-vitro studies were performed to understand mechanism of low dose radiation-induced adaptive response. Akey role of DNA damage response and antioxidant mechanismsin radioadaptive response of human cells was identified. This highlights the fallacy of using the linear no-threshold (LNT) dose-response model to extrapolate the responses observed at high doses to derive expected radiation risks at low doses. The compelling need of a rational re-evaluation of LNT paradigm that is currently adopted in radiation protection guidelines cannot be over-emphasized.
| IL-7 | | |
| Outlook on the Biological Effects of Low Dose Radiation Exposure and Challenges in the Use of Biomarkers of Risk Prediction for Health Effects | | |
P. Venkatachalam, V. Shangamitra, K. Nandhini, M. Divya, T. A. Aishwarya, Venkateswarlu Raavi1
Department of Human Genetics, Sri Ramachandra Institute of Higher Education and Research (Deemed to be University), Chennai, Tamil Nadu, 1Department of Cell Biology and Molecular Genetics, Sri Devaraj Urs Academy of Higher Education and Research (Deemed to be University), Kolar, Karnataka, India
E-mail: [email protected]
Introduction: All living organism are buildup of essential bio-molecules (nucleic acid, proteins, lipids & carbohydrates): deviation of those molecules from its normal level (biomarkers) is an indication of exposure to toxic agents and or effects of such exposure. Despite continuous refinement in the methodology towards the development of newer markers, its promise to predict risk for the long term health due to low doses of ionizing radiation exposure remains elusive. Objectives: Altered cell viability, colony forming ability, chromosome structure, and gene and protein expression are being used as biomarkers of radiation exposure (acute and chronic) as well as to predict the health effects. An ideal biomarker that has higher potential to predict risk is being the interest. Methods: We have developed a range of biomarkers dicentric chromosomes (DC), micronuclei (MN), translocation (TL), gammaH2AX (γ-H2AX), Comet assay, P53ser15 expression, gene expression changes (ATM,CDKN1A, DDB2, FDXR and PCNA) in cell lines exposed (in-vitro) low doses of radiation and in blood samples obtained from human subjects received low doses radiation during X-ray imaging. Results: Dose range (less than 1 Gy) adopted in in-vitro study induced significant changes in those biomarkers among the cell lines. In consistent with in-vitro results, a significant higher frequency of aberrations was observed in patients received radiation during X-ray based imaging (CT, interventional radiology and PET). Observed variation on the magnitude of induced DNA damages among the study subjects is suggested due to radiation sensitivity and or repair ability of subjects. Conclusion: Low doses of ionizing radiation induces significant amount of biomarker changes in the hereditary material. However, it is much more challenging to relate it to the risk for long term health effects.
| IL-8 | | |
| Radiopharmaceuticals for Imaging Bacterial Infections | | |
Archana Mukherjee1,2
1Radiopharmaceuticals Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail:[email protected]
Diagnostic imaging has an important role to play in management of focal infections i.e., fever of unknown origin (FUO), osteomyelitis, soft tissue infections (STI), bone prosthesis/device borne infections and infective endocarditis. Functional imaging techniques such as single photon emission computed tomography (SPECT) and positron emission tomography (PET) are being employed for gaining insights into molecular and cellular level information on disease progression, treatment planning and also assessing treatment response. These techniques can provide rapid three-dimensional assessment of disease burden at a very early stage. SPECT and PET are capable of specific detection of infection if a bacteria specific radiotracer is available. Molecules targeting various bacterial processes have been utilized for development of infection imaging probes. However, an ideal infection imaging agent is still not available. Therefore, need for continuous efforts to develop and improve specific agents for infection imaging has been realized. Among the promising molecules, antimicrobial peptides (AMPs) are ideal candidates for designing infection imaging agents. The talk will provide an overview of the current status of bacterial infection imaging using radiopharmaceuticals. Strategies employed to develop new SPECT and PET radiotracers and recent progress in development of ubiquicidin (UBI) based radiotracers for infection imaging will be discussed.
| IL-9 | | |
| Radiation Processing Technology for Food – Role of Radiation Physics, Chemistry and Engineering | | |
Bhaskar Sanyal
Food Technology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: A food commodity after harvest has a certain life time depending upon its nature and bioburden. Preserving food for extended periods has been one of the oldest challenges to mankind. Application of ionizing radiation such as gamma rays, X-rays, and accelerated electrons in controlled fashion has immense potential to address the challenges of food preservation. The effectiveness of processing of food by ionizing radiation depends on proper delivery of absorbed dose and its reliable measurement. Food destined for international trade, it is of utmost importance that the dosimetry techniques used for dose determination are carried out accurately and that the process is monitored in accordance with the internationally accepted procedures. The need for reliable and routine tests to determine whether or not food has been irradiated has arisen as a result of the progress made in commercialization of the food irradiation technology and increased international trade. Materials and Methods: This presentation will elaborate the role of radiation physics and chemistry in radiation measurements of absorbed dose and identification of irradiated food. In addition, a brief overview on the methodologies of dosimetry in food at commercial setup will be presented. Results: R&D activities to study suitable physical systems to measure food irradiation doses using thermoluminescence and EPR spectroscopy will be discussed. Novel approaches to identify irradiated food will also be elaborated. The deployment of radiation processing technologies also involves optimized design features of the irradiation plant to deliver desired doses ensuring operational safety. A brief description of engineering designs and technologies based on radionuclide and accelerator facilities will be presented. Conclusions: Finally, the applications of radiation technology in food and agriculture will be briefed with their present status across the country.
| IL-10 | | |
| Radiation Processing of Foods – A Success Story of DAE | | |
Sachin N. Hajare
Food Technology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
It is estimated that about 30- 40% of foods is been lost annually in India indicating the need of the preservation technology to reduce these post-harvest losses. Food Technology Division (FTD) is a pioneer institute in India working on R&D activities pertaining to radiation processing of foods, an alternative preservation technology. It helps in sprout inhibition of bulbs and tubers, hygienization of spices, insect disinfestation of cereals and pulses and delay in ripening in fruits. The division has worked on long term sprout free storage of onion and potatoes by developing precise Standard Operating Procedures (SOP) that were successfully tested on a large-scale commercial storage trials of irradiated onion (15 tons) and potato (28 tons), with the shelf life of 7.5 and 8 months, respectively. Additionally, stuffed baked food (SBF) was developed with a shelf life of 6 months at ambient temperature using radiation processing that was deployed as emergency food to the flood victims in Himachal Pradesh. Similarly, to increase the export volume of irradiated mangoes, a process including irradiation (400 Gy) was developed to extend the shelf life of Kesar mangoes to about 30 days at 10° C. Using this process, about 16 tons of Kesar mangoes were successfully shipped to Newark, USA and marketed in 2022. Further, semi commercial storage trials of irradiated (650 Gy) cereals and pulses with the shelf life of >1y were carried out. Active research in fruit preservation resulted in development of shelf stable Jamun and Strawberry products which are available on online platforms like Amazon. Similarly, research in grain technology culminated into a development of gluten free multigrain premix consisting of radiation processed dietary fibre. Based on this research, Government of India has given class-based approval in 2016 for food irradiation and about 25 radiation processing plants have been established till date.
| IL-11 | | |
| Understanding Mechanisms of Radiation Resistance in Glioblastoma for Novel Therapeutics | | |
Shilpee Dutt
Advanced Centre for Treatment, Research and Education in Cancer, Navi Mumbai, Maharashtra, India
E-mail: [email protected]
In our lab we are trying to address the fundamental issue of radiation resistance and recurrence in glioblastoma multiforme (GBM). GBM is the most aggressive and malignant primary brain tumor with median overall survival of barely 18 months. Despite aggressive radiotherapy recurrence in GBM is inevitable making GBM the most difficult brain tumor to treat. We have shown that GBM recurrence is due to the residual disease cells also called persister cells that survive radiotherapy. Therefore, it is imperative to eliminate residual/persister cells to prevent recurrence and improve patient outcome. However, due to lack of patient biopsies post-surgery and radiotherapy, the molecular mechanisms leading to persister cell survival are not known. To overcome this lacune, we have developed a cellular model from primary culture pf patient samples and PDOX (patient derived orthotopic xenograft) model in mice that mimic clinical scenario of GBM resistance and recurrence. Using these model systems, we are able to capture the rare population of residual/ persister cells that survive radiotherapy and are responsible for relapse. We take multi-pronged approach to delineate molecular mechanisms of radiation survival in persisters and recurrent cells to enable us to target these cells for GBM therapeutics. In this talk, I will discuss recent findings from our lab that describe novel mechanisms of DNA damage repair pathway modulation in residual cells that help them survive radio therapy. Inhibition of these pathways provide persister/residual cell–specific radio sensitization and expand the therapeutic window for DNA damage response–targeted agents used in combination with radiotherapy.
| IL-12 | | |
| Radioresistance of Hypoxic Tumour Cells; Lessons from Hypoxia and HIF-1 Biology | | |
Hiroshi Harada1,2
1Laboratory of Cancer Cell Biology, Graduate School of Biostudies, Kyoto University, 2Department of Genome Dynamics, Radiation Biology Center, Kyoto University, Kyoto, Japan
E-mail: [email protected]
High-precision radiation therapy has been developed and improved due to the integration of knowledge and technologies of physics, engineering, chemistry, and biology. It provided radiation oncologists with means to administer a booster dose of radiation specifically to very small target fractions in a tumour tissue and markedly decrease an excessive radiation dose to normal tissues. However, even the state-of-the-art strategies have often failed to accomplish a complete remission, and patients suffer from recurrence and/or distant metastasis of caners after treatments. To overcome the problems, it is important to elucidate biological mechanisms by which cancer cells acquire radioresistance, and consequently, survive, recur, and metastasize after radiation therapy. Tumour hypoxia has drawn much attention in radiation oncology and biology since Thomlinson and Gray reported the presence of hypoxic cells in malignant solid tumours and showed that they exert a negative impact on the outcome of radiation therapy. My laboratory has contributed to elucidate molecular mechanisms, wherein genetic alterations observed in cancer patients as well as hypoxic stimuli activate hypoxia-inducible transcription factor, HIF-1, and eventually induce tumour radioresistance. In this talk, I would like to introduce the historical background that has led to the focus on tumour hypoxia and moreover present our findings about the HIF-1-mediated molecular mechanisms behind radioresistance of hypoxic cancer cells and tumour recurrence after radiation therapy. I would like to provide rational basis for targeting HIF-1-positive hypoxic tumour cells to enhance effect of radiation therapy.
| IL-13 | | |
| Ionizing Radiation and Immune System: Low and High Dose Effects | | |
Bhavani S. Shankar
Radiation Biology and Health Sciences Division, Bioscience Group, Modular Laboratories, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Exposure to ionizing radiation at low and high doses can exert varying responses on the immune system. Radiation exposures to doses <100 mSv is important since it contributes to our understanding of biological effects of low dose radiation. Low dose ionizing radiation (LDIR) induced transient adaptive response (AR) in BALB/c mice which was due to hyper activation of MAPK signaling. Similar experiments in C57BL/6 mice indicated absence of AR despite low dose induced MAPK activation. Evaluation of upstream MAPK activators indicated that differential induction of Ca2+ and NO signaling in BALB/c and C57BL/6 mice was responsible for these effects.
On the other hand, localised radiation exposures can play a crucial role in affecting the tumor induced immunosuppression. Select doses of ionizing radiation increased immune activating potential of dendritic cells (DC) through STAT5/Zbtb46 signalling mediated by irradiation induced apoptotic bodies (AB). Splenic and bone marrow derived DC from tumour bearing mice exposed to localised irradiation showed abrogation of tumour induced immunosuppression. This phenomenon was also recapitulated in DC derived from normal mice exposed to whole body irradiation or those differentiated from in vitro irradiated progenitor cells. Further, these DCs derived from irradiated progenitors could resist the suppressive effects of tumour conditioned medium and had increased immune activating potential.
Exposure to higher doses of localized radiation increases immunosuppressive molecules like PD-1. COX-2 inhibitor NS-398 decreases tumor induced expression of PD-1 and hence was used in combination therapy along with localized radiation and anti-PD1 antibody. There was a significant reduction in tumor burden with NS-398/radiation combination or NS-398/radiation/anti-PD-1 combination when compared to either NS-398 or radiation alone. These results thus demonstrate that though localized irradiation can activate DC, it can also induce immunosuppressive molecules and use of anti-PD-1 or agents like NS-398 can decrease PD-1 expression and can potentiate radiotherapy.
| IL-14 | | |
| Targeted Alpha Radionuclide Therapy: The Fundamentals and Their Current Place in Nuclear Oncology Practice | | |
Sandip Basu1,2
1Radiation Medicine Centre, Bhabha Atomic Research Centre, Tata Memorial Centre Annexe, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Targeted Alpha Therapy (TAT) with unsealed systemic radiopharmaceuticals has generated much interest in the domain of Nuclear Oncological practice. It offers several potential advantages in patients with metastatic/advanced cancer. These include: (a) high linear energy transfer (LET) (~80 keV/μm) & relative biological effectiveness (RBE), (b) a short particle range (~50–100 μm) and consequently, (c) a high tumor cell-killing efficiency (more likely to cause double-strand breaks to DNA molecules) with lesser energy deposition and toxicity to the surrounding normal tissues. In this treatise, we shall discuss the comparative radiation biology and the cellular molecular effects of the alpha vs. the traditionally used beta radionuclides, drawing reference from their fundamental physical properties. The alpha emitters in current clinical use (viz. 225Ac, 223Ra, 211At) along with related radiopharmaceuticals (e.g. 223RaCl2,225Ac-PSMA-617, 225Ac-DOTATATE and 211At-MABG) and their present and evolving role in clinical practice will be deliberated.
| IL-15 | | |
| Role of Extracellular Vesicles and DNA Fragments in the Non-Targeted Effects of Targeted Radionuclide Therapy | | |
Jean-Pierre Pouget1,2
1Institut de Recherche en Cancérologie de Montpellier, Université de Montpellier, 2Institut Régional du Cancer de Montpellier, Montpellier, France
E-mail: [email protected]
With the development of new therapeutic radiopharmaceuticals in oncology, it is important to determine the contribution of targeted and non-targeted (bystander and systemic) effects of targeted radionuclide therapy (TRT). In the last decade, our team investigated the contribution of non-targeted cytotoxic and genotoxic effects in vitro and in vivo (WT C57BL/6J and athymic nude mice) during alpha (212Pb/212Bi, 213Bi) and Auger (125I) TRT. In vitro, we showed that bystander effects contributed to 7-36% and 27-29% cell killing during alpha RIT and Auger TRT, respectively. We demonstrated that the bystander cell response was partly mediated by lipid raft-mediated activation of p38 kinase and c-JUN N-terminal kinases (JNK). We then showed that RIT efficacy was reduced in vitro and in vivo when TRT was combined with ASMase inhibitor (imipramine) or with drugs modifying cholesterol metabolism such as filipin, methyl-beta-cyclodextrin (or pravastatin). Reactive oxygen species also played a significant role in these bystander effects. Using autoradiography and voxel dosimetry, we confirmed the occurrence of bystander effects in vivo also, during Auger and alpha TRT. We isolated extracellular vesicles from the secretome of cells exposed to TRT and showed that they were responsible for decreased in clonogenic survival decrease in vitro and for tumor growth delay in vivo after intratumoral injection. We also showed that the latter therapeutic efficacy was enhanced in immune competent mice suggesting a role of immune cells. In conclusion, we confirmed that non-targeted effects play a central role during TRT and that drugs modifying cholesterol metabolism can modify RIT efficacy.
| IL-16 | | |
| Theranosis with 177Lu in India: From Nowhere to Almost Everywhere | | |
Tapas Das
Radiopharmaceuticals Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
The word 'Theranosis' was coined to describe the process of combining the two interdependent applications, namely therapy and diagnosis, using the same agent. Since radiopharmaceuticals are employed for both diagnostic and therapeutic purposes, the concept of, 'theranosis' is particularly relevant in nuclear medicine practices. Lutetium-177 is a theranostic radionuclide, as it decays by the emission of β- particles [Eβ(max) 497 keV], which are useful in radionuclide therapy (RNT) along with gamma photons (113 and 208 keV), suitable for performing diagnostic imaging. In the last two decades, production and use of 177Lu and 177Lu-based radiopharmaceuticals have seen a phenomenal growth all over the world and the Indian contribution in this field is well-recognized. The first irradiation of natural Lu2O3 target was carried out in 2000 and subsequently irradiation of enriched (in 176Lu) Lu2O3 target was attempted in mid-2001 for the production of 177Lu with higher specific activity. 177Lu, suitable for clinical applications, is being produced and regularly supplied from BARC (through BRIT) since the end of 2006. At the same time, effort was directed to develop formulation methodologies of few state-of-the-art radiopharmaceuticals for RNT. BARC started commercial supply of 177LuCl3, as the precursor radiochemical, and along with know-how transfer for the formulation of the radiopharmaceuticals (177Lu-DOTA-TATE and 177Lu-EDTMP) at hospital end, which helped the hospitals to initiate clinical procedures by the end of the first decade of 2000 onwards. Subsequently, few more products, like 177Lu-PSMA-617, 177Lu-DOTMP and 177Lu-HA were developed and deployed to the nuclear medicine centres of India. Presently the use of 177Lu is second to 131I among all the RNT applications. Currently, BRIT supplies 177Lu, both as precursor radiochemical and as 177Lu-labeled radiopharmaceuticals, to more than 50 nuclear medicine centres of India. The talk will encompass this phenomenal growth in the use of 177Lu-based products during the last two decades in India.
| IL-17 | | |
| Optimising the Radiobiological Impact of Protons and High-LET Radiation in Head and Neck and Glioblastoma Cell Models | | |
M. R. Fabbrizi1, J. R. Hughes1, R. Aiyappa-Maudsley1, R. M. Hill1, S. Rocha2, K. Petersson3, M. A. Hill3, A. J. Chalmers4, J. L. Parsons1,5
Departments of 1Molecular and Clinical Cancer Medicine and 2Molecular Physiology and Cell Signalling, University of Liverpool, Liverpool, 3CRUK and MRC Oxford Institute for Radiation Oncology, University of Oxford, Oxford, 5The Clatterbridge Cancer Centre, NHS Foundation Trust, Bebington, England, 4Institute of Cancer Sciences, University of Glasgow, Glasgow, Scotland, UK
E-mail: [email protected]
Radiotherapy using conventional x-ray irradiation is still the most effective treatment for solid tumours of the head and neck and brain, although adverse treatment side effects are common and inherent tumour radioresistance remains a significant barrier to effective treatment. Use of alternative radiation modalities such as precision targeted proton beam therapy (PBT), and higher linear energy transfer (LET) radiation leading to an enhanced biological effect, are attractive options although there is significant biological and clinical uncertainty with these treatments and how they can be optimised for tumour treatment, which necessitates further preclinical research. We have examined the radiobiological response of tumour cell models of head and neck squamous cell carcinoma (HNSCC) and glioblastoma (GBM) to protons of increasing LET using the 60 MeV cyclotron at the Clatterbridge Cancer Centre, in comparison to conventional x-ray (photon) irradiation. We have demonstrated that relatively high-LET protons generated at the Bragg peak induce significant decreases in HNSCC and GBM cell survival post-irradiation versus low-LET protons and photons, due to increases in the complexity of the DNA damage that persists for several hours post-irradiation. Through siRNA screening, we have identified specific components of DNA repair, including poly(ADP-ribose) polymerase-1 (PARP-1), poly(ADP-ribose) glycohydrolase (PARG) and 8-oxoguanine DNA glycosylase (OGG1), that when targeted with siRNA/inhibitors can radiosensitise cells to high-LET radiation through delays in complex DNA damage repair. We are now expanding our research to investigate the impact of reduced oxygen (hypoxia) in driving increased cellular radioresistance, and how this can be overcome potentially through targeting the DNA damage response. Furthermore, we are examining the impact of ultra-high dose rate (FLASH) protons on the radiobiological response, given its significant potential in normal tissue sparing whilst maintaining tumour control.
| IL-18 | | |
| Cytoplasm Damage Induced Defensive Cellular Response in Microbeam Targeted Mammalian Cells | | |
T. Konishi, N. Autsavapromporn1, T. A. F. Tengku Ahmad2, J. Wang3
Single Cell Radiation Biology Team, National Institutes for Quantum Science and Technology, Chiba, 1Department of Radiology, Faculty of Medicine, Chiang Mai University, Chiang Mai, Thailand,3Key Laboratory of Ion Beam Bioengineering, Hefei Institute of Physical Science, Chinese Academy of Sciences, Hefei, China,2Radiation Safety and Health Division, Malaysian Nuclear Agency, Bangi, Malaysia
E-mail: [email protected]
Introduction: Primary target of radiation is the cellular DNA, but consequence of cytoplasmic damage is yet to be understood. To distinguish the cellular responses triggered from DNA damage and cytoplasmic damage, microbeam would be most effective technology. Microbeam which is a highly advanced technology that enables studies to target cells efficiently with several micrometer sized beam and desired amount of radiation. Materials and Methods: SPICE-QST microbeam provides 3.4 MeV protons beam of a size of below 2-μm in diameter size, which can be targeted to cytoplasm and/or nucleus of the mammalian cells. We performed a cytoplasm targeted irradiation (Cyto-IR) of the WI-38 cells to investigate the cytoplasmic damage response. We focused on the activation of nuclear factor (erythroid-derived 2)-like 2 (NRF2) and its antioxidative signaling pathway. Results: As a result, Cyto-IR induced mitochondria fragmentation, which accelerated the mitochondrial superoxide (MitoSOX) production. MitoSOX triggered the NRF2 nucleus translocation and upregulated the expression of its target genes, such as heme oxygenase 1. In addition, we found that cytoplasmic irradiation induced upregulation of ERK 1/2 and resulted radiation-adaptive response to later X-ray exposure. Conclusions: NRF2 antioxidative response is suggested to play a key role against DNA damage under cytoplasmic irradiation. Cytoplasmic damage triggered signaling pathway would be important in understanding the mechanism of defensive role of mammalian cells against radiation.
| IL-19 | | |
| A New Biophysical Model for Treatment Plan Optimization in Heavy-Ion Cancer Therapy | | |
F. Ballarini1,2
1Department of Physics, University of Pavia, 2National Institute of Nuclear Physics, Section of Pavia, Pavia, Italy
E-mail: [email protected]
Introduction: Heavy-ion cancer therapy requires knowledge of the ion Relative Biological Effectiveness (RBE) along the whole beam path. RBE is a complex quantity, since it depends on the particle energy – and thus its penetration depth - as well as other factors including particle type, dose, cell type and biological endpoint. Theoretical modelling is therefore mandatory especially for scanning pencil beam irradiation, for which two radiobiological models (Local Effect Model version I or LEM-I, and Microdosimetric Kinetic Model, or MKM) are currently used in clinical practice. Materials and Methods: In this work we discuss a biophysical model developed in Pavia, called BIANCA (BIophysical ANalysis of Cell death and chromosome Aberrations), which was benchmarked against several heavy-ion data sets, not only for C-ions but also for He-ions, which are now used in Heidelberg (Germany), and O-ions, which may be used in the future. Furthermore, following interface with the FLUKA radiation transport code, BIANCA was applied to re-calculate RBE distributions for C-ion patients previously irradiated at the CNAO hadrontherapy centre (Pavia, Italy), where the radiobiological optimization is currently based on LEM-I. Importantly, the BIANCA predictions were performed not only for tumour cell death (RBEsurv) but also for healthy tissue damage, represented by chromosome aberrations in peripheral blood lymphocytes (RBEab). Results: The BIANCA predictions were in good agreement with the considered data, thus allowing for model validation. Concerning the patient cases, in the target and in the entrance channel the cell survival RBE predicted by BIANCA was lower than that provided by LEM-I, whereas very similar results were found in the Organs at Risk. Furthermore, significant differences were found between RBEsurv and RBEab, also dependent on fractional dose and LET. Conclusions: Overall, the results obtained by BIANCA show that our model is a good candidate tool for the radiobiological optimization of heavy-ion treatment plans. Furthermore, this work suggests that, when performing such optimization, in healthy tissues the information on cell survival should be integrated by information more closely related to the induction of late damage, such as chromosome aberrations.
| IL-20 | | |
| Can cN3b Status in Head and Neck Squamous Cell Cancer Be Treated with Definitive Radiotherapy? A Single Institutional Experience | | |
G. Sarma, J. Nath1
All India Institute of Medical Sciences, 1Dr. B. Borooah Cancer Institute, Guwahati, Assam, India
E-mail: [email protected]
Introduction: Extranodal extension (ENE) in head and neck squamous cell carcinoma (HNSCC) is a known potential poor prognostic factor. The 8th edition of AJCC included ENE in the staging system of Head and Neck cancers. There is very less evidence to support the treatment of cN3b HNSCC with definitive radiotherapy. This study aims to assess the treatment outcome in patients of HNSCC with cN3b disease treated with definitive radiotherapy. Materials and Methods: Forty-five HNSCC patients with cN3b disease treated with definitive radiotherapy with or without concurrent chemotherapy between January 2018 to December 2018 were studied retrospectively. Results: The median age of the study patients was 60 years (40–75years). Only 35 patients (77.8%) could complete the prescribed course of treatment. The most common cause of non-completion was treatment-related toxicities. After a median follow-up period of 9.3 months (range 2–33), the median OS and PFS were 22.6 months and 7.2 months respectively. Fourteen patients (31.1%) in our study developed grade III/IV mucositis, and 11 patients (24.4%) developed severe grade III/IV dermatitis. The locoregional failure constituted 24 patients (53.3%). Conclusion: The treatment outcome of HNSCC with cN3b disease is very poor. Patient selection and a personalized approach form the cornerstone for undertaking patients with cN3b HNCC to be treated with definitive radiotherapy.
Keywords: Extranodal extension, Head and Neck Cancer, N3b, Radiotherapy
| IL-21 | | |
| Utility of Split-Course Hypofractionated Radiotherapy in Palliation for Head and Neck Cancer | | |
Krishna Sharan, Umesh Velu
Department of Radiotherapy and Oncology, Kasturba Medical College, Manipal Academy of Higher Education, Manipal, Karnataka, India
E-mail: [email protected]
Introduction: Hypofractionated radiotherapy schedules are most suited for palliation, where shortened overall treatment time carries great importance. Additionally, all efforts to minimize toxicity must be taken. Various schedules have been tried for palliation in Head-and-Neck cancers (HNC); we report our experience with a palliative split-course RT regimen of 22.5Gy/5#/5 days delivered to Gross Tumor using Volumetric Modulated Arc Therapy (VMAT), repeated again after a month for patients who had partial response and limited toxicities. Such a regimen substantially reduces treatment duration, while tailoring the treatment based on response and toxicities. Materials and Methods: HNC patients treated between 2014-2020 with the palliative split-course schedule were retrospectively identified from records; patients with incomplete details, alternative palliation schedules, and a follow-up of less than 3 months were excluded. Results: Of 144 patients, 60 patients were eligible for analysis. The mean age was 61 years (range: 36-92); 47 (78.3%) were males. 16 (26.7%) had a performance score ≥2. Pharyngeal subsites were the most frequent (46.7%). 6(10%) had progression following 1st course, and another 6(10%) had >50% reduction in volume. 30 (50%) reported good symptom relief, and 11(18.3%) reported no relief. Acute toxicities were low, and ≥grade 3 toxicities were noted in only 4 (6.7%). Conclusions: Regimens with high fractional dose for palliation can reduce treatment time and possibly improve response, and when delivered with conformal techniques, are also associated with reduced toxicity. Nevertheless, a small subset of patients obtains no benefit from even such hypofractionated schedules; studies need to be conducted towards understanding this underlying clinical radioresistance in these tumors.
| IL-22 | | |
| Investigation of Clinical role of BED in Radiotherapy Planning: The Way Forward from “One-Size-Fits-All” to Personalized Radiation Therapy Approach | | |
T. R. Verma, S. Rajurkar, S. P. Mishra1, M. L. B. Bhatt
Department of Radiotherapy, King George's Medical University, 1Department of Radiation Oncology, Dr Ram Manohar Lohia Institute of Medical Sciences, Lucknow, Uttar Pradesh, India
E-mail: [email protected]
Introduction: Although radiation oncology technology has undergone immense developments, the prediction of individual's radiotherapy responses is still one of the major issues and perhaps one of the greatest challenges in the practical implementation of personalized radiation therapy. The interaction of ionizing radiation with biological systems is considered probabilistic in nature in the sense that it can result in partial damage, sub-lethal damage and cell killing. To date, radiation biological effects have been characterised by the Biological Equivalent Dose (BED) with particular values of parameters α/β, which itself found to be varying over range. The aim of this talk is to present the roles and limitations of BED in radiotherapy planning in the context of development of radiotherapy as personalised radiation medicine treatment. Materials and Methods: The radiotherapy planning for cancer patients including Ca Breast, brain, cervix, oesophagus, larynx etc was done following the departmental/RTOG guidelines. The radiotherapy plan evaluation was performed using the dosimetric and physical dose indicators e.g. Max dose, Min dose, Mean dose, Conformity index, homogeneity etc derived from DVH. In case of interrupted treatments, BED calculation based on LQ model using the tumor specific values of α/β was done for compensating the interruption time. Results: Dosimetrical and physical dose assessment provides only volumetric dose coverage information but does not give any biological response in general and does not include tumor characteristics e.g. sensitivity, heterogeneity, hypoxia, angiogenesis, DNA repair system and microenvironment in the calculation of BED in interrupted treatment. Conclusions: The selection of the α/β value for BED calculation plays an important role in the outcome of radiotherapy. The radiotherapy plan should be evaluated not only on the basis of physical dose indicators, but also on the basis of radiotherapy response indicators e.g. Monitor unit, integral dose, overall treatment time, TCP and NTCP, etc.
| IL-23 | | |
| Mitochondrial Dynamics in Radiation Protection | | |
Damodar Gupta, Sweta Sanguri
Division of CBRN, Defence Research and Development Organization, Institute of Nuclear Medicine and Allied Sciences, New Delhi, India
E-mail: [email protected], [email protected]
Mitochondrial dynamics is vital for the structural integrity and function of mitochondria. Mitochondria coordinate with other cell organelles and plays crucial roles in wide variety of cellular functions. Emerging experimental evidence from recent years have accelerated research in understanding mitochondria as an important target for ionizing radiation (IR) and its significance of cellular response. Mitochondria constantly undergo fission and fusion. On exposure to IR, the finely regulated homeostasis between fission and fusionis disrupted resulting in mitochondrial fragmentation, cristae remodelling opening of mt-pores, resulting in apoptosis. Drp1 and Fis1 proteins play important role in mitochondrial fission. Drp1 is a cytosolic protein; it translocates to mitochondria and accumulates to the fission sites of mitochondrial outer membrane. Fis1 is a small membrane protein anchored on the outer membrane of mitochondria. Mitofusin1 (Mfn1), Mitofusin2 (Mfn2) and Optic Atrophy1 (OPA1) are important components that derive mitochondrial fusion process. All of three proteins are large GTPases of dynamin superfamily. Mfns and OPA1 coordinate to carry out outer membrane and inner membrane fusion respectively. A complex interaction between Reactive oxygen species (ROS) and mitochondrial dynamics has been reported. Dynamic change in mitochondrial morphology has been shown as a significant contributing factor to ROS overproduction in high glucose conditions. It has been established that under certain conditions, mitochondrial fragmentation and ROS may exacerbate each other to form a vicious cycle ultimately resulting in cell injury and death. IR exposure to cells has been known to disrupt fission and fusion homeostasis directly resulting in mitochondrial fragmentation and reduction in respiratory chain functions. Therefore, protection of mitochondria from IR exposure is crucial in order to minimize damage to the cells. Damage to the cells can be minimized by restoration of mitochondrial structure and functions; and therefore, the effect of radio-modifying agents that restores mitochondrial functioning is worth investigating.
| IL-24 | | |
| Organodiselenide, Novel Therapeutic Agent to Mitigate Radiation Induced Pneumonitis and Cell Migration in Lung Cancer Model Systems | | |
Amit Kunwar1,2
1Radiation and Photochemistry Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Radiotherapy the major treatment modality for cancers originating in the thorax is associated with dose limiting side effects of pneumonitis and metastasis. There is a dire need for easily administrable radiotherapy adjuvant drugs for use in clinics to prevent radiotherapy side effects and to improve the quality of life of cancer patients. In this context, a series of in house synthesized aliphatic and aromatic selenium compounds were screened for radio-modulatory effects against radiation induced pneumonitis and cell migration. Materials and Methods: The effect of test compound against radiation pneumonitis (RP) was evaluated using murine models mimicking human RP. The effect of test compound against cell migration was studied using cellular and xenograft tumor models of A549 cells. The most potent compound was investigated for mechanism of action through various biochemical assays. Results: These studies have led to the identification of diselenodipropionic acid (DSePA), as a candidate drug molecule to prevent RP. Mechanistically, it has been estabilished that oral DSePA is absorbed maximum in the lung tissue, induces seleno-protein like GPx and in turn suppresses NF-kB/IL-17/G-CSF/neutrophil axis to mitigate RP. It is also established that DSePA per se does not protect lung cancer cells from radiation toxicity instead activates the biological mechanism of cell cycle perturbation, unfolded protein response, and apoptosis in lung cancer cells through reductive stress rather than the commonly presumed concept of oxidative stress. Further, DSePA treatment also augments radiation kill by supressing DNA repair (ATM/RAD51/DNAPKs/p53) and cell migration (Akt/G-CSF/EMT) pathways. Conclusions: From in vitro and in vivo studies, DSePA appears to be a novel therapeutic agent to mitigate radiation induced pneumonitis and cell migration in lung cancer model systems.
| IL-25 | | |
| Opuntiol Prevents Photoaging of Mouse Skin via Blocking Inflammatory Responses and Collagen Degradation | | |
N. Rajendra Prasad, P. Veeramani Kandan
Department of Biochemistry and Biotechnology, Annamalai University, Chidambaram, Tamil Nadu, India
E-mail: [email protected]
Introduction: Ultraviolet radiation (UV) is a very prominent environmental toxic agent. In the present study, we investigated the potential of opuntiol, isolated from Opuntia ficus-indica, against UVA radiation-mediated inflammation and skin photoaging in experimental animals. Methodology: The skin-shaved experimental mouse was subjected to UVA exposure at the dosage of 10J/cm2 per day for ten consecutive days (cumulative UVA dose: 100J/cm2). Opuntiol (50mg/kg b.wt.) was topically applied one hour before each UVA exposure. UVA (100J/cm2) exposure induces epidermal hyperplasia and collagen disarrangement which leads to the photoaging-associated molecular changes in the mouse skin. Results: Opuntiol pretreatment prevented UVA-linked clinical macroscopic skin lesions and histological changes in the mouse skin. Further, opuntiol prevents UVA-linked dermal collagen fiber loss in the mouse skin. Short-term UVA radiation (100J/cm2) activates MAPKs through AP-1 and NF-κB p65 transcriptional pathways and subsequently induces the expression of inflammatory proteins and matrix-degrading proteinases in the mouse skin. Interestingly, opuntiol pretreatment inhibited UVA-induced activation of iNOS, VEGF, TNF-α, and COX-2 proteins and consequent activation of MMP-2, MMP-9, and MMP-12 in the mouse skin. Moreover, opuntiol was found to prevent collagen I and III breakdown in UVA radiation-exposed mouse skin. Conclusion: Thus, opuntiol protects mouse skin from UVA radiation-associated photoaging responses through inhibiting inflammatory responses, MAPK activation, and degradation of matrix collagen molecules.
| IL-26 | | |
| Radiation Injury Scenarios: Animal Models and Countermeasures | | |
Sandeep K. Shukla, Ajay K. Sharma, Aman Kalonia, Priyanka Shaw, Rishav Kumar
Division of Molecular and Radiation Biosciences, Institute of Nuclear Medicine and Allied Sciences, DRDO, New Delhi, India
E-mail: [email protected]
Radiation injury may occur in a variety of scenarios, like planned, unplanned, and natural. The injury pattern may be localized, partial, or whole-body. The extent of injury depends on a number of factors, viz. cell type, tissue, dose, and type of radiation. There are well-known instances where radiation exposure is coupled with other types of traumatic injuries. The Hiroshima and Nagasaki nuclear detonation and the Chernobyl nuclear accidents have well documented occurrences of radiation combined injuries (RCI). The combined injury may be of various types, viz. burn, wound, and chemical, that may greatly affect radiation response alone. During a radiation accident or nuclear detonation, it is highly likely that individuals will suffer burns and wounds in addition to radiation. The organ systems of an individual may respond differently due to combined effects. Even after rapid advancements in understanding of radiation injury as well as trauma, the mechanism of RCI is not fully understood. To better understand RCI, many different animal models have been used, each with specific drawbacks. It is imperative to develop suitable animal models that can imitate the RCI. The selection of an appropriate animal model will depend on the type of injury and the animal's response. The radiation countermeasure for RCI is not available at present. The therapeutic approaches for treatment of RCI are very limited and the currently available clinical practices for traumatic injuries are being utilized depending upon the nature of the injury. The present topic will cover radiation combined injury scenarios, mechanisms of injury, animal models, and promising therapeutics.
| IL-27 | | |
| Multiparametric Approach for Biodosimetry Analysis of Challenging Excess Exposures | | |
Nagesh N. Bhat
Biodosimetry Lab, Radiological Physics and Advisory Division, Health, Safety and Environment Group, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Biodosimetry assays based on cytogenetic indicators of radiation damage in human peripheral blood lymphocytes offer themselves as tools of excess exposure assessment. During investigation of excess exposures, Biodosimetry plays a major role to help ascertain the genuineness of exposure. Besides, in many radiological incidents, physical dosimeters may not be available. For effective medical management of exposed individuals and triage, dose information is essential. Cytogenetic indicators such as dicentrics, micronucleus, translocations and premature chromosome condensation have demonstrated to be useful tools for such management. These biological indicators also have limitations in complex exposure scenarios. Non-uniform fields, mixed radiation fields, extremity exposures, protracted exposures, chronic exposures, inter-individual variation of response and medical conditions such as leucopenia are some of the examples. Physical conditions of every exposure scenario are different and they offer challenges for dose assessment. No single indicator will suffice to address the needs of all these challenges. The classic Dicentric chromosome aberration (DCA) assay being highly specific to radiation, which has withstood the test of time, is regarded as the gold standard. The limitations are, stability of signal, may not work at high doses and need of skilled and highly labor intensive scoring. DCA in combination with other assays such as Fluorescent in situ hybridization assay, Premature Chromosome Condensation (PCC) assay, micronucleus assay, γ-H2AX assay and also clinical dosimetry methods help to draw scientific conclusions. When metaphase and culture-based assays fail as in severe leukopenia and apoptosis due to high doses, mitotic fusion based PCC offers detection of multiple signals such as translocations, dicentrics and rings besides quantification of breaks with FISH staining. Interpretation of biodosimetry results with multiple parameters to address such scenario is considered. Statistical uncertainties and dose estimation uncertainties are considered while assigning doses for regulatory requirements. The multi-parametric approach has helped to resolved many complex scenarios of radiation exposures. Biodosimetry lab of HS&EG, BARC being central lab in India, has established state of the art facilities, automation tools and SoPs for addressing the requirements of Biodosimetry analysis and developed many indigenous facilities for achieving high-throughput analysis.
| IL-28 | | |
| Effects of Spaceflight on Biological Systems | | |
Sharmila Bhattacharya
Biological and Physical Sciences Division, NASA Headquarters, Washington DC, USA
E-mail: [email protected]
Space biology investigations help elucidate the effects of spaceflight on biological systems. Spaceflight exposes living organisms to environmental stressors that are not generally experienced on Earth. For instance, a stay on the International Space Station or a journey to the Moon or Mars results in a microgravity environment for an organism, while a stay on the Moon imposes a gravitational force that is one-sixth that of Earth's gravity. Altered gravity environments are known to affect the physiology, behavior, and molecular responses of biological systems. In addition, the levels of ionizing radiation are significantly elevated in the spaceflight environment, and long-duration human spaceflight missions necessitate research on the biological consequences of exposure to radiation, especially in conjunction with altered gravity and other environmental stressors. Dr. Bhattacharya will discuss the use of model organisms to elucidate the molecular, physiological, and behavioral changes experienced by biological systems during exposure to spaceflight and in ground-based analogs of spaceflight.
| IL-29 | | |
| The Modulating Effect of Oxidative Metabolism and Oxygen Partial Pressure on Propagation of Radiation Bystander Effects | | |
G. Gonon1,2, S. M. de Toledo2, V. Perumal3,2, J-P Jay-Gerin4, E. I. Azzam2,5
1Institut de Radioprotection et de Sûreté Nucléaire, PSE-SANTE/SERAMED/LRAcc, Fontenay-Aux-Roses, France,2Department of Radiology, Rutgers New Jersey Medical School, Newark, New Jersey, 3Department of Human Genetics, Sri Ramachandra Institute of Higher Education and Research (Deemed to be University), Chennai, Tamil Nadu, India,4Département de Médecine Nucléaire Et De Radiobiologie, Faculté de Médecine et Des Sciences De La Santé, Université de Sherbrooke, Sherbrooke, 5Radiobiology and Health Branch, Isotopes, Radiobiology and Environment Directorate, Canadian Nuclear Laboratories, Chalk River, Ontario, Canada
E-mail: [email protected]
Introduction: Redox modulated pathways play important roles in out-of-field effects of ionizing radiation. We investigated how the redox environment impacts the propagation of stressful effects from irradiated to bystander cells.
Materials and Methods:
- Normal human fibroblasts that have incorporated [3H]-thymidine were intimately co-cultured with bystander cells during 24 h in a strategy that allowed isolation of bystander cells with high purity. The antioxidant glutathione peroxidase (GPX) was maintained either at wild-type conditions or overexpressed in the bystanders
- Confluent cell cultures pre-treated with t-butyl-hydroperoxide (t-BOOH), an oxidizing agent, or maintained in low oxygen pressure environment approaching in vivo conditions were exposed alongside respective controls to low fluences of 3.7 MeV α particles.
Results: Following 24 h of coculture with [3H]-thymidine-labelled cells, levels of stress-responsive p21Waf1, p-Hdm2, and connexin43 proteins were increased in bystander cells expressing wild-type GPX relative to respective controls. These levels were significantly attenuated when GPX was ectopically overexpressed in the bystanders, demonstrating by direct approach the involvement of oxidative metabolism.
- By exposing confluent cell cultures to 3.7 MeV α particles wherein only ~2 % of cells are traversed through the nucleus by a particle track, increases in chromosomal damage 3 h post-exposure were greater than expected (p<0.001) and further enhanced in presence of t-BOOH (p<0.05). While maintaining and irradiating cell cultures at low oxygen pressure (3.3 or 6.7 mmHg) still supported the participation of bystander cells in responses assessed by chromosomal damage and stress-responsive protein levels (p<0.001), the effects were attenuated compared to ambient pO2 (141 mm Hg) (p<0.05).
