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Purpose: Track structure Monte Carlo (MC) codes have achieved successful outcomes in the quantitative investigation of radiation-induced initial DNA damage. The aim of the present study is to extend a Geant4-DNA radiobiological application by incorporating a feature allowing for the prediction of DNA rejoining kinetics and corresponding cell surviving fraction along time after irradiation, for a Chinese hamster V79 cell line, which is one of the most popular and widely investigated cell lines in radiobiology. Methods: We implemented the Two-Lesion Kinetics (TLK) model, originally proposed by Stewart, which allows for simulations to calculate residual DNA damage and surviving fraction along time via the number of initial DNA damage and its complexity as inputs. Results: By optimizing the model parameters of the TLK model in accordance to the experimental data on V79, we were able to predict both DNA rejoining kinetics at low linear energy transfers (LET) and cell surviving fraction. Conclusion: This is the