Conclusions: Together, the results show that bystander effects are attenuated at below ambient pO2 and when metabolic oxidative stress is reduced but increased when the basal redox environment tilts towards oxidizing conditions. They are consistent with bystander effects being independent of radiation dose rate.
| IL-30 | | |
| Non-Targeted Effect of Pelvic Irradiation on the Brain Through the Gut-Brain Axis | | |
B. S. Venkidesh, Thokur Sreepathy Murali1, Kamalesh Dattaram Mumbrekar
Departments of Radiation Biology and Toxicology and 1Biotechnology, Manipal School of Life Sciences, Manipal Academy of Higher Education, Manipal, Karnataka, India
E-mail: [email protected]
Introduction: Pelvic irradiation has been proven effective in treating a variety of malignancies, including cervical, prostate, and colorectal cancers. One of the unwanted consequences of pelvic radiotherapy is gut microbial dysbiosis, which is correlated with the severity of radiation-induced complications.This could further influence the communication between the gut and the central nervous system in a bidirectional manner.Earlier studies indicate that gut dysbiosis is linked to various neurodegenerative diseases, and bacterial supplementation might help mitigate the disease conditions. Materials and Methods: A single dose of 6Gy of pelvic radiation was given to a 3-4month old Sprague Dawley rat. At regular intervals, fecal samples were collected and processed to determine the microbial abundance using 16S rRNA sequencing. Further, behavioral, histological, and hippocampalgene expression analyses were performed to determine the indirect effect of microbial dysbiosis on the brain. Results: Characteristic changes in the microbial diversity wereobserved at each time point after the pelvic irradiation. There was a significant reduction in the locomotory effect and anxiety level in the open field test of behavior analysis. Further, a significant reduction of the neuronal cells, matured neurons, and an increase in the reactive astrocytes confirmed that pelvic radiation causes neuroinflammation. Finally, the gene expression study showed reduced neuronal plasticity. This study showed that pelvic radiation-induced gut dysbiosis changes the behavior, intestinal morphology, integrity, and brain neuronal maturation, lowering the neuronal plasticity expression levels. Conclusion: Overall, pelvic radiation-induced dysbiosis in the gut microbiota, in turn, led to behavioral changes and neuroinflammation through gut-brain axis interaction.
Acknowledgment: The authors would like to thank the Manipal Academy of Higher Education (MAHE), Manipal for support and infrastructure facilities.
| IL-31 | | |
| Radiation Exposure Assessment: Tools and Techniques | | |
Ajay K. Sharma, Sandeep K. Shukla, Sukhvir Singh, Pradeep Goswami
Institute of Nuclear Medicine and Allied Sciences, DRDO, New Delhi, India
E-mail: [email protected]
There is plentiful likelihood of radiation exposure during radiation emergency operations, accidental or intentional release from nuclear reactor or detonation of radiological dispersal or improvised nuclear device. Radiation exposure assessment is the key criterion to guide the remedial action required to prevent acute injury and also to minimize the potential long term health effects. A number of tools and techniques have been developed for radiation exposure assessment. Biological assessment includes the gold standard of counting the dicentrics, lymphocytes count and also biomarkers such as g-H2Ax, FLt-3, p53 and many more. Physical system includes various dosimetry devices which are developed based on physical phenomena of ionization of material. Now a day's some computer or mobile app based systems are also being developed for the assessment of radiation dosimetry purposes. Each of these has got some advantages and disadvantages over the other. INMAS is also working in this area for quite some time and have developed a smart phone based radiation alert system that utilizes the CMOS sensor present in most of the smart phones. Gamma Radiation generates distinct pattern which are captured as photo frames. The image processing algorithm has been used for detection of spot/artifact produced after gamma radiation exposure. This technology has got immense potential to be used for detection of radiation under various categories like nuclear emergency, radio-isotopes facilities, nuclear medicine, radiation screening at security checkup point for civil and army installations etc. The topic will cover different radiation exposure assessment tools and techniques with their importance.
| IL-32 | | |
| Fluorescence in situ Hybridization Assay for Interphase Chromosomal Aberrationbased Biological Dosimetry | | |
Prabodha Kumar Meher1, Lovia Lundholm,1Andrzej Wojcik1,2
1Department of Molecular Biosciences, Center for Radiation Protection Research, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden, 2Institute of Biology, Jan Kochanowski University, Kielce, Poland
E-mail: [email protected]
Metaphase spreads stained with Giemsa or painted with chromosome specific probes by Fluorescence in situ Hybridization (FISH) have been in use since long for retrospective radiation dose assessment (biological dosimetry). However, this analysis is time consuming. Similarly, analyzing radiation induced chromosomal damage in G0/G1 cells or non-dividing cells by premature chromosome condensation (PCC) is difficult and time consuming. Following large scale radiological emergencies, the time required for analysis is more important than precision of dose estimate. Painting of whole chromosomes using chromosome specific probes in interphase nuclei by the FISH technique will eliminate the time required for cell culture and allow a fast dose estimate. In order to test the applicability of interphase FISH for quick biological dosimetry, whole blood from a healthy donor was irradiated with 8 Gy of gamma radiation. Irradiated whole blood was kept for 2 h at 37°C to allow DNA repair and thereafter processed for FISH with probes specific for chromosome 1 and 2. Damaged chromosomal fragments, distinguished by extra color domains, were observed in interphase nuclei of lymphocytes irradiated with 8 Gy. These fragments were efficiently detected and quantified by the FISH technique utilizing both confocal and single plane fluorescence microscopy. Furthermore, a clear dose response curve for interphase fragments was achieved following exposure to 0, 1, 2, 4 and 8 Gy of gamma radiation. These results demonstrate interphase FISH as a promising test for biodosimetry and for studying cytogenetic effects of radiation in non-dividing cells.
| IL-33 | | |
| Biodosimetric Potential of Acrocentric Chromosome Associations against Gamma Irradiation in Human Lymphocytes | | |
Ravindra M. Samartha
Department of Research, ICMR-Bhopal Memorial Hospital and Research Centre, Government of India, Bhopal, Madhya Pradesh, India
E-mail: [email protected]
Introduction: Acrocentric chromosome associations (ACA) have been widely studied for their incidence in diverse health conditions. Epidemiological findings suggest that low radiation exposure can be a possible cause of nondisjunction in humans. Therefore, frequency of ACA was studied to determine the possible dose-response relationship of gamma irradiation in human lymphocytes. Materials and Methods: Peripheral blood collected from three healthy donors was irradiated with 0, 0.1, 0.25, 0.5, 0.75, 1, 2, 3, 4, and 5 Gy gamma radiation. Chromosomal preparations were made after 48 hrs culture as per the standard guidelines. The experiment was repeated three times, with different donor each time. Results: The ACA frequency in irradiated lymphocytes showed an increase with the dose of radiation. The D-G type of association was most prominent and showed a dose-dependent significant increase in frequency. The dose response of ACA frequencies against radiation dose was found to be linear [ACA Frequency = 0.2923 (± 0.0276) + 0.1846 (± 0.0307) X D and correlation coefficient, r = 0.9442]. As expected dicentric chromosome (DC) frequencies followed the linear quadratic fit model [DC frequency = 0.0015 (±0.0013) + 0.0220 (± 0.0059) X D + 0.0215 (± 0.0018) X D2 and correlation coefficient, r = 0.9982]. The correlation curve was prepared for ACA frequencies versus DC frequencies, resulting in a regression equation as y = 1.130x + 0.4051 (R2 = 0.7408; p = 0.0014). Conclusion: The results of the present study showed an increase in frequencies of ACA in irradiated lymphocytes with an increase in radiation dose thus, ACA may serve as a candidate cytogenetic biomarker for radiation biodosimetry. In microscopic analysis, when compared to DC scoring, ACA scoring requires a lower number of total cells, which may be considered an advantage.
| IL-34 | | |
| Genomic Perspective: Radiation Resistance in Deinococcus Radiodurans | | |
Swathi Kota
Molecular Biology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Deinococcus radiodurans, a non-pathogenic bacterium, is characterized for its extraordinary resistance to various DNA damaging agents including ionizing radiation. Though strong anti-oxidant mechanisms and efficient double strand break repair pathways have been implicated in its radioresistance, the exact molecular mechanisms are not understood. Amongst many, we tried to understand the impact of the two important genomic features on radioresistance of this bacterium (i) its GC rich genome with >69% of Guanine and (ii) multipartite genome and ploidy. The organisms harbouring GC rich genomes are known to exist in different forms other than B-form DNA, especially in G-quadruplex (G4 motif) structures. We found that GC-rich genome of this bacterium harbours numerous potential G4 DNA motifs. Stabilization of G-quadruplex structures with G4 DNA binding ligands during post irradiation recovery phase could sensitize this bacterium to ionizing radiation. Putative G4 DNA motifs could fold as G-quadruplexes in vitro and high throughput transcriptomics as a function of G4 drugs and gamma radiation suggest the role of G4 DNA structure dynamics on DNA damage responsive gene expression. Another interesting cytogenetic feature of this bacterium is multipartite genome and polyploidy. The genome consists of two chromosomes and two plasmids of 6-8 copies during the exponential phase. We have shown that the genome segregation mechanisms of chromosome I and secondary genome elements (chromosome II and mega plasmid) are independent and secondary genome elements are dispensable for normal growth but indispensable for extreme radioresistance. Results supporting the involvement of these genomic features in the radioresistance will be presented.
| IL-35 | | |
| Genetic Enhancement of Crop Plants Using Radiation-Induced Mutagenesis | | |
Suvendu Mondal1,2, Archana Rai1,2, Anand M. Badigannavar1,2, B. K. Das1,2, J. Souframanien1,2, J. G. Manjaya1,2, P. K. Mukherjee1,2
1Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Training School Complex, Mumbai, Maharashtra, India
E-mail: [email protected]
The transition of the study of effects of radiation on plants to its utilization in developing high yielding crop cultivars has made a new dimension of radiation research. The biomarker for radiation injury in plants serves as a good indicator for radiation fallout in any specific area. Ionizing radiation was used in agriculture for the creation of desirable mutations in plant genome. Most of the ionizing radiation causes direct damage through energy deposition on genomic DNA and indirect damage to the nucleotide base through generation of potent radicals or reactive oxygen species. Thus, ionizing radiation can generate large spectrum of DNA damages like single base substitution, small deletion/insertion, large deletion, inversion and translocation. Creation and fixation of such alternations in crop genome through mutation breeding activities at BARC have resulted in the development 56 high yielding crop varieties with desired traits like higher yield, early maturity, semi-dwarf, seed color, disease and pest resistance, abiotic stress tolerance, large seed size, improved seed quality etc. These available mutant germplasms are now being utilized in mutagenomics study to identify the nature of mutations, and causal genes. Present talk will highlight the development of superior crop cultivars through radiation research and exploitation of available mutants in molecular genetics study.
| IL-36 | | |
| Induced Mutagenesis: An Important Tool for Genetic Improvement of Ornamental Flower Crops | | |
Sudhir Singh
Plant Biotechnology and Secondary Metabolites Section, Nuclear Agriculture and Biotechnology Division, Mumbai, Maharashtra, India
E-mail: [email protected]
Genetic variability is the most important factor to determine the success of a plant breeding program for crop improvement. For modern and industrialized floriculture, there is always demand and necessity for new crop varieties. Induced mutagenesis through the application of physical and/or chemical mutagens remains an important tool of ornamental and floricultural plant breeding. The possibilities for creating different genotypes are infinite, and induced mutagenesis can assist us in improving a crop through mutant induction. Among more than 2300 officially released mutant varieties worldwide so far, 566 represent ornamental plants (http://www-mvd.iaea.org). Some of the selected traits of the mutant ornamental plants are flower colour, flower morphology, plant architecture, compact growth, flower type, variegated leaves and disease resistance.We developedseveral stable mutants in Chrysanthemum, Carnation and Gladiolus through induced mutagenesis in combination with biotechnological approaches. This indicates that induced mutagenesis can be an important tool for creating novel genotypes in vegetatively propagated ornamental crops.
| IL-37 | | |
| Strategy for Uranium Projects in India | | |
R. Gupta, A. K. Sarangi
Uranium Corporation of India Ltd
E-mail: [email protected]
The mining and processing of uranium ore in India started after the formation of Uranium Corporation of India Ltd. (UCIL) in October 1967. Since then, the Company has grown manifold supporting the uranium need of the country. The Company is now operating six underground mines, one opencast mine and two processing plants in the Singhbhum region in Jharkhand. One more underground mine and processing plant are in operation at Tummalapalle in Andhra Pradesh. A large uranium province has already been identified by AMD adjacent to Tummalapalle which host about 60% of Indian uranium resource. Known deposits in Singhbhum in Jharkhand host about 20% of country's resource. Both the areas await efforts to expand and / or open production facilities. In addition, the areas around Rohil in Rajasthan, Gogi in Karnataka, Lambapur-Pedagattu in Telengana and Mahadek basin in Meghalaya host substantial resource with potential for new set-ups. India's known uranium deposits are of low grade and small to medium in size. The large low-grade resource around Tummalapalle need out-of-box considerations in mine development strategy which may include contract mining for development and bulk production, adopting cutting edge technology for early production, tie-ups with machinery manufacturers, maximising the use of deslimed tailings for mine back filling, minimising the process loss etc. Potential deposits in Jharkhand and Rajasthan may call for distinctive mine layout and development approach for early production, expansion of existing plants to accommodate additional ore and / or setting of new plant for cluster of small deposits and extraction of all valuable by-products for commercial benefit to the project along with conservation of valuable mineral wealth. Other greenfield areas need long-term project development strategy considering the potential negative public opinion against uranium mining. Coordinating with AMD to educate people on benefits of uranium mining, site-specific proposals for potential Project Affected People (PAP), neighbourhood infrastructure development work before the start of project, involving State Govt and local bodies as stake holders etc will help to garner good-will for uranium mining. Further, initiating the process for regulatory clearances including land acquisition, preparation of critical agreements / contracts, identification of contractors / suppliers, development of flowsheet for new deposits etc during exploration will help in early commissioning of the project.
| IL-38 | | |
| Microbial Responses to Uranium Exposure and the Underlying Mechanisms for Uranium Tolerance | | |
Celin Acharya1,2
1Molecular Biology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Uranium (U) is a naturally occurring radionuclide. The levels of U contamination in the environment as a result of accidental release from anthropogenic nuclear activities could present major health concerns for human and environmental health. The toxic effects of U are associated with its bioavailability and speciation. Prokaryotes have been present on earth for over 3 billion years and are considered to be the most successful life forms. While U is physiologically irrelevant to the microbial cells, the ever-changing environmental landscape resulting from elevated concentrations of U has facilitated the evolution of adaptive responses in bacterial strains native to uranium enriched environments. Such bacteria effectively interact with U through direct or indirect mechanisms which results in mobilization or immobilization of U in the environment. My talk will present an overview of our laboratory studies on some of these mechanisms demonstrated by the microbes isolated from the marine environment and uranium rich deposit. These uranyl-microbe interactions importantly form the basis for developing strategies for management of uranium contamination.
| AARR Award Recipient Lecture (AARRL) | | |
| AARRL-S1 | | |
| Association of Low-Dose Ionizing Radiation with Cancers: Chinese Medical X-Ray Workers Cohort Study | | |
Qiang Liu, Yeqing Gu, Jinhan Wang, Yan Wang, Chang Xu, Liqing Du, Kaihua Ji, Ningning He, Manman Zhang, Huijuan Song
Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin, China
E-mail: [email protected]
Introduction: As well known to all, the dose-response relationship between cancers and protracted low-dose rate exposure to ionizing radiation is still uncertain. The aim of the present study was to estimate quantified relationships between low-dose radiation exposures and site-specific solid cancers among Chinese medical X-ray workers. Materials and Methods: The Chinese medical X-ray worker study(CMXW) included 27,011 medical X-ray workers employed at major hospitals in 24 provinces in China from 1950 to 1980, and a control population of 25,782 physicians matched by hospital, who were unexposed to X-ray equipment. Poisson regression models were used to estimate the excess relative risk (ERR) and excess absolute risk (EAR) for incidence of solid cancers and leukemia associated with cumulative dose. Results: A total of 1,643 cases of solid cancer were identified in 1.45 million person-years offollow-up. Among X-ray workers, the average cumulative colon and red bone marrow dose was 0.084Gy and 0.046Gy, respectively. The estimated ERR/Gy for all solid cancer was 0.87(95% CI: 0.48, 1.45), and the EAR was 22 per 104PY-Gy (95% CI: 14, 32).The most common solid cancers were lung, liver, and stomach cancer. We found a positive relationship between 5-year lagged organ-specific dose and liver (ERR/Gy = 1.48; 95%CI, 0.40-2.83) and esophagus cancers (ERR/Gy = 18.1; 95%CI, 6.25-39.1). No significant relationship between cumulative organ-specific doses and other cancers. Moreover, the results showed a statistically significant EAR for lung, liver, thyroid, and skin cancers.We found a positive relationship between 2-year lagged cumulative red bone marrow dose and risk of leukemia excluding chronic lymphocytic leukemia (CLL) (ERR = 0.66 per 100 mGy, 90% CI: 0.09, 1.53; EAR = 0.29 per 104PY-100 mGy, 90% CI: 0.07, 0.56). Conclusions: The present study provides strong evidence of the dose-response association between cumulative organ-specific dose and incidence of site-specific cancers. These findings provided more useful insights into the risks of site-specific cancers from protracted low-dose rate exposure to ionizing radiation.
| AARRL-S2 | | |
| Radiation Biology, Medicine and Chromosome | | |
S. Tashiro
Department of Cellular Biology, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima, Japan
E-mail: [email protected]
My first encounter with radiation biology was a lecture on atomic-bomb survivor leukemia when I was a student in medical school. Since then, I have been interested in the mechanism of chromosome translocations. Chromosomal DNA, on which human genetic information is inscribed, is constantly damaged by radiation exposure from the natural world and ultraviolet rays of sunlight, etc. Repair errors of DNA damage can cause chromosomal aberrations, genetic mutations, and other alterations of genetic information, leading to leukemia, solid tumors, and other diseases known to be caused by radiation exposure to the human body. DNA repair system and higher order nuclear architecture play important roles in chromosome translocations. RAD51 is a key factor involved in the recombinational repair of DNA double strand breaks. We first identified that RAD51 forms nuclear foci at sites containing DNA damage by using a UVA microirradiation system. We then studied the mechanisms of RAD51 focus formation using bio-imaging techniques. We could show that the radiation-induced focus formation of RAD51 is regulated by the multi-layer systems, including histone modifications/exchange, nuclear-cytoplasm shuttling and SUMO modification systems. By applying knowledges and techniques we had obtained through DNA repair research, we established the PNA-FISH technique for the high-throughput chromosome analysis in biological dosimetry. Applying this technique, we could detect the chromosome abnormalities induced by a standard dose CT scan, but not by a low-dose CT scan, used for the lung cancer screening in high risk population, suggesting that application of low-dose CT for lung cancer screening is justified from a biological point of view. We also found the possibility that chromosome aberrations could be a maker to predict side effects of radiation therapy for esophageal cancer. We are going to expand the application of such new biodosimetry techniques for the better management of medical exposure.
| AARRL-S3 | | |
| The Efforts of Medical Physicists for Ending the SARS-CoV-2 Pandemic Crisis | | |
S. M. Javad Mortazavi, Alireza Mortazavi1
Department of Medical Physics and Engineering, School of Medicine, Shiraz University of Medical Sciences, 1School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
E-mail: [email protected]
As of October 15, 2022, SARS-CoV-2, the RNA coronavirus that caused the pandemic, has affected over 624 million people globally, with more than 6.57 million reported deaths. During the COVID-19 pandemic, many physicists around the globe, used their expertise to manage the crisis. A team led by Dr. Joseph John Bevelacqua, Dr. Alireza Mortazavi, and I introduced the use of low-dose radiation therapy (LDRT) for COVID-19 pneumonia patients in early April 2020. What makes our contribution to COVID-19 science so important is not only proposing a treatment method, but also warning the scientists and health policymakers about the hazards of treatment methods such as antivirals that exert selective pressure on the virus and drive it to mutations and the emergence of new variants. At that time scientists were not seriously worried about the emergence of new variants. Our early report received global attention, especially for noting the key disadvantages of antiviral drugs that exert selective pressure on the virus. The radiation doses that can be used in LDRT are much lower than the dose required for the inactivation of the virus. Therefore, LDRT could not accelerate the evolution of the virus through mutations. The overall outcome of the several clinical trials conducted on LDRT for COVID so far, are promising. Given this consideration, despite early debates, now the therapeutic advantages of LDRT for COVID are well documented. In summary, the continuous exposure of the virus to different stressors creates selective pressure and paves the road for viral evolution. LDRT as an absolute selective pressure-free treatment has shown to reveal promising results in clinical trials conducted so far.
| AARRL-YS1 | | |
| Using Nano-Metal Organic Framework to SequesterUranium In Vivo | | |
Juan Diwu
State Key Laboratory of Radiation Medicine and Protection, School for Radiological and Interdisciplinary Sciences (RAD-X) and Collaborative Innovation Center of Radiation Medicine of Jiangsu Higher Education Institutions, Soochow University, Suzhou, China
E-mail: [email protected]
Introduction: In vivo sequestration of uranium is critical for the nuclear and radiation safety, since uranium isotopes play an indispensable role in the development of nuclear industry and exhibit both chemo- and radio-toxicity. In this work, functional nano MOF materials are proposed to decorporate uranium in vivo. Materials and Methods: A zirconium-based organic framework (noted as UiO-66-(COOH)4-180) with uncoordinated carboxyl groups was synthesized by solvothermal method and post-synthetic exchange method. The binding ability and selectivity of UiO-66-(COOH)4-180 toward uranium were evaluated by chemicaladsorption experiments. The cytotoxicity, biodistribution, metabolism, and uranium decorporation efficacy of UiO-66-(COOH)4-180 were comprehensively evaluated by in vitro and in vivo studies. Results: Theuranyl adsorption experiments show that UiO-66-(COOH)4-180 reached the adsorption equilibrium within 1 min in water and removed more than 65% of uranyl inFBS within 5 min. Moreover, in the presence of other endogenous metal cations, the uranyl removal rate was still nearly 100%. The cytotoxicity of UiO-66-(COOH)4-180 was on the similar level of the clinically approved drug, ZnNa3-DTPA. And biodistribution and metabolism studies show that UiO-66-(COOH)4-180 could selectively accumulate in kidneys where uranium mainly deposits.The uranium decorporation experiments in vivo show that UiO-66-(COOH)4-180 could successfully reduce the amounts of uranyl deposited in kidneysand femurs by up to 55.4% and 36.5%, respectively, while DTPA only exhibited limited effect. Conclusions: Due to the pre-organization and preferential orientation of the carboxyl group in the channel, which forms a coordinated trap to combine with the uranyl units, UiO-66-(COOH)4-180 exhibits the fastest uranium uptake kinetics and high selectivity toward uranyl. In vivo assays suggest that UiO-66-(COOH)4-180 could prevent the uranyl deposition, and compared with ZnNa3-DTPA and BHC which both contain carboxylic functional groups,UiO-66-(COOH)4-180 significantly improves the uranium decorporation efficiency.The rational design and development of these novel nanomaterials is essential for emergency treatment of individuals internally contaminated by radioactive nuclides.
| AARRL-YS2 | | |
| The Tissue-Sparing Effect of Microbeam Radiotherapy for Preserving Male Fertility | | |
Hisanori Fukunaga1,2
1Faculty of Health Sciences, 2Center for Environmental and Health Sciences, Hokkaido University, Sapporo, Japan
E-mail: [email protected]
Introduction: The delivery of a single high-dose fraction of radiation to a large treatment area divided into several smaller fields has been the basis of microbeam radiotherapy (MRT). MRT has demonstrated an effective tissue-sparing effect (TSE) in various tissue types and species after spatially fractionated irradiation at the microscale level. However, the underlying biological mechanism of this process remains obscure. Thus, we hypothesized that the TSE of MRT on the testes helps preserve male fertility while allowing for the delivery of high radiation doses to the tumor. Materials and Methods: We used high-precision MRT techniques and a unique ex vivo testicular tissue culture to monitor the progress of spermatogenesis and to easily assess radiation-induced biological effects. In addition, a transgenic mouse model expressing acrosome-green fluorescent protein was used as a meiosis-specific biomarker. Results: We demonstrated the significant testicular TSE of high-precision MRT for maintaining spermatogenesis. Our analysis results also clarified that the survival and potential migration steps of non-irradiated germ stem cells in irradiated testicular tissues are required to ensure an effective TSE for spermatogenesis. Conclusions: Our findings suggest the clinical potential of MRT for achieving better outcomes while preserving male fertility during and after radiotherapy.
| AARRL-YS3 | | |
| Tumor-Targeted Magneto-Liposomes Induce Radio-Sensitization in Cancer Cells via Induction of Enhanced DNA Damage and Activation of JNK Pro-Apoptotic Signalling Axis | | |
Neena G. Shetake, Amit Kumar, B. N. Pandey
Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Radiotherapy is regarded as one of the most important modalities of cancer treatment, but its therapeutic efficacy is hampered by its non-specificity and the associated normal tissue damage.These limitations can be overcome by development of nanoparticles for targeted delivery of anti-cancer drugs/radio-sensitizers. Materials and Methods: Liposomes and IONPs were synthesized by thin film hydration and chemical co-precipitation methods, respectively. The targeted nano-formulation (T-LMD) was characterized by DLS and cryo-TEM, followed by studying the endocytic uptake by confocal microscopy. The radio-sensitization efficacy of T-LMD was determined by MTT and clonogenic assays. Mechanism of radio-sensitization was studied by Western blotting, immuno-cytochemistry and flow cytometry. Results: Cyclic RGD labeled liposomes (T-LMD) co-encapsulated with IONPs and doxorubicin (Dox) were synthesized and showed an average size of 166 ± 42 nm. Blocking the αvβ3 integrin receptor with cyclic RGD peptide and co-localization studies using early endosomal marker (EEA-1), validated the receptor mediated uptake of T-LMD in U373MG (human brain cancer) cells. T-LMD showed a dose dependent increase in radio-sensitization in WEHI-164 and U373MG cells as studied by MTT and cell cycle analysis. γ-H2Ax assay suggested a significant increase in the magnitude of DNA double strand breaks (DSBs) after treatment with T-LMD±2 Gy. Interestingly, T-LMD also induced a slower repair kinetics of DSBs as compared to only 2 Gy treatment. Mechanism of chemo-radio-sensitization was found to involve activation of JNK mediated pro-apoptotic signaling axis and delayed repair of DNA double strand breaks. Real time imaging of ICG labeled T-LMD suggested ~6-18 folds higher tumor accumulation of T-LMD as compared to off-target organs (kidney, liver, spleen, intestine, lungs and heart). Moreover, T-LMD showed significantly higher combinatorial tumor therapy efficacy as compared to LippodTM (clinical liposomal Dox). Conclusions: T-LMD showed targeted delivery of Dox and enhancement of radio-therapy efficacy, suggesting its potential for targeted multi-modal cancertherapy.
| SRR Young Scientist Award Presentation (SRR-YSA) | | |
| SRR-YSA1 | | |
| Estimation of Radium Content Its Effect and Radon Exhalation Rates in Soil Samples of in Uttarakhand Himalaya | | |
Manjulata Yadav, R. C. Ramola1
Department of Physics, Government Degree College Nainidanda, 1Department of Physics, H. N. B. Garhwal University, Badshahi Thaul Campus, Srinagar, Uttarakhand, India
E-mail:[email protected]
The radon emanation is associated with the presence of radium and its ultimate precursor uranium in the ground. Although these elements occur in virtually all types of rocks and soil, their amounts vary with the specific site and geological materials. Each decay of a 226Ra nucleus produces a 222Rn atom. If the radon is produced close to the earth surface, it can escape rather than remain trapped in the ground. The radon emanation rate varies from place to place to differences in radium concentration and soil permeability. The total amount of radon emanation per unit area and per unit time is called the exhalation. In this paper we calculate the value of radon and its effect in human being. The values of radon exhalation rate and radium contents in soil samples of Tehri were found to vary from 0.042×10-5 Bq.kg-1h-1 to 1.49×10-5 Bq.kg-1h-1 with an average of 0.74 ×10-5 Bq.kg-1h-1 and from 31±6 Bq.kg-1 to 67±8 Bq.kg-1 with an average of 46 Bq.kg-1, respectively. The observed significant values of radium and radon mass exhalation rate are due to the presence of uranium bearing minerals in the area.
| SRR-YSA2 | | |
| Radioimmunotherapeutic Agents for Targeting Human Epidermal Growth Factor Receptor 2 Overexpressing Cancers | | |
Rohit Sharma1,2, Archana Mukherjee1,2
1Radiopharmaceuticals Division, BARC, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Development of radiolabeled antibody-based radiopharmaceuticals has seen a renaissance in the last decade. Monoclonal antibodies (mAbs) and their fragments are gaining attention especially in oncology as molecules specific to the cell surface antigens overexpressed on tumors, immune cells and also for targets in the tumor microenvironment have been developed which can be radiolabeled for targeting cancers. Trastuzumab and pertuzumab are FDA approved monoclonal antibodies targeting HER2 receptors overexpressed on breast cancers. Lu-177 (β-max498 KeV, t1/26.7 d) is a promising radioisotope for development of radioimmunotherapeutic agents. It has several advantages over other radionuclides for radioimmunotherapy (RIT) including relatively long physical half-life which appropriately matches with the biological half-life of intact mAbs. Materials and Methods: To demonstrate the potential of177Lu labeled trastuzumab/pertuzumab and their F(ab')2 fragments for targeting HER2 receptors, p-SCN-Bn-CHX-A”-DTPA chelator was conjugated to antibodies and its fragments and subsequently labelling with177Lu was optimized. The radiochemical purity (RCP) labeled antibody were assessed by chromatographic technique (SE-HPLC & PC).The radioimmunoconjugates were used for in-vitro binding affinity and specificity studies in SK-BR3, SK-OV-3 and MDA-MB-231 cell lines while in-vivo studies were conducted in tumor bearing female SCID mice. Results: 177Lu-labeled formulations could be prepared with >95% radiochemical purity (% RCP) as ascertained by chromatography techniques. In-vitro binding studies revealed high affinity and specificity of the formulations towards HER2 receptors. Strong synergy in binding of radiolabeled pertuzumab in presence of cold trastuzumab was observed. SPECT imaging in tumor bearing mice revealed specific tumor uptake and prolonged retention of the radioformulationsin HER2 positive tumors. In-vivo synergy in binding and uptake was also confirmed by SPECT imaging studies. Conclusions: The results of the studies revealed that these radioformulations could serve as effective RIT agents for management of HER2 overexpressing cancers. Further studies are underway to obtain the regulatory clearance for translating177Lu labeled formulations to clinics.
| SRR-YSA3 | | |
| A Study of Alpha Particle Track Profiles on CR-39 from Beach Sand By Laser Induced Microscopy | | |
S. S. Chavan, H. K. Bagla
Department of Nuclear and Radiochemistry, K. C. College, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected]
Pure radioactivity in soil and beach sand rises from 238U, 232Th and natural 40K. The vaporous radioactive isotope of radon through characteristic natural sources incorporates a significant portion in the total major natural resources of exposure to human beings. Alpha particle radioactivity in the form of tracks on CR-39 as Solid State Nuclear Track Detector (SSNTD) has been evaluated. CR-39 detectors were exposed to different Beach sand samples collected from east coastal India, containing Thorium as an alpha source. CR-39 detector with size 1X 1.5 cm2 was exposed in about 1 g of each sand sample and 10 mL of each sea water sample in different containers. After exposure, chemical etching was done with 6 M NaOH and newly introduced etchant with addition of 5% Tetraethyl ammonium bromide with 6 M NaOH at 60°C for different etching time.The Atomic Force Microscope and Laser Induced Microscope has been employed to study the surface of CR-39 detector. Track density (Td) and track diameter for sand samples were calculated. The improved resolution of Laser Induced Microscopy allows for more detailed studies of the tracks formed by ionising particles. 238U &232Th presence by EDXRF and their content were measured by ICP-MS. The output of the newly developed etching technique appears extremely better for the visual track appearance, track density and track diameter profile. The various track profiles from sand samples using advanced microscopy results have been specified that the natural alpha radionuclides were present in the samples. However, the activity levels in few sand samples were significantly higher than the other collected sand sample. 238U &232Th contents of collected beach sand sample from selected area was found much less than the permissible limit which is safe from the health hazard point of view.
| SRR-YSA4 | | |
| Studies on Removal of Lead from Water Using Radiation Grafted Adsorbent | | |
Swarnima Rawat1,2, N. Misra1, A. Singhadeb1, S. A. Shelkar1, N. K. Goel1, A. R. Khaire1, M. Ali3, V. Kumar1,2
1Radiation Technology Development Division, BARC, 3Chemical Engineering Division, BARC, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected]
Contamination of water with lead (Pb) has arisen as a major concern, since Pb is reported to adversely affect kidney, liver, nervous system and reproductive system. As per WHO guidelines, maximum permissible limit for lead in drinking water is 10 ppb. Therefore, need for efficient removal of Pb from water using green methodologies is paramount. In this work, a low cost, efficient and biodegradable radiation functionalized polymer based anionic adsorbent has been fabricated by radiation grafting of 4-Vinyl benzene sulfonic acid sodium salt (VBSA) on cotton cellulose using gamma radiation. This anionic adsorbent (VBSA-g- Cellulose) despite functioning on the basis of ion exchange has an edge over conventional ion exchange resins due to its biodegradability, cost-effectiveness and green approach of fabrication with only water being used as green solvent. The adsorbent was characterized using techniques, such as grafting yield estimation, FTIR, TGA and SEM.VBSA-g- Cellulose adsorbent was subsequently used for removal of Pb from water. Pb concentration in water was estimated using ICP-OES. Adsorption isotherm was established by carrying out equilibrium adsorption studies. Adsorption isotherm data was analysed by linear and non-linear fitting of different isotherm models (Langmuir, Freundlich, etc.) to find the best fit model. Langmuir Isotherm model was found to best fit the adsorption data. The adsorption capacity of the adsorbent for Pb was found out to be ~98 mg/g. Additionally, different kinetic models (pseudo first and second order) were also used to analyse the kinetic data. The adsorption process was found to follow pseudo second order kinetics. The radiation grafted adsorbent could be regenerated using suitable eluent and recycled for several cycles.
| Oral Presentation (OP) | | |
| O-1.1 | | |
| BCCIP Mediated Replication Stress in Glioma Stem Cells Drives Their Radiation Resistance | | |
Bhawna Singh1,2, Shilpee Dutt1,2
1Shilpee Dutt laboratory, Tata Memorial Centre, Advanced Centre for Treatment, Research and Education in Cancer, Navi Mumbai, 2Homi Bhabha National Institute, Training School Complex, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Radiation therapy acts by damaging DNA, increasing DNA replication stress (RS) leading to apoptosis in cancer cells. Radio-resistance in glioblastoma (GBM) is attributed to GBMstem-like cells (GSCs) that survive radiotherapy by overcoming radiation-induced RS. However, lack of molecular understanding of RS management in GSCs prevent the exploitation of this pathway to target GSCs. Therefore, we sought to identify key players involved in RS pathway in GSCs. Materials and Methods: GSCs were isolated from GBM patient samples and cell lines. Stemness was characterized by neurosphere formation, surface markers and gene expression. RS was determined by DNA fibre analysis, proximity ligation assay (PLA), immunoblotting, immunofluorescence and comet assays. Knockdowns and molecular biology experiments were performed for functional studies. Results: Stem cells were enriched from GBM patient samples and cell lines by neurosphere cultures and characterized using OCT4, SOX2 and Nestin markers at protein and RNA levels.We found significant GSC enrichment in neurospheres. DNA fibre assay and recruitment of replication proteins (pRPA, MRE11, RAD51, γH2AX) revealed high intrinsic RS and DNA damage in GSCs compared to non-GSCs. BRCA2 and CDKN1A-interacting protein (BCCIP) was identified as a BRCA2 and RAD51 binding protein that we hypothesized plays a key role in RS. Indeed, PLA and colocalization with RAD51 showed BCCIP recruitment at replication forks. siRNA-mediated knockdown of BCCIP significantly increased stalled replication fork frequency, origin firing, reduced replication recovery, and fork speed that resulted in DNA double-strand break formation. Furthermore, BCCIP at replication fork is required to prevent nucleolytic degradation of nascent DNA strands by MRE11. Conclusions: We identified BCCIP as a novel protein, crucial for maintenance of replication fork stability, replication stress response efficiency, and genome instability in GSCs. Thus, combining BCCIP inhibition with radiation therapy is a promising GSC-specific therapeutic approach to improve patient outcomes in GBM.
| O-1.2 | | |
| Role of Wharton's Jelly Mesenchymal Stem Cells and their Soluble Factors in Therapeutic Radioprotection | | |
Dharmendra Kumar Maurya1,2
1Radiation Biology and Health Sciences Division, Bio-Science Group, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected]
Radiation exposure can cause toxicity to sensitive tissues and organs leading to Acute Radiation Syndrome (ARS). During accidental radiation exposure, severity of ARS varies with the absorbed dose. The treatment modalities for management of ARS are very limited and ineffective. Cell-based therapy specially using stem cells and their soluble factors are gaining wide attention in the field of regenerative medicine to treat various pathological conditions like hereditary disorders, ageing and degenerative diseases. Therefore, we hypothesized that stem cells and their secreted factors may be useful for amelioration of ARS. We have employed human Wharton's Jelly Mesenchymal Stem Cells (hWJ-MSCs) for the therapeutic management of radiation injuries in mice. It was found that hWJ-MSCs and their conditioned media exhibit significant therapeutic radioprotective potential against radiation induced hematopoietic toxicity and gastro-intestinal injury. These cells preferentially home to the radiosensitive tissues like spleen, bone marrow and small intestine of irradiated mice and secrete various soluble mediators which can repair/regenerate the damaged tissue. The transplanted hWJ-MSCs produce human cytokines and induce the production of mouse cytokines in the irradiated mice. Among these, WJ-MSCs derived human IL-6 and G-CSF were found to play a causal role in radioprotection. Further, it was found that radioprotection offered by hWJ-MSC-conditioned media was abrogated when it was neutralized with G-CSF antibody indicating the role of the G-CSF in WJ-MSC-conditioned media mediated radioprotection. Soluble mediators secreted in mice after WJ-MSCs infusion showed very complex protein-protein-interactions and these interactions could be responsible for specific consequences in radiosensitive and radio-resistant tissues. In conclusion, systemic infusion of hWJ-MSCs and their conditioned media will have significant potential for treating accidentallyexposed radiation victims by helping in the regeneration and repair of damaged tissues and organs.
| O-1.3 | | |
| Investigation of DNA Double-Strand Break Repair in Mitochondria under Various Stress Conditions | | |
Diksha Rathore, Sathees C. Raghavan
Department of Biochemistry, Indian Institute of Science, Bengaluru, Karnataka, India
E-mail: [email protected]
Mitochondrial DNA is continuously exposed to various endogenous and exogenous agents that cause DNA damage. Base Excision repair (BER) is one of the well-established DNA repair pathways in mitochondria. Besides, mismatch repair, homologous recombination (HR), and micro-homology mediated end joining (MMEJ) etc have also been reported. However, Nonhomologous end joining (NHEJ) is undetectable in mitochondria. In the present study, we aim to investigate impact of various stress conditions on DNA repair proteins in mitochondria. Firstly, we have investigated impact on translocation of DNA repair proteins to mitochondria following exposure to ionizing radiation (IR) or Menadione, a chemical that induces ROS in mitochondria. To study this, HeLa cells were exposed with Menadione (50 μM, 1 h) or IR (2, and 5 Gy), and the effect was assayed either by immunofluorescence or western blot analysis. Results revealed that treatment with Menadione increased the translocation of DNA repair proteins such as DNA LigaseIII, MRE11, RAD51, PARP etc. The functional significance of increased translocation of these proteins into mitochondria was investigated by preparing mitochondrial extract (ME) with and without Menadione or IR treatment. DSB repair efficacy of the extracts were evaluated and compared using HR, NHEJ and MMEJ assays. A significant increase in the efficacy of MMEJ and HR was observed when DNA substrates bearing DSBs were incubated with mitochondrial extract that was prepared from Menadione treated cells. Impact of exposure of IR in mitochondrial DNA repair will also be discussed. In summary, exposure to the DNA damaging agents results in the translocation of proteins to mitochondria leading to the elevated repair.
| O-1.4 | | |
| Oncogenic Role of TCTP in Radioresistance and Poor Outcome in Oral Cancers | | |
Dipti Sharma1,2*, Sagar N. Pawar1,#, Prasad Sulkshane1,2,3,#, Rohit Waghole1,$, Yasser Mohd1,2,4,$, Sushil S. Pawar5, Sadhana Kannan6, Nazia Chaudhary2,7, Nikhil Gadewal8, Jai Prakash Agarwal9, Tanuja Teni1,2
1Teni Lab, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, 6Clinical Research Secretariat, Advanced Centre for Treatment, Research and Education in Cancer, TMC, 7Cell and Tumor Biology, Advanced Centre for Treatment, Research and Education in Cancer, TMC, 8Bioinformatics Centre, Advanced Centre for Treatment, Research and Education in Cancer, TMC, Navi Mumbai, 5Department of Oral Pathology and Microbiology, KBH Dental College and Hospital, Nashik, 2Homi Bhabha National Institute, 9Department of Radiation Oncology, Tata Memorial Hospital, Mumbai, Maharashtra, India, 3Glickman Laboratory, Technion-Israel Institute of Technology, Haifa, Israel, 4Institute of Pathology, Greifswald University Medicine, Greifswald, Germany
E-mail: [email protected]
#These authors contributed equally. $These authors contributed equally.
Introduction: Radiotherapy resistance is a major hurdle in oral cancer treatment leading to poor outcomes. To understand radioresistance associated molecular perturbations, we previously established two radioresistant sublines from parental oral cancer cell lines and identified Translationally Controlled Tumour Protein (TCTP) as commonly differentially over expressed protein. The current study thus aimed to assess the role of TCTP in radioresistance and patient outcomes in oral cancers. Materials and Methods: TCTP expression was assessed by Immunohistochemistry and Immunoblotting in oral cancer cells and tissues. Coimmunoprecipitation, Immunofluorescence, molecular docking & simulation was performed to assess the interaction of TCTP with DNA damage repair proteins. Clonogenic assay, Soft-agar assay and Matrigel invasion assay was used to assess radioresistance, anchorage-independent growth and invasive abilities of the cells respectively. Results: TCTP levels were significantly (p<0.0001) elevated in oral premalignant lesions and oral tumours versus normal mucosa. Additionally, oral cancer patients with elevated TCTP showed significantly (p<0.05) poor Recurrence Free Survival (RFS) and poor overall survival. Additionally, TCTP levels were further upregulated in oral cancer cells upon exposure to Ionizing Radiation (IR) and remained elevated during the establishment of radioresistant cells. Interestingly, elevated TCTP levels in treatment-naive oral tumours from patients treated with curative radiotherapy exhibited significant (p<0.01) association with unfavourable treatment response and poor RFS (p<0.0001), implying TCTP to be a radioresistance-related factor. We found TCTP colocalized and was present in complex with pATM and γH2AX in response to IR. TCTP knockdown resulted in delayed phosphorylation of ATM and decreased levels γH2AX. Docking and simulation analysis indicated TCTP stabilises ATM-H2AX complex. TCTP downregulation in radioresistant cells resulted in radio-sensitisation, decreased anchorage-independent growth and decreased invasion. Conclusions: Our studies demonstrate oncogenic role of TCTP in oral carcinogenesis partly via stabilisation of ATM-H2AX complex, thereby enhancing DNA damage response. Thus our studies suggest TCTP as potential radioresistance and prognostic marker for oral cancer patient outcome.
| O-1.5 | | |
| Cancer Risks after Low-Dose Exposure to Ionizing Radiation: IARC Perspectives | | |
Evgenia Ostroumova, Ljubica Zupunski, Joachim Schüz
International Agency for Research on Cancer-WHO, Lyon, France
E-mail: [email protected]
International Agency for Research on Cancer (IARC) is a specialized institution of the World Health Organization focused on prevention, early detection, and causative cancer research. Environmental and Lifestyle Epidemiology Branch (ENV) of IARC is at the forefront of identifying modifiable cancer risk factors and protective factors to support the IARC vision of a world where fewer people develop cancer. Environmental factors are considered by IARC and ENV to comprise potentially carcinogenic substances in any medium – including soil, water, food, and air – that expose humans in their workplace, at home, and in the general environment. Among these factors are all types of radiation (ionizing, and non-ionizing). Lifestyle factors are habits or behaviours that are due to individual choice and to life circumstances within a socioeconomic and cultural context. The scope of ENV's ionizing radiation research includes long-term cancer risks in various groups of populations, namely in utero exposed, offspring to exposed parents, residents of radioactively contaminated territories, occupationally exposed professionals, or patients who underwent medical diagnostic irradiation (computer tomography). ENV scientists actively collaborate with international experts in dosimetry, radiobiology, clinical medicine, epidemiology, and risk communication to study the consequences of the Chernobyl radioactive fallout, nuclear weapon tests in the Semipalatinsk nuclear test site in Kazakhstan, and radioactive contamination of the Techa River from activities of the “Mayak” plutonium production association in the Southern Urals of Russia. Special attention is given to cancer research in relation to uranium exposure from gold mine tailings in the South Africa residents living in the proximity to the tailings. ENV and IARC act as an observer and play an active role in the meetings and task group activities carried out by the United Nations Scientific Committee on Effects of Atomic Radiation (UNSCEAR) and the International Commission on Radiological Protection (ICRP).
| O-1.6 | | |
| DNA Methylation Profile of Repetitive Element, DNA Damage Response and Repair Genes in Human Blood Cells from Normal and High Level Natural Radiation Areas of Kerala Coast, India | | |
Rashmi Priya, D. C. Soren, Birajalaxmi Das
Low Level Radiation Research Section, Radiation Biology and Health Sciences Division, Bio Science Group, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected]
Introduction: Biological effect of chronic low dose ionizing radiation was evaluated at molecular level on human population residing in normal and high level natural radiation areas (N &HLNRA) of Kerala coast. The level of radiation dose in Kerala coast varies from <1.0 mGy to 45 mGy/year. Studies conducted in this population showed reduction of DNA damage, efficient repair and active role of DNA damage response (DDR), DNA repair, Methylation and immune response genes in HLNRA. The objective of the present study is to find out the effect of chronic low dose radiation on DNA methylation profile of DDR, repair and repetitive element in blood cells of individuals from N& HLNRA population. Methodology: Blood sample were collected from 42 healthy individuals from NLNRA (N=15), HLNRA Group I (1.51 -10.0 mGy/year, N= 18) and HLNRA Group II (>10.0 mGy/year, N=9). Genomic DNA isolated from peripheral blood mononuclear cells (PBMCs) was bisulphite converted and DNA methylation status was determined for repetitive element (LINE1) and promotor methylation status of ATM, RAD23B, MLH1, MGMT APC, DNMT3A, MRE11A, XPC, BRCA1, PARP1, MTHFRusing Methyl sensitive high melting (MS-HRM) method.Gene expression of selected DDR and repair genes are also evaluated. Results: LINE1, MTHFR and TNFα showed maximum % of methylation at basal level in NLNRA and HLNRA. BRCA1 and RAD23B genes showed hyper-methylation among HLNRA group II. ATM, APC and PARP1 showed similar pattern among NLNRA and HLNRA individuals. MLH1, XPC and MGMT remained un-methylated. Gene expression profile did not show significant changes at majority of the genes studied in HLNRA. Conclusion: Increased Promotor methylation observed at some of the DNA repair genes i.e., BRCA1 and RAD23B could be indicative of epigenetic signatures due to chronic low dose radiation. However, further studies with larger sample size are required to draw firm conclusion.
| O-1.7 | | |
| Investigating the Effects of Low Dose Radiation Exposure on Brain in a Mouse Model | | |
Rekha Koravadi Narasimhamurthy1, Bola Sadashiva Satish Rao1,2, G. Gireesh3, Kamalesh Dattaram Mumbrekar1
Departments of 1Radiation Biology and Toxicology and 3Cell and Molecular Biology, Manipal School of Life Sciences, Manipal Academy of Higher Education, 2Directorate of Research, Manipal Academy of Higher Education, Manipal, Karnataka, India
E-mail: [email protected]
Introduction: Radiological interventions have remained the most popular medical approach in terms of diagnosis and treatment. However, the early and late neurological consequences of radiation exposure, especially at low doses, on the brain remain ambiguous and controversial, thereby necessitating thorough investigation. Materials and Methods: One month oldC57/BL male mice were irradiated with a whole-body single dose of 0.5 Gy X-ray and assessed for acute and late neurological consequences. After 14 days or 6-month post exposure, animals were subjected to behavioural analysis to assess changes in exploratory behaviour, anxiety, learning and recognition memory. Further the animals were sacrificed and brain tissue was collected for various histological, biochemical and next generation sequencing based transcriptomic analysis. Results: Histological analysis revealed a significant reduction in neuronal cell survival, while increased microgliosis and astrogliosis were also observed as indicated by Iba-1 and GFAP protein expression. Further, early exposure showed immediate and substantial reduction in the enzyme acetylcholinesterase. Golgi-cox staining also revealed a significant decrease in the dendritic spine density along with a decreased trend in the overall neurite path length and branching pattern. Hippocampal transcriptome profile revealed a number of differentially expressed genes that play a role most prominently in immune system, signal transduction, sensory perception, metabolism of proteins, cellular responses to stimuli and cell cycle. However no significant changes were observed in early and late behavioural analysis. Conclusions: Low dose radiation causes significant histological and gene expression and other molecular changes by affecting several signalling cascades within the brain that severely hamper the neuronal homeostasis. Further metabolomic approach could potentially improve our overall understanding of the repercussions of exposure to low dose radiation.
Funding and Acknowledgements: This work was supported by The Science and Engineering Research Board [Grant No: 637 ECR/2017/001239/LS]. The authors would like to thank the Director, Manipal School of Life Sciences and Manipal Academy of Higher Education (MAHE), for support and infrastructure facilities. RKN would like to thank MAHE, Manipal, for the Dr. T.M.A Pai fellowship and KSTePS, Govt of Karnataka, for the scholarship.
| O-1.8 | | |
| Role of Genetic Disease Factor APTX in DNA Double Strand break Repair | | |
Rikiya Imamura1,2, Mizuki Saito1, Mikio Shimada1,3, Masamichi Ishiai2, Yoshihisa Matsumoto1,3
1Department of Transdisciplinary Science and Engineering, School of Environment and Society, Tokyo Institute of Technology, 2National Cancer Center Research Institute, 3Laboratory for Zero-Carbon Energy, Institute of Innovative Research, Tokyo Institute of Technology, Tokyo, Japan
E-mail: [email protected]
Introduction: Aprataxin (APTX) has an enzymatic activity removing adenosine monophosphate (AMP) from DNA-5' end during abortive ligation by DNA ligases. The mutations on APTX cause the genetic diseases such as Ataxia-Oculomotor Apraxia 1 (AOA1). In addition, APTX physically binds to XRCC1 and XRCC4, suggesting its implication in base excision repair (BER) and DNA double-strand break (DSB) repair via non-homologous end joining (NHEJ) pathway. Even though the role of APTX in BER is demonstrated by recent studies, there have been few evidences for its involvement in DSB repair. In this study, we aimed to reveal the role of APTX in DSB repair using the human cells. Materials and Methods: We established APTX knock-out (APTX-/-) cell using CRISPR-Cas9 genome editing system and GFP-tagged APTX (GFP-APTX) stably expressing cell by random integration in human osteosarcoma cell U2OS background. After exposure of the wild type (WT) and APTX-/- U2OS cells to ionizing radiation (IR), survival rate was measured by colony formation assay and DSB markers, i.e., gamma-H2AX and 53BP1 foci, were examined by immunofluorescence. XRCC1 or XRCC4 was depleted by RNA-interference. The recruitment of GFP-APTX to DNA damage was analysed by laser micro-irradiation and live-cell imaging using confocal microscope. Results: Survival rate decreased in APTX-/- cell compared with WT cell after IR treatment. In immunofluorescence, higher number of gamma-H2AX foci were retained in APTX-/- cell than in WT cell, but 53BP1 foci were not discernibly different between APTX-/- cell and WT cell after IR treatment. Although the recruitment of GFP-APTX to DNA damage was suppressed by XRCC1 depletion, but was not appreciably affected by XRCC4 depletion. Conclusions: Our results showed that the interaction between APTX and XRCC4 has at most little contribution to conventional NHEJ and suggested a novel role of APTX in DSB repair, which is distinct from XRCC4-dependent NHEJ.
| O-1.9 | | |
| A Single-Institution Retrospective Analysis of Outcomes for Locally Advanced Rectal Cancer Treated with Neoadjuvant Chemoradiotherapy | | |
Kannan Venkatesan, Vivek Anand, Ranjeet Bajpai, Rohit Santosh Kabre1, Vinay Babu, Ajay Kolse2, Sudesh Deshpande3
Departments of Radiation Oncology and 3P. D. Hinduja National Hospital and Medical Research Center, 2Holy Spirit Hospital, Mumbai, Maharashtra, 1Department of Radiation Oncology, HCG Cancer Center, Vadodara, Gujrat, India
E-mail: [email protected]
Introduction: Neoadjuvant chemoradiotherapy (NACTRT) is an established standard of care for the management of locally advanced rectal cancer (LARC). A retrospective analysis was conducted to assess the results of treatment and predictors of overall survival (OS) for patients treated in a tertiary care hospital in India from 2013 to 2019. Materials and Methods: Biopsy-proven 79 patients of LARC were included in the analysis. Clinical and treatment parameters were assessed. Prognostic factors associated with OS and pathologic response were studied. Treatment toxicities and sphincter preservation rates were documented. Results: There were 55(69.6%) patients with Stage III rectal cancer. Majority of the patients, i.e., 74 (93.7%), received long-course radiation therapy by intensity-modulated radiation therapy–volumetric modulated arc therapy technique after computed tomography + magnetic resonance imaging fusion-based simulation. Pathological complete response (pCR) was achieved in 20 (25.3%) while 45 (57%) showed tumor downstaging post-NACTRT. Time interval (TI) to surgery of ≤8 weeks was significantly associated with higher pCR and tumor downstaging. Actuarial 5-year OS rate was 64.4% and local recurrence-free survival (LRFS) rate of 72.2%. Both OS and LRFS were significantly higher in patients who showed pCR and tumor downstaging. Long-term gastrointestinal and genitourinary toxicities were seen in 11 (15%) and 15 (21.5%) patients, respectively. Conclusions: This study reaffirms the efficacy of preoperative NACTRT as an upfront standard of care in LARC in the Indian population. TI to surgery to the tune of ≤8 weeks yielded optimal pCR and tumor downstaging which translated into significantly better survival.
| O-1.10 | | |
| Repurposing Aspirin to Reverse Radioresistance in Cervical Cancer by Disrupting GADD45α-AURKA Interplay | | |
Salini Das, Dilip Kumar Ray1, Manisha Vernekar2, Sutapa Mukherjee
Departments of Environmental Carcinogenesis and Toxicology, 1Medical Physics and 2Gynecological Oncology, Chittaranjan National Cancer Institute, Kolkata, West Bengal, India
E-mail: [email protected], [email protected]
Introduction: Cervical cancer recurrence is one of the burning issues against the therapeutic success. Development of radioresistance is the predominant reason of disease relapse in cervical cancer. Therefore targeting the molecular interplay that imparts radioresistance is crucial for better treatment results. Cell cycle arrest and DNA damage response are two key strategies of radiation induced killing of cells and thereby altered expression and activities of molecules involved in these pathways lead to development of radioresistance. Radiosensitizers are therefore used to target such mechanisms and restoring sensitivity towards radiation. The aim of the study is to investigate the contributory role of Mitotic Ser-Thr Kinase (AURKA/Aurora Kinase A) and DNA damage sensor protein GADD45α in imparting radioresistance in cervical cancer and targeting this interplay using low dose Aspirin to reverse radioresistance. Materials and Methods: Cervical squamous cell carcinoma cell line SiHa was taken to develop a radioresistant subline SiHa/RR. Molecular interactions between GADD45α and AURKA was examined by western blot, Flow cytometry, semi quantitative RT-PCR, Immunofluorescence, Ectopic overexpression and inhibition analysis. These results were further validated in patient derived cervical biopsy samples for assessing their expressions in patients of advanced cervical carcinoma stages in pre and post radiated conditions. Intervention using 5μM Aspirin in SiHa and SiHa/RR along with patient derived primary cells were performed to target the interaction and restore radiosensitivity. Results: Elevated expression of AURKA and lowered expression of GADD45α was found in SiHa/RR and post radiated patient samples. Ectopic overexpression of AURKA showed radioresistant phenotype in parental SiHa while overexpression of GADD45α showed radiosensitized SiHa/RR. Inhibition of AURKA in SiHa/RR also reflected improved response patterns upon radiation exposure. These results were further justified by treating patient derived primary cells with Aspirin followed by irradiation. Conclusion: Aspirin can sensitize radioresistant cervical cancer cells by targeting the interplay of GADD45α-AURKA.
| O2: Radiation Physics/Chemistry, Food Irradiation, Nuclear Agriculture and Environment | | |
| O-2.1 | | |
| DNA Damage by Proton and Alpha – Geant4-Dna Monte Carlo Study | | |
Arghya Chattaraj1,2, T. Palani Selvam1,2
1Radiological Physics and Advisory Division, Health Safety and Environment Group, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: In radiobiology, radiation induced DNA strand breaks such as Single Strand Breaks (SSB) and Double Strand Breaks (DSB) result in mortality or mutagenesis depending on the damage quality and the cell repair ability. The energy deposited by the ionising radiation crossing the cells is directly or indirectly responsible for the SSB and DSB. The present study is aimed at calculating DSB/SSB ratio for proton and Alpha particles of different energy using Monte Carlo methods. Materials and Methods: In the present study, Geant4-dna track structure model was used to simulate the proton and alpha tracks within a 1 mn the present study, Geant4-dna track structure model was used to simulate the proton and alpha tracks within a 1 SB) result in mortality or mutagenesis depending on the damage quality and the cell repair ability. The energy deposited by the ionising radihm uses the distribution of energy deposits by ionising particles and a damage probability function that depends on the amount of total energy deposits. Protons and alpha particles with energy 0.5 – 10 MeV are considered in this study. Results: For both proton and alpha particles, absorbed dose to water and the number of DSBs decrease with the increase of their energy. However, in the investigated energy range, number of SSBs decrease with energy for protons and the trend is reverse for alpha. When energy changes from 0.5 MeV – 10 MeV, the number of SSBs and DSBs vary from 37.03 – 4.89 and 6.06 – 0.27, respectively for protons and from 40.9 – 60.62 and 45.41 – 15.22, respectively for alpha particles. DSB/SSB ratio is in the range of 0.16 – 0.06 and 1.11 – 0.15 for protons and alpha particles for the above-mentioned energy ranges. Conclusions: The higher DSB/SSB ratio of alpha is indicative of higher cell damage capability of alpha than proton.
| O-2.2 | | |
| Assessment of Transportation of Alpha-Radioactivity from Different Types of Fertilizers to Agriculture Field Soils and Rice Crops in West Bengal State, India | | |
Biswajit Das1,2, Argha Deb2,3
1Department of Physics, Vidyasagar University, Midnapore, 2School of Studies in Environmental Radiation and Archaeological Sciences, Jadavpur University, 3Department of Physics, Jadavpur University, Kolkata, West Bengal, India
E-mail: [email protected]
Introduction: Activity concentrations of radionuclides present in agriculture soils may be enhanced from the extensive use of different types of fertilizers in agriculture fields. The present work deals with the alpha radioactivity measurements in the fertilized soils and different paddy plant's parts grown in ten different agriculture fields located in two areas of Belda and Kalyani in two districts of Paschim Medinipur and Nadia in West Bengal state, India. The plants were grown under actual field condition using different amounts of organic and inorganic fertilizers. Materials and Methods: The alpha radioactivity has been measured by track-etch technique using CR-39 solid-state nuclear track detectors (SSNTDs). The CR-39 SSNTDs has been calibrated experimentally by using a distributed reference source material of IAEA-448 soil. Results: The transfer factor of alpha radioactivity (TFAR) for the agriculture soils of five paddy fields located in Belda has been found to vary from 1.31 to 1.77 for the application of organic and inorganic fertilizers. The TFAR for rice crops cultivated in these fields has been found to vary from to . The TFAR for the agriculture soils of other five agriculture paddy fields located in Kalyani has been found to vary from 1.26 to 1.80 for the application of inorganic fertilizers. The TFAR for rice crops cultivated in these fields has been found to vary from to . Conclusions: The alpha radioactivity of the fertilized soils and rice crops is higher in case of paddy plants grown with the use of phosphorus based inorganic fertilizers than with the use of organic fertilizers and non-phosphorus based inorganic fertilizers. The TFAR for agriculture soils to plants varies also with the species of plants. Novelty of the present study relies on the measured alpha radioactivity values and the evaluated CF values for CR-39 SSNTDs.
| O-2.3 | | |
| Gamma Radiation Mediated Fabrication of An Amidoxime Based Biodegradable Cellulosic Adsorbent for Effective Removal of Uranium from Ground Water | | |
Nilanjal Misra1, S. Shelkar1, N. K. Goel1, S. Rawat1,2, M. Anitha3, Virendra Kumar1,2
1Radiation Technology Development Division, BARC, 2Homi Bhabha National Institute, 3Materials Processing and Corrosion Engineering Division, BARC, Mumbai, Maharashtra, India
E-mail: [email protected]
Contamination of groundwater by heavy metals, such as As, Pb, Th, U etc. is a cause of serious concern for government agencies worldwide. U, in particular, besides being radioactive, has equally damaging chemical effects with studies revealing it to be a primary cause for Nephritis (kidney damage). To address this issue, a novel biodegradable cellulosic adsorbent has been fabricated through valorization of cotton cellulose fabric to selectively remove U from ground water. 60Co gamma radiation mediated simultaneous irradiation grafting process was employed to functionalize cotton cellulose fabric with Polyacrylonitrile, followed by chemical treatment to convert the nitrile groups to amidoxime groups, well known for selective removal of uranium. Grafting parameters were optimized to arrive at a grafting yield of 30%. Samples were characterized by SEM, FTIR, DSC and TGA analysis techniques to confirm the introduction of amidoxime functional groups onto cellulose surface. Adsorption studies carried out under batch and column processes using aqueous solutions of U ion yielded good uptake profile with the uptake capacity peaking at ~60 mg/g adsorbent, determined via ICP-OES analysis. The adsorbent viability is further enhanced though regeneration achieved using an optimized eluent system that ensured reusability for multiple cycles with minimal attrition losses. The developed biodegradable cellulose-based adsorbent system, therefore, has potential applications as an effective U remediation strategy to provide safe drinking water in areas afflicted by U contamination.
| O-2.4 | | |
| Varietal Improvement of Chrysanthemum through Induced Mutagenesis | | |
Ramesh K. Satdive, Sudhir Singh, Shraddha Singh, Himanshu Mishra1, Jagadeesh Chitti1, Suresh N. Manvatkar1, Chandrakant Salunkhe1
Plant Biotechnology and Secondary Metabolites Section, Nuclear Agriculture and Biotechnology Division, 1Landscape and Cosmetics Maintenance Section, A and SE Division, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Genetic variation is necessary for any plant breeding program for crop improvement. The rate of spontaneous mutations that occur naturally due to various radiations, cosmic rays, and several radioactive elements is low. Such mutations can also be induced artificially by several physical agents like gamma-rays and X-rays, as well as by chemical agents. For modern and industrialized floriculture, there is always demand and necessity for new ornamental crop varieties. The possibilities for creating different forms and improving ornamentals are infinite, and induced mutagenesis can assist us in improving a crop through mutant induction. The objective of the present study was to create variation in Chrysanthemum by gamma radiation and improve traits such as flower colour, morphology, type, plant architecture, variegated leaves, and disease resistance. Chrysanthemum (locally known as 'Shrevanti' or 'Guldaudi') is one of the most popular flowers in India. It is used both as a commercial and popular exhibition/cut flower crop. It has earned tremendous popularity in the floriculture industry due its wide range of flower colour, form, and excellent keeping quality. Materials and Methods: We selected eight popular varieties of Chrysanthemum for the studies, including Purnima, Pusa Aditya, Shweta, Thaichin, Sharadhar etc. For irradiation dose (GR50) optimization, rooted cuttings of the selected varieties were exposed to the different doses (0-100 Gy) of gamma radiation. Further, two hundred rooted cuttings of the selected Chrysanthemum varieties were treated with the optimized gamma-irradiation dose (10 and 20 Gy along with control) and were planted on raised beds. The various parameters, such as height, colour variation, number of flowers/branches etc were recorded at regular intervals. Results: The optimized irradiation doses for the selected Chrysanthemum varieties 10-20 Gy. We screened twelve promising stable mutants in the selected varieties. Conclusion: We could develop twelve stable mutants in Chrysanthemum, indicating that induced mutagenesis can be an important tool for creating novel genotypes in vegetatively propagated ornamental crops.
| O-2.5 | | |
| Genetic Improvement in Plant Architecture, Maturity Duration and Agronomic Traits of Three Traditional Rice Landraces Through Gamma Ray Based Induced Mutagenesis | | |
Richa Sao,Parmeshwar K. Sahu, Suvendu Mondal1, B. K. Das1, Deepak Sharma
Department of Genetics and Plant Breeding, Indira Gandhi Krishi Vishwavidyalaya, Raipur, Chhattisgarh, 1Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Rice landraces are the important source of several valuable genes for crop improvement except with few lacunae viz., poor yield potential, tall plant stature, late maturity duration which restrict their commercial cultivation. Radiation induced mutation breeding offers simple, fast and efficient way to rectify these major defects without altering their original identity. Materials and Methods: The present study deployed the radiation (gamma rays @ 300Gy) induced mutation breeding for improvement and revival of three traditional rice landraces viz., Samundchini, Vishnubhog and Jhilli. Desirable mutants identified were evaluated based on 13 agro-morphological and 16 grain quality traits in M4, M5 and M6 generations along with their parents and three checks. Further SSR markers based genomic similarity and genetic diversity study was also preformed. Results: All the mutants of the three landraces showed reduction in maturity duration and plant height compared with their parents. Furthermore, two, six and three mutants of Samundchini, Vishnubhog and Jhilli have increased yield potential over their parents, respectively. Interestingly, Samundchini Mutant-18-1 (22.45%), Vishnubhog Mutant-74-6 (36.87%) and Jhilli Mutant-13-5 (25.96%) showed highest yield advantage over their parents. Further, ANOVA based on RCBD revealed ample variations among the genotypes for studied traits. In addition, all the traits consistently showed high to moderate PCV and GCV and slightly difference between them in all the three generations. Moreover, in association analysis, the traits viz. fertile spikelets/panicle, panicle length, total tillers/plant, spikelet fertility percent and 100 seed weight with grain yield/plant whereas the traits, hulling (%) and milling (%) with HRR (%) were consistently showed high direct effects and significant positive correlation. SSR marker based genome similarity in rice mutants and corresponding parents ranged from 95.60% to 71.70% (Vishnubhog); 95.62% to 89.10% (Samundchini) and 95.62% to 80.40% (Jhilli), indicating the trueness of the mutants. Moreover, UPGMA algorithm and Gower distance based dendogram; neighbor joining tree and PCA scatter diagram assured that mutants were grouped with their respective parents and fell into separate cluster showing high similarity between mutants and parents and dissimilarity among the 24 genotypes. Conclusions: Results revealed that irradiation could generate a considerable amount of genetic variability, and provide new avenues for crop improvement and diversification.
| O-2.6 | | |
| Antimutagenic and Anti-proliferative Activities of Ethoxy-Substituted Phylloquinone (Vitamin K1 Derivative) from Spinach and Effect of Radiation Processing | | |
Sanjeev Kumar, Jyoti Tripathi, Dharmendra K. Maurya1, Satyendra Gautam
Food Technology Division, Bhabha Atomic Research Centre, 1Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Genotoxic agents present in the environment often pose a threat to the genomic stability resulting in various life threatening diseases including neoplastic induction. In the current study, a novel antimutagenicity compound was identified from spinach and characterized as 2-ethoxy-3-(3,7,11,15-tetramethylhexadec-2-ethyl) naphthaquinone -1,4-dione (ethoxy-substituted phylloquinone; ESP). ESP was further analyzed for anti-proliferative activity and underlying mechanism. Materials and Methods: Antimutagenicity assay was performed using rifampicin resistance assay in Escherichia coli MG1655 and thymidine kinase gene mutation assay in human lymphoblast (TK+/-) cell line. Lung cancer (A549) cell line was used for antiproliferative activity evaluation. For molecular elucidation of underlying mechanism, proteomics analysis such as 2-D gel electrophoresis and SWATH-MS analysis was performed. Also, antimutagenicity in single gene knock-out strains of E. coli was performed and findings was validate in TK+/- using RT-PCR. Results: ESP, a novel compound from spinach was found to have high antimutagenicity and antiproliferative activity. DAPI competitive binding assay and molecular docking showed H-bonding and hydrophobic interactions between ESP and B-DNA dodecamer residues at minor groove. In E. coli genes such as tnaA (tryptophanase) and dgcP (diguanylate cyclase) were upregulated in presence of ESP which have been reported to be involved in indole and cyclic-di-GMP biosynthesis, respectively, and cell division retardation. Thus, providing additional time for DNA repair leading to antimutagenicity. In case of TK+/- cell line system also, ADCY genes (adenylate cyclase), a functional analogue of dgcP gene, were found to be up-regulated. Antiproliferative effect in lung cancer (A549) cells was possibly due to down-regulation of proteins involved in EGFR signaling and NEDDyylation and up-regulation of certain tumor suppressor (STAT1 and NDRG1) proteins. Treatment of spinach powder with gamma radiation (5-20 kGy) from cobalt (Co-60) enhanced the extractability of ESP up to 4.4-fold at the highest dose of 20 kGy while maintaining its bioactivities. Conclusions: Current findings thus indicate that ESP could be a novel therapeutic agent to combat mutagenic assault as well as proliferation of cancerous cells.
| O-2.7 | | |
| Micropropagation and Determining Gamma Ray Dosage Sensitivity in Red Banana Variety (Musa acuminata) | | |
S. V. Sawardekar, S. H. Sherkar, R. S. Deshpande
Plant Biotechnology Centre, Dr. Balasaheb Sawant Konkan Krishi Vidyapeeth, Dapoli, Maharashtra, India
E-mail: [email protected]
Red banana, also known as lal-velchi, is one of the favourite banana cultivar of Konkan region in Maharashtra. Induced mutation technique is particularly important for bananas as there is limited sexual reproduction and parthenocarpic fruits. First and most important step in mutagenic treatment is to estimation of the most appropriate mutagenic treatment. In this study, in vitro shoots of red banana were treated with gamma radiation (60Cobalt), utilized as a mutagenic agent. The objective was to determine the gamma ray lethal dose (LD) in red banana variety. The axenic cultures of passage 3-4 were irradiated with doses of 10, 20, 30, 40, 50, 60, 70, 80, 90 and 100 Gray (Gy). The in vitro cultures were evaluated for per cent survival, multiplication rate, length of shoots, number of leaves and length of leaves. Based on the values, the lethaldose 50% (LD50) was determined as 20 Gy. Lower doses (10 Gy and 20 Gy) enhanced shoot multiplication ratio and no multiplication was seen in shoots irradiated with 30 Gy and above. The survival rate of individual shoot declined with increase in irradiation dose indicating an inverse relationship between the two parameters. The results revealed that a significant reduction in the survival percentage, shoot length and number of shoots with increasing levels of gamma irradiation doses. After the increasing levels of gamma irradiation doses more than 30 Gy, all irradiated shoots turned brown and dead. Also during in vitro propagation of irradiated shoots (LD50 = @20 Gy), medium supplemented with 6 mg/l 6-benzylaminopurine (BAP) + 20 mg Casein hydrolysate + 30 mg/l Adenine sulphate showed maximum rate of multiplication rate (1:1.8). Shoots were multiplied and rooted on different rooting medium. Maximum rooting responses observed in medium supplemented with 1.5mg/l indole-3-butaric acid (IBA) + 0.5mg/l Naphthalene acetic acid (NAA). Hence, this study presents the in vitro propagation of gamma irradiated cultures and its usefulness for the selection of desirable mutants.
| O-2.8 | | |
| Effect of FtsK Dependent Site-Specific Recombination on the Radioresistance of Deinococcus Radiodurans | | |
Shruti Mishra1,2, Hari S. Misra1,2, Swathi Kota1,2
1Molecular Biology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail:[email protected]
Deinococcus radiodurans, an extremely radioresistant coccus bacterium, withstands around 15 kGy dose of radiation, i.e. about 3000 times the radiation tolerance of humans. Its highly efficient DNA damage repair mechanism and unique arrangement of the genome are some of the factors responsible for this extraordinary radioresistant phenotype. This bacterium contains a polyploid multipartite genome, which is tightly packaged in the form of doughnut shaped toroidal structure. How this unique genome organization is maintained is still unknown. In other bacterial systems, chromosome dimers are formed after replication due to homologous recombination between the sister chromosomes. In D. radiodurans, the polyploidy increases the chances of the formation of chromosome dimers. So, how these dimers are resolved and whether that has any significance to radiation resistance in this bacterium are the most intriguing questions. The chromosome dimer resolution system involves the function of site-specific tyrosine recombinases (Xer) that could function independently or by the activation from the FtsK protein. Bioinformatics analysis has revealed that D. radiodurans encodes FtsK and six putative tyrosine recombinases indicating the existence of a functional site-specific recombination (SSR) system. We have earlier demonstrated the functional activity of drFtsK in-vitro. We created different domain disruption mutants of ftsK and these mutants showed many abnormalities at the nucleoid as well as the cellular levels. Mutants also exhibited increased generation time than the wild type in normal condition and post gamma irradiation. Trans-expression of the active tyrosine recombinase system (XerCDs) from E. coli in D. radiodurans resulted in a drastic change in nucleoid compactness and arrangement. This suggests the activation of SSR system of E. coli by drFtsK. In-silico data revealed that only one putative Xer (dr_0513) showed interaction with the γ-domain of drFtsK. Our findings highlighting the role of SSR in deinococcalnucleoid structure and radioresistance would be presented.
| O-2.9 | | |
| Radiation Dosimetry Assessment of 177Lu-CXCR4 Antagonist for Effective Dose Calculation using OLINDA/EXM 2.0 Computed Software | | |
Tamanna Lakhanpal, B. R. Mittal, Jaya Shukla, Yogesh Rathore, Rajender Kumar, Komalpreet Kaur, Pankaj Malhotra1, Gaurav Prakash1, Alka Khadwal1, Charanpreet Singh1, Munish Kumar
Departments of Nuclear Medicine and PET Centre and 1Clinical Haematology and Medical Oncology, Post Graduate Institute of Medical Education and Research, Chandigarh, India
E-mail: [email protected], [email protected]
Introduction: Radiation dosimetry refers to the quantitative radiation absorbed dose estimation for internal emitters (beta or gamma emitters). For radionuclide therapies, it plays a pivotal role in the estimation of the critical organ, residence time and specific organ absorbed dose calculation. In addition, it has provided patient stratification towards the best treatment by avoiding any undue radiation exposure and preventing organ toxicities. In this context, radiation dosimetry was performed using OILNDA/EXM 2.0 computed software in patients with lymphoma using a radiolabelled 177Lu-CXCR4 antagonist. Materials and Methods: Optimization and standardization and of radiolabelling parameters such as reaction volume, time, pH and temperature were done for 177Lu-CXCR4 antagonist. Quality control checks were performed such as radionuclide purity, radiochemical purity, sterility, serum stability, pyrogenicity, etc. Standardization of the dosimetry procedure was carried out for patients with lymphoma for calculation of critical organ and residence time in various organs using Organ Level INternal Dose Assessment (OLINDA/EXM 2.0) computed software. Results: No radionuclide and radiochemical impurities were found in the in-house synthesized 177Lu-CXCR4 antagonist. In addition, the radiopharmaceutical preparation was sterile and pyrogen free. Stability in PBS buffer and human serum was inferred up to 21 days. The mean residence time of radiolabelled 177Lu-CXCR4 antagonist in the different organs of interest were 9.9 ± 0.03 h in kidneys, 1.09 ± 0.01 h in the liver, and 0.05 ± 0.006 h in spleen tissues. The OLINDA/EXM 2.0 effective dose calculated was maximum for kidneys (8.78 mSv/ MBq) indicating kidneys are the critical organ. Conclusion: Kidneys are the critical organ for177Lu-CXCR4 antagonist determined using OILNDA/EXM 2.0 computed software.
| O-2.10 | | |
| Assessment of Natural Radiation Levels Due to 222Rn, 220Rn and Progeny in Indoor Environment of Outer Himalayan Region, India | | |
Taufiq Ahamad, O. P. Nautiyal1, Manish Joshi2, A. S. Rana, Prakhar Singh3, A. A. Bourai4
Department of Physics, Shri Guru Ram Rai PG College, 1Uttarakhand Science Education and Research Centre, Dehradun, 3Department of Physics, Gurukul Kangri University, Haridwar, 4Department of Physics, H.N.B. Garhwal University, Tehri Garhwal, Uttarakhand, 2Radiological Physics and Advisory Division, Bhabha Atomic Research Center, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: The study of measuring natural radiation levels resulting from natural radioactivity due to the concentration of radon (222Rn), thoron (220Rn) and progeny is important as they are the significant contributor to background radiation dose to inhabitants. The time integrated passive measurements were carried out in indoor environment of dwellings in the Champawat District of outer Kumaun Himalayan belt, Uttarakhand, India with the aim of investigating significant health risk to the inhabitants in the study area. Materials and Methods: The measurements were performed using single entry pinhole dosimeters and progeny sensors (DTPS/DRPS) with the recently developed LR-115 detector-based solid state nuclear track detectors to measure indoor levels of 222Rn, 220Rn and their progeny concentrations, respectively. Further, the corresponding indoor gamma level was also measured in respective dwellings to assess indoor gamma dose rate. Results: The estimated value of annual indoor 222Rn concentration lies in the ranges from 13.40 to 210.62 Bq/m3 whereas thoron concentration ranges from 19.72 to 179.0 Bq/m3 with the annual average of 85.0± 43.07 Bq/m3 and 84.46 ± 36.89 Bq/m3 respectively. The observed value of gamma dose rate in indoors vary from 0.08 μSv/h to 0.32 μSv/h with an average of 0.18 ± 0.06 μSv/h. Conclusions: The annual average value of indoor radon was found to be greater than both the worldwide (40 Bq/m3) and national (42 Bq/m3) averages, but indoor radon concentration falls within reference limit of 200-300 Bq/m3 and in the same way, the annual average value of indoor thoron was found to be much higher than the worldwide and national averages of 10 Bq/m3 and 12.2 Bq/m3, respectively. The obtained results strengthen the current database as no study has been performed earlier in the present study area and the results so obtained are consistent with recent research done in the Indian Himalayan areas.
Keywords: Gamma level, LR-115 detector, Natural radiation, Pinhole Dosimeter, Progeny Sensors, Radon/Thoron and progeny
| Poster Presentation (P) | | |
| Topic 1 DNA Damage Response and Repair | | |
| P-1.1 | | |
| A Novel Dual Inhibitor of Topoisomerase 1 and PARP1 for Efficient Cancer Sensitization | | |
Ananda Guha Majumdar1,2, Papiya Dey1,2, Shikha Shree1,2, Birija Sankar Patro1,2
1Bio-Organic Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected], [email protected]
Topoisomerase 1 (TOP1) is an essential enzyme in humans which mediates relaxation of DNA supercoils at the interface of critical processes like DNA replication and transcription. TOP1 performs DNA relaxation through a cleavage-rotation-religation cycle, involving a transient TOP1-DNA covalent complex (TOP1cc) as an obligate intermediate with a nanosecond-scale half-life. Clinically approved TOP1 inhibitors act through the stabilization of these TOP1ccs, thus converting them into long-lived, stable TOP1ccs, which act as sources of toxic double stranded DNA breaks. Poly-ADP-Ribose Polymerase 1 (PARP1) plays a key role in inducing resistance to TOP1 inhibitors through multiple mechanisms: removal/repair of TOP1ccs through promoting endonucleolytic/proteosomal degradation of TOP1cc, replication fork reversal and nucleoplasmic sequestration of TOP1. Consequently, combinatorial inhibition of TOP1 and PARP1 is an emerging concept and certain combinations are under clinical trial for augmenting cancer therapy. However, combinatorial therapies are often limited by toxicity and incompatible pharmacokinetics and pharmacodynamics of individual drugs. It is imperative to develop a novel single molecule which can impart dual inhibition of TOP1 and PARP1. In this regard, we designed and synthesized a series of 11 compounds combining critical part pharmacophores of naphthalimides (widely known investigational Topoisomerase inhibitors) and Olaparib (clinically approved PARP inhibitor), and evaluated them for their cytotoxic effects. The initial cytotoxicity screen revealed a candidate molecule (PAR-3), which showed significantly higher cytotoxic potential compared to Olaparib. In vitro assays revealed that PAR-3 inhibits TOP1 catalytic activity, coupled with a modest inhibition of PARP1 activity. PAR-3 was shown to stabilize TOP1ccs inside cells, resulting in accumulation of DNA damage. PAR-3-induced stabilization of TOP1ccs resulted in significantly lower induction of total cellular PARylation, as compared to camptothecin, a classical TOP1 inhibitor. Taken together, our work presents a proof-of-concept for an effective dual inhibitor of TOP1 and PARP1, which may act as a foundation for development of more efficient molecules in future.
| P-1.2 | | |
| BRD4 Degradation Sensitizes Breast Cancer Cells to Gamma Radiation by Attenuating DNA Damage Response | | |
Anusree Dey1,2, S. Uppal1,2
1Molecular Biology Division, Bhabha Atomic Research Centre, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Bromodomain Protein 4 (BRD4) is a member of the Bromodomain and Extra terminal domain (BET) family of proteins containing dual bromodomains which bind to acetylated histones in chromatin. BRD4 performs diverse biological functions including regulation of transcription and alternative splicing, epigenetic remodeling, stem cell maintenance, etc. BRD4 overexpression has been linked with several types of cancers. Being a druggable target, BRD4 is an attractive candidate for exploiting its potential in cancer therapeutics. Breast cancer is a highly complex and aggressive disease. Though radiotherapy is one of the primary treatment modules, high incidences of radio-resistance affect its therapeutic potential. Our present work shows that degradation of BRD4 using small molecule degrader sensitizes breast cancer cells to gamma radiation, as determined by clonogenic assay. Notably, BRD4 depletion leads to accumulation of more DNA double strand breaks in gamma radiation exposed breast cancer cells, as observed using comet assay and gamma H2AX foci immunostaining. Moreover, this increased DNA damage levels seems to be contributed by both enhanced DNA damage as well as reduced DNA repair in irradiated BRD4 depleted cells. Further, we also found that depletion of BRD4 leads to inhibition of DNA replication suggesting a critical role for BRD4 in DNA replication. In line with this, we have shown that BRD4 interacts with RPA2 and PCNA, two key proteins involved in the DNA replication as well as DNA damage repair, using in-situ Proximity Ligation Assay (PLA). More importantly, based on the whole genome transcriptome analysis, we found that BRD4 regulates transcription and splicing patterns of key DNA damage response genes. Our results show that BRD4 depletion leads to radio-sensitization of breast cancer cells, which may be attributed to accumulation of DNA damage and DNA replication arrest in the cells. Taken together, our results suggest a role for BRD4 in radiation induced DNA damage response in breast cancer cells.
| P-1.3 | | |
| GCN5 as a Master Regulator of DNA Double Strand Break Repair Mediates GBM Radio Resistance | | |
Debashmita Sarkar1,2, Shilpee Dutt1,2
1Shilpee Dutt laboratory, Tata Memorial Centre, Advanced Centre for Treatment, Research and Education in Cancer, Navi Mumbai, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Modulation of the DNA damage response pathways in Glioblastoma (GBM) by novel mechanisms distinct from genotoxic responses in normal cells pose challenge to current treatment modalities. Here we interrogated the mechanism of Double Strand Break (DSB) repair regulation by GCN5, a histone acetyl transferase for understanding the differential repair response of GBM to radiotherapy. Materials and Methods: DSBs in glioblastoma cells were induced by X-rays using linear accelerator. Functional studies were done post pharmacological inhibition using Butyrolactone-3 and siRNA, shRNA mediated knockdown of GCN5 by immunoblotting, real time qPCR, immunofluorescence, immunoprecipitation. DSB repair pathways were studied by comet assay and HR-NHEJ reporter assay. Results: We found that post irradiation in U87MG and LN229 cells, there was 30-50 fold upregulation of GCN5 in transcript level along with its rapid recruitment to DSBs via PARP1 mediated parylation. Interestingly, GCN5 depletion when combined with IR radio sensitized GBM cells. This was due to persistent DNA damage as observed by neutral comet assay and unresolved Ɣ-H2AX foci as observed by confocal microscopy upon GCN5 knockdown. Moreover, genetic perturbation of GCN5led to significant decrease in expression, activation and DSB recruitment of DNAPK and ATM, the key kinases of NHEJ and HR repair pathways respectively. Additionally, mass spectrometric analysis, immunoprecipitation and colocalization studies revealed physical interaction of GCN5 with ATM and DNAPK. Finally, we demonstrated that in irradiated cells, both HR and NHEJ repair efficiencies as assessed by HR-NHEJ in vivo reporter assays were significantly reduced upon GCN5 knockdown. Conclusion: Taken together, we identified GCN5 as an atypical protein that is recruited to double strand breaks via PARP1.GCN5 augment survival of GBM cells under genotoxic stress by regulating both HR and NHEJ repair via controlling ATM and DNAPK signaling respectively. Therefore, this study identifies GCN5 as a novel molecular target in GBM therapeutics.
| P-1.4 | | |
| Inhibition of RECQL5 Function Evokes NHEJ Response and Sensitivity of HR-Proficient Breast Cancers to PARP Inhibitor Treatment | | |
Krupa Thankam Philip1,2, Kartik Dutta1,2 Usha Yadav2,3, Saikat Chakraborty1,2, Birija Sankar Patro1,2
1Bio-Organic Division, 3Radiation Physics and Advisory Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected], [email protected], [email protected]
PARP inhibitors (PARPi) are clinically approved for the treatment of BRCA-mutated hereditary breast and ovarian cancers with homologous recombination (HR) deficiency based on synthetic lethality concept. PARP inhibition leads to replication associated DNA damages that are repaired by HR-dependent mechanisms. If not repaired, it ultimately leads to accumulation of double strand breaks (DSBs) and cell death. However, ~90% breast cancers are BRCA wildtype (HR-proficient) and hence show de novo resistance to PARPi. Hence it is imperative to explore newer targets in HR-proficient hereditary breast and ovarian cancers to generate synthetic lethality in PARPi exposure, which may create a major new treatment opportunity. RECQL5 physically interacts and disrupts RAD51 from pre-synaptic filaments, aiding HR resolution, replication fork protection and preventing illegitimate recombination. In the current investigation, we show that targeted inhibition of HR by stabilization of RAD51-RECQL5 complex by a pharmacological inhibitor of RECQL5 (4a) in the presence of PARPi [talazoparib (BMN673)] leads to abolition of functional HR with uncontrolled activation of NHEJ pathway. Concomitant treatment with PARPi and 4a generates copious amounts of replication stress, prolonged cell cycle arrest, extensive DSB formation, changes in nuclear morphology, chromosomal aberrations and sensitization of HR-proficient breast cancers. Imperatively, 4a was ineffective against normal mammary epithelial cells, which expresses low RECQL5 vis-à -vis breast cancer cells. Moreover, functional inhibition of RECQL5 inhibits metastatic potential of breast cancer cells in response to PARPi. Together, our results demonstrate that RECQL5 in HR-proficient cancers can be targeted by 4a, and this strategy may be useful in expanding the treatment horizon of PARP inhibitors.
| P-1.5 | | |
| Investigation of Double Strand DNA Break Repair of Mycobacteria Using CRISPR/Cas9 | | |
Megha Sodani1,2, Chitra S. Misra3, Devashish Rath2,3, Savita Kulkarni1,2
1Radiation Medicine Centre, Bhabha Atomic Research Centre, Tata Memorial Centre Annex, 2Homi Bhabha National Institute, 3Applied Genomics Section, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: CRISPR/Cas9 can function as anti-microbial by introducing specific double strand break (DSB) in the genome. DSBs in the genome are fatal unless repaired by systems such as non-homologous end joining (NHEJ) or homology directed repair (HDR). Mycobacterium is one of the few bacteria which possess NHEJ in addition to HDR. We investigated the ability of Mycobacterium smegmatis to repair the breaks caused by Cas9. Materials and Methods: Cas9 and a mutated derivative, Cas9n (Cas9-nickase) were codon-optimized, cloned and inserted into the M.smegmatis genome using an integrative plasmid. sgRNA cassettes engineered to target different loci (recA, ilvH, argF, and an intergenic region on the chromosome and gfp on multi plasmid copy, pMN437) were constructed and cloned in a plasmid and transformed in the above hosts. The survival outcomes were evaluated. Result: Expression of Cas9 along with cognate sgRNA resulted in high killing efficiency with a 1000-fold decrease in viable bacterial counts. The percentage of decrease in survival was independent of the locus targeted. However, in case of Cas9n no decrease in bacterial counts as compared to the untargeted control was observed. This implies that while M. smegmatis repairs single strand breaks efficiently, it has poor ability to repair DSBs caused by Cas9. To leverage this shortcoming, CRISPR system was used to target drug resistance in this microbe. In a mixed population of antibiotic-resistant and sensitive mycobacterial cells, selectively targeting a plasmid-borne factor, pMN437, that confers hygromycin resistance, turned the entire population sensitive to the drug. Conclusion: Survival studies after introduction of ds DNA breaks using CRISPR-Cas9 system indicated poor DBS DNA repair and no evidence for participation of NHEJ machineryin Mycobacterium. This makes the CRISPR-Cas9 system an effective anti-mycobacterial agent that could also target drug resistance in this pathogenic bacterium.
| P-1.6 | | |
| Transcript Profile of Fanconi Anemia (FA) DNA Repair Pathway Genes with Chronic and Acute Low Dose Ionizing Radiation in Human Peripheral Blood Mononuclear Cells | | |
S. Nishad1,2,Anu Ghosh1,2
1Animal House Facility and Radiation Signalling Section, Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Low dose ionizing radiation (IR) exposure induces a complex network of cellular and molecular responses which are not fully understood. Our previously generated proteome profiles of human samples from high level natural background radiation areas (HLNRA) of Kerala, India showed over-expression of FA pathway. This pathway coordinates repair of DNA strand breaks through proteins of all the major DNA repair pathways. The present study analysed transcript profiles of FA proteins to understand their role in radiation stress. Materials and Methods: The blood samples (n=15) collected from HLNRA were stratified into Group II(1.5–5.0 mGy/y) and Group III (5.01–14.0 mGy/y) based on the annual dose received by the individuals. The samples from normal level natural radiation areas (NLNRA) were classified as Group I (≤1.5 mGy/y). The samples (n=8) for acute in vitro irradiation studies were collected from Mumbai, India. The SYBR GREEN chemistry based RT-qPCR was used to study mRNA expression. Results: Transcript profiles of FA proteins showed up-regulation with chronic IR in both Group II and III HLNRA individuals, as compared to NLNRA. FA genes BLM, FANCA, FANCI, FANCM, FANCQ and MLH1 showed up-regulation (≥ 1.4-fold) in Group II individuals. The Group III individuals showed similar patterns of expression for all the genes, except FANCQ. Principal component analysis (PCA) explained ~97% of the data variance and could distinctly differentiate the groups according to annual radiation dose. Two dose points (300 mGy and 1 Gy) were selected for acute in vitro studies and expression levels were analysed at 5 min post irradiation. At high dose of 1 Gy, the samples showed up-regulation for BLM, FANCA, FANCD1, FANCD2, FANCF, FANCG, FANCM, FANCT, FANCP, FANCN, FANCB and MLH1. At low dose of 300 mGy, the average expression pattern showed up-regulation for all the above genes, except BLM, FANCA, FANCM, FANCT and FANCN. PCA clearly clustered (data variance ~62%) individuals based on radiation dose. Conclusions: The up-regulation of FA core pathway genes with chronic/acute low dose IR indicates its active pro-survival role in radiation response.
| P-1.7 | | |
| Elucidating the Effect of Caffeine on NHEJ Repair Pathway | | |
Susmita Kumari, Sathees C. Raghavan
Department of Biochemistry, Indian Institute of Science, Bengaluru, Karnataka, India
E-mail: [email protected]
Genome stability and maintenance of its integrity is of utmost importance for the survival of any organism. Human genome is under constant threat by several endogenous and exogenous factors. In order to maintain genomic stability, cells have evolved different DNA repair pathways. Caffeine, an analogue of adenosine, is one of the commonly consumed stimulants across the world. It has been reported that caffeine inhibits the protein kinase activity of ATM and ATR (PI3KK family) although certain other studies show that caffeine failed to affect ATM autophosphorylation, which is a crucial step in ATM activation. Further, Caffeine treatment disrupted the formation of Rad51 filaments and thereby inhibiting the repair by HR. Thus, although certain studies suggest that Caffeine could play a role in DNA repair, its effect on NHEJ and MMEJ are poorly understood. It is also unclear, whether such an effect could contribute towards oncogenesis/cancer therapy. Using an array of biophysical and biochemical methods, we show that incubation with caffeine lead to significant reduction in DNA end joining when oligomeric DNA substrate harbouring DSBs were incubated with cell-free extract prepared from rat testes. The observed inhibition was consistent when joining was performed using sticky, blunt and non-compatible ends. Further, its impact on MMEJ is being evaluated. Using immunofluorescence assay, we show that there is a dose dependent increase in DNA breaks and cell death in mammalian cells. Studies using mice model showed that upon caffeine treatment (50 mg/kg body weight), there was a significant regression of tumor development as compared to the control. In summary, it appears that intake of Caffeine at certain concentration may lead to inhibition of DNA repair leading to accumulation of DNA breaks in cancer cells resulting in tumor regression.
| P-1.8 | | |
| A Novel Role of RECQL5, a DNA Repair Protein in Mitochondrial Dynamics Associated Radio-Resistance in Cancer | | |
Saikat Chakraborty1,2, Shikha Shree1,2, Birija Sankar Patro1,2
1Bio-Organic Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected], [email protected]
Clinically radiotherapy is successfully used for the treatment of different types of cancer. However, the clinical outcomes are refractory due to de novo and acquired radio-resistance in cancers. It is essential to understand the molecular mechanism(s) of radio-resistance of cancer for improving the clinical outcomes of radiotherapy. The most important target of ionizing radiation is DNA and repair of IR-induced DNA damages are extensively reported to be associated with radio-resistance of cancer. In recent past, an emerging role of mitochondria and metabolism is advocated in radio-resistance of cancer. However, the precise mechanism of regulation of mitochondrial dynamics and its role in radio-resistance is not yet known. RECQL helicase family proteins (RECQL1, BLM, WRN, RECQL4 and RECQL5) play crucial role in various DNA repair process (BER, NER, NHEJ and HRR). We and others have reported that RECQL1, BLM and WRN deficient cells show marginal or no sensitivity towards IR treatment. Interestingly, we observed a novel phenomenon that RECQL5 deficient cancers are radio-resistant in nature. Although, homologous recombination repair (HRR) is hyper-activated in RECQL5-deficient cancer cells, the kinetics of repair of IR-induced DSBs are found to be grossly unaffected. Interestingly, we found that residual unrepaired DSBs are almost similar in RECQL5-KO cancers cells and RECQL5-proficient cancers, suggesting differential nuclear DNA repair might not account for radio-resistance of RECQL5-deficient cancers. Intriguingly, RECQL5 deficiency led to significant changes in mitochondrial dynamics and stress induced mitochondrial hyperfusion phenotype (SIMH). A detailed role of RECQL5 in SIMH and mitophagy in radio-sensitivity will be presented in the conference.
| P-1.9 | | |
| Replication Dysfunction in Conjunction with PARP Inhibition Leads to Contextual Lethality in Cancers | | |
B. Ganesh Pai1,2, Kshama Kundu1,2, Papiya Dey1,2, Birija Sankar Patro1,2
1Bio-Organic Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected], [email protected], [email protected]
Pharmacological inhibitors of Poly (ADP-ribose) polymerase (PARPi) are effective against homologous repair deficient cancers, based on the synthetic lethality concept. This mode of action was limited to its role in DNA repair. Recent reports have unravelled that PARPi resistance is replication-associated and that replication-dysfunction is a major vulnerability of cancers to PARPi-therapy. Therefore, effective combinatorial approaches using inducers of replication-distress to target PARPi-resistant tumours are warranted. In this regard, we screened a library of stilbenes, known to induce replication-dysfunction, for RAD51-downregulation and as a consequence, homologous recombination (HR). From the screen, 4,4'-dihydroxystilbene (DHS) emerged as a potent molecule for RAD51 down regulation. DHS induced extensive synergistic cell death in ovarian cancer (OC) cells when combined with talazoparib, a third-generation PARPi. Mechanistically, we observed extensive DNA-DSB induction under talazoparib plus DHS treatment. An unbiased whole transcriptome (RNA-Seq) analysis of SK-OV-3 cells showed that DHS downregulated HR factors, presumably leading to induction of the observed synergy in cell death. OC cells challenged with talazoparib were found to rely heavily on RAD51-dependent HR or alternate mechanisms, while DHS significantly attenuated the talazoparib induced RAD51 foci formation and DR-GFP based HR repair. Interestingly, flow cytometry, click-IT and immunofluorescence microscopy-based analyses revealed that the DNA damage induced by talazoparib plus DHS were predominantly in the S phase of the cell cycle, indicating replication dysfunction. DHS contributed extensively in trapping of PARP on chromatin when combined with talazoparib. This further led to the observed replication stress/dysfunction in cancer cells. Further, we found that the inhibition of back-up Okazaki fragment processing by enhanced 53BP1 levels and non-suppression of replication gaps in the presence of talazoparib plus DHS led are key factors contributing to contextual synthetic lethality. Taken together, we present a novel replication-dysfunction mediated approach for enhancing PARPi sensitivity in cancers.
| Topic 2. Low and High LET Radiation Biology | | |
| P-2.1 | | |
| Investigating the Hematopoietic Response to Ionizing Radiation using Drosophila Larval Lymph Gland | | |
Jagdish Gopal Paithankar, Mohamed Sabeelil Islam, N. Karunakara1, K. Bhasker Shenoy2, Anurag Sharma
Nitte (Deemed to be University), Nitte University Centre for Science Education and Research, Division of Environmental Health and Toxicology, 1CARRT, Mangalore University, 2Department of Applied Zoology, Mangalore University, Mangaluru, Karnataka, India
E-mail: [email protected]
Introduction: The health consequences of ionizing radiation (IR) exposure are greatly concerned. Hematopoietic stem cells (HSCs) in the bone marrow provide blood cells throughout the lifetime. Due to continuous blood cell production and the high rate of cell division, hematopoiesis is susceptible to any stress, including IR. In this study, IR-induced hematopoietic response was investigated using the Drosophila hematopoietic organ, the lymph gland. The Drosophila lymph gland is comprised of well distinct regions. A pool of progenitor cells is in the inner medullary zone, and differentiated hemocytes are in the outer cortical zone. It also consists of a posterior signaling center which acts as a regulatory niche. At the onset of pupariation, the lymph gland bursts and release the progenitor and differentiated hemocyte in the circulation. Materials and Methods: Drosophila flies/larvae (wild type/transgenic) were maintained in standard conditions. Radiation treatment (4-12 Gy) was given to the third instar larvae using a Cobalt-60 irradiator. The lymph gland was dissected post-IR exposure, and the hematopoietic response, such as progenitor and differentiated hemocytes status, structural damage to the organ, ROS generation, and cell death, were studied. Results: Structural defects such as premature lymph gland bursting were observed at higher IR doses. However, significant variations in the blood cell populations, along with elevated ROS levels in the progenitors and the differentiated hemocyte population, were observed at lower doses. It has been established that niche plays a vital role in controlling the hematopoietic program in stressful situations. Interestingly, we did not follow any role of the hematopoietic niche in IR-induced hematopoietic injury. Conclusions: We found that lower doses negatively impact the hematopoietic progenitors/differentiated hemocytesin the lymph gland. However, higher doses promote premature lymph gland bursting and release of the hemocyte in the circulation, which might be a defence mechanism against IR exposure.
| P-2.2 | | |
| Effect of Gamma Radiations on Burn Wound Healing in Mouse Model | | |
Lipsy Goyal1,2, Amarjit S. Naura2, Anshoo Gautam1
1Institute of Nuclear Medicine and Allied Sciences, Defence Research and Development Organization, New Delhi, 2Department of Biochemistry, Panjab University, Chandigarh, India
E-mail: [email protected]
Introduction: Combined radiation and burn injury is defined as thermal trauma along with radiation injury. It can occur in people exposed to nuclear explosions, nuclear accidents or radiological terrorist attacks. Previous studies had reported the delay in irradiated wounds as radiation results in fibroblast injury as well as endothelial cell necrosis. Here, we studied the effect of gamma radiations on burn wound healing in comparison of single injuries. Materials and Methods: To investigate the effect of radiation on the healing of burn wounds, C57BL/6 male mice were irradiated with a selected dose, using a60Co γ-irradiator, along with 5% of whole body full thickness burn. Wound contraction, wound size and eschar fall were evaluated in combined injury burn wounds. Skin histology along with hydroxyproline estimation was performed. Various oxidative stress markers were studied. Body weight change, food and water uptake were recorded daily. Results: Combined burn wound had shown a delayed wound healing, when compared to burn alone group. Further, wound skin histology had shown a higher inflammation in combined burn wound even after 14 days of injury. Decreased hydroxyproline levels were observed in combined injury group. Biochemical parameters resulted in increased oxidative stress in combined injury animals as compared to individual injuries. Our data showed a significant reduction in body weight in combined injury group along with decreased uptake of food and water, when compared with individual injuries. Conclusion: In this study, we observed that gamma irradiated burn wounds show delayed healing as compared to burn alone wounds.
| P-2.3 | | |
| Effects of RF-EMR (2.45 GHz) on Mouse Leydig Cell (TM3): In-Vitro Study | | |
Pooja Jangid1, Rajeev Singh1,2
1Department of environmental studies, Satyawati College, University of Delhi, 2Department of Environmental Science, Jamia Millia Islamia University, New Delhi, India
E-mail: [email protected], [email protected]
Introduction: The threat of male infertility became a big issue for the human population as modern technologies emerged as a source of radiofrequency radiation (RFR) and studies have shown that these radiations are lethal for human health. These technologies have become an essential part of our everyday life. Therefore, it is necessary to find its effects on reproduction. The current study concentrated on the biological consequences of electromagnetic radiation exposure on mouse Leydig (TM3) cell in a time-dependent way. Materials and Methods: The cells were subjected to 2.45 GHz non-ionizing electromagnetic radiation in the exposure system at various time intervals (15, 30, 45, 60, 90, and 120 minutes), and cell were assessed for viability, apoptosis, ROS, and testosterone levels. Results: Cell viability was not significantly (p<0.05) affected by EMF exposure via the Exposure system. In the irradiated group, there was a significant increase in apoptotic cells as compared to the control group. At 60, 90, and 120 minutes, the irradiation group had significantly lower testosterone levels than the control group. At all of the time periods following irradiation through exposure system, the levels of ROS did not significantly alter. Conclusions: In conclusion, the current study shows that exposure to radiofrequency electromagnetic radiation emitted from exposure systems can cause adverse effects on the male fertility by impairing Leydig cell (TM3) function, like decreased testosterone production and increased cell apoptosis.
| P-2.4 | | |
| Epithelial Mesenchymal Transition Associated Changes in A549 Cells Irradiated with Low Dose of Alpha Particle Irradiation | | |
R. Vasumathy1,2, B. N. Pandey1,2
1Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: High LET radiotherapy is widely considered approach to efficiently kill tumor cells with minimal damage to normal tissues. However, the effects of high LET radiation at very low doses like 1.36cGy is not much discussed in literature. In our previous work we observed increased proliferation of A549 cells at low dose (1.36cGy) of alpha particle irradiation associated with Connexin 43 and Survivin proteins. These results established the need to see if any EMT associated changes are observed in the cells irradiated with 1.36cGy of alpha particle radiation. Materials and Methods: Cell adhesion and migration of A549 cells irradiated with 1.36cGy alpha particle irradiation were evaluated by crystal violet assay and wound closure assay respectively. Expression E-cadherin and N-cadherin after irradiation was assayed by western blotting. Levels of TGF1-ß,a crucial molecule involved in EMT of cancer cells and IL-1ß which is known to augment effects of TGF1-ß were assayed in cell culture supernatants of irradiated A549 cells. Levels of p-AKT was assayed in alpha particle irradiated A549 cells by western blotting. Results: Alpha particle irradiated A549 cells (1.36 cGy) showed EMT associated changes like decreased adhesion (~13%) and increased migration (~13%) than that of control. Also the hallmark of EMT, cadherin switching was evidenced in alpha particle irradiated cells by decrease in levels of epithelial marker, E-cadherin (0.74± 0.04 fold) and increase in levels of mesenchymal marker, N-cadherin (1.5 ± 0.01 fold).There was a 2.7 fold increase in TGF-ß1 levels and 1.2 fold increase in IL-1ß levels in the cell culture supernatants of irradiated cells. Irradiated A549 cells also showed significantly higher phosphorylation of AKT (~2.8 fold) than that of control after 48h of irradiation which is known to be upregulated by TGF1-ß during EMT. Conclusions: A549 cells irradiated with low dose of alpha particle irradiation (1.36cGy) is shown to induce EMT associated changes.
| P-2.5 | | |
| Role of Kinetin Intervention in Ameliorating the Behavioral Changes in Swiss Albino Mice Exposed to Gamma Radiation | | |
R. Vishakh, N. Suchetha Kumari1, Shailaja S. Moodithaya2
Central Research Laboratory, KS Hegde Medical Academy, Nitte (Deemed to be University), Departments of 1Biochemistry and 2Physiology, KS Hegde Medical Academy, Nitte (Deemed to be University), Mangaluru, Karanataka, India
E-mail: [email protected]
Introduction: Studies confirm that, no dose of radiation exposure is safe to the human exposure. Behavioural changes especially the cognition and anxiety are considered to be the important problems faced due to the radiation induced neuronal damage especially in the hippocampal region. Exposure of radiation will lead to the increased production of ferrous ions which will combine with the hydrogen peroxide formed to oxidative stress may lead to the production of superoxide radicles which will further damage the neurons. Thus in the present study we aimed to evaluate the radiation induced changes in the iron status and its role in the behavioural changes in mice model. Materials and Methods: The study was performed after clearance from institutional animal ethics committee. The work was carried out at KS Hegde Medical Academy, Mangalore and radiation facility of CAART of Mangalore University was utilized. Free iron in serum was estimated spectrophotometrically, the Ferritin and transferrin levels were estimated using commercially available kits. Open field behaviour test and radial maze test (8 armed) was performed to assess anxiety and learning ability in Swiss albino mice. Results: LD50/30 was found to be 6.26 Gy at the dose rate of 6Gy/minute. So, a sublethal dose for 2Gy was used for further studies. A significant decrease in the learning ability and increased anxiety in the radiation control group which significantly improved in Kinetin treatment group. But no significant difference in the levels of ferritin, transferrin, and free iron in the radiation control group. Conclusions: The kinetin intervention reduces the anxiety and improves the learning ability in mice. But, according to the results it was not due to changes in iron status. Further study are needed to find the mechanism behind the same.
Keywords: Behaviour studies, Iron profiling, ionizing radiation, Radioprotector
| Topic 3 Tumor Hypoxia and Chemo-Radioresistance | | |
| P-3.1 | | |
| Monoterpene Indole Alkaloid Derivatives Overcome P-gp Mediated Multidrug Resistance in Cancer Cells | | |
B. Pradhapsingh, N. Rajendra Prasad
Department of Biochemistry and Biotechnology, Annamalai University, Chidambaram, Tamilnadu, India
E-mail: [email protected]
Introduction: Multidrug resistance (MDR) remains a major challenge in cancer chemotherapy, mainly due to the overexpression of ABC drug-efflux transporters, namely P-glycoprotein (P-gp/ABCB1). In a previous study, N-alkylated derivatives of monoterpene indole alkaloids, isolated from the African medicinal plant Tabernaemontana elegans (Apocynaceae family), were found to be P-gp inhibitors. This study aimed to further investigate selected compounds as MDR reversers, using as model resistant human cervical carcinoma cells (KB-ChR-8-5 cells) overexpressing P-gp. Methods: Three N-alkylated derivatives of the indole alkaloid tabernaemontanine, bearing N-(para-bromobenzyl) (NBBT) N-(para-methyl benzyl) (NMBT) and N-(para-methoxyphenethyl) (NMPT) moieties, were investigated for their ability to reverse P-gp mediated MDR in KB-ChR-8-5 cells, by the calcein-AM drug efflux assay and their binding affinity were analyzed by molecular docking methods. Compound NBBT was further investigated for its ability to modulate P-gp expression by immunocytochemistry. Combination assays with doxorubicin and its ability to induce apoptosis, using the acridine orange/ethidium bromide assay, as well as cell migration assays were also performed. Results: In the functional assay, NBBT exhibited the highest P-gp inhibitory activity in a dose-dependent manner and was further investigated. It significantly decreased P-gp overexpression by inactivating nuclear translocation of the NF-κB p-50 subunit. Doxorubicin showed 6.3-fold resistance in KB-ChR-8-5 cells compared with its parental KB-3-1 cell line. However, NBBT significantly reduced doxorubicin fold resistance to 1.7, 1.3, and 0.4 with 1, 3, and 6 μM of NBBT treatment, respectively. Moreover, NBBT showed strong synergism with doxorubicin for all the concentrations studied in the drug-resistant cells. Furthermore, NBBT plus doxorubicin combination treatment showed increased apoptotic incidence and decreased cell migration compared with doxorubicin alone treatment. Conclusion: The present findings suggest that NBBT could be a lead candidate for the reversal of P-gp- mediated MDR in cancer cells.
| P-3.2 | | |
| Enhanced Delivery of Quercetin and Doxorubicin Using β-Cyclodextrin Polymer to Combat P-Glycoprotein-Mediated multidrug-Resistant Cancer | | |
Charan Singh Pawar, N. Rajendra Prasad, Priya Yadav
Department of Biochemistry and Biotechnology, Annamalai University, Chidambaram, Tamil Nadu, India
E-mail: [email protected]
Introduction: Cyclodextrins (CDs), carbohydrate-based polymers have recently been considered an effective drug-delivery agent due to their cone-shaped configuration which consists of a hydrophobic cavity and hydrophilic matrix. Drug delivery of quercetin (QCT) and doxorubicin (DOX) by β-cyclodextrin polymer(β-CDP) has the potential to bypass P-glycoprotein (P-gp) mediated multidrug resistance (MDR) in cancer cells by inhibiting the activity of ABC-transporter efflux pumps. QCT is a flavonoid used as a P-gp inhibitor to inhibit the excess efflux of DOX from MDR cancer cells. DOX, is an anticancer drug, which is used to treat various types of cancers. We prepared β-cyclodextrin polymer co-loaded quercetin and doxorubicin nanocarrier (β-CDP/QD NCs) to reverse MDR in P-gp overexpressing KB-ChR 8-5 cancer cells. Materials and Methods: The β-CDP/QD NCs were prepared by the freeze-dried method and characterized by various microscopic and spectroscopic techniques. The P-gp overexpressing KB-ChR 8-5 cancer cells were treated with β-CDP, QCT, DOX, and β-CDP/QD NCs. The treatment was done to examine the cell cytotoxicity, drug internalization, nuclear fragmentation, and apoptosis changes in KB-ChR 8-5 cancer cells. Results: The microscopic and spectroscopic techniques confirmed the successful encapsulation of QCT and DOX in the β-CDP. The β-CDP/QD NCs show more potency against P-gp overexpressing KB-ChR 8-5 than β-CDP, QCT, and DOX. The NCs induce enhanced apoptotic morphological changes, nuclear damage, and alteration in mitochondria. Thus, the present results suggest that β-CDP/QD NCs inhibit the proliferation of MDR cancer cells by enhancing apoptotic cell death. Conclusions: The synthesized β-CDP/QD NCs significantly inhibits the growth of P-gp overexpressing KB-ChR 8-5 cancer cells by sustained release of both QCT and DOX in the drug resistant cancer cells.
| P-3.3 | | |
| Alterations in Transcriptome and Proteome in Isogenic Radioresistant Lung Cancer Cells after Fractionated Irradiation | | |
Himanshi Narang1,2, S. Nishad1, Anu Ghosh1,2
1Animal House Facility and Radiation Signalling Section, Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Fractionated radiation therapy remains the most prominent method for treating cancer; the clinical outcome however is limited by the development of radioresistance over the course of treatment. To further improve the therapeutic ratio, better insight into mechanisms operative in development of radioresistance is needed. Present study was designed to study the molecular changes in lung cancer cells irradiated with multiple doses of irradiation over a period of month to simulate radiotherapy schedule under clinical setting and compare these changes vis-a-vis unirradiated and/or single irradiated lung cancer cells. Materials and Methods:A549 lung cancer cells were irradiated with multiple fractions of radiation to generate the radioresistant isogenic cell line. Total ten fractions of 2 Gy were delivered over a period of five weeks. Cells were then allowed to recover for a week and were termed 10F-A549. A549 and 10F-A549 cells were analysed for their radioresistance, DNA repair kinetics and molecular changes including proteome and DNA repair transcriptome with or without irradiation to understand the transient and persistent molecular changes after irradiation. Results: Radioresistant cells subjected to multiple fractions of irradiation showed less DNA damage and faster repair upon challenging dose. Proteomic studies revealed alterations of many pathways in 10FA549 cells including energy metabolism pathways and cytoskeletal organization.LC-MS/MS identified 1345 proteins in single 2Gy γ-irradiated A549 (R-A549) cells lines and 1195 proteins in multiple γ-irradiated (2Gy x 10) A549 (C-10F-A549) cells. The LFQ analysis identified differential alteration (fold change ≥1.5 or ≤ 0.67) of 696 proteins (491 up and 205 down) in R-A549 cell lines while 786 proteins were modulated (363 up and 423 down) in multiple γ-irradiated A549 (10F-A549) cells. At transcription levels, many DNA repair genes showed modulation of expression with respect to irradiation. Conclusion: Radioresistant cells obtained by multiple radiation exposure simulating protracted clinical schedule of radiotherapy revealed alterations in many pathways relevant or non relevant to radiation. These findings can have important clinical implications in terms of finding promising targets for modulation for desired cellular responses.
| P-3.4 | | |
| Ovarian Cancer – Evolution of Natural Products against Radiation therapy? | | |
Komal Jha, Chitra Gupta
Department of Chemistry, Bundelkhand University, Jhansi, Uttar Pradesh, India
E-mail: [email protected], [email protected]
Ovarian cancer ranks fifth amongst the gynaecological cancer makes it fatal and causes more chances of deaths in women from all over the world. Epithelial ovarian cancer consists of surgical removal including abdominal hysterectomy and directing of six cycles of intravenous chemotherapy. According to American cancer society statistic 2022 in U.S 1,918,030 new cancer cases and 609,360 deaths are approximately can be reported. Due to lower chances of detecting it earlier this cancer becomes almost untreatable which in turn makes it more difficult to treat. Earlier detection and advances in screening of cancer can provide numerous chances for treatment and usage of more natural products like quercetin, paclitaxel, vincristine, etoposide…etc which strengthen's our immune system and does not lead to radiation therapy for this cancer. In this review we want to study new natural products usage with chemotherapy which are non-toxic with less or no side-effects and has truncated chances of relapses.
Keywords: Natural Products, Non-toxic, Ovarian Cancer, Radiation therapy, Relapses
| P-3.5 | | |
| ER Stress and GCN5 as Novel Molecular Targets for GBM Senotherapy | | |
Madhura Ketkar1,2, Sanket Desai2,3, Amit Dutt2,3, Shilpee Dutt1,2
1Shilpee Dutt Laboratory, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, 3Integrated Cancer Genomics Laboratory, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, Navi Mumbai, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: We have shown that radiation therapy induces senescence (TIS) in residual cells of glioblastoma (GBM) which, unfortunately, is reversible leading GBM relapse. Here, we aim to understand the mechanisms of senescence reversal in GBM residual cells that can be exploited for senotherapy. Materials and Methods: Residual senescent (RS) and residual senescence escaper (RSE) cells were captured from radiation survival models generated from GBM primary patient samples and cell lines. RNA sequencing of 40 samples (parent, RS, RSE and relapse) was done to identify candidate genes/pathways involved in TIS reversal. Real time-qPCR, Immunoblotting, SA-Beta-Galactosidase assays, genetic and pharmacological perturbation of genes were performed for mechanistic studies. Results: Post lethal dose of radiation to cell lines and primary cultures of GBM patient samples, surviving 10% residual cells showed senescence phenotype (TIS) assessed by high beta-galactosidase activity, high expression of p21, p16 and SASPs (senescence associated secretory phenotype). Unfortunately, TIS reversed forming relapse. RNA sequencing analysis of parent, RS, RSE and relapse cells showed significant enrichment (mRNA and protein levels) of Cytokine signaling and Endoplasmic Reticulum stress related pathways in the RS cells. However, these pathways showed significantly reduced expression in RSE population. Pharmacological inhibition of UPR in the residual senescent cells led to senescence maintenance and prevented relapse. Independently, we found GCN5 a histone acetyltransferase knockdown also induced senescence in GBM cells. Notably, this unique senescence was characterized by irreversibility, p16high, p21high, Beta galhigh and SASPlow phenotype. GCN5 knockdown did not elicit ER stress/UPR. Additionally, conditioned media from radiation induced senescent cells but not from GCN5 knockdown induced senescent cells could induce senescence in parent glioblastoma. Indicating that secretome plays a role in maintenance of senescence and may decide the reversibility of phenotype. Conclusions: We have identified GCN5 as novel target for senotherapy in glioblastoma and ER stress/UPR pathway as molecular switch for reversal of radiation induced senescence in GBM.
| P-3.6 | | |
| Transcriptomics Analysis of Radio-resistant Lung Cancer Cell lines and Respective Tumorospheres for Identification of Cancer Stem Cells and Gene Signatures to Predict Radiotherapy Response in Lung Adenocarcinoma Patients from TCGA Database | | |
Murali M. S. Balla1,2, Pooja Patheja1,2, Dhruv Das2,3, Vanshika Gupta4, Harish Shrikrishna Bharambe5, Shalaka Masurkar5, Nikhil Gadewal5, Neelam Vishwanath Shirsat5, B. N. Pandey1,2
1Radiation Biology and Health Sciences Division, Bio-Science Group, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, 3Applied Genomics Section, Bio-Science Group, Bhabha Atomic Research Centre, Mumbai, 5Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, Navi Mumbai, Maharashtra, 4DRDE, DRDO, Ministry of Defence, Government of India, Gwalior, Madhya Pradesh, India
E-mail: [email protected]
Introduction: Radiotherapy is one of the main strategies in the treatment of non small cell lung cancer patients. However, poor prognosis of radiotherapy is one of the major challenges, which was addressed in this study through transcriptomics analysis. Materials and Methods: Radioresistant lung cancer cells and respective tumorospheres obtained from A549 cells were characterized for radio-resistance and enrichment of cancer stem cells followed by transcriptomics analysis. Data obtained was evaluated to select genes which are significantly correlated with lung cancer patient data from the TCGA database. Machine learning tools were further applied on patient data from TCGA database to predict therapy response. Results: The gene expression profiles of radioresistant cell lines and respective spheroids, showed 450 and 481 genes differentially regulated compared to respective controls. Further analysis showed that 122 genes were commonly expressed between the groups, 313 and 342 genes were specific to radioresistant cell lines and spheroid cultures, respectively. These genes were then correlated with the gene expression profiles of the lung adenocarcinoma cancer patients' samples obtained from the TCGA database. Binary logistic regression of 120 genes resulted in 7 genes (RIMS2, KCNH3, LHFPL4, L1CAM, SYTL3, NR3C2 and HS3ST1) which showed significant association with the radiotherapy response of patients. When logistic regression machine learning classification model was trained on 80% of the patient data, the model accuracy was found to be 82% with AUC=0.85±0.12. Finally, when nomogram analysis was performed it obtained a positive likelihood ratio of 5.41 suggesting the promising prognostic and predictive nature of these gene signatures to know the radiotherapy response of the lung cancer patients. Conclusions: The predictive ability of cancer stem markers and gene signatures obtained from transcriptomics analysis hold potential to predict the radiotherapy outcome in lung adenocarcinoma cancer patients.
| P-3.7 | | |
| Modulation of Epithelial to Mesenchymal Transition and Cancer Stem Cells by Radiation and Cisplatin in Breast Cancer Cells | | |
Priya Kundu1,2, Bhavani S. Shankar1,2
1Immunology Section, Radiation Biology and Health Sciences Division, Bioscience Group, Modular Laboratories, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Radiotherapy and chemotherapy are the two major treatment modalities in breast cancer. Relapse, resistance and metastasis are the major challenges following chemo / radio therapies for which epithelial to mesenchymal transition (EMT) and cancer stem cells(CSCs) are primarily responsible. This study aims to understand the relationship between EMT and CSCs and elucidate the underlying signalling mechanisms involved following treatment of breast cancer cells with radiation and cisplatin. Materials and Methods: MCF7 breast cancer cells were treated with radiation (6 Gy) and cisplatin (5μM) followed by recovery period for 5 days. Epithelial - mesenchymal markers, EMT transcription factors, CSC markers and transcription factors were assessed by RT-PCR and flow cytometry. Differentially regulated genes were studied by EMT array (84 genes). Cytokines were assessed by RT-PCR/ ELISA and signalling pathways by RT-PCR/ western blot. Results: Radiation and cisplatin treatment induced hybrid EMT in breast cancer cells as indicated by increased epithelial (E-cadherin, Desmoplakin and Claudin-1) and mesenchymal (Vimentin, Fibronectin and N-cadherin) markers along with morphological changes. EMT array of radiation and cisplatin treated cells indicated upregulation of transcription factors Slug and STAT3. There was an enrichment in breast cancer stem cells following radiation and cisplatin as seen by markers CD44+CD24-/low %, aldehyde dehydrogenase and CSC transcription factors (Oct4, Sox2 and Nanog). There was an increased TGF-β / SMAD3 as well as IL-6 / STAT3 signalling pathway. These pathways co-operate through Notch signalling to result in upregulation of Snail / Slug expression. Conclusions: Radiation and cisplatin treatment in breast cancer cells upregulated TGF-β / SMAD3 and IL-6 / STAT3 signalling pathways. SMAD3 / STAT3 signalling in turn upregulated Snail / Slug transcription factors that can drive EMT and CSCs. Notch signalling may be an intermediary in this signalling network.
| P-3.8 | | |
| Establishment and Characterization of a Clinically Relevant In Vitro Chemo-Radioresistant Cervical Cancer Model to Understand the Mechanistics of Therapy Resistance | | |
Reshma Reddy1,2, Vagmi Gaiwak1,2, Jayant Sastri Goda1,2, Tanuja Teni1,2
1Tata Memorial Centre-Advanced Centre for Treatment, Research and Education in Cancer, Navi Mumbai, 2Homi Bhaba National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
In locally advanced cervical cancer (LACC) patients, concurrent chemo-radiotherapy (CCRT) is the mainstay of treatment wherein 30 - 40% patients exhibit relapse within first 2-3 years of treatment, indicating acquirement of resistance to combination (chemo and radio) therapy. However, there is a lack of robust model system to comprehend the molecular events arising post combination therapy contributing to chemo-radioresistance. Further, about 99.7% cervical lesions show presence of human papilloma virus (HPV) and persistent infection with “high-risk” subtypes like 16, 18, 31, 33 and 45 has been demonstrated in promotion of both pre-invasive and invasive cervical cancers. Previous studies from our lab have demonstrated that significantly high persistent HPV 16/18 DNA viral load in immediate post treatment period in LACC treated with CCRT correlated with high loco-regional and overall relapses, indicating a possible association of high persistent HPV 16/18DNA viral load with chemo-radioresistance. Thus, it is important to understand the association of HPV infection with chemo-radioresistance. With regard to this, in the present study we attempt to establish an in vitro chemo-radioresistant cervical cancer model simulating the clinical treatment regimen. Briefly, HPV 16 positive SiHa cells were treated with fractionated radiotherapy concurrently with cisplatin. Following generation of the resistant line, its cellular, molecular, phenotypic and transcriptomic characterization was performed at early, mid and late interval resistant cells Delayed doubling time, increased colony formation post cisplatin and radiation exposure, increased migration, elevated expression of anti-apoptotic, drug influx/efflux markers and HPV onco-proteins were observed in the resistant cells compared to parent. Ultrastructural alterations like enlarged cell and nucleus, spherical mitochondria, and enlarged golgi stacks were observed in resistant cells. To conclude, the study will provide an in-depth understanding of sequential molecular events occurring during the acquirement of chemo-radioresistance and can be utilized for development of new therapeutic strategies that may have translational value.
| P-3.9 | | |
| Mitochondrial Redox Pathway Activation and Enhanced Chromatin Condensation of Radiation-Resistant Breast Cancer Cells in Hypoxic Tumor Microenvironment | | |
Sukanya Rauniyar1,2, Kshama Pansare3, Bharat Khade1, Chilakapati Murali Krishna2,3, Sanjay Gupta1,2,4
1Epigenetics and Chromatin Biology Group, Gupta Laboratory, Cancer Research Institute, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, 3Chilakapati Lab Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, 4Epigenetics and Chromatin Biology Group, Gupta Laboratory, Cancer Research Institute, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, Navi Mumbai, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: In solid tumors, hypoxia associates with failure of anticancer treatment strategies. Moreover, intermittent reoxygenation of hypoxic cells potentially affects biological outcomes in clinics. Hypoxia associated epigenetic alterations are responsible for differential response to therapies. Therefore, present study highlights the alterations in epigenome, their biological effect and potential therapeutic targets during hypoxia-reoxygenation events in parental and therapy-resistant breast cancer cells. Materials and Methods: Parental and radio-resistant breast cancer cells were exposed to hypoxia-reoxygenation for deciphering the alterations in histone post-translational modifications, cellular and molecular pathways associated with energy metabolism using Raman spectroscopy, electron microscopy, confocal microscopy, and functional assays. In vivo studies were carried out to understand the importance of chromatin modifiers as radio-sensitizer along with conventional radiotherapy. Results: In response to hypoxia-reoxygenation, the parental and radio-resistant cells showed ultrastructural mitochondrial aberrations like cristae disintegration, swelling along with changes in their structural and functional proteins. Incoherence, the increase of total and mitochondrial ROS with enhanced γH2AX foci and decreased pATR levels were observed in parental cells compared to radio-resistant cells. These cells have increased survival potential due to increased levels of pAKT and pERK1/2. The nuclear periphery has increased levels of H3K9me3 and H3K27me3 with elevated migratory potential in hypoxia-reoxygenated cells compared to parental cells.The combinatorial treatment of parental and resistant cells with class 1 HDAC inhibitor followed by radiotherapy to hypoxic and reoxygenated resistant cells leads to enhanced cell death and regression of tumor. Conclusion: Mitochondrial alterations, ROS signalling, cell survival associated with chromatin condensation play an essential role in cell proliferation and better survival of radio-resistant cells compared to parental cells under hypoxia and reoxygenation. Therefore, the targeting chromatin condensation helps in achieving enhanced therapeutic efficacy.
| Topic 4. Combinatorial and Heavy Ion Radiotherapy | | |
| P-4.1 | | |
| Di-Selenide Decorated Gold-Nanocomposite for the Chemo/Radiotherapy of Glioblastoma Multiforme | | |
J. Aishwarya1,2, Beena G. Singh2,3, Ram P. Das2,3, Vikram Gota1,2, Amit Kunwar2,3
1Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, Navi Mumbai,2Homi Bhabha National Institute, 3Radiation and Photochemistry Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Glioblastoma multiforme (GBM) is the most aggressive malignant brain tumour. Gold nano-conjugates of suitable shape and size have been shown to effectively permeate the blood BBB and are also reported for radio-sensitizing activity. Moreover, organo-diselenides (-Se-Se-) are well reported to exhibit potent anti-cancer and radio-sensitization activities through redox modulation. Therefore, the objective of the present study was to design a diselenide decorated gold nanoparticles and evaluate its chemotherapeutic and radio-sensitizing activities. Materials and Methods: The physiochemical characterisation of nanocomposite was performed using various spectroscopic techniques. The cytotoxicity of the nanoconjugate in the presence and absence of radiation was evaluated in cellular models of GBM by clonogenic assays. Results: Au nanoparticles of spherical morphology and of size in the range of 15 nm (confirmed by TEM) were synthesized using Turkevich method. In order to decorate these nanoparticles with diselenide, an organodiselenide (R-Se-Se-R) was covalently linked to FDA approved biopolymer, gelatin, to prepare G-Se-Se-G polymer. Subsequently G-Se-Se-G was used as a stabilizing agent to replace citrate from the surface of gold nanoparticle and to obtain [email protected] nanoparticles. The composition or assemble of nanoparticles was confirmed by Raman spectra, XPS, FTIR, Cyclic Voltammetry and CD. The MTT and clonogenic assays confirmed the chemotherapeutic potential of G-Se-Se-G polymer and [email protected] nanoparticles in Glioma cell lines with IC50 values in the range of 0.001 to 0.1 μg Se equivalent. Further, bare gelatin coated Au nanoparticles exhibited radio-sensitization activity with a dose modification factor of 0.76. The radio-sensitizing ability of [email protected] formulation is under evaluation and will be presented in the conference. Conclusions: The reaction conditions have been optimised to synthesize the size selective Au-nanoconjugates. The preliminary cellular studies indicated both chemotherapeutic and radio sensitizing activities of Au-nanoconjugates.
| P-4.2 | | |
| A Bodipy-Based Novel Fluorescent Probe for Targeting Endoplasmic Reticulum of Pancreatic Ductal Adenocarcinoma for Photodynamic Therapy | | |
Nitish Chauhan1,2, Mrunesh Koli1,2, Tapan Kumar Ghanty1,2, Soumyaditya Mula1,2, Birija Sankar Patro1,2
1Bio-Organic Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected], [email protected]
To date, pancreatic adenocarcinoma (PDAC) is a devastating disease for which the incidence rate is close to the mortality rate. The survival rate has evolved only 2–5% in 45 years, highlighting the failure of current therapies. Recently, photodynamic therapy (PDT), based on the use of an adapted photosensitizer (PS) showed great promise for PDAC treatment. However, potent photosensitizers with efficient targeting potential to PDAC is not available for PDAC therapy. Considering the fact that PDACs are heavily relying on endoplasmic reticulum (ER) for hormone and protein synthesis, the current project was focussed on to design and develop ER targeting photosensitisers. In this regard, an efficient synthesis route was developed for making bis-chromophoric dyes (compound 3 and 7) by orthogonically connecting BODIPY and naphthalene moieties (known to localize to ER). Interestingly, we have found the efficient intersystem crossing of compound 7 than compound 3. Further our extensive photochemical study showed that compound 7 efficiently forms transient triplet state and generates copious amount of singlet oxygen, which is one of the paramount features of photo-dynamic therapy (PDT). In biological system, compound 7 showed excellent photodynamic effects to cause robust killing of multiple pancreatic ductal carcinoma cells. Intriguingly, we observed rapid uptake and localization of compound 7 in endoplasmic reticulum of PDAC cells. Interestingly, live cell time series experiment with compound 7 showed simultaneous dual fluorescence of green in ER and red in lipid droplet respectively in temporal fashion, owing to its differential fluorescence properties in different polar medium. Mechanistically, compound 7 targets ER of the PDAC cells and causes ER stress, which ultimately leads to robust photosensitization of PDAC cells in the presence of visible light. Therefore, this BODIPY-based novel fluorescent probe have theragnostic application for Pancreatic Ductal Carcinoma diagnosis and treatment.
| P-4.3 | | |
| Synergistic Interaction of Heavy Metal Salts with Hyperthermia or Ionizing Radiation | | |
M. S. Tolkaeva, V. G. Petin
A. Tsyb Medical Radiological Research Center – Branch of the National Medical Research Radiological Center of the Ministry of Health of the Russian Federation, Obninsk, Russia
E-mail:[email protected]
Introduction:The study of the patterns of manifestation of synergism with the interpretation of the results obtained after the simultaneous action of hyperthermia or ionizing radiation with various salts of heavy metals is an urgent problem of modern radiobiology and ecology. The combined use of radiation therapy or hyperthermia with various chemical preparations containing heavy metals leads to an increase in the efficiency of cell inactivation, and makes it possible to reduce the applied doses and concentrations inside the tumoursand the whole organism. However, the synergy of heavy metals with various environmental agentsmayresult to the formation of an additional negative impact on living organisms. Materials and Methods: Diploid yeast cells (strain XS800) were used in the experiments. To quantify synergistic effect, cell survival was determined after individual effects of heavy metal salts, hyperthermia, ionizing radiation, and their simultaneous action. The following heavy metal salts were used: lead iodide, potassium dichromate, cisplatin, copper sulfate, zinc sulfate. Results: In all cases, a synergistic interaction was observed – the experimental survival curves after combined exposures were lower than expected ones for independent addition of theeffects from each agent. The dependences of the synergistic enhancement ratio on the concentration of the chemical agent are dome-shaped, in which the synergy first increases, reaching a maximum at the optimal ratio of the applicable agents, after which it gradually decreases. To interpret the results obtained, we use a mathematical model of synergism, according to which the synergistic effect is due to the formation of additional effective damage and thereby increase the positive effect of agents under combined effects. Conclusions: These data indicate the universality of the manifestation of synergistic effects and demonstrate the possibility of mutual enhancement of the agents used in combined treatment and the harmful effects of environmental agents.
| P-4.4 | | |
| Synthesis and Characterization of Silver Nanoparticles Synthesis using Mentha Piperita and Evaluation of Its Anticancer Properties in In -Vitro Studies on Skin Cancer Cell Line A431 | | |
P. Kaushik, A. Yadav, A. Sharma1, M. K. Sharma2
Department of Zoology, Centre for Advanced Studies, University of Rajasthan, 1Department of Zoology, Government College, Jaipur, 2Department of Zoology, SPC Government College, Ajmer, Rajasthan, India
E-mail: [email protected]
Introduction: Mentha is a genus belonging to the family of Lamiaceae. Mentha taxonomy is highly complicated and includes about 42 species and 15 hybrids, with hundreds of cultivars. Mint is herbaceous, perennial aromatic herbs that are cultivated for their essential oils used both for medicinal and aromatic purposes. Mentha piperita species are rich in polyphenols, caffeic acid, caftaric acid, cinnamic acid, ferulic acid, and oleanolic acid. Recent advances in nanoscience and nanotechnology radically changed the way we diagnose, treat, and prevent various diseases in all aspects of human life. Silver nanoparticles (AgNPs) are one of the most vital and fascinating nanomaterials among several metallic nanoparticles that are involved in biomedical applications. AgNPs have been focused on potential applications in cancer therapy. Materials and Methods: In the present study green nanoparticles are were synthesized using aqueous extract of Mentha piperita leaf. The AgNPs were characterized using Ultra-Violet Visible (UV-VIS) spectroscopy, Fourier Transform Infrared (FTIR) spectroscopy and Scanning Electron Microscopy (SEM) to study the shape and size of the AgNPs. The anti-cancer activity of AgNPs have been evaluated against skin cancer cell line (A431). Results: The preliminary results of the study suggest control of survival and proliferation rate of cancer cells using trypan blue viability test and reduction in metastasis potential by inhibition of cell migration rate as studied by in-vitro scratch test. Conclusion: The antiproliferative and antimetastatic activities of green AgNP using Mentha piperita leaves provides an insight into its potential application in anticancer therapeutic research. Further experimentation is in pipeline in our laboratory to deduce the anti-cancer efficacy and mechanism.
| P-4.5 | | |
| Development of Zinc Gallate Based Formulations for Detection, Targeted 10B Delivery and Prognosis in Cancer Treatmentbyboron Neutron Capture Therapy | | |
Prasad P. Phadnis1,2
1Chemistry Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Cancer research is directed for non-invasive and multimodal utility for diagnosis and therapy as well as evaluation of treatment success to avoid recurrence of cancer and metastasis. Hence, we have developed biocompatible, tumor targeting nanoformulations consisting of zinc gallate (ZnGa2O4:0.5%Cr) based persistent luminescence nanoparticles (PLNPs) loaded with 10B(OH)3 and further conjugated with pH-(low)-insertion peptide (pHLIP) which has affinity for local acidity of cancerous cells. The formulations exhibited ability for detection of cancerous tissues and effective treatment by boron neutron capture therapy (BNCT) as well as facilitated in vivo bioimaging. Materials and Methods: The chemicals used for preparation of nanoformulations viz., zinc nitrate, gallium nitrate, chromium nitrate and chemicals for functionalization like TEOS, CTAB, APTMS, HBTU, EDAC as well as amino acids for synthesis of peptide were procured from Sigma-Aldrich, India. The ZnGa2O4:(0.05%)Cr PLNPs were synthesized and characterized by powder XRD and TEM analyses. The PLNPs exhibited emission centered at 690 nm for ~24 h up on exposure to UV or LED. They were functionalized with silica and conjugated with 10B(OH)3 and further with pHLIP to obtain the test formulations which were evaluated for cellular internalization and biocompatibility and against melanoma and fibrosarcoma induced in C57BL/6 and BALB/c mice, respectively. It was evaluated for therapeutic utility by i.v. injection of formulation followed by exposure of tumor region of mice with epithermal neutrons having an energy of 3 mev and aflux of 2 × 105 neutrons·cm−2·s−1 for 20 min and treatment repeated once after 2 days. The treatment was coupled with in vivo imaging and ex vivo studies on different organs of mice by photon imager. The treatment was followed by monitoring the tumor volume for next 15 days. Further, histopathological studies were performed on tissues of diverse organs and tumors. Results: The evaluation of formulations against tumor in C57BL/6, BALB/c mice by BNCT revealed ~80-85% reduction in tumor volume after exposure to neutrons and monitored the treatment for15 days. The histopathological studies indicated no signs of any damage to the vital organs for both control and treated mice proving non-toxicity of formulations and hence biocompatibility. Conclusions: The results have exhibited multimodal utility and potency of formulations, encouraging for their further evaluations anticipating their clinical use for BNCT assisted with bioimaging.
| P-4.6 | | |
| Role of ATR in Cold Atmospheric Plasma Irradiation Induced Bystander Signaling in A549 Lung Adenocarcinoma Cells | | |
Somnath Ghosh1, Anu Ghosh1,2
1Animal House Facility and Radiation Signaling Section, Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: The radiation-induced bystander effect is defined by the presence of biological effects in cells that were not themselves in the field of irradiation. Plasma, as the fourth state of matter contains numerous active components including photons, free radicals, mild heat, electrons and ions. Cold atmospheric plasma (CAP) has been widely applied in biology and medicine including cancer treatments. However, contributions of CAP induced bystander effect have not been studied. Materials and Methods: Lung adenocarcinoma A549 cells were exposed to 5 min CAP irradiation. The medium from the irradiated cells was transferred to non-irradiated A549 cells to establish the contribution of the CAP induced bystander effect in the survival of the neighbouring cells. Results: Plasma irradiated and bystander cells showed decreased cell survival (as seen with clonogenic assay) and increased DNA damage (as seen with y-H2AX foci) as compared to non-irradiated A549 cells. Plasma-induced bystander effects may be mediated through soluble factors released by irradiated cells. DNA damage response proteins (ATM and ATR) studied in plasma irradiated cells and bystander cells, The A549 cells were divided into two groups; one was treated with ATR inhibitor (NU6027) and the other with ATM inhibitor (KU-55933). Inhibition of ATR in A549 cells completely inhibited DNA damage response signaling in bystander A549 cells. On the other hand inhibition of ATM in A549 cells had no effect in inhibition of bystander effect in bystander A549 cells. Conclusions: DNA damage response signaling was found to be mediated through ATR, but not ATM, in plasma irradiated and bystander A549 cells.
| Topic 5 Radiosensitizers and Radio-Protectors | | |
| P-5.1 | | |
| Binding Studies of Trichostatin-A, a Potential Radiomitigator, to the Bovine Serum Albumin by Multispectroscopic Techniques | | |
Anu Prakash, Paban K. Agrawala
Department of CBRN, Institute of Nuclear Medicine and Allied Sciences, New Delhi, India
E-mail: [email protected]
Introduction: Trichostatin A (TSA), a possible radiomitigator, inhibits mammalian histone deacetylase enzyme family preferentially class I & II. One of the most key aspects in the process of developing new therapeutics is determining the nature of the interaction between the drug and the protein. Since bovine serum albumin are the carrier proteins, these kinds of investigations have been a fascinating area. They offer details on the structural characteristics, dose efficacy, bioavalibilty, & toxicity. This study's objectives were to better understand the mechanisms underlying TSA and BSA interaction. Materials and Methods: Bovine serum albumin, Trichostatin A, Phosphate buffer saline, pH 7.4. For UV & Fluorescence study, BSA was titrated with different conc. of TSA and all the samples were scanned from 200 and 400nnm and 305-400 (excitation at 295nm) respectively. Florescence readings were taken at different temperature. In the same way as the circular dichroism investigation was carried out, the BSA was scanned using various TSA concentrations, and an MRE plot was obtained. Results: In UV & fluorescence, the absorption peak of BSA shifted to a shorter wavelength. Fluorescence and FRET study revealed TSA-BSA undergo static quenching. Negative G0 value showed the spontaneous binding between drug and ligand. Far-UV CD spectra of BSA showed that TSA has the potential to modify the secondary structure of BSA in a concentration-dependent fashion. Conclusions: This study revealed the strong binding between TSA-BSA and complex formed was through static quenching process. At higher concentration of TSA there was change in BSA secondary structure.
| P-5.2 | | |
| Nrf-2 Inhibition Leads to Radio-Sensitization of Human Lung Cancer Cells via Induction of Oxidative Stress, Reduced DNA Damage Repair and Ferroptosis | | |
Archita Rai1,2, Raghavendra S. Patwardhan1, Pradnya K. Pachpatil2,3, Sejal Patwardhan2,4, Santosh K. Sandur1,2
1Radiation Biology and Health Sciences Division, Bio-science Group, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, 3Bio-Organic Division, Bio-science Group, Bhabha Atomic Research Centre, Mumbai,4Advanced Centre for Treatment, Research and Education in Cancer - Tata Memorial Centre, Navi Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Ionizing radiation exerts its cytotoxicity primarily through the generation of reactive oxygen species (ROS). Nrf-2responds to oxidative stress through co-ordinated induction of battery of antioxidant genes by binding to antioxidant response element (ARE). Constitutive activation of Nrf-2-ARE pathway leads to radio-resistance. Pharmacologic inhibition of Nrf-2combined with radio/chemo-therapy is emerging as a promising strategy for better therapeutic gain. Materials and Methods: Effect of Clobetasol Propionate (CP), a known Nrf-2 inhibitor, on radiation induced cytotoxicity in cancer cells was studied using clonogenic, propidium iodide and MTT assays. Molecular mechanism of CP mediated radio-sensitization was investigated by using TEM, Western blotting, immunofluorescence assay and free iron measurement. Results: Treatment of A549 cells with CP prior to radiation exposure led to radio-sensitization. Combination of CP and ionizing radiation (IR) resulted in elevated ROS levels and reduced DNA damage repair. Transient over-expression of Nrf-2 abolished CP mediated radio-sensitization confirming the involvement of Nrf-2.Combination of CP with IR led to loss of mitochondrial membrane potential. Visualization of mitochondrial ultrastructure under TEM, revealed swollen and ruptured mitochondria along with loss of cristae which are strongly indicative of ferroptosis. Combination of CP with IR resulted in release of higher levels of free iron compared to those treated with either CP or radiation alone, which was abrogated by using an iron chelator, deferoxamine (DFO). Treatment of cells with DFO or Liproxstatin-1 (Ferroptosis inhibitor) prior to CP treatment abolished CP mediated radio-sensitization indicating a pertinent role of ferroptosis. Here, we show that pharmacologic inhibition of Nrf-2 leads to enhanced iron release, oxidative stress, reduced DNA damage repair and subsequent cell death through ferroptosis in radio-resistant human lung cancer cells. Conclusions: Our studies clearly demonstrate that Nrf-2 could be a potential target during radiotherapy of cancers that overexpress this pro-survival transcription factor.
| P-5.3 | | |
| Diselenodipropionic Acid Potentiates Radiation Toxicity in A549 Cells: Insight into Mechanism of Action of Combinatorial Treatment | | |
V. V. Gandhi, K. A. Gandhi1, M. Nayak2,3, A. Kunwar2,3
Department of Radiation Oncology, Thomas Jefferson University, Philadelphia, United States, 1Department of Clinical Pharmacology, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, Navi Mumbai, 2Radiation and Photochemistry Division, Bhabha Atomic Research Centre, 3Homi Bhabha National Institute, Anushaktinagar, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Diselenodipropionic acid (DSePA), is a multifunctional organoselenium compound. Recently, it is reported for chemotherapeutic activity against lung cancer cells. Since, chemotherapy and radiotherapy are used in combination for effective management of cancer in general, it is important to understand the response of combinatorial treatment of DSePA and radiation on viability of lung cancer cells and the associated mechanisms. The present study was conducted to address these queries. Materials and Methods: The effect of combinatorial treatment on the viability of A549 cells was studied by clonogenic and AnnexinV-PI assays. The mechanism of action was investigated by monitoring the DNA repair kinetics and expression of various signalling proteins related to DNA damage response and cell migration pathways by RT-PCR, flow cytometry and biochemical assay. The in vivo efficacy of combinatorial treatment was evaluated using A549 xenograft tumor in NOD-SCID mice. Results: The investigations revealed that combinatorial treatment (2 Gy γ-radiation + 5 μM DSePA) led although exhibited reductive environment (marked by decrease in ROS and increase of GSH/GSSG) at early time point (2 - 6 h post-radiation) induced significantly higher extent of apoptosis as compared to radiation and DSePA treatments separately. Mechanistically, it was established that DSePA significantly delayed the repair of radiation induced DNA damage by suppressing DNA repair (ATM/ DNAPKs/p53) pathway. Additionally, DSePA treatment also prevented radiation induced cell migration by suppressing Akt/G-CSF/epithelial mesenchymal transition (EMT) pathway. Finally, in vivo studies also confirmed the potency of combinatorial treatment (2 Gy x 4 and DSePA (0.25 mg/kg body weight daily)) for reducing the growth of A549-derived xenograft. Conclusions: In conclusion, DSePA showed synergism with radiation toxicity in A549 cells by inhibiting DNA repair and cell migration.
| P-5.4 | | |
| Radio-Sensitizing Efficacy of DHS against Multiple Pancreatic Ductal Carcinoma Cells | | |
Krishna Mohan Repaka1,2,3, Mrityunjay Tyagi2,3, Birija Sankar Patro2,3
1Radiopharmaceutical Program, Board of Radiation and Isotope Technology, 2Bio-Organic Division, Bhabha Atomic Research Centre, 3Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Despite several advances in therapeutic management of pancreatic cancer, the survival outcomes are abysmally low owing to late detection, extreme metastatic properties and drug resistance. Pancreatic cancer stem cells (PCSCs) contribute to the tumorigenesis, progression, and chemoresistance rendering insights into its malignancy. Previously, DHS (trans-4, 4'-dihydroxystilbene), a Resveratrol analogue reported10 times more potencyin killing various cancer cells. Therefore, in the current study we investigated the potential effect of combining DHS and standard IR in pancreatic cancer to eradicate both cancer cells and CSCs thereby enhancing the therapeutic efficacy.Initially, radio-sensitizing efficacy of DHS was studied employing clonogenic and sub-G1 assays. The impact of combination therapy on Cancer stem like cells (CSLC)was elucidated by side population assay while the self-renewal capacity of PCSCs was evaluated by its pancreatospheres generation ability. Furthermore, the specific nature of PCSCs was studied through flow cytometric analysis of its molecular markers viz., CD44/CD24.Combination effects on migration potentials of PDAC cells were studied using scratch assays and the EMT marker gene expressions through RT-PCR analysis. Involvement of transcription factor stargeting PCSCs was elucidated through western blot analysis.We observed that DHS at 5-10 μM is a potent radiosensitizer in vitro. Combination of DHS and radiation inhibits the proliferation of both non-stem and stem like cells. There was significant reduction in CSLC population (quantified as pancreatosphere) in combination of DHS and IR. We also observed decrease in invasion and metastasis properties of DHS and IR treated pancreatic cancer cells. Mechanistically, our results showed that combination of DHS and IR reduces the levels of STAT3, Twist and Slug, responsible for regulating both CSLC and metastasis properties of cancer cells. Importantly, the radiosensitizing effect of DHS is mediated through modulating NF-κB and STAT3 protein levels.In summary, the invitro results of this study firmly establishes that DHS in combination with standard IR can effectively target hard-to-treat, chemo- and radioresistant stem-like PDAC cells. The anti-cancer activity of the combination was found to be based on inducing down regulation of the NF-κB, STAT3-stemness gene, Slug and Snail pathway.
| P-5.5 | | |
| Radioprotective and Cytotoxic Functions of Nanoparticles, Metal Complexes and Alkene Derivative | | |
R. Gangabhagirathi1,2, S. T. Mehetre2,3
1Bio Organic Division, 2Homi Bhabha National Institute, 3Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Most synthetic molecules currently under study for clinical and commercial applications exhibit little to extreme undesirable consequences on the organisms in addition to specific effects. These effects are determined by the nature and dosage of the compound. Nanoparticles and metal complexes have been put to use here for utilizing their potential to kill eukaryotic and prokaryotic cells in vitro culture systems. In addition, radioprotective properties of an alkene derivative molecule (AD) have been investigated in depth here and subsequently one may visualize its potential to serve these requirements. Materials and Methods: Different biochemical assays, SOD/SDH activity and GSH were measured using spectrophoto/fluorometric methods.SDH was estimated using PMS/DCIP, electron acceptors. Prokaryotic and eukaryotic cell culture technique: includes antioxidant enzyme assay, cytotoxicity assays. Results: Metallic nanoparticles exerted cytotoxic action on eukaryotic cells. Different species of EColi showed increase in zone of inhibition with increasing concentration of metal. Significant cytotoxic effect of synthetic Platinum complex on eukaryotic cells was also observed. Following exposure of lipid bilayers preparations to different gamma radiation doses, different types of damages were observed. Presence of AD lead to significant inhibition of radiation induced changes. Peroxidised products reduced while loss of SDH/SOD activities were reduced. Conclusions: Cytotoxic potency of the nanoparticles increased gradually as the net content of metal component increased.Moreover, significant reduction in radiation induced loss of glutathione, activity of SDH and SOD was exerted by AD, emphasizing its radio-protecting role. Further exploration using other systems would get an insight into its versatile role and side effects if any. Because of their instant therapeutic effect, synthetic preparations are even now preferred over plant preparations.
| P-5.6 | | |
| Reactive Oxygen Species in Diseases: The Radiation Viewpoint | | |
Rajesh Kumar Gandhirajan
Department of Human Genetics, Faculty of Biomedical Sciences, Technology and Research, Sri Ramachandra University, Chennai, Tamil Nadu, India
E-mail: [email protected]
Reactive Oxygen and Nitrogen Species (RONS) in biological systems has been well elucidated over the past decades. Oxidation and reduction reactions in the tissues is essential for daily metabolic activities, but at the same time loss of balance in these systems lead to aging and development of various diseased states. Hence identification of disease and tissue specific molecular ROS determinants are critical to understand pathogenesis of the disease. ROS plays a predominant role in pathogenesis of acute inflammation, ischemia/reperfusion injuries, neurodegenerative disorders, atherosclerosis, diabetes and cancer. With the recent advances in understanding the molecular nature of diseases, ROS pathways need to be exploited for designing novel treatment strategies in multiple disease models. During sepsis, acute lung injury (ALI) results from activation of innate immune cells and endothelial cells by endotoxins, leading to systemic inflammation through proinflammatory cytokine overproduction, oxidative stress, and intracellular Ca2+ overload. Inhibition of membrane bound NADPH oxidase 2 (NOX2) or Stromal interaction molecule 1 (STIM1) inhibited endothelial damage both in vitro and in vivo. Furthermore, conditional endothelial-STIM1 knockout mice and pharmacological inhibitor BTP2 protects the mice from acute lung injury. ROS can also alter the structure and function of protein by post translation modifications. Due to evolutionarily conserved nature of cysteine residues, they are the excellent target of s-glutathionylation. Hippo pathway transcriptional co-activators YAP/TAZ differ in their presence of cysteine residues. S-glutathionylation of TAZ at C315 stabilizes the protein during oxidative stress, whereas YAP lacking any cysteine undergoes proteasomal degradation. TAZ s-glutathionylation, was observed in mouse kidneys following ischemia/reperfusion injury indicative of major role in tissue regeneration/remodelling following oxidative stress. However, cancer cells, display a differential redox status in comparison to healthy cells. This makes cancer vulnerable to pro-oxidant therapies. Ionizing radiation generates reactive oxygen and nitrogen species and selectively kills cancer cells in vitro and in vivo. However, a subset of cancer cells is resistant to ionizing radiation due to altered redox homeostasis. We highlight recent studies that contributes to radioresistance and strategies to circumvent in laboratory and the clinic. With these recent advances in identification of disease specific ROS pathways and tools to manipulate ROS locally and systemically, there is an exciting opportunity to develop novel radiation based pre-clinical therapies tailored to several disease states.
| P-5.7 | | |
| Selective Protection of Normal Lymphocytes Against Ionizing Radiation Induced Apoptosis by Withaferin A Via Activation of ERK/Nrf-2/HO-1 Axis | | |
Shivani R. Nandha1,2, H. N. Bhilwade1, Deepak Sharma1,2, Rahul Checker1,2
1Radiation Biology and Health Sciences Division, Bio-science Group, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected]
Introduction: Ionizing radiation (IR) finds application in diverse fields including medicine, agriculture, power generation, research etc. Occupational, unplanned, or accidental exposure to IR can have detrimental effects on the overall health of individuals. Further, IR-induced normal tissue toxicity is a major hurdle in effective anti-cancer radiotherapy. Thus, there is a need to develop efficacious radioprotectors. In this direction, we have studied the radioprotective potential of WithaferinA (WFA), the biologically active withanolide, obtained from the plant Withaniasomnifera. Materials and Methods: Radiation-induced cell death and apoptosis was assessed by PI, DNA fragmentation and caspase-3 assays. Mitochondrial ROS, total intracellular ROS levels and GSH levels were measured using fluorescent probes. In vivo radioprotective effects of WFA were studied in wild type and Nrf-2 knockout mice. Results: WFA offered selective protection to normal murine splenic lymphocytes and bone-marrow cellsbut not tumour cells, against radiation-induced apoptosis. WFA inhibited IR-induced DNA fragmentation, DNA damage and caspase-3 activity.WFA offered protection to lymphocytes against IR-induced apoptosis even when administered 2 hrs post-radiation exposure. WFA treatment reduced intracellular ROS and GSH levels and WFA mediated radioprotection was abrogated by thiol-based antioxidants highlighting the critical role of cellular thiols in the observed radioprotection.WFA administration to mice led to reduction in the frequency of WBI-induced mnPCEs with concomitant increase in the PCEs/NCEs ratio.WFA induced Nrf-2 pathway and the expression of cytoprotective genes such as catalase, hemeoxygenase-1, peroxiredoxin-2, superoxide dismutase via ERK activation.WFA was able to rescue mice against lethal dose of WBI (8.5 Gy), however, it failed to prevent WBI-induced mortality in Nrf-2 KO mice. These results provide evidence for the pivotal involvement of Nrf-2 in the radioprotective effects of WFA. Conclusion: WFA provides selective radioprotection to normal cells via modulation of cellular redox status and induction of the pro-survival and cytoprotective Nrf-2 pathway.
| P-5.8 | | |
| Protective Efficacy of Resveratrol against ROS Induced Cytotoxic and Genotoxic Effects in Human Blood Lymphocytes | | |
Sonia Velarsan, Jacob Moses, J. Vijayalakshmi
Department of Human Genetics, Sri Ramachandra Institute of Higher Education and Research, Chennai, Tamil Nadu, India
E-mail: [email protected]
Introduction: When human cells are exposed to ionizing radiation, Reactive Oxygen Species (ROS) are generated by radiolysis of water. This indirect method, which accounts for approximately 80% of DNA damages. The aim of the study was to evaluate the protective efficacy of resveratrol (RSV) mediated cytotoxic/genotoxic effect of ROS induced in PBL. Materials and Methods: Heparinized blood samples were collected from two healthy volunteers (n=2), exposed to various concentrations of RSV for two hours and then exposed to the chemotherapeutic drugs to induce ROS generation. MTT assay and DPPH assays were performed to measure the antioxidant efficacy of RSV. Micronuclei frequency was analysed to evaluate genotoxicity effects. Results: Drug treatment decreased the number of viable cells as indicated by the MTT assay, whereas when treated with RSV it showed an increase in the number of viable cells (p<0.05). In the DPPH assay, the inhibition percentage increased significantly as the RSV concentrations increased. The NDI showed significant decrease when treated with drugs but RSV treated cells upon exposure remained undamaged at large, an increase in the frequency of micronuclei when treated with drugs was found with declined steadily as the concentration of RSV increased. The NDCI showed a decrease in cells treated with drugs along with RSV which was found to be statistically significant (p<0.05). Therefore, the present study indicates that RSV has an antioxidant potential which can quench the ROS activity. Conclusions: The findings suggest that RSV, a natural antioxidant offers protection against ROS and do not exert any genotoxicity in human In future, RSV can be used as a radioprotective supplement to the humans exposed to ionizing radiation to reduce the frequency of DNA damage in normal cells.
Keywords: Antioxidant Assay, DP, Human peripheral blood lymphocytes, Micronucleus, Resveratrol, PH Assay
| P-5.9 | | |
| Application of Non-Ionizing Radiation Therapy in Combination with Suitable Photo-Sensitizers for Localized Cancer Treatment: Potential and Limitations | | |
Bidisha Maiti, Oyendrila Ghosh, Supratim Ghosh
Department of Anti-Cancer Drug Development and Chemotherapy, Chittaranjan National Cancer Institute, Kolkata, West Bangal, India
E-mail: [email protected]
Introduction: Non-ionizing radiation therapy or photo-therapy is one of the emerging approaches for treating localized malignancies in a selective manner. Radiations of visible or near-infrared (NIR) wavelength can produce reactive oxygen species (ROS) or hyperthermia in presence of appropriate sensitizers. Materials and Methods: One of the photo-sensitizers having absorbance maxima ~ 415 nm was synthesized by noncovalently assembling meso-tetra-4-pyridyl porphine with single stranded DNA (Porphyrin:DNA photo-sensitizer)and the other one, having absorbance maxima ~ 810 nm was synthesized by reducing gold chloride (HAuCl4), followed by conjugation of ss-DNA via Au-S bonding (GNR:DNA photo-sensitizer). Activity was evaluated in vitro as well as in vivo. Results: Porphyrin:DNA photo-sensitizer is capable of producing ROS and cell membrane lipid peroxidation, upon 420 nm laser irradiation. The therapeutic potential was evaluated in xenografted tumor, via intratumoral injection, followed by radiation with 420 nm laser. Results demonstrated that tumor volume was reduced for the short term but regrowth was observed.GNR:DNA photo-sensitizer is capable of producing notable hyperthermia, in vitro. In vivo evaluation on tumor xenografted mouse via tail vain injection was performed in combination with 808 nm laser irradiation. Results demonstrated that reduction in tumor volume is not significant. However, the treatment caused significant hematological toxicity. Only808 nm laser irradiation without photo-sensitizer also caused reduction in RBC as well as WBC count, most likely due to nonspecific ROS production. Molecule like curcumin, having ROS quenching as well as immune-boosting potential significantly reduced hematological toxicity and enhanced anti-tumor activity. Conclusions: Non-ionizing radiation in combination with appropriate sensitizer is capable of producing ROS or hyperthermia; those can eliminate malignant cells. Strategies for selective applications are required to achieve therapeutic efficacy. Combined application of appropriate natural products can reduce nonspecific toxicity and enhance anti-cancer activity, including enhancement of immune system for long term disease management.
| P-5.10 | | |
| Radioprotective Potential of Bacterial Supplementation against Gastrointestinal and Nervous System in a Mouse Model | | |
B. S. Venkidesh1, S. R. Shankar1, Rekha Koravadi Narasimhamurthy1, Satish Bola Sadashiva Rao1,2, Kamalesh Dattaram Mumbrekar1
1Department of Radiation Biology and Toxicology, Manipal School of Life Sciences, Manipal Academy of Higher Education, 2Manipal Academy of Higher Education, Manipal, Karnataka, India
E-mail: [email protected]
Introduction: Radiation resultant gut dysbiosis has been observed in cancer patients receiving pelvic radiotherapy. Nearly 90% of cancer patients receiving pelvic radiation exhibit gastrointestinal (GI) damage, most of which are linked to gut dysbiosis and include gastritis and bloody diarrhea.Furthermore, it has been established that an axis interconnects the gut and the brain, and microbial composition plays a significant role in this two-way communication. Probiotics have been demonstrated to effectively reduce oxidative stress in the intestine, protect the integrity of the mucosa, and prevent gastrointestinal toxicity. They have also shown to prevent neurotoxicity. Materials and Methods: Male adult C57BL/6 mice were given bacterial supplements containing 9 types of bacteria that are typically found in the gut orally for 14 daysand a single dose of 4 Gy whole-body irradiation given on the 7th day. Histological analysis of the brain and intestine was performed to measure gastrointestinal and neuronal damage. Results: Bacterial supplementation reduced the neuronal death and neuroinflammation in the hippocampus and had a radioprotective effect on intestinal architecture. Conclusions: The bacterial supplementation could protect the neuron in the hippocampal region and decrease neuroinflammation by reducing the number of microglial cells. Therefore, bacterial supplementation can be used to mitigate the radiotherapy induced adverse effect on gastrointestinal tract and the nervous system.
Funding and Acknowledgements: This work was supported by the MAHE intra mural grant (2019).The authors would like to thank Manipal Academy of Higher Education (MAHE), for support and infrastructure facilities. VBS would like to thank MAHE, Manipal, for the Dr.T.M.A Pai fellowship.
| Topic 6: Radiologial Imaging and Nuclear Medicine | | |
| P-6.1 | | |
| Novel DNA-Based Aptamers against Thyroid Hormones (T4 and T3) are Being Developed with the Goal of Using Them in Radiopharmaceuticals | | |
Chandan Kumar, Rani Gnanasekar1, Vijay B. Kadwad1
Radiopharmaceuticals Division, Bhabha Atomic Research Centre, Mumbai, 1Board of Radiation and Isotope Technology Vashi, Navi Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Aptamers are nucleic acids akin to antibodies for target recognition. It became popular in recent decades due to the ease of chemical synthesis, modification, and manipulation. Additionally, the aptamer has great chemical stability, low molecular weight (10–14 kDa), and minimal immunogenicity. These characteristics make aptamer an interesting and effective probe for the recognition of molecular signatures for diagnosis and therapy. Materials and Methods: The SELEX process was employed to select aptamers against the thyroid hormones thyroxine (T4) and triiodothyronine (T3). In a nutshell, DNA aptamers were screened against T4 and T3 for eight rounds of selection. The final DNA pool was sequenced using NGS. Out of 2 million sequences, only 20 sequences were chosen after analysis of the NGS data. Selected sequences were synthesized and confirmed for precise binding with Iodine-125-labeled T4, T3, and T2 (diiodothyronine) molecules. Results: It was discovered that just a few of the 20 sequences demonstrated substantial binding. Additionally, it was noted that sequences preferentially binding to T4 do not bind to T3, and vice versa. Conclusions: The distinctive DNA sequences that specifically bind T4 and T3 molecules may be exploited to develop radioimmunoassays or other appropriate assays for monitoring thyroid diseases
| P-6.2 | | |
| Toxicological Profiling of DHPN and Identification of Its Target Organ Using Nuclear Imaging in Wistar Rats | | |
Chandrakala Gholve, Yogita Shete, Sutapa Rakshit, Savita Kulkarni
Radiation Medicine Centre, BARC, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: N-Bis (2-hydroxypropyl) nitrosamine is a nitrosamine chemically genotoxic and carcinogenic in nature. Evaluation of DHPN's effect on vital organs was done by histopathology and nuclear imaging technique to find the effects at the cellular level. Materials and Methods: A total of 18 male (270-280 g) 8-week-old healthy inbred Wistar strain rats were grouped into 2 groups (DHPN treatment, n=12) and (control, n=6). Group 1 was injected with DHPN, at a dose rate of 83 mg/animal by intraperitoneal route, fortnightly, for 110 days, whereas Group 2 received only saline. Blood samples were collected for analysis and all vital organs were collected for histopathology. 18F-FDG PET-CT and 99mTechnetium pertechnetate scintigraphy imaging was performed for target organ. Results: At the end of 16 weeks, the body weight in the DHPN-treated group (532.6 ± 33.1 g) was significantly lower than in the control group (582.5 ± 17.67 g) (P< 0.05). Haematology showed lowered hemogram with an increase in neutrophils in treated animals. The gross morphology of all vital organs was normal, with no discoloration of colour, texture, and weight difference in the organs. Microscopic lesions showed degeneration, preneoplastic and neoplastic alteration in the brain, spleen, kidney, liver, lung and thyroid. Liver and kidney showed mild to moderate degrative changes with basophilic foci indicative of preneoplastic alternation. Broncho-alveolar carcinoma with pneumonic changes was seen in the lung. Thyroid showed hyperplastic to neoplastic changes in all the animals with distance metastasis. 18F-FDG and 99mTc-pertechnetate scans indicate thyroid carcinoma. Minute alternation can be detected by imaging techniques in a lesser number of animals. Conclusions: Our study reports that DHPN promotes thyroid proliferation and increases susceptibility to thyroid cancer followed by lung, kidney and liver. Hence, the thyroid gland was the target organ identified by nuclear imaging techniques in the identification of initial changes and further supported by histopathology.
| P-6.3 | | |
| Indigenous Designing of Second Stage Semi-Automated Solid Target Irradiation Assembly Compatible with PETtrace-800 | | |
Kanchan Kushwaha, Tejinder P. Sabharwal1, Sunil Kamble1, Kavindra Pathak1, N. K. Prasad1, Manish Kumar1, Savita Kulkarni
Radiation Medicine Centre, BARC, 1Electromagnetic Application and Instrumentation Division, BARC, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Medical cyclotron facility (PETtrace-800) at Radiation Medicine Centre, BARC is capable to provide 16.5 MeV proton beam. The facility has only liquid and gas targets. A first stage solid target assembly was developed and tested indigenously. Materials and Methods: In the first stage design, the attenuation disc serves the purpose to attenuates the energy of incident proton beam, from 16.5 MeV to required value, depending on its thickness. In this work, design of the first stage solid target assembly has been modified with the provision of changeable attenuation disc such that the attenuation disc of different thickness can be mounted in to the solid target assembly without dissembling it. Results: The first stage design is completely modified with an additional cross roller guide assembly that can be manually pushed in or out. This guide assembly holds the changeable attenuation disc sandwiched between two sturdy plates. To replace the attenuation disc simply the guide tray is pushed out without disturbing the assembly and the attenuation disc is changed and pushed back in. The heat generated in attenuation disc will be removed by chilled water. This arrangement enables the production of more than one radioisotope in the same target assembly, without disassembling the target assembly. Conclusion: Based on this design the fabrication will be carried out. This second stage solid target assembly will be more useful in research & development of new radioisotopes and corresponding radiopharmaceuticals.
| P-6.4 | | |
| Utility of striatal to occipital ratios for differentiating between various causes of Parkinsonism and healthy and non-dopamine deficient symptomatic subjects from Parkinsonian patients using hybrid imaging using F18-DOPA | | |
Keerti Sitani, M. Sandhya
1National Institute of Mental Health and Neuro Sciences, Bengaluru, Karnataka, India
E-mail: [email protected]
Introduction: F18-DOPA can be used to differentiate non-dopamine deficient symptomatic subjects from Parkinsonian patients and various etiologies from one another which otherwise can be challenging. Striatal-to-occipital ratio's (SOR) are currently used for the same. Materials and Methods: 60 patients were retrospectively analyzed, of which 36 were idiopathic Parkinson's disease, 5 atypical Parkinson's disease and 20 normal on imaging. All of them underwent PET/MRI post F18-DOPA injection, 90 minutes after injection. Visual assessment was done and point region of interest was drawn at the least radiotracer concentration, the regions chosen were bilateral occipital cortex, caudate head, anterior putamen, posterior putamen, periaqueductal gray matter and red nucleus. Ratios were taken for bilateral caudate head, anterior putamen, posterior putamen, periaqueductal gray and red nucleus to occipital. Results: Of 36 with idiopathic Parkinson's disease, 15 had left sided involvement, 4 with bilateral (2 showing left> right and 2 showing right> left) and 16 showing right sided involvement, cases with right sided involvement had a right posterior putamen: occipital ratio<left and vice versa. 8 out of them also had concurrent caudate hypometabolism (3 right and 3 left).5 with atypical Parkinson's disease, two having unilateral right caudate nucleus hypometabolism (right caudate nucleus: occipital ratio < left)one having bilateral caudate nucleus hypometabolism and two having left sided caudate nucleus involvement (left caudate nucleus : occipital ratio< right).20 patients turned out to be normal on scan, out of them 5 were suspected of drug induced Parkinsonism, 9 suspected of essential tremors, 3 presented with slowness of gait and three presented with nonspecific symptoms. The tests of significance applied were student paired t-test to compare the groups with normal and IPD and the p values came out to be <0.05. Conclusions: It can be concluded that, caudate nucleus and putamen: occipital ratios can be used for differentiating between various causes of Parkinsonism.
| P-6.5 | | |
| Bio-Distribution of 1-[18F] Fluoro-Isopropyl-L-Tyrosine (1-[18F]FPT) and Its Comparison with [18F]FET as a PET Imaging Agent for Brain Tumor | | |
N. Lakshminarayanan, Abhinav Bajpai, Avik Chakraborthy, Yogita Pawar, P. R. Chaudhary1, Bhabhani Mohanty1, Savita Kulkarni
Radiation Medicine Centre, Tata Memorial Hospital, BARC, Mumbai, 1Advanced Centre for Treatment Research and Education in Cancer, Navi Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: [18F]labeled amino acids are used as PET imaging agent especially for brain tumors. The most commonly used PET imaging agent for this purpose is [18F]FET, which is a fluoroethyl analogue of L-tyrosine. In present study, we have studied bio-distribution and microPET imaging of 1-[18F]FPT and compared its uptake with [18F]FETin C57BL/6 mice induced with B16-F10 melanoma. Materials and Methods: Precursor synthesis and radiosynthesis of 1-[18F] FPT and [18F]FET were performed as per the methodologies depicted in our earlier publications (1,2). C57BL/6 mice were procured from BARC Animal House, after the approval from BAEC. About 105 cells of B16-F10 melanoma cell lines were injected in the shoulder of C57BL/6 mice. Tumor uptake studies were performed after the tumor was developed to a palpable size (5 mm diameter). Uptake studies were performed in various time points viz. 30, 60, 90 and 120 min. Results:The chemicals and radiochemical yields of both 1-[18F]FPT and [18F]FET were good and comparable. Tumor uptake studies of 1-[18F]FPT showed tumor to brain ratio of 2.4% and maximum tumor uptake was observed at 60 min. Tumor to brain ratio of [18F]FET was found to be 2.6% with the maximum uptake at 60 min. The tumor to brain ratio was found to be fairly constant till 120 min, with maximum clearance through urine. Conclusions: Tumor uptake studies showed good tumor to brain ratio of 2.4 % and maximum tumor uptake at 60 min. The results are comparable and in good agreement with the most commonly used [18F]FET. With the above results it can be concluded that 1-[18F]FPT can be a potential PET brain tumor imaging agent.
| P-6.6 | | |
| A Liposomal Nanoformulation for Immunotargeting and Monitoring Drug Delivery by SPECT Imaging | | |
Shishu Kant Suman1,2, Archana Mukherjee1,2
1Radiopharmaceuticals Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Smart nanoformulations for targeted delivery and quantitation of drug delivered at the target site are desired for the management of cancers. Herein, we report the formulation and preclinical evaluation of 99mTc labeled liposomes loaded with doxorubicin and decorated with antibodies for targeting HER2 receptor overexpressing cancers. Materials and Methods: Liposomes were prepared by ethanol injection method. Active pH gradient method was employed for loading drug, doxorubicin into the liposomes. Doxorubicin loaded liposomes (Lip-Dox) were conjugated withHER2 receptor targeting monoclonal antibody trastuzumab (Lip-Ab-Dox) and radiolabeled with 99mTc. The stability of 99mTc-Lip-Ab-Dox and 99mTc-Lip-Dox was studied. For ascertaining the affinity of 99mTc-Lip-Ab-Dox against HER2 receptor, internalization studies were carried out inHER2 positive SK-BR-3, SK-OV-3 and HER2 negative MDA-MB-231 cell lines. In vivo comparative evaluation of 99mTc-Lip-Ab-Dox and 99mTc-Lip-Dox in SK-BR-3 xenograft bearing SCID mice was also performed. Results: Liposomes of size 84.2 ± 2.9 nm could be prepared and were remotely loaded with Dox (encapsulation efficiency : 26 ± 6%). 99mTc labelling of Lip-Ab-Dox and Lip-Dox was achieved with 100% radiochemical purity (RCP) and the radioformulations were found to be stable in saline and serum at 37°C and 4°C up to 24 h. Significantly high internalization of 99mTc-Lip-Ab-Dox compared to 99mTc-Lip-Dox was observed in HER2 positive cells. Confocal microscopy data corroborated the findings of internalization assay. In HER2 negative cells, insignificant uptake and internalization was observed with both the formulations. 99mTc-Lip-Ab-Dox showed two-fold higher uptake inSK-BR-3tumor in SCID mice at 6 h and 24 h compared to99mTc-Lip-Dox as ascertained by SPECT imaging and biodistribution studies. Conclusions: A smart liposomal nanoformulation for immunotargeting and monitoring of drug delivery of chemotherapeutic drugs using SPECT imaging was successfully formulated. Encouraging results were observed in preclinical studies with the 99mTc labeled immunotargeted liposomal formulation.
| P-6.7 | | |
| Nuclear Imaging Technique to Study the Radiation Response in Cell Organelles of Salivary Gland | | |
Y. H. Shete, U. S. Bhartiya, J. Joseph, Y. S. Raut, S. Awasare, D. Singh1, A. A. Damle, S. P. Kulkarni
Radiation Medicine Centre, Bhabha Atomic Research Centre, 1National Institute for Research in Reproductive Health, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Different organs have different sensitivities to ionizing radiation. Cell and cell organelles show reversible and irreversible structural and functional changes in response to ionizing radiation and to the amount of dose delivered at site. Salivary gland is the most sensitive organ for ionizing radiation followed by bone marrow. The radiation response in cell organelles was studied in rabbit parotid salivary glands by Nuclear imaging and the histopathological changes were assessed to understand the morphological cell death and cell organelles effect at ultrastructure level. Materials and Methods: The study comprised of male New Zealand Rabbit (n=6). Rabbits were exposed to 131Iodine radiation (1GBq) as an internal dose. Scintigraphy imaging was done for salivary gland by 99mTechnetium scan. Parotid salivary gland was collected for histology, special staining and electron microscopy analysis. Results: Rabbit salivary gland basal scan showed reduced uptake in treated animals as compared to control. Histopathology showed multifocal areas of lipomatosis and basophilic cellular degeneration in acini. Variable basophilic intracellular granules were seen in treated animals as compared to control. The signs of radiation-induced damage were different for different cell organelle reflected by basophilic degeneration and infiltration of connective tissue within cells. was further supported by a special stain for calcium deposition. The changes seen in mitochondria, endoplasmic reticulum, Golgi-complex and lysosome were considered as direct intracellular effects of irradiation. These findings were further corroborated by the calcium infiltration, increased glycolipids, damaged aggregated nucleic acids and proteins as indicated by histology. Conclusion: Radiation response in cell organelles was variable and Nuclear Imaging technique was useful in imaging the functional damage. The altered uptake and cell morphological changes induced by radiation were further supported and concurred by histopathology and TEM analysis.
| Topic 7. Clinical and Space Radiation Biology | | |
| P-7.1 | | |
| Identification of Gene Features for Prediction of Primary Site in CUP-NETs Through Machine Learning | | |
M. K. Padwal1,2, S. Basu2,3, B. Basu 1,2
1Molecular Biology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, 3Radiation Medicine Centre, Bhabha Atomic Research Centre, Tata Memorial Hospital Annexe, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Neuroendocrine tumors (NETs) are rare tumors derived from the neuroendocrine cells and are often detected at the metastasis stage. The primary site identification is essential for the clinical management and treatment of the cancers. RNA sequencing datasets from the cancerous tissues hold the secrets that are relevant to the genesis and the dynamics of the disease. Machine learning facilitates comprehension of the high-dimensional complex RNA-seq data and recovery of most relevant gene features. Materials and Methods: RNA-seq datasets from the pancreatic (pNETs) and small intestinal (siNETs) were retrieved from the NCBI database (n = 186). The libraries were checked for read quality and aligned to human genome (hg38) after adaptors removal. Reads were quantified using STAR, normalized using the VST and sequencing batch corrected using SVA. Further, the samples were partitioned into train (70%) and test (30%) datasets. Gene features associated with the primary site were selected using mRMRe algorithm. Performance of selected features was evaluated using five different machine learning algorithms (RF, XGB, GBM, SVM and k-NN). Differential expression analyses were performed using DESeq2. Results: A set of 12 genes were identified as important features for prediction of primary site. Three best performing models (RF, XGB and SVM) showed 100% accuracy in the training dataset and 81.25% accuracy in the test dataset. Splitting of the genes into smaller feature sets improved the prediction accuracy to 100% as evaluated by 2 RF, 8 GBM, and 3 XGB models. All identified gene features were differentially expressed between the NETs of pancreatic and small intestinal origin, with tissue specific expression reported for a few features. Conclusions: A set of 13 gene features could accurately discriminate pNETs from siNETs, based on RNA-seq data. Identified gene features could complement clinical management of NETs patients with unknown primary.
| P-7.2 | | |
| Stereotactic Radiosurgery for Vestibular Schwannomas with Cyberknife: A Single Institution Experience, Preliminary Analysis | | |
B. Patneedi, D. Saran1, S. Roy2, S. Rai3, S. Vangipuram4, S. Shrivastava3
Assam Cancer Care Foundation, Guwahati, Assam, 1Medanta Medicity, New Delhi,2Therapist Cancer Center, 3HCG Cancer Center, Mumbai, Maharashtra, 4Apollo Hospitals, Chennai, Tamil Nadu, India
E-mail: [email protected]
Introduction: Vestibular schwannoma is benign tumors of vestibular nerve which usually presents with progressive hearing loss. The treatment options include observation, surgery and radiosurgery. Cyberknife is a dedicated robotic radiosurgery platform with real time tumor tracking and flexible planning. In this study we evaluated Dosimetric parameters of critical neurological structures, target volume coverage during Cyberknife radiosurgery for vestibular schwannoma. Materials and Methods: Thirteen patients of vestibular schwannomas who underwent Stereotactic radiosurgery with Cyberknife from 2019-2022 were included in this study. All patients underwent planning Brain CT scans plain and contrast and a contrast enhanced Brain MRI scan. A dose of 12-14Gy in one fraction was prescribed to the GTV. Target coverage, conformity index, gradient index, and doses tobrainstem, cochlea, whole brain were calculated. Statistical analysis was done using SPSS software version 28. Results: Of thirteen patients, fifty four percent were males and 46% were females with median age of 55 years. Fifty-four percent patients were Koos Grade I and Grade II and III were seen in 38.7% & 7.7% respectively. The left and right sided lesions were equally distributed. The prescribed dose range was 12-14Gy in single fraction. The mean target coverage 94.10% (91-98%), conformity index 1.19 (1.05-1.5), gradient index 4.4 (2.99-8.7). The mean tumor volume was 2.02cc (0.19-6.1cc). Brainstem Dmax was 10.8Gy (1.49-13.83), ipsilateral temporal lobe Dmax 11Gy (6.4-15.5), Cochlea mean dose was 4.7Gy (3.9-9.7). All the patients are in follow up and are controlled till last follow up. Conclusion: Our single institution preliminary analysis of Cyberknife radiosurgery for vestibular schwannoma shows promising dosimetric and clinical results. It validates that Cyberknife Stereotactic radiosurgery is safe and amenable treatment option owing to the non-invasive nature of treatment and good clinical outcomes. Further long-term follow up and large cohort size is required to evaluate disease control and side effects in our set-up.
| P-7.3 | | |
| Regional Variation of Cancer Incidence in Nepal | | |
Rudra Prasad Khanal, Ishwar Koirala, Kanchan P. Adhikari1
Department of Physics, Tribhuvan University, 1NAMS, Bir Hospital, Tribhuvan University, Kirtipur, Nepal
E-mail: [email protected]
Introduction: Non-communicable disease, such as cancer has spread all over the world for some last decades. Even though the advancement of technology, it has been a burden for every country. In the context of Nepal, 10 to 15 thousand new cancer incidences are being registered in different hospitals for treatment. Since the date of starting nuclear medicine at Bir Hospital in 1998, cancer patients are getting treatment regularly. According to the data of the population-based cancer registry, approximately 60% of the population having a middle-class income is being affected by cancer in Nepal. Materials and Methods: The study is aimed to find out the particular place where the population density of new cancer incidence is highest in Nepal and to inform the concerned regulatory body that is working for cancer screening and early detection for the proper treatment from the beginning. To find out the places where the population density of new cancer incidence is high, all the data of cancer patients were collected from five different renowned hospitals and also from the population-based cancer registry center and then analyzed the data. The history of cancer patients was studied from 2003 to 2020, but here the data are analyzed from 2015 to 2020 only, to find the latest trend of cancer incidence. Results: In the five major hospitals in Nepal, the total new cancer incidence was 61783 from 2015 to 2020. Out of those 34617 were female and 27176 were male. This research shows that female cancer patients were more every year. In the male, lung cancer patients were more than cancer of other organs, but in females, the number of breast cancer patients was greatest. The age-adjusted mortality rate for males in Kathmandu valley was 36.3 and for females was 27.0 per 100,000 population. The cancer incidence and mortality rate were slightly lesser in other districts of Nepal. This rate increased with the increase in the age of people. Over 60 years, cancer incidence and mortality rates have been found to increase rapidly. Conclusion: This research supports conducting the program of cancer screening and early detection at Kathmandu valley with high priority and then Morang, Rukum, SSDM, etc. to control cancer.
| P-7.4 | | |
| Organ Doses and Effective Dose to Astronauts at Low Earth Orbit and Its Variation Due to Spacecraft Wall Materials | | |
Sandipan Dawn1,2, A. K. Bakshi1,2, B. K. Sapra1,2
1Radiological Physics and Advisory Division, BARC, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
As a part of 'Gaganyaan' missions, India is planning to send astronauts to Low Earth Orbits (LEO) (~400 km a.s.l). Astronauts at LEO are exposed to significant radiation field due to the Galactic Cosmic Rays (GCR) consisting of 87% proton, 12% 4He, and 1% heavy high energy ions (high LET) with peak energies around 1 GeV/u. We have used FLUKA and HZETRAN space radiation transport codes with Badhwar-O'Neil 2014 model to evaluate the effect of Aluminium (Al) shielding on energy spectra of GCR and organ doses, effective dose equivalents received by the astronauts. Organ doses and total effective dose have been evaluated using Computerised Anatomical Male (CAM) phantom. It is observed that the fluence of low LET ions such as proton, 7Li, 4He are not attenuated by the spacecraft wall Al shielding. In addition, additional low energy peaks (1-10 MeV) appear in the fluence spectra due to spallation and Intra-Nuclear Cascade (INC) reactions. Additional pions, muons and neutrons are also produced which can contribute to the additional doses. Without any effective shielding, effective dose received by the astronauts during a moderate solar activity is 0.5 mSv/day. Skin and retina doses can be up to 0.8 mSv/day whereas max lung doses are around 0.5 mSv/day. For a high shielding thickness of 7.5 cm, effective dose is 0.37 mSv/day. But the skin dose reduces to 0.32 mSv/day. Doses to other sensitive organs such as lung, brain, kidney also varies within 0.3 to 0.4 mSv/day. The result shows that spacecraft aluminium wall is not very effective shielding against GCR. On an average, astronauts receive 0.3-0.5 mSv/day effective doses during space missions with moderate solar activities. Organs like skin, retina, and eye lens can receive a high dose equivalent of 0.7- 0.8 mS/day.
| P-7.5 | | |
| Split X-Jaw Techniques of Volumetric Modulated Arc Radiotherapy in Nasopharyngeal Cancer: A Comparative Dosimetric Study | | |
Jyotiman Nath, Gautam Sarma1, Shachindra Goswami, Shashi Bhushan Sharma, Pranjal Goswami, Apurba Kumar Kalita
Department of Radiation Oncology, Dr B Borooah Cancer Institute, 1Department of Radiation Oncology, All India Institute of Medical Sciences, Guwahati, Assam, India
E-mail: [email protected]
Introduction: The multi-leaf collimators on the Varian linear accelerator move on a carriage with a maximum leaf span of 15 cm. Therefore, treatment of larger planning target volumes like in nasopharyngeal cancer with traditional open and limited Jaw technique yields compromised dose distribution. The current study aims to compare the split X-Jaw planning technique of VMAT with the traditional open and limited jaw techniques of VAMT in Nasopharyngeal carcinoma treatment. Materials and Methods: Computed tomography data sets of 10 nasopharynx cancer patients were enrolled for the study. Only PTVs requiring a field size larger than 18 cm in the x-jaw position were considered. For each case, three separate treatment plans were generated viz. open, limited, and split x-jaw planning techniques with similar planning objectives. Results: The target dose coverage with split x-jaw VMAT was superior to both open and limited jaw planning techniques with a statistically significant difference in the intermediate dose PTV (PTV59.4), p<0.05. Comparable results were obtained in terms of OAR sparing in all the techniques, however, the dose to the spinal cord and larynx was significantly lower with the split technique (p<0.05). Conclusions: The split x-jaw planning technique of VMAT can be adapted for larger PTVs requiring an x-jaw of more than 15 cm. The only concern with this technique is the increased MU.
| P-7.6 | | |
| Development of High Sensitive Quantification Method for Cardiovascular Marker “C-Reactive Protein” | | |
Tanhaji S. Ghodke, Vijay Kadwad1, N. Karunakara, K. Bhasker Shenoy
Department of Applied Zoology/CARRT, Mangalore University, Konaje, Karnataka, 1Board of Radiation and Isotope Technology, Navi Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: In 1930, Tillet and Francis while working with Streptococcus pneumonia, accidentally discovered a protein named as C-reactive protein (CRP) which is an important inflammatory marker. Since last couple of decades its evolved as potent cardiovascular disease marker. CRP normal level in healthy population is below 1 mg/L. However, in cardiovascular diseases, moderate risk level is reported less than 2-3 mg/L and more than 3mg/Lis increasing the chances of the cardiovascular diseases. Conventional agglutination, turbidometry and nephelometry are not sensitive to quantify the CRP. Hence, development of high sensitive radioimmunoassay method is proposed to fulfil current need. Materials and Methods: Polyclonal anti-CRP antibody raised in rabbit and CRP for standard preparation Bovine serum albumin (BSA) andNa 125I from BARC. Radiolabelling of 125I to CRP was carried out by using minor modification in chloramine-T method. Radioiodination yield, specific activity, and radiochemical purity were analysed by the paper electrophoresis methods. % Binding for assay standards was obtained by counts per minutes (CPM) using gamma counter. Results:Iodination yield of tracer was ~81.4%. Along with this specific activity was calculated and found to be 18 ±1μCi/μg. Radiochemical purity was analysed by the paper electrophoresis for every week for two months and it is found to be ~98%. Table 1 presents that the standard curve was calculated with the standards concentration (X-axis) vs %B/B0 (Y-axis). Conclusions: A standard curve is obtained and lyophilised standards were found to be stable for 1 years. Radiolabelled chemical purity was <98 % for the two months and Prepared CRP standards are ranged from 150-4800ng/mL.
| Topic 8. Biological Dosimetry and Radiation Biomarker | | |
| P-8.1 | | |
| Molecular Spectrum of Somatic Mutations at HPRT Locus in the Normal and High-Level Natural Radiation Areas of Kerala Coast, India | | |
Anila Gopinathan1,2, P. R. Vivek Kumar1, G. Jaikrishan1, Birajalaxmi Das2,3
1Low Level Radiation Research Laboratory, Low Level Radiation Research Section, Radiation Biology Health Sciences Division, Bio-Science Group, Bhabha Atomic Research Centre, Kollam, Kerala, 2Homi Bhabha National Institute, 3LLRRS, RB and HSD, Bio-Science Group, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected]
Introduction: High background radiation areas of Kerala in the south west coast of India with natural deposits of monazite beach sand provide opportunity to study the effect of chronic low dose ionizing radiation exposure in humans. Analysis of molecular nature of somatic mutations at the reporter gene locus such as HPRT (Hypoxanthine guanine phosphoribosyl transferase) deletion spectrum may have the potential as a radiation biomarker. Our study compares the molecular spectrum of HPRT mutations between individuals from normal and high-level natural radiation areas (N & HLNRA) of Kerala. Materials and Methods: A T-lymphocyte cloning assay was carried out in peripheral blood samples of 10 individuals (5 NLNRA and 5 HLNRA) to isolate HPRT mutant lymphocyte colonies. Out of the total 69 HPRT mutant colonies studied, 31 and 38 mutant colonies belonged to NLNRA and HLNRA, respectively. A multiplex-PCR of the genomic DNA from mutant colonies resulted in the simultaneous amplification of exons 1-9 of the HPRT gene. Results: Mutants were classified as (i) total deletion (mutants with no amplification of exons 1-9 of the HPRT gene), (ii) terminal deletion (deletion mutation involving either exons 1or), (iii) intragenic deletion (deletion mutation involving exons 2-8) (iv) band shift (change in the size of exon amplification product), (v) intact pattern or point mutation (mutants with exons 1-9 of expected size, but may have changes too small to be detected by m-PCR). Comparison of the deletion mutation spectrum in N & HLNRA individuals revealed 9.7% total deletions in NLNRA and 7.9% in HLNRA, terminal deletion was 6.5% in NLNRA, and none was observed in HLNRA. Band shift mutations were 3.2% in NLNRA and 5.3% in HLNRA, and intragenic deletions constituted 5.3% in HLNRA individuals, whereas none in NLNRA. Conclusions: Percentage of various deletion mutations observed at the HPRT locus in NLNRA and HLNRA individuals was statistically comparable.
| P-8.2 | | |
| Establishment of Dose Response Relationship for X-Rays Induced Micronucleus Formation in Human Lymphocytes, Employing Giemsa and Acridine Orange Staining | | |
Jacob Moses1, Satish Srinivasan2, Rajesh Chaurasia1,3, Sonia Velarsan1,3, J. Vijayalakshmi1
1Department of Human Genetics, Sri Ramachandra Institute of Higher Education and Research,2Department of Radiation Oncology, Sri Ramachandra Medical Centre,Chennai, Tamil Nadu, 3Radiological Physics and Advisory Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: The CBMN assay in lymphocytes is one of the best standardized and validated techniques for individual/population, radiation dose assessment. This method has been proposed as an alternative to the dicentric chromosome assay in biodosimetry as it is efficient and easy to conduct. Nevertheless, for application as a bio-dosimetry tool in large-scale radiological accidents, the CBMN assay needs further strategies to speed up the analysis. Materials and Methods: Blood-samples of two volunteers were obtained and irradiated with X-ray (598 MU/min)(0-5Gy), to construct dose-response curves employing manual scoring mode by Giemsa and Acridine orange staining. Results: For the construction of dose-response curves linear quadratic model was used, Y = αD + βD2 +C with the 95% confidence limit using Dose Estimate Software (version 5.2). The equations obtained for the two donors of Giemsa was Y = 0.0737± (0.006) D + 0.0189± (0.0016) D2 + 0.0071± (0.0017) and which iscompared to the two donors of AO was found to be Y = 0.1389± (0.0147) D + 0.0236± (0.0037) D2 + 0.0124± (0.0042) and the Correlation coefficient was performed using IBM- SPSS (version 26.0) between the two yields of MN and was found to be R2 = 0.997 suggesting a very strong statistical corelation. A paired t-test was also performed between the yields of MN, the result p = 0.020 showed statistical significance indicating the agreement between the curves. It was found that the yield of MN obtained using AO was higher over Giemsa. Conclusion: Specificity and sensitivity of MN, detection and quantification increased with the use of AO over Giemsa. It could be due to high specificity and fluorescence intensity of AO. Also, analysis time was significantly reduced.
| P-8.3 | | |
| Influence of Inflammation on Gene Expression: From a Biodosimetry Point-of-View | | |
K. Nandhini, Teena Koshy, K. Satish Srinivas1, P. M. Venkata Sai2, Venkateswarlu Raavi3, Emanuel Bhaskar4, Swathy Moorthy4, P. Venkatachalam
Departments of Human Genetics, 1Radiation Oncology, 2Radiology and Imaging Sciences and 4General Medicine, Sri Ramachandra Institute of Higher Education and Research (Deemed to be University), Chennai, Tamil Nadu, 3Department of Cell Biology and Molecular Genetics, Sri Devaraj Urs Academy of Higher Education and Research (Deemed to be University), Kolar, Karnataka, India
E-mail: [email protected]
Introduction: Development of a gene signature has been configured as devices suitable for low-cost, point-of-care measurements making it an appealing strategy for rapid radiation triage and dosimetry. One of the main difficulties using gene expression as a biomarker of exposure is its highly dynamic and transient nature. In view of the significance of acute and chronic inflammation on certain gene expression and its relevance on biodosimetry, we have investigated CDKN1A and FDXR expression in the regional population and in-vitro as well as in known inflammation condition to establish a baseline expression level and adopt it for meaningful and reliable radiological triage. Materials and Methods: Blood samples from healthy volunteers (N=12) were collected and processed, of which two samples were exposed to various concentrations of lipopolysaccharide LPS (0.5, 8.0μL/mL) to induce acute inflammation. Sample from subjects with diabetics/hypertension as chronic inflammation conditions (N=22) were collected. RNA was isolated by TRIzol method and quantification of gene expression was performed using TaqMan gene expression assay kit. ΔΔCT method was adopted for the relative gene expression. Results: The acute inflammation induced by LPS under in-vitro, increased expression of CDKN1A (ΔΔCT=2.81-16.62) and FDXR (ΔΔCT=4.27-132.06) in a concentration dependent manner. However, at higher concentrations (8μg/ml), an attenuated fold-change when compared to that of unexposed control sample. Similar to the in-vitro, subjects with chronic inflammation also were found to have 0.56- and 0.97-fold increase in the expression of CDKN1A and FDXR respectively in inflammatory patients. Conclusions: The study results suggest that the inflammatory condition needs to be considered at times of radiation emergency and triage of exposed populations based on gene expression biomarker.
| P-8.4 | | |
| Correlation of Tracks Formed in CR39 and Induction of γH2AX in A549 Cells After Alpha Particle Irradiation | | |
R.Vasumathy1,2, Jyothish Babu3, Sanjay Shinde1, B. N. Pandey1,2
1Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, 3Electromagnetic Application and Instrumentation Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: A computer-controlled, automated, alpha-particle (Am241) irradiator (BARC Bio Alpha) was designed and developed for radiobiological experiments. Present study was aimed to correlate the tracks formed through CR39 dosimetry and induction of γH2AX as DNA double strand marker in A549 cells after alpha particle irradiation at various distances from the exit window of the alpha irradiator. Materials and Methods: SRIM code analysis was carried out to calculate the energy of alpha particle in air at different heights from the exit window. CR-39 discs were irradiated for a time period of 40 min at different distances from the exit window (0.05 cm to 3.5 cm with an increment of 0.5 cm) using known height spacers. After irradiation, CR-39 detectors were chemically etched to visualise the alpha particle tracks using microscopy and to count using Image J software. Human lung adenocarcinoma A549 cells were irradiated for same the time and heights followed by determination of γH2AX expression analysis after 30 min of irradiation by flow cytometry. Results: The 3.86 MeV energy of the alpha particle at 0.05 cm distance from the exit window was reduced to 0.197 MeV at 3.5 cm as determined by SRIM code analysis. Number of tracks formed in CR39 was decreased from 13946 ± 1890.6 (at 0.05 cm) to 11.6 ± 1.8 (at 3.5 cm) with increase in distance from the exit window. In alpha particle irradiated A549 cells, there was an increase in γH2AX positive cells in irradiated than that of control cells. There was a significant decrease (~40 %) in γH2AX positive cells with increase in distance from the exit window from 0.05 cm to 3.5 cm. Pearson correlation coefficient was found to be 0.96 suggesting a strong and positive correlation between number of CR39 tracks and γH2AX positive cells in alpha particle irradiated cells. However, γH2AX showed significant sensitivity to determine double strand break at lower energy than CR39 detectors.
Conclusions: The study showed correlation and sensitivity of physical and biological techniques after alpha particle irradiation.
| P-8.5 | | |
| Is Co-Localization of γH2AX and 53BP1 Follow 1:1 Correlation: Investigation in Human Lymphocytes Challenged with 60Co-γ-Rays | | |
Ranjana S. Pathak1,2, R. K. Chaurasia2,3, Anjana Goel1, S. C. Tripathi1
1Department of Biotechnology, Institute of Applied Science and Humanities, GLA University, Mathura, Uttar Pradesh,2Radiological Physics and Advisory Division, Bhbaha Atomic Research Centre, 3Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction:Since long DSB repair proteins γH2AX and 53BP1, are considered as sensitive marker of DSB quantification and dose estimation. Presently, co-localization of g2AX and 53BP1, are considered as sensitive marker of DSB quantification and dose estimation. Presently, co-localization of found to be 0.96 suggesting a strong and positive correlation between number of CR39 tracks and at 0ction of the dose. Present study has demonstrated foci build up and decay kinetics of γH2AX and 53BP1 along their co-localization up to 4 hours after irradiation. Materials and Methods: Human PBMCs were isolated and irradiated with 1 Gy 60Co-γ-rays, and incubated in ambient conditions up to 4 hours. Intermittently, PBMCs were fixed and immunofluorescence stained to visualize γH2AX and 53BP1 foci, along their co-localization. Quantifications were carried out, in 3D mode, using Leica confocal microscope. Results: γH2AX and 53BP1 foci decay kinetics has extensively been explored by earlier researchers but the available data demonstrates ambiguity on their co-localization, in post irradiation repair kinetics. In this work, we have measured γH2AX and 53BP1 foci build up and their decay kinetics, up to 4 hours, after irradiation. It was observed that foci build up follow rapid exponential growth and decay follows exponential decay pattern. Kinetics of co-localization was measured in the same time frame, it was observed that co-localization of foci were not following 1:1 correlation. Since, γH2AX and 53BP1 foci were not same in number, and their decay kinetics too were not following exactly same pattern of foci buildup and decay, this could be the reason for not following 1:1 co-localization pattern. Conclusions: Co-localization of γH2AX and 53BP1 foci, in foci build up and decay kinetics, is not following complete (100%) colocalization. Difference in number of foci formed and their foci build up and decay kinetics could be the probable reason for this.
| P-8.6 | | |
| Exploring The Role of miRNAs as Radiomodifyers | | |
Sandeep Mallya, Soumya Amara, Kamalesh D. Mumbrekar1
Department of Bioinformatics, Manipal School of Life Sciences, Manipal Academy of Higher Education, Manipal,1Department of Radiation Biology and Toxicology, Manipal School of Life Sciences, Manipal Academy of Higher Education, Manipal, Karnataka, India
E-mail: [email protected]
Introduction: DNA repair mechanism is essential to maintain the integrity of the genome. Any aberration of this mechanism leads to the increased incidence of various pathological conditions including cancer. Mutation in or downregulation of the DNA repair genes can lead to accumulation of chromosomal aberrations. These mutations may affect both the oncogenes as well as the tumour suppressing genes, resulting in cancer. Therefore, the expression level of DNA repair genes is critical both for cancer therapy and tumorigenesis. MicroRNAs (miRNAs) typically play a role in decreased gene expression, and it is seen that miRNAs are misexpressed in cancer cells. Materials and Methods: We explored the TCGA data for mRNA and miRNA expression. miRNA candidates were obtained by identifying the target DNA repair genes that were differentially expressed in several cancers, as well as identifying the miRNAs that targeted multiple DNA repair genes. Results: We identified the miRNA associated DNA repair genes and checked for their corelation, clinical response and their survival. Conclusions: Overexpression of miRNAs could downregulate DNA repair genes and may sensitize cancer cells to radiation and chemotherapeutic agents. Alternatively, tumours that deplete the expression of these miRNAs may develop resistance to conventional cancer therapy.
Acknowledgement: We thank the Manipal School of Life Sciences, MAHE for providing the infrastructure necessary for the study.
| P-8.7 | | |
| Evaluation of Micronuclei Frequency in Patients Receiving Therapeutic 131I Exposure | | |
U. S. Bhartiya, Kamaldeep, N. N. Bhat1, G. Malhotra, S. Jaiswal2, S. P. Kulkarni, A. K. Tyagi
Radiation Medicine Centre, BARC, 1Environmental and Bio-Dosimetry Section, RP and AD, Bhabha Atomic Research Centre, 2Department of Biotechnology, Institute of Science, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: 131I therapy is widely used in patients of differentiated thyroid carcinoma for the ablation of remnant thyroid tissue post thyroidectomy as well as metastasis. These patients are in hypothyroid state with consequent reduction in renal clearance of 131I thereby increasing the extent of whole body exposure. Materials and Methods: Present study was carried out to assess the DNA damage due to the therapeutic 131I exposure at 48h by measuring micronuclei (MN) frequency in peripheral blood lymphocytes (PBLs) in patients of differentiated thyroid cancer. The study group consisted of 11 patients of differentiated thyroid carcinoma.In this study, peripheral blood is considered as representation of whole body but not necessarily the representation of tissues which are likely to accumulate iodine such as thyroid or region of tissues with metastasized nodules. The blood samples of the patients were collected pre and post 131I therapy (1.7-7.4 GBq) and processed for PBL MN frequency by cytokinesis-block MN assay. Results: 131I exposed patients demonstrated significant (<0.001) increase in the MN frequency at 48h after therapeutic 131I exposure compared to their basal MN index (Pre 9.6±4.0 /1000BN cells Post 19.3± 6.6 /1000BN cells). The observed increase in the micronuclei index did not correlate with the administered 131I activity. Further, significant individual variation of MN frequency is observed for a given activity administered. These observations point out influence of possible clinical and patient specific conditions such as thyroid uptake/activity at the time of administration, which may influence the whole body dose. Conclusions: The study has revealed significant DNA damage in PBLs as indicated by increased PBL MN frequency post 131I therapy, which can be considered as indicator of whole body dose. However marked individual variations were observed in the DNA damage response to 131I therapy.
| Topic 9. Non-targeted Effects of Radiation and Genomic Instability | | |
| P-9.1 | | |
| A Pilot Study to Evaluate Radiation-Induced Adaptive Response and Its Implication on Cancer Radiation Therapy | | |
T. A. Aishwarya, K. Sathish Srinivas1, P. Venkatachalam
Department of Human Genetics, Faculty of Biomedical Sciences and Technology, Sri Ramachandra Institute of Higher Education and Research (Deemed to be University), 1Department of Radiation Oncology, SRMC and RI, Sri Ramachandra Institute of Higher Education and Research (Deemed to be University), Chennai, Tamil Nadu, India
E-mail: [email protected]
Introduction: Radiation-induced adaptive response (RIAR) is a phenomenon where prior exposure to low priming dose (PD) reduces biological effects of subsequent exposure to high challenging dose of radiation. While, existence of RIAR has been demonstrated in cell lines and animal models with radiation protection purpose, translation into clinical scenario is emerging but remains uncertain. Materials and Methods: Blood samples from healthy donors (n=3) were irradiated in-vitro to 0.1 Gy (dose received during PET/CT imaging (PD)), 2 Gy (dose fraction received during radiation therapy (CD)), then to 0.1 Gy+2 Gy X-rays (PD+CD). Blood samples were also collected from cancer patients (n=6) before and after completion of PET/CT procedure; a portion was further irradiated ex-vivo to 2 Gy. Cell proliferation was measured as Mitotic Index (MI) and induced damages as chromosomal aberration, micronucleus formation. The phenomenon was also examined upon exposure to Bleomycin(PD=80, CD=140, PD+CD=80+140 μg/ml) using trypan blue & MTT assay. Results: MI was 6.96% and 2% in the healthy subjects and cancer patients respectively. Mean aberration frequency obtained was 0.316±0.047 and 0.76±0.23 and 0.142±0.011 and 0.141±0.016 for CA and MN for respective subjects. Among study subjects, four showed trend of RIAR measured using CA (lower in PD+CD exposed lymphocytes than exposed to CD alone). Contrarily, one subject observed aberration frequency between two irradiation conditions (after PD+CD, CD alone) did not show any difference. Whereas an attenuated MN formation, increased MI were observed in samples exposed to PD+CD than CD alone in all subjects but were non-significant; lymphocytes exposed to BLM(PD+CD) showed attenuated MN formation than CD treatment alone. Conclusions: Ex-vivo X-irradiation to PBL from cancer patients with clinically relevant dose demonstrated the existence of RIAR and suggests that this phenomenon may have an impact on RT outcome. Confirmation requires additional samples and data from tumour tissue for its translation application.
| P-9.2 | | |
| Multi-Omics Approach Evaluating Pathways after Radiation Exposure | | |
Kiran Maan1,2, Radhika Bakhshi1, Poonam Rana2
1Shaheed Rajguru College of Applied Sciences for Women, University of Delhi, 2Institute of Nuclear Medicine and Allied Sciences-DRDO, New Delhi, India
E-mail: [email protected]
Introduction: The event of an intentional or nuclear accident results in exposure to a mass population. The use of a multi-omics approach may be able to address the pathophysiology involved after radiation exposure and can aid in triage management. years have seen a significant increase in the use of omics techniques like transcriptomics and metabolomics to help comprehend underlying pathways and identify biomarkers. In this study, we employed a combined multi-omics strategy based on transcriptome analysis, together with metabolomics and lipidomics of blood samples from murine exposed to total body irradiation. Materials and Methods: C57BL6 mice (n = 8) were exposed to 7.5 Gy dose (Gamma radiation; 1 Gy/min dose-rate) and blood samples were collected 24 h post-irradiation. Enrichment and mRNA library were prepared for Illumina followed by quality assessment and processing. LC-MS acquisition was performed using BeH Amide columnfor metabolomics and CSH C18 column for lipidomics with ESI probe in both modes using UHPLC-5600+tripletof. Data were preprocessed and multivariate analysis was done using Metaboanalyst software. Univariate analysis and joint-pathway analysis of deregulated genes (DEGs) along with metabolites/lipids were performed either using metaboanalyst or BioPAN software. Results: Transcriptomics and metabolomics data displayed demarcation between the 7.5 Gy group from controls using multivariate analysis. Deregulated amino acids, various PC, PE, and carnitine were observed along with many deregulated genes (e.g., NOS, ELOVL5, ELOVL6, FADS2, Hmgcs2, Oxct2a). Joint pathway analysis and STITCH interaction showed radiation exposure resulted in changes in immunological processes, histidine metabolism, carbohydrate, lipid metabolism, amino acid, nucleotide metabolism, and fatty acid metabolism. Elicited immune response was also observed as DEGs were enriched in immunological functions by Gene Ontology. Conclusions: The quest for important insights into the crosstalk between various biomolecules is being driven by the integration of multi-omics.
| P-9.3 | | |
| Effects of Radiation in Combination with Immune Adjuvant CpG-ODN in Non-Targeted Mouse Fibrosarcoma Tumors | | |
Pooja K. Melwani1,2, Hansa D. Yadav1, Murali MS Balla1,2, B. N. Pandey1,2
1Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Radiotherapy remains a keystone for the treatment of several types of tumors. It has been shown in clinical cases and animal models that ionizing radiation apart from killing directly irradiated cells affects growth of tumor cells at distant sites (abscopal effect). This effect is mediated by radiation-induced enhanced immunogenic cell death and activation of immune system at irradiated sites. Use of immune adjuvants such as CpG-ODN has been shown to elicit radiotherapy induced tumor regression in animal models, however its effect on distant tumors is unknown. Materials and Methods: Fibrosarcoma models in immune competent BALB/c mice were generated by transplanting WEHI-164 cells in both the hind legs of mice. On 8th day, only one of tumor was irradiated locally while rest of the body was kept shielded. CpG-ODN (synthetic oligodeoxynucleotides containing unmethylated CpG motifs) was injected intratumorally in irradiated tumor. Tumor growth in both the legs was monitored. Mice were sacrificed on day 22, serum and tumor samples were collected for cytokine and immuno-histochemistry (IHC) analysis, respectively. Results: Irradiation resulted in significant decrease in tumor volume in directly irradiated tumors. However, significant decrease in size of shielded tumors was also observed at 5 Gy than control and 2 Gy group. Serum analysis showed significant increase in levels of IFN-γ and TNF-α, while IHC showed increase in lymphocyte infiltration in 5 Gy irradiated group compared to control suggesting an increase in inflammation and immune response in the irradiated mice. Treatment with 5 Gy + ODN resulted in further decrease in tumor growth in treated and shielded legs. IHC also showed decreased vasculature and more apoptosis/necrosis in tumors obtained from both legs in this group. Interestingly, 3 out of 8 mice in 5 Gy + ODN group showed complete tumor regression upto 90 days. Further challenge with tumor cells in the treated mice led to rejection of tumors, suggesting development of anti-tumor immunity in these mice. Conclusions: Radiation along with immune adjuvants CpG-ODN offers promising therapeutic strategy in fibrosarcoma mouse model.
| P-9.4 | | |
| Radiation Induced Changes in DNA Methylation and Its Association with Leukemia | | |
T. Haritwal, P. K. Agrawala
Department of Radiation Genetics and Epigenetics, Institute of Nuclear Medicine and Allied Sciences, DRDO, New Delhi, India
E-mail: [email protected]
Introduction: Secondary cancers caused by radiation exposure during radiotherapy in the course of cancer treatment have emerged as a clinically serious problem. hematopoietic stem cells (HSCs) have the possibility of turning into cancer when subjected to whole-body radiation therapy during cancer treatment.Radiation exposure has genotoxic effects but in addition it has epigenotoxic effects too.DNA methylation is necessary for appropriate regulation of the expression of genetic material. Trichostatin A has shown histone deacetylase inhibitor activity in various cancerous cell lines.In this study, we investigated the radiation induced changes in DNA methylation, its association with cancer and effects of Trichostatin A on DNA methylation. Materials and Methods: For that purpose, we have exposed C57BL/6J male mice to 3 Gy of gamma radiation and Trichostatin A was administered 1 and 24h post irradiation as radiomitigator. Then methylated DNA was assessed in blood, bone marrow and Spleen 24h post irradiation directly by quantifying total methylated DNA, DNMT activity and histone H4 modifications. Results: Hypomethylation was observed in all these tissues, moreover the DNMT activity and histone H4 modification has shown the similar results. There may be a genetic connection between radiation-induced hypomethylation and susceptibility to acute myeloid leukaemia. Conclusion: At present, there is no experimental evidence that the modifications in DNA methylation during the development of mouse hemopoietic cells also apply to other tissues and tumours in mice or humans.These findings suggest that radiation-induced DNA hypomethylation influences the progression of leukemogenesis but it further needs to be investigated.
| Topic 10. Radionuclide Effects and Decorporation | | |
| P-10.1 | | |
| Response of Chryseobacterium sp. PMSZPI to Uranium Exposure | | |
Devanshi Khare1,2, Celin Acharya1,2
1Molecular Biology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected]
Heavy metals and radionuclides dispersed in sub-surface environment resulting from anthropogenic activities cause a challenge to organisms living in different habitats. Some organisms have adapted to these conditions by modulating their biology. One such bacterium, Chryseobacterium sp. PMSZPI, isolated from uranium (U) mine, could tolerate high concentrations of uranium and other heavy metals. Mapping the cellular and molecular responses that is instigated by toxic metal exposure will facilitate the mechanistic understanding of adaptive responses of PMSZPI. Genome sequencing of ~ 3.2 Mb of Chryseobacterium sp. PMSZPI and comparative genomics with uranium sensitive strains revealed the presence of genes predominantly involved in metal efflux by transporters, phosphatases involved in precipitation, gliding motility and biofilm formation. To ascertain the pathways involved in conferring tolerance to uranium and other heavy metals in this organism, we performed unbiased RNA-seq analysis by paired-end sequencing on Illumina Novaseq 6000 platform with challenging concentration of uranium (300 μM) under short term and long-term exposure conditions. Within 30 min of exposure, a total of (483) significantly differentially expressed genes (DEGs) were observed (~288 upregulated and ~195 downregulated transcripts). After 24 h, ~ (330) DEGs were observed (~199 upregulated and ~131 downregulated). Our transcriptome data indicated antioxidant response in PMSZPI by induction in expression of genes including catalase, thioredoxins, superoxidase dismutase and thiol peroxidase. Many membrane proteins like ABC transporters and P-type ATPases were upregulated possibly to enhance the efflux of heavy metals. DNA binding response regulators were upregulated under U challenge which might be involved in activation or repression of their target genes on receiving metabolic/environmental signals. We observed downregulation of genes encoding 30S and 50S ribosomal proteins suggesting utilization of energy for stress response instead of metabolic processes. Overall, we attempted to emphasize the significance of stress and adaptive responses of PMSZPI at sublethal concentration of uranium.
| P-10.2 | | |
| Thorium Decorporation Efficacy of a Novel Rationally Designed Multidentatecatecholamide Derivative for Management of Internal Contamination | | |
Manjoor Ali, Sourav K. Das, Ritesh Ruhela1, D. K. Singh1, B. N. Pandey, Amit Kumar
Radiation Biology and Health Sciences Division, 1Materials Processing and Corrosion Engineering Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]; [email protected]
Introduction: For management of internal contamination in human with radioactive tetravalent actinides (e.g. Thorium, Plutonium), DTPA (diethylenetriammine pentaacetate) is the only FDA-approved agent. However it has several major limitations (efficacy from organs, toxicity, etc). In the present study, a multidentatecatecholamide derivative (TDA) has been rationally designed, synthesized and evaluated in cell and animal models for Thorium decorporation. Materials and Methods: Thorium decorporation efficacy of TDA was determined using various in-vitro/ex-vivo assays viz. hemolysis assay, RBC binding assay, whole blood decorporation assay and human liver cell binding assay. Finally, animal experiments were performed using metabolic cages to assess the Th decorporation potential of TDA. Results: TDA was found to prevent Th-induced hemolysis in a concentration-dependent manner. Typically, it was found to be ~2-10folds more effective than the DTPA to prevent hemolysis, depending on the treatment conditions. In-vitro Th decorporation assays using human erythrocytes, liver and lung cells showed significantly higher potential of TDA than the DTPA at equimolar concentration. Further, ICP-MS analysis of mice tissues/organs showed that TDA treatment (30-120 μmole/kg) was more effective in removing Th from major vital organs (such as liver, spleen, skeleton) as compare to only Th or Th+DTPA treatment. Serum biochemical analysis revealed that TDA treatment significantly mitigated Th-induced alteration in liver function markers. Conclusions: Our data suggested significant potential of TDA for thorium decorporation from major target organs and mitigation of thorium toxicity.
| P-10.3 | | |
| Mechanism of Thorium-232 Toxicity in Human Lung Cells: Role of DNA Damage Response, Redox Alteration and HSPs | | |
Sourav Kumar Das1,2, Manjoor Ali1, Neena G. Shetake1, B. N. Pandey1,2, Amit Kumar1,2
1Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: sour[email protected], [email protected], [email protected]
Introduction: Inhalation represents the most prevalent route of exposure with Thorium-232 compounds (Th-nitrate/dioxide) in occupational scenario. Our results highlight the mechanism of Th response in normal human alveolar epithelial cells (WI26), exposed to Th-nitrate or colloidal Th-dioxide. Materials and Methods: Effects of Th-nitrate/dioxide (1–100 μg/ml, 24–72 h) on WI26 were assessed in terms of changes in morphology, proliferation, plasma membrane integrity (LDH leakage) and mitochondrial activity (MTT assay). Mechanism of Th internalization and associated intracellular changes were studied by TEM, IF, ICP-MS and other cell biology assays. Results: Cell count, MTT and LDH leakage assays showed ThO2was more toxic than the Th-nitrate to WI26 cells. IC50 (72h) of ThO2 and Th-nitrate determined by cell count was found to be 5 μg/ml and 25 μg/ml, respectively. However IC50 (72h) by MTT assay was found to be 10 μg/ml (ThO2) and 25 μg/ml (Th-nitrate). ThO2 treated cells showed ~3 fold higher leakage of LDH than the Th-nitrate-treated cells. TEM and IF analysis suggested that Th-dioxide followed a clathrin/caveolin- mediated endocytosis, however, membrane perforation seemed to be the mode of Th internalization in cells exposed to Th-nitrate. ThO2 treatment resulted in significant increase in expression of early endosomal antigen (EEA) marker, suggesting the involvement of endocytosis mechanism. ICP-MS analysis showed a significantly higher uptake of Th in cells treated with Th-dioxide than with Th-nitrate. Our results highlighted Th-induced ROS and their possible site of generation, which was found to induce gamma-H2AX foci.The levels of HSP70/90 were also found to be affected by Th-dioxide/ Th-nitrate. Conclusion: Present study revealed the effect of chemical form (oxide/nitrate) on the magnitude of Th-toxicity and in the mechanism of its internalization (endocytosis/perforation). ATM (DDR mediator)and HSP90 were found to play a key role in Th-induced cytotoxicity in normal human alveolar epithelial cells (WI26).
| Topic 11. Radiation Effects on Micro-organisms and Plants | | |
| P-11.1 | | |
| A Snapshot at Protein Complexes engaged in Double Strand Break Repair in Deinococcus Radiodurans | | |
Aman Kumar Ujaoney1, Mahesh Kumar Padwal1,2, Bhakti Basu1,2
1Molecular Biology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Deinococcus radiodurans is well known for its DNA repair proficiency. The survival of this bacterium is governed by an efficient macromolecular repair processes including DNA. DNA metabolism which includes its repair following any stress can occur by multiple pathways some of which are unique to this bacterium. RecA, DdrA, Ssb and DdrB (bait proteins) are important enzymes and accessory proteins of many such pathways which mostly function as transient or stable complexes with their partner proteins during DNA repair processes. Materials and Methods: Polyclonal antibodies were generated against DNA repair bait proteins. The 6 kGy irradiated Deinococcal cells were chemically cross-linked invivo with Lomants reagent, mid-way through post-irradiation recovery. Polyclonal antibodies against Ssb, RecA, DdrA and DdrB were used to immuno-precipitate cross-linked protein complexes. The interacting proteins in the interactomes were identified by LC-MS/MS analyses. Bioinformatics was applied to extract significant interactions. Binary interactions were confirmed by pull-down assays. Results and Conclusions: This work focused on protein complexes associated with the bait proteins Ssb, RecA, DdrA and DdrB. The invivo cross-linked interactomes were overlapping and comprised of 399 proteins belonging to various functional categories including DNA repair,transcription, cell division and oxidative stress alleviation. Binary interactions were confirmed for proteins involved in homology-dependent DSB repair. A total of 27 binary interactions were confirmed in our study, of which 25 interactions were reported for the first time. Unique aspects of DNA repair discovered in this comprehensive study are (a) Ssb protein acts as DNA repair protein sponge as it showed maximum interactions, (b) DdrB protein acts at the intersect of strand annealing and strand exchange pathways, and (c) DNA repair proteins interact with cell division proteins. Overall, the study unravels novel ways in which various DNA repair pathways cross-talk and contribute to the radiophily of D. radiodurans.
| P-11.2 | | |
| Response of Deinococcus Radiodurans to Directly Damaging Genotoxic Stressors is Regulated through RDR Regulon | | |
Narasimha Anaganti1,2, Bhakti Basu1,2
1Molecular Biology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: The Deinococcus radiodurans bacterium is highly resistantto very high dosesof ionizing radiation, UV and many genotoxic chemical reagents. The organism responds to the radiation stress through the coordinated expression of genes belonging to Radiation and Desiccation Response (RDR) regulon operated through cis-acting sequence RDRM (Radiation Desiccation Response Motif), trans-acting repressor DdrO and protease PprI (also called IrrE). The role of the RDR regulon in other DNA damaging stresses such as UV rays, chemical mutagens, DNA intercalating agents, oxidative stress, metal stress, etc. is explored by monitoring activation of key RDRM-promoters tagged to GFP reporter. Materials and Methods: The bacterial strains used were E. coli (DH5α) and D. radioduransR1. The cloning and over expression were done using standard molecular biology techniques. The D. radiodurans clones carrying pKG with different deinococcal RDRM-promoters (PgyrB, PddrB and P1143) were subjected to various genotoxic stresses and the promoter activity was monitored through Gfp fluorescence for 6h during post stress. DdrO abundance in different stresses was detected by immunoblotting using anti-DdrO antibodies. Results: The present study evaluated whether RDR regulon controls the response of D. radiodurans to various DNA damaging stressors, which it is resistant to, such as UV rays, mitomycin C (MMC), methyl methanesulfonate (MMS), ethidiumbromide (EtBr), ROS generating stresses (H2O2, MV and metal ions). Activation of 3 RDR regulon genes (ddrB, gyrB and DR1143) was studied by taggingtheir promoter sequences with a highly sensitive GFP reporter. Here we demonstrated that all the DNAdamaging stressors elicited activation of RDR regulon of D. radiodurans in a dose-dependent and RDRM-/IrrE-dependent manner. However, ROS-mediated indirect effects [induced by hydrogen peroxide (H2O2),methyl viologen (MV), heavy metal/metalloid (zinc or tellurite), etc.] did not activate RDR regulon. The level of RDRM-promoter activation was inversely proportional to the cellular abundance of repressor DdrO. Conclusions: The stresses that caused direct DNA damage or distortion in DNA structure were found to be regulated through RDR regulon. The ROS generating stresses that cause indirect damage to DNA did not elicit RDR regulon genes in D. radiodurans.
| P-11.3 | | |
| Natural Transformation Specific DrDprA protein Function as Molecular Switch for RecA Dependent and Independent Repair in the Radiation-Resistant Bacterium Deinococcus Radiodurans | | |
Dhirendra Kumar Sharma1,2, Hari Sharan Misra1,2, Yogendra S. Rajpurohit1,2
1Homi Bhabha National Institute, 2Molecular Biology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
The uptake of environmental DNA (eDNA) by certain bacteria and its integration into their genome creates genetic diversity and new phenotypes. DNA processing protein A (DprA) is part of a multiprotein complex and facilitates natural transformation (NT) phenotype in most bacteria. The Deinococcus radiodurans, an extremely radioresistant bacterium, is efficient in NT, and its genome encodes nearly all the components of the natural competence complex. The DprA of this bacterium (DrDprA) has characteristics of DprA proteins as known in other bacteria. Further, the mechanisms underlying its roles in the transformation of eDNA and regulation of RecA dependent and independent repair in D. radiodurans were investigated. DrDprA is found to be a unique domain organization implicating some unique functions compared with DprA of other bacteria. In vitro studies showed that the purified recombinant DrDprA binds to both ssDNA and dsDNA with nearly equal affinity and protects ssDNA from nucleolytic degradation. DrDprA showed a strong interaction with DrRecA indicating its role in RecA catalyzed functions in vivo. Mutational studies identified amino acid residues responsible for its oligomerization, interaction with DrRecA, and DNA binding characteristics of DrDprA. Further, we demonstrated that both oligomerization and DNA binding properties of DrDprA are integral to its support in DrRecA catalyzed strand exchange reaction (SER) in vitro. The D. radiodurans cells lacking the dprA gene showed DNA double-strand break (DSB) repair defect and radiation sensitivity at higher doses of ɤ-radiation. The DSB repair kinetics with thymine analog suggested that deletion of dprA gene from D. radiodurans genome promote the DSB repair by single-strand annealing (SSA) instead of extended synthesis-dependent strand annealing (ESDSA) repair. Together, these results suggested that DrDprA is largely structurally conserved with DprA homologs but showed some unique structure-function features to support DrRecA functions. Moreover, observed in vivo data suggested that DrDprA has a key role in selection of DSB repair scheme in the repair of damaged genome. A comprehensive study discussing DrDprA biochemical characterization and DSB repair roles in gamma radiation resistance and DSB repair will be presented.
| P-11.4 | | |
| Microrhizome Induction in Turmeric Using irradiation for Improved Yield and Curcumin Content | | |
Suchita N. Kamble, Ramesh K. Satdive, Sudhir Singh, C. Jagadeesh1, Suresh N. Manvatkar1, Chandrakant Salunkhe1
Plant Biotechnology and Secondary Metabolites Section, Nuclear Agriculture and Biotechnology Division,1Landscape and Cosmetics Maintenance Section, A and SE Division, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Turmeric (Curcuma longa L.) is an economically and medicinally important spice crop. Curcumin, a major bioactive compound present in its rhizome imparts therapeutic properties to turmeric. Due to huge demand, development of novel turmeric cultivars with high yield and curcumin content is essential. In vitro mutagenesis has proved to be a valuable method for improving vegetatively propagated crops like turmeric. The present study was conducted to induce microrhizome in turmeric shoot cultures using gamma radiation and field assessment of these plantlets was carried out for improved yield and curcumin content. Materials and Methods: In vitro shoot cultures of turmeric were exposed to different doses of gamma radiation (0,5,10,15,20,25,30,35,40,45 and 50Gy). Further, shoot cultures of turmeric were exposed with the optimized gamma-irradiation dose. A detailed comparative field evaluation of microrhizome-producing plants, non-irradiated in vitro plants and conventional rhizome-raised plants was carried out for improvedyield and curcumin content. Results: In vitro turmeric shoots exposed at 20-25Gy produced maximum microrhizomes after 30 days of irradiation. Shoots producing microrhizomes were superior in height, produced more leaves with enlarged size compared to the control. Field assessment performance of microrhizome producing plants in terms of yield and curcumin content was better than the other two. These plants produced higher number of mother rhizomes(8-10) per plant with average yield of 2.1kg/plant, non-irradiated plants produced (5-6) mother rhizomes with average yield 1.2 kg/plant and rhizome raised plants produced (5-7) mother rhizomes with an average yield of 1.5 kg /plant respectively. Curcumin content was also more in microrhizomes producing plants as compared to the other two. Conclusions: Microrhizome induction and growth rate of in vitro plantlets producing microrhizomes indicate that gamma irradiation at 20-25Gy effectively improves productivity in turmeric. Better recovery of curcumin vis-à -vis rhizome weight was the main advantage of in vitro microrhizome induction.
| P-11.5 | | |
| Screening of Emerging Pathogens of Microbial Safety Relevance From Sea Food and Chicken and Assessment of their Sensitivity Towards Gamma Radiation | | |
H. D. Khade
Food Technology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Sea food export has increased significantly in current decades and this has been becoming a main source of foreign exchange for the country. However stringent hygienic practices have to be implemented for domestic consumption as well international trade. Presence of new emerging pathogens in the supply chain could be introduced as process contamination. This may lead to microbial spoilage as well as health safety issues to the consumers. Processing by gamma radiation can mitigate those concerns to a significant level. Current R and D work aims to screen emerging pathogens of microbial safety relevance and their control by gamma radiation processing. Materials and Methods: Fresh chicken carcasses and sea food (Prawns and fish) samples were collected from various retail outlets of Mumbai city. Enrichment of bacterial pathogens was carried out in selective broth containing antibiotics mainly Vancomycin, Cephatoxime and Nalidixic acid. After enrichment loop full of broth was inoculated on selective media containing antibiotics. The isolates obtained after enrichment were Subjected to 16S rRNA gene amplification and sequencing was carried out for identification. Radiation sensitivity in terms of D10 value for all emerging pathogens was elucidated. Results: Two isolates showed 86.01 and 85.78% similarity with Myroid pelogicus and Myroid albus, respectively when sequence homology search was carried out. Other isolates showed similarity of 98.32, 93.0% and 92.31 % with Proteus terrae, Enterobacterial spps and Pseudomonas jaseenji respectively. D10 value of Myroid pelogicus /Myroid albus Proteus terrae, Enterobacterial spps and Pseudomonas jaseenji was 85, 125, 85 and 72 Gy respectively. Conclusion: Gamma radiation can be applied to ensure safety of sea food in context of presence of these various emerging pathogens. Prominent emerging pathogenic microorganisms was observed in sea food and chicken as confirmed by sequencing. Processing by gamma radiation was found to be effective in hygienizing the samples and ensure their microbial safety.
| P-11.6 | | |
| Proteomic Profiling and Metabolomic Profiling of Radiodurans Response to Thioredoxin Reductase Inhibitor and Ionizing Radiation Treatment | | |
M. Sudharsan, N. Rajendra Prasad
Department of Biochemistry and Biotechnology, Annamalai University, Chidambaram, Tamil Nadu, India
E-mail: [email protected]
Introduction: The gram-positive bacterium Deinococcus radiodurans can survive under extreme conditions like ionizing radiation, ultraviolet, vacuum, and desiccation. This study illustrates the importance of the thioredoxin reductase (TrxR) antioxidant system in the survival of D. radiodurans against ionization radiation. The thioredoxin reductase/thioredoxin (TrxR/Trx) system is a major thiol-based antioxidant system it maintains cellular redox homeostasis. Materials and Methods: 10 μM of TrxR inhibitors such as ebselen, auranofin, and epigallocatechin gallate (EGCG) were used for the inhibition of the TrxR antioxidant system. We have analyzed the binding affinity for TrxR inhibitor towards TrxR protein, TrxR activity, intracellular redox ratio, cell survival, membrane damage, antioxidant enzymes (catalase and SOD) activity, generation of intracellular ROS (Reactive oxygen species), and protein carbonylation level. Also, we have analyzed the level of DNA damage and expression of DNA damage repair genes such as DdrA, IrrE, and PprA. Moreover, we have analyzed the intracellular metabolomic and proteomic changes upon TrxR inhibitors (10 μM) and ionizing radiation (4 kGy) treatment. Results: The results show that TrxR inhibitors pre-treatment affects the survival against ionizing radiation (4 kGy) by altering the activities of antioxidant enzymes and elevating intracellular ROS and protein carbonylation levels. The level of DNA damage has increased in the TrxR inhibitor and radiation treated group compared to control and radiation control. The TrxR inhibitors increase membrane damage towards ionizing radiation by altering the fatty acid and lipid composition of the cell wall. The metabolite analysis demonstrates disturbance in the cellular redox status affects the metabolites synthesis mainly intermediates of carbohydrate and fatty acid metabolism. The proteomic result confers the changes in the cellular metabolism and also TrxR inhibition affects the expression of stress response proteins. Conclusion: Thus, the TrxR antioxidant system is vital for the radiation survival of D. radiodurans.
| P-11.7 | | |
| Impact of Stimulation through Gamma Irradiation on Seedlings of Lathyrus (Lathyrus sativusL.) | | |
Mayuri Sahu, Shubha Banerjee1,2, Ajay Tiwari,S. J. Jambhulkar1,2
Departments of Genetics and Plant Breeding and 1Plant Molecular Biology, Indira Gandhi Krishi Vishwavidyalaya, Raipur, Chhattisgarh, 2Board of Research in Nuclear Sciences, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
A study was undertaken on induced mutagenesis with two varieties of lathyrus, Mahateora and Prateek. The mutagen used was Gamma rays (Control, 75 Gy, 100 Gy, 150 Gy, 200 Gy, 250 Gy, 300 Gy, 350 Gy, 400 Gy, 450 Gy, 500 Gy, 550 Gy 600 Gy and 75 Gy + EMS at Bhabha Atomic and Research Centre, Mumbai and field studies carried under AICRP on forage crop and utilization, Research cum Instructional Farm, Department of Genetics and Plant Breeding, College of Agriculture, Indira Gandhi KrishiVishwavidalaya, Raipur, (C.G.). Whereas lab evaluation was done at Chhattisgarh States Seed Certification agency. A difference was observed between the varieties Mahateora and Prateek in the degree of tolerance to the mutagens. Radiation dose of 350 Gy proved most lethal and reduced the percentage of survived plants in both varieties. The gamma irradiation has a (GR 50) are positive effect on biological traits of lathyrus varieties. In contrast, gamma rays had some stimulatory effects at laboratory condition on germination %, seedling height (mm), plumule length (mm) and radicle length (mm). Field condition date are recorded on plant survival % in 30 days, 60 days and at the time of crop harvesting. The variety Mahateora and Prateekin have to be found more sensitive to gamma radiation. The results indicated that low radiation (100 Gy and 150 Gy) and medium radiation (300 Gy and 350 Gy) doses have small biological effects on lathyrus in both condition in M1generation respectively.
| P-11.8 | | |
| A Study of Bacteria and Fungi Growth on E-Beam Irradiated Catheter | | |
S. A. Shaikh, S. S. Chavan, V. C. Mali, A. K. Pandey, H. K. Bagla
Department of Nuclear and Radiochemistry, Kishinchand Chellaram College, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected]
Ionizing radiations are considered as the potential method for the bulk and safe sterilization of the medical products. However, higher dose of the radiations may also damage the materials leading to the potentials sites for the fungi and bacteria growth. In the present work, Latex Foley balloon, as a urinary catheter was irradiated with e-beam and subjected to microbial studies using ISO standards and methods. The samples were irradiated with dose of 100 kGy by e-beam (Accelerator ILU) at BRIT, Mumbai, of 3.50 MeV energy set and pulse current of 250 mA. Catheters were irradiated with a dose of 5kGy/pass. It was observed that Coliforms and E.coli did not grow on either on unirradiated or irradiated samples. However, bacteria and fungi (Yeast and Mold) did exhibit higher growth on the electron beam irradiated samples with respect to unirradiated catheters samples. The total Yeast and Mould count of 35 CFU/g was observed on 100 kGy sample when tested as per IS 5403-2005 with respect to unirradiated sample (5 CFU/g). Similarly, total bacterial count of <10 CFU/g on irradiated sample was observed when tested as per ISO 11737-1:2018 as compared to total bacterial count of 5 CFU/g on unirradiated sample. The higher growth of bacteria and fungi was attributed to radiation damage on the catheter caused by higher energy electron beam. The catheter samples were also irradiated to 50 kGy and 75 kGy to understand the maximum at which fungi growth is comparable to unirradiated sample. The total Yeast and Mould counts on the catheters irradiated with 75 kGy and 50 kGy were < 10 CFU/g and 5 CFU/g, respectively. Thus, it suggests that the e-beam irradiation of catheters up to 50 kGy did not damage the catheter to form the damage sites that promotes the bacteria and fungi growth.
| P-11.9 | | |
| Improvement of Floral Traits in Carnation using Induced Mutagenesis Approach | | |
Shraddha Singh1,2, Ramesh Satdive1, Sudhir Singh1,2, Himanshu Mishra3, C. K. Salunkhe3, S. R. Dhiman4
1Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, 3Landscape and Cosmetics Maintenance Section, A and SE Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, 4Dr. Y. S. Parmar University of Horticulture and Forestry, Solan, Himachal Pradesh, India
E-mail: [email protected]
Introduction: Carnation (Dianthus caryophyllus L., family Caryophyllaceae) is among the most popular commercial cut flowers of the world. It is preferred for commercial purpose because of its excellent keeping quality, withstand in long distance transportation and remarkable quality to rehydrate. Though, the potential for this crop is high, there are limited local varieties available in India. Thus, there is an urgent need for development of improved varieties in terms of floral characteristics and higher productivity. Induced mutagenesis is a promising way in creating genetic variability in the target plants, and in turn developing novel genotypes with desired traits. Materials and Methods: Rooted plantlets of carnation (var. Bizet) were irradiated with different doses of gamma rays (0-100 Gy) for selection of an optimized dose. Later, carnation plantlets (1000) were irradiated with selected doses of gamma rays (10 and 15 Gy). Five putative mutants were screened, showing different flower colour from the parent plant. These putative mutants were propagated and stability of the traits was assessed through further generations. Detailed field evaluation of confirmed mutants is being carried out and data related to different vegetative, floral and yield characters has been recorded. Results: The stability of the traits in three putative mutants was confirmed in second generation (M1V2). These confirmed mutants have striking colour difference than their parentand have potential to be identified as new varieties. These mutants have been multiplied and their performance has been evaluated in field. Performance of all the three mutants has been found at par with the parent. Conclusions: We developed three promising stable mutants of Carnation (var. Bizet) and these findings may pave way for release of new Carnation varieties.
| P-11.10 | | |
| Understanding Mechanism of Enzymes Involved in Oxidative Stress Protection in Prokaryotes through X-Ray Crystallography | | |
Subhash C. Bihani1,2
1Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Structural biology is the study of three dimensional structures of biomacromolecules using an integrative approach comprising computational biology, X-ray crystallography, and biochemical & biophysical studies. X-ray crystallography is particularly suitable to study catalytic mechanism of enzymes. X-ray crystal structures of reaction intermediates represents snapshots of different stages of reaction pathway and provide unparalleled information about the enzyme mechanism. FrnE is a disulfide oxidoreductase involved in oxidative stress protection and radiation resistance in Deinococcus radiodurans. The structure of FrnE provided evidence of a new catalytic mechanism involving active site cysteines and an additional pair of C-term cysteines. Many structures of FrnE with cysteine residues in different oxidation states were determined. These structures along with bioinformatics, biochemical and biophysical studies established FrnE as a novel cytoplasmic disulfide oxidoreductase in prokaryotes. Further, the structural basis for the roles of RecA (DNA repair enzyme) and RqkA (a serine/threonine kinase) in radioresistance of Deinococcus radiodurans were also elucidated. KatB is a manganese catalase from N2 fixing cyanobacteria, Anabaena and is crucial for protection from salt and desiccation induced oxidative stress. Structure-function studies on KatB is among the most extensive on manganese catalases. The crystal structures showed that its active site has key differences from other known manganese catalases. This study established presence of two distinct active site configuration in manganese catalases representing two different phylogenetic groups. The structure can serve as a template for the design of synthetic mimetics for therapeutic purpose and for rational protein engineering. Further work using mutagenesis, biochemical, and physiological studies provided molecular basis for the role of non-active site residues in the activity and stability of these enzymes.
| Topic 12 Non-Ionizing Radiation and Hot topics in Biology | | |
| P-12.1 | | |
| A Novel Method for the Isolation of Cell-Free DNA (cfDNA) from Cell-Conditioned Media | | |
Dhruv Das1,2, Rajani Kant Chittela1,2, AVSSN Rao1
1Applied Genomics Section, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: The clinical utility of cell-free DNA (cfDNA) as non-invasive or minimally invasive diagnostic, prognostic, and theragnostic biomarkers has been under critical evaluation in recent years.Understanding the origin and biological significance of cfDNA may determine its future value as a biomarker. The possible sources of cfDNA are apoptosis, necrosis, and/or active release of cellular DNA into the cell free milieu. Understanding cfDNA biology requires the assistance of in-vitro cell culture models. However, the quantity and quality of extracted cfDNA from cell-conditioned media(CCM) and their biological interpretation will depend on the methods used for extraction and quantification.Most of the kits available in market are targeted towards the extraction of cfDNA from the plasma samples. Here, we intend to develop a cfDNA extraction method from CCM and compare the cfDNA yields with different quantification techniques. Materials and Methods: In this study, we devised an organic solvent-free cfDNA extraction method from CCM and compared it with three other published methods and four commercially available kits. Quantification of cfDNA has been performed using quantitative Real-Time PCR (qRT-PCR) targeting two different loci, andFluorescence-based Qubit assay. Results: The cfDNA yield from CCM provided by the developed method was found to be either higher or comparable with the other methods used. The absolute concentration of the extracted cfDNA varies with the target used for qPCR assay; however, the relative trend remains similar for both qPCR assays. Conclusions: We developed a cost-effective cfDNA extraction protocol for CCM with minimal hands-on time; low sample volume that can be utilized for understanding the biology of cfDNA using in-vitro cell culture models. This method also can be applied to the serum samples from mice where amount of the sample is a major limitation.
| P-12.2 | | |
| Immune Health and Environment: Communicating the Risks of Non-Ionizing Electromagnetic Radiation Exposure | | |
H. Yadav1, R. Singh1,2
1Department of Environmental Studies, Satyawati College, University of Delhi, 2Department of Environmental Science, Jamia Millia Islamia University, New Delhi, India
E-mail: [email protected], [email protected]
Introduction: Non-ionizing radiation is penetrating the population at a growing rate, whether from environmental or medical diagnostic sources. This is a significant epidemiological problem on a global scale, and it is important to fully understand how non-ionizing radiation functions and the possible long-term health effects. The immune system provides internal protection and promotes prolonged health and survival. Despite the fact that a number of research on the health impacts of RFR have been published, no adequate consensus has been achieved. This study sought to ascertain if exposure to non-ionizing electromagnetic radiation from mobile phones would have a cell-specific impact on innate immune responses. Materials and Methods: THP-1 Monocytic cells were exposed to non-ionizing electromagnetic radiation from mobile phones under controlled conditions for several time periods (15, 30, 45, 60, 90, and 120 minutes). Each channel represented a 5 MHz wide frequency band with the middle frequency of 900, 1900, or 2318 MHz. The cells were then processed for various immunological assays. Results: In comparison to the control, the non-ionizing electromagnetic radiation significantly increased the generation of reactive oxygen species particularly superoxide and nitrite production. In contrast, non-ionizing electromagnetic radiation decreased cellular phagocytic activity. Additionally, changes in Nf-kB levels, and pro-inflammatory cytokine levels were seen. With greater exposure time, DNA damage increased slightly. Cell viability was not greatly impacted. Conclusions: The data indicated immune-modulatory role of non-ionizing electromagnetic radiation.
| P-12.3 | | |
| Anticancer Activity of Camel Milk against Human Skin Cancer (A431) Cell Line | | |
N. Jain, S. Keelka, P. Jain, P. Kaushik, M. K. Sharma1
Department of Zoology, Centre for Advanced Studies, University of Rajasthan, Jaipur,1Department of Zoology, SPC Government College, Ajmer, Rajasthan, India
E-mail: [email protected]
Introduction: Camel milk is one of the most functional dietary supplements with profound therapeutic properties against various human ailments like hepatitis C, psoriasis, tuberculosis, diabetes, gallstones, jaundice, autism, nervous and respiratory disorders and predominantly cancer. The exceptional potency of camel milk to cure malignancies has been explored in various studies such as in the liver, breast, colorectal and esophageal cancer cell lines. Materials and Methods: The present study investigated the effect and the underlying mechanisms of camel milk from domestic camels (Camelus dromedaries) of the Bikaneri breed, on the proliferation of human skin cancer cells using an in vitro model of the human Squamous epithelium skin cancer (A431) cell line. Results: According to our findings, the Trypan Blue Exclusion Assay showed that A431 cells had a decreased rate of survival and proliferation after being incubated with camel milk in a concentration- and time-dependent manner as compared to control cells. Moreover, the migration property of tumour cells that contributes to its metastatic potential was also investigated by In Vitro Scratch Assay, which exhibited a reduction of movement in the milk treated sample compared to the (untreated) control both in terms of dose and time. Conclusions: These results showed that camel milk is potent for inhibition of cell growth and also effective in inhibition of migration properties of cancer cell lines. Further studies are in progress to analyze the anti-cancer properties and its underlying mechanism.
| P-12.4 | | |
| Screening of Potential Inhibitors of SARS-CoV-2 Main Protease | | |
Preeti Tripathi1, Amit Das1,2
1Protein Crystallography Section, Radiation Biology and Health Science Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institue, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: COVID-19 pandemic caused by SARS-CoV-2 (Severe acute respiratory syndrome virus) has caused about 65 lakhs mortalities worldwide. Main Protease of SARS-CoV-2 virus (Mpro) is 34kDa protease involved in viral polyprotein processing leading to functional viral proteins. Therefore, inhibitor binding studies and crystal structures of Mpro with inhibitors will enable structure-based inhibitor design. There is an urgent need to develop more non-covalent inhibitors of Mpro. Materials and Methods: Recombinant His6-tagged Mpro was expressed in Escherichia coli BL21 (DE3) cells and purified by Ni-affinity chromatography.The protein was characterized using various biophysical techniques like SDS-PAGE, circular dichroism, DLS, gel-filtration and fluorescence. The Mpro enzymatic assays were performed using FRET-labelled peptide substrate with competing inhibitors. Molecular docking was done using softwares like SeeSAR, AutoDock and LeadIT. The compounds showing highest binding affinities were procured and used in Mpro FRET assays. Several crystallization screens like PACT, MD-105, etc. were used for crystallization of Mpro using Mosquito dispenser. Few crystals were obtained using 10mg/ml Mpro and crystallization conditions were optimized to obtain better crystals to prepare Mpro-inhibitor complexes. Results: SARS-CoV-2 Mpro purification was optimized and biophysical characterization was done. A single gel- filtration peak indicated Mpro is a dimer and reasonably pure. A FRET substrate was used to study the enzyme activity. The enzyme assay condition was optimized to perform a high throughput screening of compounds. For FRET Substrate, the Km was found to be 43.07μM. Two (Compound 1 and 2) were found to inhibit Mpro activity significantly. The reduction in enzyme activity by compound 1 and 2 was 17.8% and 9.1 % respectively. The Ki value for Mpro inhibition by compound 2 was found to be 7.38μM. Crystallization setup was made and multiple plate like crystalswere obtained.The X-ray diffraction data of SARS-CoV2 Mpro (native) and soaked with substrate were collected at 100K using our in-house MetalJet X ray diffraction system. Conclusions: The protein was successfully purified and characterised and binding of compound 1 and 2 was studied biochemically and biophysically.
| P-12.5 | | |
| Characterization of Novel Potential Inhibitor of SARS-CoV-2 PLpro | | |
Rimanshee Arya1,2, Vishal Prashar1, Mukesh Kumar1,2
1Protein Crystallography Section, Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Due to evolution of mutations in SARS-CoV-2 leading to global health crisis, it is important to develop effective antivirals against SARS-CoV-2 to contain the current COVID-19 pandemic and future coronavirus outbreaks. Targeting SARS-CoV-2 papain-like protease (PLpro) for drug development is promising due to its dual role in promoting viral replication and dysregulating innate immune responses of host proteins. Materials and Methods: Here, we screened a library of compounds to find potential inhibitors of SARS-CoV-2 PLpro using high-throughput biochemical assay. The potential inhibitors were characterized, and their binding parameters were obtained using various biophysical techniques. Protein was crystallized and X-ray diffraction data was collected. In vitro antiviral activity of the potential inhibitor against SARS-CoV-2 was analysed in Vero E6 cells using varying inhibitor concentration. We finally examined the in vivo antiviral potential of our compound against SARS-CoV-2 in Syrian hamsters at three different doses for a period of five days. Results: Of all the compounds tested, we found that, C#65 inhibits PLpro in the low micromolar range with Ki and IC50 values of 16 μM and 30 μM, respectively. KD value obtained from ITC was found to be 21 μM. We are working on the structure of C#65-protein complex to find the exact binding site of C#65. In vitro assays showed the antiviral potential of C#65 against SARS-CoV-2 with IC50 of 50 μM. In vivo efficacy was studied in Syrian hamsters and our data show that C#65 reduced the viral load significantly in the swab sample. However, there was no significant reduction in viral load obtained in lung tissue sample. Conclusions: From our biochemical analysis we infer that C#65 is a specific inhibitor of PLpro. It kills the virus in the throat due to its direct contact but didn't reach lungs at required concentrations to kill the virus. Highly polar nature of C#65 and poor cell permeability makes C#65 significantly less toxic and may be developed as an oral gargle/nasal cleaning agent to limit SARS-CoV-2 infection.
| P-12.6 | | |
| Triphenyl-Phosphonium Conjugated Diarylheptanoid, A New Semsynthetic Structural Class Membrane Targeting Antibiotic | | |
Shweta Kumari, Sundarraj Jayakumar, Gagan D. Gupta, Subhash C. Bihani, Santosh K. Sandur, Vinay Kumar
Radiation Biology and Health Sciences Division, BARC, Mumbai, Maharashtra, India
E-mail: [email protected]
Antibiotic resistance is one of the most serious Threat to public health due to limited therapeutic options. Bactericidal agents with polypharmacological profiles or targeting bacterial membrane have lower propensity for resistance development. Mitocurcumin (MitoC) is a novel compound synthesized by triphenyl-phosphonium conjugation with curcumin. Here, we have demonstrated the antibacterial properties of MitoC that is a novel structure class compound. It is a broad spectrum antibiotic and displays efficient bactericidal activity against multidrug resistant Staphylococcus aureus and Streptococcus pneumoniae (MIC values in 1.5-12.5 μM range), and coagulase-negative staphylococci does not show resistance development against MitoC. Even at sub-MIC values, MitoC displays bactericidal properties. Bactericidal action involves rapid disruption of bacterial membrane potential when interacting with the phospholipid. Scanning electron microscope images of MitoC treated cells show structural deformations in terms of shrinking, loss of turgidity and formation of blisters and bubbles on their surface. Liposome based studies and MIC values against TolC deletion mutant (ΔtolC; outer membrane protein) of E. coli suggest that extensive membrane damage is the primary mechanism of bactericidal activity. Thus MitoC represents an excellent alternative for development of new generation bactericidal molecules that may be slow to develop resistance.
| Theme 13 Radiation Physics and Chemistry | | |
| P-13.1 | | |
| Exploring Crystal and Electronic Structures of A2BIO6(A = Ba, Sr, Ca; B = Ag, Na, Au) Double Perovskite Oxides for Radiation Detection | | |
A. Alloffy Mary, R. Vidya
Department of Medical Physics, Anna University, Chennai, Tamil Nadu, India
E-mail: [email protected]
In recent years hybrid perovskites consisting of organic molecules and inorganic elements have been attracting much attention due to their potential applications in solar cells and high-energy radiation detectors. Particularly, MAPbI3 (MA-Methylammonium) is under focus for its favorable X-ray detecting properties like high stopping power, large mobility-lifetime product, and high resistance. However, its low structural stability prevents its practical usage. Hence we are exploring novel oxide materials in double perovskite structure by combining the extraordinary electronic properties of halide perovskites with the strong stability of oxide perovskites. In this work we have performed first-principles Density-Functional Theory (DFT) based calculations using the Vienna Ab-initio Simulation Package (VASP) to explore the properties of different A2BIO6(A = Ba, Sr, Ca; B = Ag, Na, Au) compounds. We report the crystal structure, electronic band, density of states, and optical properties. The crystal structures are optimized with energy and force minimizations using plane wave cutoff energy of 500 eV. To the best of our knowledge, no computational study has been hitherto reported on the monoclinic phase of these structures. The calculated formation energy is used to predict the thermodynamic stability of cubic and monoclinic structures. The most stable structure at equilibrium and high-pressure conditions is established. These compounds exhibit semiconductivity and the band dispersion features are analyzed in detail. The charge density and electron localization function plots provide insights on the nature of bonding. The obtained properties are used to explore application of these materials for high-energy radiation detection.
| P-13.2 | | |
| An Exposure to Natural Background Radiation in Eastern Nepal | | |
Arun Kumar Shrestha1,2, Sonu Limbu1, Narayan Baral1, Manish Magar1, Arbin Limbu1, Devendra Raj Upadhyay2, Ram Saran Karki2, Ganesh Kumar Shrestha1, Buddha Ram Shah3, Ram Prasad Koirala4
1Tribhuvan University, 2Department of Physics, Tribhuvan University, Kirtipur, 3Nepal Academy of Science and Technology, Lalitpur, 4M. M. A.M.C, Tribhuvan University, Biratnagar, Nepal
E-mail: [email protected]
Introduction: Terrestrial and cosmic radiation are the major contributors to natural background radiation. On accounting it, cosmic radiation exposure is around 0.39 mSv/y, terrestrial radiation is 0.48 mSv/y, radiation from water and food is 0.29 mSv/y, radiation fromthe air is 1.26 mSv/y and artificial radiation is approximately 0.5mSv/year. As a result, the average human receives roughly 2.4mSv/year from natural sources and 0.5mSv/y from man-made sources. Monitoring the natural background radiation is important to locate the high background area. The main objective of the work is to find the average background radiation in the Morang district and to observe the variation of cosmic radiation at high altitudes. Materials and Methods: In this study, background radiation was measured in 17 different municipalities of Morang with the help of a GM counter of model GMC-300E plus. Overall, 41measurements had been carried out for 10 minutes in every 20 seconds interval time. Results: The result showed that the annual effective dose of Morang was 0.24±0.02 and was below the recommended value of 1 mSv/y set by ICRP for public health. Similarly, the variation of background radiation with an altitude from 381 to 2550m showed an increasing trend. The best-fitted line depicted that background radiation increased by 16% with 1000m and it was slightly higher than the literature's result of 10-12%. The comparative study showed that the effective dose (0.24) in the study area was relatively low as compared to other places and the highest dose (0.81) was monitored in Pokhara Valley. Conclusions: The background radiation in the Morang district has been evaluated and found to have 0.24±0.02 below the recommended value. Similarly, there is an increasing trend of radiation with altitude.
| P-13.3 | | |
| A Simple Air Ionization Chamber for Gamma-Ray Dosimetry | | |
Ava Zarif Sanayei, Mustafa Farjad-Fard, Leyli Ebrahimi, Mohammad-Reza Mohammadian-Behbahani, Sedigheh Sina
School of Mechanical Engineering, Shiraz University, Shiraz, Iran
E-mail: [email protected]
A simple current-mode air ionization chamber is constructed and tested for gamma-ray dosimetry. The dosimeter consists of a generic metal cylinder with a conducting wire installed inside. A 9V battery supplies the potential difference between the cylinder and the wire, sweeping the produced charge carriers during irradiation. The charge carrier motion inside the cylinder induces a weak current signal, which is then amplified by a BC517 NPN Darlington transistor. The amplified signal then passes through the high internal resistance of a voltmeter, which provides a measurable voltage of the order of mV on its display. Experimental tests were accomplished by using a 137Cs source of activity 49.5 mCi: a SSDL-certified calibrated source with known air-kerma rate (AKR) values at several distances from the source. The dosimeter was placed at 4 distances 30, 60, 100 and 125 cm from the source head and for each case, the voltmeter readings were recorded in three repetitions. For each distance, the mean and standard deviation of the measured voltages were calculated: 115±5.57 mV at 30 cm distance (AKR = 1296.86 μGy/h), 42±1 mV at 60 cm distance (AKR = 319.63 μGy/h), 16±1 mV at 100 cm distance (AKR = 114.71 μGy/h) and 12±1 mV at 125 cm distance (AKR = 72.21 μGy/h). A calibration curve was then established, which shows a quite appropriate linear relation (R2=0.99) between voltmeter reading and AKR.
| P-13.4 | | |
| Optimisation of the Dead Layer of An Aged P-Type HPGe Detector used in Environmental Measurements with Computational Modelling | | |
M. Margret, S. Chandrasekaran, C. V. Srinivas, B. Venkatraman1
Environmental Safety Division, Indira Gandhi Centre for Atomic Research, 1Indira Gandhi Centre for Atomic Research, Chennai, Tamil Nadu, India
E-mail: [email protected]
The gamma spectrometric laboratory of Environmental Assessment division, IGCAR, Kalpakkam, uses P-type high-purity germanium (HPGe) detector for the assessment of environmental radioisotopes due to their high resolution. The Monte Carlo (MC) simulations are used to determine the full energy peak (FEP) efficiency of the detector. But studies reveal that the simulated results are not completely in line with the experimental data. An exact knowledge of the detector geometry is required to eradicate the erroneous results of FEP efficiency calculated by the MC simulations. The manufacturer can provide the exact geometry of the detector that has been procured recently. But the detector that is being used in our laboratory had been procured in the year 2007. In such case of aged detectors, the dimension of the dead layer (DL) at the germanium surface does not match with the manufacturer's information. An initial MC detector model has been developed with the DL value mentioned by the manufacturer (1 mm) for the determination of FEP efficiency. It has been observed that the computed FEP efficiency has been decreased to a maximum of 17%. From the simulations, it is evident that the reduction in FEP efficiency is due to the increase of the thickness of the dead layer of the detector that is not wrapped up by the manufacturer. With the progressive increase of the germanium dead layer thicknesses in the detector model, a good conformity has been obtained between the simulated and measured efficiencies for 2 mm thickness, in comparison to 1 mm provided by the detector manufacturer. The variation in the relative deviation between the experimental and the simulation for 2 mm DL thickness lies within + 6% for the wide range of gamma energies in the case of natural uranium and thorium series.
| P-13.5 | | |
| Over-Response Correction of SNC 350p Roos and PTW Markus Chambers in Surface Dose and Build Up Region Dosimetry of 6MV Photon Beam | | |
Mukesh Kumar Zope, Deepali Bhaskar Patil, P. A. Aslam1, George Basil2
Department of Medical Physics, State Cancer Institute, IGIMS, Patna, Bihar,1Kiran Multi Super Speciality Hospital, Surat,2Regional Cancer Care Canter and Cancer Hospital, Siddhpur, Gujarat, India
E-mail: [email protected]
Introduction: The purpose of this study was to evaluate the over-response correction with and without applying Rawlinson Equation in surface dose and PDD in build-up region for various field sizes due to the construction and design effect of Markus and Roos fixed separation parallel-plate ionization chambers. Materials and Methods: The SNC 350P Roos and PTW Markus parallel plate ionization chambers where used to measure the PDD along the central axis.The dose measurements were made in solid water phantom at the surface and build up region, of 6 MV flat photon beam from a Varian True beam linear accelerator for field sizes 10 x10, 20x 20 and 30 x 30 cm2at SAD 100cm. The entrance windows thicknesses (the effective point of measurement) for Markus and Roos chambers are 0.03 mm and 1.07 mm, respectively. The entrance window of SNC350p Roos chamber is far away from the surface and the entrance window of Markus chamber is near to the surface, and measurements should be taken at the surface, therefore the PDDs curves are shifted 1.07 mm for Roos and 0.03 mm for Markus chamber. Results:The PSD obtained using Markus chamber before and after applying the Rawlinson's equation for over response correction for filed sizes 10 x 10, 20 x 20 and 30 x 30 cm2 were 35.109%, 42.92%, 50.72% and 28.37%, 36.192%, 44.05% respectively. These values of PSDs measured using Roos chamber for filed sizes 10 x 10, 20 x 20 and 30 x 30 cm2 were 41.61%, 50.5%, 58.035%, and 33.961%, 42.871%, 50.38%, respectively before and after applying over response correction. Conclusion:The results of this study indicate that for the 6MV flattened beam there was an increase in the PSD by using Roos chamber for all field dimensions, while this value was lesser by using Markus chamber. The structure of ionization chamber is essential for measurement of accurate PSD and PDD in buildup region. Both Markus and Roos chambers have various preferable characteristics, e.g. Roos gives less value of over response correction than Markus. The thickness of entrance window for Markus is too small compared to Roos (1.07 mm for Roos and 0.03 mm for Markus). The PSD for Roos chamber was higher when compared with Markus.
| P-13.6 | | |
| Radiation Induced Grafting of Acrylic Acid onto Cellulose: An Iterative Procedure | | |
Sanju Francis, Shubhangi Shelkar, Swarnima Rawat
Radiation Technology Development Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
In this study, we have designed, developed and fabricated filter paper discs for the detection of chromium (Cr) in ground water. This was accomplished by radiation induced grafting of acrylic acid onto the surface of filter paper.Radiation induced grafting is a validated and versatile technique for imparting functionality to a polymer substrate thereby broadening its applications. However, there is an upper limit to the extent of grafting that can be achieved, mainly dictated by the monomer concentration, radiation dose, competing polymerization, additives like Mohr's salt, mineral acids etc. Irrespective of the substrate and composition of the grafting solution, after some irradiationdose, a saturation point of grafting is reached. Preliminary investigations revealed that the intensity of colour developed on the paper disc is proportional to the extent of acrylic acid grafting, therefore higher degree of grafting was required which was not achievable by the conventional method. We could find a solution to this; we could get higher degree of grafting by employing an iteration of the conventional grafting process. This acid functionalized paperwas further treated with a colour reagent, which forms a coordination complex with Cr and the paper subsequently develops colour. Unlike other substrates, the use of extraneous mineral acids is not necessary for our substrate and therefore, the problems associated with acid handling can be circumvented. The colour and its intensity produced on the paper were discernible and distinguishable to the naked eye, even at a very low concentration of 1 ppm.
| P-13.7 | | |
| Establishment of Ionometric Reference Standard for Strength Determination of High Dose Rate 60Co Brachytherapy Sources | | |
Sudhir Kumar, Rahul K. Chaudhary, S. D. Sharma, B. K. Sapra
Radiological Physics and Advisory Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: High accuracy of radiation dose determination is crucial prior to radiotherapy treatment for achieving optimal therapeutic ratio for cancer patient. Therefore, the strength of brachytherapy source should be verified independently before using for treatment as per national/international requirements. So far, reference standard for standardization of high dose rate (HDR) 60Co brachytherapy source directly in terms of Reference Air Kerma Rate (RAKR) is not available globally. To fulfill this requirement, an ionometric reference standard was developed and established to calibrate the hospital well-type ionization chamber (WTIC). Materials and Methods: To establish a newly developed cylindrical graphite ionization chamber of sensitive volume 223.5 cm3 as reference standard, experiments were performed at ten different radiotherapy centres across the country with HDR 60Co source for evaluating the basic characteristics such as charge leakage, precision, linearity, stability, charge collection efficiency (f) and inverse-square law validity (ISLV). RAKR of HDR 60Co sources was determined from the measured signal (charge/current) using the formalism developed based on Bragg-Gray and large detector cavity theories. Once RAKR was known, standardized 60Co source was used to calibrate a WTIC. Results: Charge leakage, precision, linearity, stability (24 hrs),f and ISLV of developed chamber were found equal to 0.01%, 0.03%, 0.02%, 0.05%, 99.9% and ±0.8% respectively. Experimentally determined RAKR of HDR 60Co source using this chamber was found to be in good agreement with the vendor quoted RAKR values (within 2%) having measurement traceability with PTB, Germany. Calibration coefficient(CF) obtained using standardized HDR 60Co sources for seven WTICswas also found in good agreement with the vendor quoted CF (within 1%). Conclusions: Performance of this chamber was found satisfactory as per design and qualify to be areference standard for measurement of RAKR of HDR 60Cobrachytherapy sources. Ionization chamber is now ready to calibrate the WTICs of the hospitals.
| P-13.8 | | |
| Testing the Physical Characteristics of Therapeutic Ion Beams using the CR-39 Detector | | |
Wei hai Zhuo1,2, Bo Chen1, Shiyan Yang2, Jingfang Zhao3
1Institute of Radiation Medicine, Fudan University, 2Key Lab of Nuclear Physics and Ion-Beam Application, Fudan University, 3Shanghai Proton and Heavy Ion Centre, Shanghai, China
E-mail: [email protected]
Introduction: Carbon ion beam therapy is one of the most advanced technologies, and has become a hot spot of external radiation therapy. Deeper understanding of the physical characteristics of the therapeutic carbon ion beam is helpful for testing the quality control of the treatment planning system. Materials and Methods: The physical characteristics of the therapeutic carbon ion beams were simulated using the MonteCarlo (MC) toolkit Geant4.Several characteristics of the carbon ion beams in terms of depth dose profile, spot size and LET spectrum were verified by using the self-developed techniques based on the solid-state nuclear detector (CR-39). Results: The measured dose profile in depth is consistent with the result of the Monte Carlo simulation. It indicates that the CR-39 can be used as a flux detector to measure carbon ion beams at different depths.The measured spot size in front of the Bragg peak is in good agreement with the simulation results, it shows that the beam spot size can be accurately measured by the CR-39 in micron accuracy.The measured LET spectrum distribution is basically consistent with the simulation results. The maximum relative error between the measured and simulated results at the peak of the carbon ion LET spectrum is less than 10%. The LET spectrum distribution can be used to discriminate the primary ions and secondary fragments. The width of Bragg peak and the distribution the LETd will be broadened by using a Rifi. The lower the energy of incident carbon ions, the stronger the ability of the RiFi to modulate the dose and LET spectrum distributions. Conclusions: The CR-39 is a useful detector in study of the physical characteristics of carbon ion beams in micron scale, and it is promising in testing of several quality indexes of the TPS.
| Theme 14 Food Irradiation and Nuclear Agriculture | | |
| P-14.1 | | |
| Study on Effects of Electron Beam Irradiation on Antioxidant Property of Commercially Available Honey | | |
A. N. Khan, S. Sayed, A. K. Pandey, H. K. Bagla
Department of Nuclear and Radiochemistry, Kishinchand Chellaram College, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Honey has strong antimicrobial activity however; it is likely to have few microorganisms and needs to be sterilizedbefore use. Sterilization by ionizing radiations offers several advantages. Therefore, the effects of electron beam irradiation on commercially available honey have been studied in the present work. Materials and Methods: Honey was irradiated with electron beam having specifications: E beam Energy (Set): 3.5 MeV, E-beam Current (Pulse Current) 250mA, E-Beam Average Current: 1mA, Pulse Repetition Rate: 10 Pulses/ Second (10Hz), Conveyer Speed 3cm/second, dose per pass: 5 kGy per pass. The samples were irradiated for dose 10, 20, 30 and 40 kGy. The doses given to the samples during e-beam irradiations were measured using radiometric B3 film along with sample. After irradiations, the samples were measured for pH, refractive index (representing reducing sugars), color (flavonoids) and free radical scavenging activities by DPPH method (antioxidant property) using protocols reported in the literature. Results: The pH of honey was 4.14, which was decreased to 3.5 at 40 kGy, due to radiolysis induced monitor increase of free aminoacids. The color intensity is related to pigments which are known to have antioxidant properties. ABS450 was increased from 351 to 405 i.e. honey became darker after irradiation indicating increase of phenolic contents. The free radical scavenging activity was found to increase up to 30 kGy, and it did not increase further. Increase in antioxidant activity could be attributed to increase in phenolic contents as observed in the color change. The refractive index decreased slightly up to 30 kGy, and thereafter significant decrease was observed at 40 kGy indicating decrease in the reducing sugar content. Conclusions: In conclusion, electron beam irradiation up to 30 kGy increased the antioxidant property without significantly affecting the other component of honey studied in the present work.
| P-14.2 | | |
| Futuristic Aspects of Food Irradiation – Is it a Boon or a Bane…? | | |
Aditi Shakya, Chitra Gupta
Department of Chemistry, Bundelkhand University, Jhansi, Uttar Pradesh, India
E-mail: [email protected], [email protected]
Food irraidaition is a technology that improves safety and extends the shelf life of food commodities by reducing or eradicts microorganism (bacteria, yeasts, moulds, fungi) and insects. In this process approved food is treated with ionising energy, gamma rays and electrons beams. The scientific basis and technological adaptation of the process have been well established. The potential of this technology has been well percieved in food borne disease. The International bodies including Food and Agriculture Organisation (FAO), International Atomic Energy Agency (IAEA), Codex Alimentarius Commission (CAC) investigate projects on food irraidaition. The World Health Organisation (WHO) reviewed over 500 studies and concluded that food irradiation possess no toxicological, microbiological or nutrition problems. Mainly this review article discuss pros and also throws light on public fear that irradiation process will have a radioactive influence on food products.
Keywords: Eradicts Microorganism, food irradiation, food safety
| P-14.3 | | |
| Screening of Gamma-Irradiated M2 Mutants of Lentil (Lens Culinaris Medik.) for Enhanced Phosphorus Acquisition Efficiency | | |
Biswajit Pramanik, Sandip Debnath
Department of Genetics and Plant Breeding, Palli Siksha Bhavana (Institute of Agriculture), Visva-Bharati, Sriniketan, West Bengal, India
E-mail: [email protected]
Introduction: Acidic soil generally lacks available phosphorus (P) for plants, which eventually results in the deficiency of this particular macromolecule in plant body. Due to the deficiency, a typical symptom can be observed in lentil, where accumulation of anthocyanin takes place at a greater concentration in those P-deficient plants. Therefore, this study focuses of the enhancement of P acquisition by lentil (Lens culinaris Medik.) plants (cv. Moitree) derived from the gamma-irradiated seeds, in M2 generation. Materials and Methods: First, M2 population was developed from the gamma-irradiated M1-derived seeds of lentil cv. Moitree. After proper selection in the M2 generation, based on the morphological traits, and floral and pod developmental habits, the mutants were rigorously tested individually to estimate the P percentage in the plant tissues. Results: Consequently, P percentage in plant tissues of the gamma-ray-induced mutants was found to be on the higher side (up to 200% increase) than that of the control. Moreover, the root proliferation of the mutants was detected to be escalatedup to five times too. This experiment also proves the impact of P for better root proliferation and growth in lentil. Conclusions: This study, therefore, provides a preliminary step towards the upliftment of P uptake efficiency in lentil, even in acidic soil, which must be carried forward further to the subsequent generations towards variety development.
Acknowledgment: The research was supported by YSRA-BRNS project entitled “Physical mutation to improve the P acquisition along with nodulation efficiency in lentil”, (No. 55/14/17/2020-BRNS/10360 Dated 10/12/2020) from Department of Atomic Energy, BARC, Mumbai, India.
Keywords: Acidic soil, lentil (Lens culinaris Medik.), M2 generation, Moitree, phosphorus, root proliferation
| P-14.4 | | |
| UV-C Treatment: An Emerging Technology for Shelf-Life Extension of Beetroot Juice | | |
Charu Bisht, Neha Rawat, S. K. Sharma
Department of Food Science and Technology, COA, GBPUAT, Pantnagar, Uttarakhand, India
E-mail: [email protected]
Introduction: The aim of the present study was to investigate the efficacy of UV-C, thermal and chemical treatments on the shelf life of beetroot juice. Beetroot juice provides nutrients such as potassium, magnesium, folic acid, iron etc. that are significant for normal metabolism of the body. Various preservative methods employed such as chemical, thermal and non- thermal influence the stability of juices. Materials and Methods:Beetroot juice was exposed to three different treatments i.e. UV-C, pasteurization and addition of sodium benzoate, that could directly or indirectly effect the physicochemical properties and stability at different storage temperatures. After the treatments, all beetroot juice samples were stored at varied temperatures (4° C, 25 ° C, and 40 °C) for 14 days. Results: Parameters that were taken into account were: betalain content, pH, acidity, TSS, total phenolic content, colour, microbiological and sensory attributes. The shelf life of the beetroot juice was increased by all the three treatments. Comparison was made between the three preservative treatments by analysing different parameters. Significant changes occurred in the investigated physicochemical and sensory parameters during thermal treatment whereas insignificant changes occurred, when juice was exposed to U-C radiations. Conclusion: The results obtained, support the use of UV-C technology for retainment of colour and other physicochemical properties. Hence, non- thermal treatment can be an alternative for food preservation without destruction of nutrients.
| P-14.5 | | |
| Effect of Gamma Irradiation as Pre-Treatment for Bioethanol Production from Green Coconut Coir Juice | | |
Chaturbhuj K. Saurabh, Bhaskar Sanyal
Food Technology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Bioethanol demand increases rapidly in India due to its 10% blending with petrol and by 2025 20% blending is a set target. However, due to insufficient availability of ethanol only around 50% of petrol is blended. An alternate source from second-generation (2G) biomass like fresh coconut shell for bioethanol production is necessary since first-generation biomass (paddy and sugarcane) collectively consumes 70% of India's irrigation water. In the present study, moisture entrapped in tender coconut shells, which is an absolute agricultural waste, was extracted as crude juice for 2G bioethanol production via fermentation using gamma irradiation as a pre-treatment. Materials and Methods: Extracted samples were exposed to gamma radiation using a 60Co gamma chamber having a dose rate of 5.32 kGy/h (GC–5000, BRIT, India) at room temperature. Sucrose, D–glucose, D–fructose, D–xylose, and ethanol were analyzed by enzymatic based methods. Fermentation of pre-treated coir juice was done using locally available brewer's yeast. Results: Obtained crude juice had 43.09 ± 4.56 g/L of total fermentable sugar (sucrose + D–fructose + D–glucose). The total sugar of the control juice increased to a maximum value of 53.88 ± 4.02 g/L after 2 kGy of γ-irradiation. No effect of pre-treatment on D–xylose was observed. Fermentation of the unirradiated sample resulted in 25.08 ± 2.04 mL/L of ethanol which further increased to a value of 34.28 ± 2.6 mL/L after 2 kGy. Conclusions: Fresh coir crude juice produced ethanol with minimal processing, however, ionizing irradiation as a pre-treatment method increase free sugars in the sample thus surge in ethanol synthesis was observed. This is one of the very first reports on the production of bioethanol from fresh coconut coir juice with and without any pre-treatments.
| P-14.6 | | |
| Generation of a Novel Mutation using Electron Beam Irradiation to Enhance the per se Yielding Ability in Field Corn (Zea Mays L.) | | |
Ganapati Mukri, N. Hemalatha, Chandra Prabha, Suvendu Mondal1,2, Jayant S. Bhat3, Dhandapani Raju, R. N. Gadag, Kumari Shilpa, K. V. Gowtham
ICAR-Indian Agricultural Research Institute, New Delhi,1Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, 3ICAR-IARI, Regional Research Centre, Dharwad, Karnataka, India
E-mail: [email protected], [email protected]
Introduction: Maize is the third most important cereal which supports the food security of India. Genetic enhancement of inbred lines followed by the development of hybrids based on their combining ability is the common strategy used in maize improvement. Though natural variability does exist in maize, the application of mutagen will give an additional opportunity for the genetic enhancement/creation of variability. Materials and Methods: In the present investigation a field corn inbred line, PML 93, a medium maturing inbred line having good general combining ability was irradiated with a 200 Gy Electron beam. The M1 generation was raised during kharif 2021 at ICAR-IARI, Delhi. Individual plants were self-pollinated and M2 population was planted at IARI-Regional Centre, Dharwad during rabi 2021-22. Results: Out of 1600 individuals, a total of 42 distinct mutants were isolated along with 128 variable plant types. Various morphological traits viz., leaf angle, leaf width, flowering date, and tassel density were the prime considerations for the identification and selection. These selected plant types were advanced to the next generation and planted at ICAR-IARI during kharif 2022. By virtue of genetic control of the different traits, plants showed segregation and a total of 286 mutant plants were observed in M3 generation. These plants were individually characterized and a total of 17 distinct classes of plant phenotypes were observed at the vegetative stage. Conclusions: Mutagenesis with electron beam irradiation generated genetic variability in PML 93, a field corn inbred line. The reproductive stage evaluation will further clearly classify the mutants into different novel phenotypes. After stabilizing these mutants, novel variations can be used for the genetic enhancement of the maize breeding program.
| P-14.7 | | |
| Optimization of Gamma Radiation Dose for Reduction of Salmonella Typhimurium LT2 from Betel Leaves | | |
Indu Pant1,2, Ravindranath Shashidhar1,2
1Food Technology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Betel leaves are known as paan leaves in hindi. These are cultivated in states of West Bengal, Assam, Maharashtra, Bihar, and Karnataka. It serves as traditional mouth sweetener, freshener and symbol of hospitality in Indian culture. In India, there are reports of betel leaves contaminated with Salmonella from the water which is used to soak betel leaves. These leaves are sensitive to heat and sodium hypochlorite treatment. Therefore, in this study, we have analysed the efficacy of gamma radiation to eradicate Salmonella from betel leaf surface. The D10 value of Salmonella Typhimurium was 210 Gy on betel leaf. Based on D10 value 1 and 2 kGy doses were evaluated to eliminate 5 log CFU/g of inoculated S. Typhimurium. Betel leaves were irradiated using GC5000 and microbial survival after radiation was estimated by enrichment in Lactose broth and plating on XLD agar plates. The 1 kGy was effective in reducing 3 log and 2 kGy was enough to eliminate 5 log CFU/g. The recovery of the inoculated pathogen was evaluated during 10 days of storage at 10° C. The pathogen was not recovered during storage. The sensory, texture and colour analysis was carried out for both the doses. There were changes in greenness of the betel leaves due to storage but no significant changes in greenness was noted due to radiation treatment. Similarly, no significant changes in sensory parameter were observed. At 1kGy there was no textural changes were observed, however, at 2kGy dose there was a change in textural properties of the leaves during storage. Earlier studies from India indicated that maximum salmonella load does not exceed 102 CFU/g. Therefore, this study suggests that a dose of 1kGy can be used for eliminating Salmonella from the betel leaves and ensuring the safety.
| P-14.8 | | |
| Effect of Gamma Irradiation on the Green Leafy Volatiles of Minimally Processed Cauliflower during Storage | | |
Jasraj Vaishnav1,2, Bibhuti B. Mishra1,2, Prasad S. Variyar1,2
1Food Technology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Fresh vegetables are known for a characteristic green odor termed as green leaf volatiles (GLVs) attributed to the release of C6 aldehydes and alcohols and their corresponding esters. Unsaturated fatty acids liberated from plastid membranes have been demonstrated to be the precursors in the formation of these compounds. The released fatty acids are acted upon by lipoxygenase (LOX) and further by hydroperoxide lyase (HPL) of the lipoxygenase pathway to form C6 aldehydes and alcohols. Since gamma irradiation (2 kGy) was reported to extend shelf life of minimally processed cauliflower florets, its effect on volatile flavor compounds of lipid origin (GLVs) was investigated in current study. Materials and Methods: The gamma irradiation of minimally processed cauliflower florets was performed using Cobalt-60 (Co60) based food package irradiator at FTD, BARC. For analysis of C6 volatiles, cauliflower florets were homogenized with water and supernatant collected. The volatiles were extracted using HS-SPME and subsequent analysis was performed using GC-MS. The LOX and HPL enzyme activity was determined by spectrophotometric method using standard substrates. Activity of enzyme was determined from change in substrate concentration (μM/min). Results:An enhanced accumulation of trans-hex-2-enal (2.77-fold) and hexanal (1.48-fold) was observed after irradiation, which indicated favorable increase in cauliflower aroma. We further investigated the mechanism of enhanced formation of GLVs in the irradiated samples. No changes in the activity of lipoxygenase (LOX) and hydroperoxylyase (HPL) were observed after radiation processing. Higher content of free linolenic acid could have resulted in a greater substrate availability for LOX and HPL resulting in greater release of GLVs in cauliflower. Conclusions:Higher content of linolenic acid in fatty acid profile of gamma irradiated cauliflower confirmed an increased liberation of precursor linolenic acid under radiation stress causing enhancement in GLVs content, which indicated better aroma characteristics. Besides being highly effective method of ensuring food safety and extending shelf life, radiation processing also improved aroma quality in cauliflower.
| P-14.9 | | |
| Effect of Gamma Radiation on Flavour Metabolites of Fresh Onion (Allium Cepa) | | |
Kavita1,2, Vanshika Adiani1,2, Bibhuti B. Mishra1,2
1Food Technology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: The sprouting is one of the major reason of postharvest losses in onion and gamma irradiation is an established practice for inhibition of sprouting. In current study, for understanding longer storage effect of gamma irradiation, the flavour metabolites profile and biochemical markers were evaluated during ambient temperature (28±2°C) storage for period of more than 3 months. Materials and Methods: The gamma irradiation of onion (Agrifound light red variety) was performed with absorbed doses up to 200 Gy using Cobalt-60 (Co60) based food package irradiator at FTD, BARC. After radiation treatment, onion packets were stored at ambient (28 ±2°C)temperature in multiple sets for 105 days. Non-irradiated fresh samples were kept in parallel as control.The volatile compounds in onion were analyzed by gas chromatography–mass spectrometry (GC–MS) using head space analysis. The biochemical parameters were evaluated using spectrophotometric methods. Results: The GC/MS analysis showed sulfur containing metabolites as primary volatiles among total of 75 identified compounds which included different hydrocarbons, alcohols, aldehydes, esters, furans, and ketones. The most prevalent sulphur compounds observed in both irradiated and non-irradiated onions were dimethyl trisulfide, methyl propyl trisulfide, dipropyl disulphide, methyl-trans-propenyl-disulfane, propenyl-propyl-disulfane, ethyl butenal, nonane, decane, and beta ocimene. The relative quantification showed that content of sulfur compounds (known for pungency) increased during storage in irradiated (100 and 200 Gy) onion, which was reported to have disease preventive potentials.In addition, the effect of irradiation (up to 200 Gy absorbed dose) on flavonoids, phenolics, ascorbic acid and pyruvic acid contents was found to be insignificant. However, during storage an increase in flavonoids and phenolics content was observed in radiation processed onion, which has advantages due to known nutritional benefits. Conclusions: The findings showed enhancement of pungency in both control and irradiated onion during storage. Gamma irradiation also resulted minor increase in flavonoids and phenolics content, which is advantageous due to reported health benefits.
| P-14.10 | | |
| Peaceful Applications of Radiation Techniques in Crop Improvement and Food Processing | | |
Parmeshwar K. Sahu,Richa Sao, Raviraj Singh Patel, Samrath Baghel, Ishu Kumar Khute, B. K. Das1, Deepak Sharma
Department of Genetics and Plant Breeding, Indira Gandhi Krishi Vishwavidyalaya, Raipur, Chhattisgarh, 1Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Over the time, human population is growing significantly and arable lands are decreasing which may threaten the human civilization on earth. To meet the global food requirements, higher productivity with sustainable utilization should be the prime objective. Several techniques have been developed for increasing the food productions and their sustainable utilization. Interestingly, nuclear radiation based techniques are important among them. Nuclear techniques are not only acknowledged for developing atomic weapons but also well-known for their peaceful application in the field of agriculture and food processing. Materials and Methods: Now days, radiation techniques are being utilized for crop improvement; extension of shelf life of fruits, vegetables and RTE/ RTC food products; disinfection of food grains and spices; control of fungi & bacterial infestation in food products etc. Moreover, they are also useful in making colorful diamonds which cost higher than original. Results: In this way, excellent work for the improvement and revival of traditional landraces through radiation induced mutation breeding was performed. More than 50 rice mutant varieties have been developed from more than 90 traditional rice landraces by using radiations. Of which five rice mutants viz., TCDM-1, Vikram-TCR, CG Jawaphool Trombay, TCVM and TCSM have been released by SVRC and notified by Government of India. In addition to this, with the help of nuclear techniques, flower colour have been changed in many varieties of chrysanthemum, gladioli and tube rose; sexual morphology has been changed in spine gourd; disease resistance has been developed in wheat; seed size and leaf width of Lathyrus has been increased; oil content of safflower variety has been increased etc. Besides the crop improvement, more than 25 nutritionally enriched ready to eat/ ready to cook food products have been developed from IGKV developed bio-fortified crop varieties with extended shelf life which will be channelized in mid-day meal scheme of state government. Conclusions: In this way, radiations can be utilized peacefully for plant architect improvement, food processing, developing ready to eat products, enhancing food quality and many more. This would be beneficial for many farmers, self help groups, industries and researchers.
| P-14.11 | | |
| Improvement and Revival of Traditional Rice Landraces of Chhattisgarh through Radiation Induced Mutation Breeding Under the Joint Collaboration of IGKV, Raipur and BARC, Mumbai | | |
Parmeshwar K. Sahu,Richa Sao, Raviraj Singh Patel, Samrath Baghel, Ishu Kumar Khute, Satyapal Singh,B. K. Das1, Deepak Sharma
Department of Genetics and Plant Breeding, Indira Gandhi Krishi Vishwavidyalaya, Raipur, Chhattisgarh,1Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Chhattisgarh is popular as'Rice Bowl of India' due to its rich heritage of rice landraces. More than 23250 rice germplasm have been recorded in the state which is conserved at IGKV, Raipur. Interestingly, several traditional rice landraces have premium grain quality, special aroma, therapeutical properties, nutritional values etc. Unfortunately, many of these rice landraces have not been grown in farmers' field due to poor yield potential (2-3 t/ha), late maturity duration (150-175 days) and tall plant height (150-180cm) and therefore they are at edge of getting extinct. To overcome this, mutation breeding could offer a simple and quick approach for improvement of these landraces through radiation induced mutagenesis without altering their original features. Materials and Methods: IGKV, Raipur and BARC, Mumbai started a joint program for the improvement and revival of about 100 traditional rice varieties of Chhattisgarh through radiation (gamma rays, electron beam, X-rays, proton beam and thermal neutrons) induced mutation breeding in 2013. The major aim of this collaborative work is to develop dwarf to semi-dwarf (100-120 cm), early to medium maturity duration (105-125 days) and good yield potential (4.5 to 6.5 t/ha) mutants by maintaining their original grain quality features. Results: Till now, five rice mutant varieties viz., TCDM-1, Vikram-TCR, CG Jawaphool Trombay, TCVM and TCSM have been released and notified by Government of India. TCDM-1, CG Jawaphool Trombay and TCVM are aromatic fine grain mutant varieties whereas Vikram-TCR and TCSM have non- aromatic but high yield potential. More than 50 stable rice mutants are under evaluation at station, state and national multi-location trials. In addition, about 550 mutant lines in the background of 35 rice landraces are in various mutagenic stages of confirmation, stabilization and evaluation. Interestingly, the mean percentage reduction in plant height was 7% to 55% and days to maturity was 6% to 40% whereas the mean percentage increase in grain yield per plant was 10% to 65% in various rice mutants with respect to parent, which indicated the potential of radiation induced mutation breeding. Currently, it is the World's largest active mutation breeding program for the improvement and revival of traditional rice landraces.Conclusions: The success of this collaborative work indicated that radiation induced mutation breeding is one of the most viable tools for improvement and revival of the traditional rice landraces and could be applied in other crops also to improve one or two major drawbacks.
| P-14.12 | | |
| Identification of QTL for Large Seed Trait of An Electron Beam Induced Mutant in Groundnut (Arachis hypogaea L.) | | |
Poonam G. Bhad1,2, Suvendu Mondal1,2, Anand M. Badigannavar1,2
1Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Seed size is key agronomic factor and has been utilised for improvement of pod yield and confectionary trait in groundnut. Seed size is a polygenic trait and little is known about the mechanism of groundnut seed size development. A large seed mutant, TG 89 with 54% greater seed size was identified through an electron beam mutagenesis of a popular groundnut variety,TG26. Present study was undertaken to identify genomic locations for large seed trait in groundnut. Materials and Methods: Large seed mutant TG 89 was hybridized with a normal seed genotype ICGV 15007. A F2 mapping population with 124 plants was utilized for phenotyping for seed size. About 1431 Transposable element markers (TE) and simple sequence repeat (SSR) markers were employed for genotyping of this population. The linkage mapping and QTL identification was carried out using QTL IciMapping ver 4.1. Results: Out of 1431 molecular markers screened, 83 markers were found polymorphic between TG 89 and ICGV 15007. Bulk segregant analysis with these markers revealed that three markers (Ah1TC3A12, TE 333 and TE 278) had a putative association with the mutant large seed trait. In order to map the genomic regions associated with seed size, F2 mapping population was genotyped with 64 polymorphic TE and 19 SSR markers. These markers were mapped on to the 14 linkage groups covering map distance of 1053 cM. The QTL analysis revealed a major QTL for the mutant large seed trait between flanking markers AhTE333 and AhTE810 in Arahy.05 chromosome with 13% PVE for the seed size. This QTL was located within map interval of 4.7 cM that corresponds to 16.6 Mbp of genomic fragment which harbours an ortholog of BIG SEEDS 1 gene. Quantitative RT PCR revealed down-regulation of this BIG SEEDS 1 gene in the mutant indicating probable loss of function mutation in this gene. Conclusions: Currentstudyidentified a major QTL on Arahy.05 chromosome and provided a valuable information for candidate gene identification for large seed trait in groundnut.
| P-14.13 | | |
| Food Irradiations: Emerging Technologies with Future Aspects | | |
Rupal Jain, Chitra Gupta
Department of Chemistry, Bundelkhand University, Jhansi, Uttar Pradesh, India
E-mail: [email protected], [email protected]
Food Irradiations Technologies are Non-thermal, Energy efficient, non-Chemical and Physical method of Food Preservation in which Food is Irradiated to various Ionizing and Non-Ionizing Radiations. Many cases from ancient to modern era arise due to food borne diseases, Every year 600 million cases and 420,000 deaths of food borne are recorded by WHO(World Health Organization) which causes huge economic burden and environmental problems related to disposing the waste of food materials and harmful chemical preservatives. These methods encompass drying, storing, pickling, fermentation, pasteurization and among the emerging technologies. The Spotlight is Radiaton Technologies. An Ultramodern route to enhancing the quality, reduce waste and energy consumption in Food Irradiations. The aim of this review is to use irradiations techniques more effeciently for microbiological safety, minimal biochemical changes, promoting the maintenance of nutritional and stimulating quality of products.
Keywords: Food Irradiations, Ionizing-Radiations, Non- Ionizing Radiation
| P-14.14 | | |
| Microbial Safety Assurance of Phyllanthus Niruri Herbal by Radiation Processing and Underlying Molecular Mechanism Contributing To Hepatoprotective Property Against Anti-Malarial drug Mefloquine | | |
S. Saxena, D. Narayan1,S. Gautam2
Food Technology Division, BARC, 1A and SED, BARC, 2Food Technology Division, BARC, Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Introduction: Phyllanthus niruri is an herb of profound ethno-medicinal importance. However, the dried herbal is susceptible to microbial spoilage during post harvest processing and storage affecting its microbiological safety, shelf life and marketability. The objective of this investigation was to elucidate the effect of gamma radiation processing on the microbial safety and functionality of this herbal in terms of antimutagenic & hepatoprotective potential followed by understanding the molecular basis responsible for the hepatoprotective property of P. niruri against mefloquine (antimalarial drug belonging to 8-amino-quinolines). Material and Methods: P.niruri was subjected to radiation processing (10 kGy) followed by solid phase extraction of the soluble phenolics. Antimutagenicity was assessed by E. coli based rpoB/ RifR assay system and hepatoprotective activity were elucidated in HepG2 cell line against mefloquine. Oxidative stress was quantified based upon H2DCFDA assay. SWATH-MS analysis was carried out to investigate the protein profile in the cell line system under various treatment conditions. Results: Radiation processing ensured microbial safety and resulted in value addition through increase in phenolics content for root, stem and leaf to 2.8, 2.2 and 1.6 times, respectively leading to variation among the functional attributes. Herbal displayed strong antimutagenicity (52%) against EMS induced mutagenesis which enhanced (74%) upon irradiation. Root extract conferred strong hepatoprotective property and radiation processing led to prominent increase in this bioactivity against mefloquine. H2DCFDA assay confirmed mefloquine induced oxidative stress (by 2.6 fold) which was significantly mitigated by P. niruri root extract. Conclusions: To understand the molecular mechanism responsible for the P. niruri mediated hepatoprotection, SWATH-MS based proteomic analysis was performed. This confirmed up-regulation of proteins involved in cellular detoxification as well as in the maintenance of the cellular redox homeostasis. Predominantly the enzymes involved in ROS detoxification were prominently up-regulated by P. niruri aq. root extract including catalase, superoxide dismutase and glutathione S-transferase. Current findings thus provide credible scientific evidence ascertaining the dual functional role of radiation treatment.
| P-14.15 | | |
| Applied Mutagenesis to Develop YVMV Resistance in Okra (Abelmoschus Esculentus L. Moench) | | |
Soham Hazra, Shouvik Gorai1, Arup Chattopadhyay2, Md Nasim Ali3, Sanjay Jambhulkar4, Anirban Maji1
School of Agricultural Sciences, JIS University, Kolkata, Departments of 1Genetics and Plant Breeding, 2Vegetable Science and 3Agricultural Biotechnology, Bidhan Chandra Krishi Viswavidyalaya, Nadia, West Bengal, 4Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India
E-mail: [email protected]
Wide variability does not exist in okra [Abelmoschus esculentus (L.) Moench], requiring induction of genetic variability. The present investigation was outlined for applied mutagenesis in the variety Pusa Sawani having excellent fruit quality but highly susceptible to YVMV disease. Seed germination/seedling emergence and seedling growth were evaluated under laboratory and field conditions by imposing 300 to 1000 Gy gamma radiation to determine median lethal (LD50) and median growth reduction (GR50) doses with Co60gamma rays. The LD50 and GR50 doses varied for both growing conditions. The LD50 and GR50 doses of gamma radiation determined under field condition exhibited close correspondence and presented a reliable picture regarding lethality and injury in the M1 generation. The R2 for the linear model for seedling emergence, shoot and root length, fresh and dry weight of seedlings under field condition were 0.908, 0.902, 0.911, 0.983 and 0.972, respectively. Radiation dose should be optimized through use of LD50 and GR50 under field condition and the dose between 350 Gy and 450 Gy was optimum. Two selected putative mutant families in the M5 generationviz., 350//10///3-9////28 and 450//66///2-4////39 showed resistance to yellow vein mosaic virus disease. Five characters viz., plant height (cm), branches per plant, fruit diameter (cm), fruit weight (g) and trichome density of fruits differed significantly among the two putative mutants and Pusa Sawani. Content of ascorbic acid and total phenol and activity of two enzymes viz, peroxidase and polyphenol oxidase in leaf was high the two promising putative mutants which deserve due attention to be developed as a YVMV resistant variety or to be utilized as resistant genetic resources in future breeding programme of okra for resistance to YVMV disease.
| Theme 15 Radiation Technology for Environmental Protection | | |
| P-15.1 | | |
| Synthesis of Radiation Polymerized Bio-polymer based Superabsorbent Hydrogel | | |
Debaparna Dutta, Kuber C. Bhainsa, Tapan K. Ghanty1,2
Nuclear Agriculture and Biotechnology Division, BSG, Bhabha Atomic Research Centre, 1Bioscience Group, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected], [email protected], [email protected]
Climate change together with irregular rain fall has significant effect on crop production, especially in semi-arid and arid region of our country compromising food security. Hence, in order to address the problem of water scarcity and drought, Bioscience Group, BARC, is working on development of superabsorbent polymeric hydrogel with high water absorption capacity. Hydrogels are cross-linked three-dimensional polymeric network systems which can absorb large amount of water without dissolving in water. It has been widely used to improve the water holding capacity of different soils and hence reduce frequency of irrigation, prevent soil erosion, soil compaction and leaching thereby mitigating water stress conditions. In this study, hydrogel has been prepared using naturally available bio-polymers with inherent eco-friendly properties. Further, polymerisation process involved use of cold ionising energy, i.e., gamma radiation. The characterisation study of the prepared hydrogels was carried out using Thermo-gravimetric analyser, FTIR and SEM. Further, absorption equilibrium and release kinetics of the prepared hydrogel was investigated in varying pH conditions. The prepared hydrogel exhibited more than 500 times water absorption capacity suggesting its applicability in field condition for growing plants with less water.
| P-15.2 | | |
| Radiation Grafting as an Effective Tool for Tailoring Enzyme Immobilization Templates | | |
Shubhangi Shelkar1, Nilanjal Misra1, Swarnima Rawat1,2, N. K. Goel1, Virendra Kumar1,2
1Radiation Technology Development Division, Bhabha Atomic Research Centre, 2Homi Bhabha National Institute, Mumbai, Maharashtra, India
E-mail: [email protected]
Enzyme immobilization is an innovative strategy widely used to enhance the stability and commercial viability of industrially and biocatalytically relevant enzymes. Of the numerous strategies adopted for enzyme immobilization onto suitable templates, such as physical entrapment, encapsulation, covalent bonding etc., immobilization onto radiation grafted surface-functionalized polymeric substrate is an attractive proposition, since it is a solvent free, single step covalent bonding approach. In this work, 2,3-Epoxy propyl methacrylate (EPMA) monomer has been grafted ontopolymers PE-PP and PP tubes in benign solvent media to generate functional templates for effective immobilization of enzymes: horseradish peroxidase (HRP) and lipase, respectively. Parameters: radiation dose, monomer concentration, solvent polarity, etc., were investigated to optimize grafting parameters, followed by one-step, room temperature covalent coupling of the enzyme. These immobilized enzyme systems have been observed to demonstrate storage and thermal stability compared to their free counterparts, and have been effectively demonstrated to be useful in applications such as dye degradation, and pesticide estimation in aqueous media.
| P-15.3 | | |
| Study of Radiometric and Physico-Chemical Parameters of Groundwater in Damak Municipality, Nepal | | |
Arun Kumar Shrestha1,2, Ganesh Kumar Shrestha1, Buddha Ram Shah3, Ram Prasad Koirala4
1Tribhuvan University, 2Department of Physics, Tribhuvan University, Kirtipur, 3Nepal Academy of Science and Technology, Lalitpur, 4M. M. A.M.C, Tribhuvan University, Biratnagar, Nepal
E-mail: [email protected]
Introduction: Groundwater is the main source of drinking water in rural as well as urban areas of Damak. It is always assumed to have fresh but once it is contaminated, persists for a long period of time due to the slow movement in the aquifer. So it is very important to determine the suitability of drinking water by measuring the radiometric and physicochemical parameters. The main aim of the work is to evaluate the annual effective dose by ingestion and the water quality index using physicochemical parameters of water of all wards of Damak Municipality for the suitability of drinking purposes. Materials and Methods: Sixteen physicochemical parameters were tested in Nepal Batawaraniya Sewa Kendra, Biratnagar during the monsoon and post-monsoon period to evaluate the WQI for suitability for drinking purposes. Activity concentrations of radionuclides such as 226Ra, 232Th, and 40K were also measured using a NaI gamma spectrometer in NAST to evaluate the annual effective dose for infants, children, and adults. Results: Among 10 water samples, one sample fell in the category of the 'good', six samples in 'poor', two samples in the 'very poor', and one sample in the 'unsuitable for drinking purpose' during the monsoon period. However, during the post-monsoon period, two samples fell in the category of the 'good', six samples in the 'poor' category, one sample in the 'very poor' category, and one sample in the 'unsuitable for drinking purpose' category. Similarly, average activity concentrations of 226Ra, 232Th, and 40K in the groundwater of Damak were Bq/L, Bq/L, and Bq/L respectively. The average annual effective doses due to ingestion of these waters were μSv/yr, μSv/yr, and μSv/yr for infants, children, and adults respectively. Conclusion: The average annual effective dose from all radionuclides in drinking water is far below the recommended reference level therefore it is recommended that the water is safe for human consumption from radiation. However, physicochemical parameters show that water is not so good for drinking due to the high content of iron and EC and low value of DO.
| P-15.4 | | |
| Soft and Flexible Radiation Shields Based on PVA/PEG Blend Loaded with Bismuth (III) Oxide | | |
Srilakshmi Prabhu, S. G. Bubbly, S. B. Gudennavar
Department of Physics and Electronics, CHRIST (Deemed to be University), Bengaluru, Karnataka, India
E-mail: [email protected]
Introduction: Biodegradable and biocompatible composites based on blending of poly (vinyl alcohol) (PVA) and polyethylene glycol (PEG) loaded with bismuth oxide (Bi2O3) particles was developed as novel materials for shielding high energy X-rays and γ-rays produced in medical and industrial sector. Materials and Methods: PVA/PEG composites containing varying concentration (0 – 30 wt%) of Bi2O3 fillers were synthesized using an easy, low cost, and sustainable approach, which is the solution casting technique. The extent of uniformity in filler distribution, the factor which significantly influences the radiation shielding efficiency, was studied using scanning electron microscopy (SEM). NaI (Tl) and CdTe detector spectrometers were used to study the radiation attenuation performance of composites in the wide energy range of medical and industrial interest (30 – 1332 keV) using 137Cs, 60Co, 22Na and 133Ba sources. Results: The needle-shaped α-Bi2O3 fillers were found to be distributed homogenously throughout the PVA/PEG matrix at all loadings leaving no voids or interstitial spaces. 30 wt% Bi2O3 containing PVA/PEG composites showed highest peak degradation temperature (~ 330 °C) and tensile strength (~120 MPa). Also, these composites exhibited good X-ray and γ-ray shielding properties in the examined energy region. Conclusions: Unlike high modulus polymers such as polyethylene, polyimide or epoxy, PVA/PEG composites were found to be highly flexible (>50% strain), biocompatible and environmentally friendly. Owing to good shielding ability in the energy region of medical interest, these composites can undoubtedly be potential radioprotective materials.
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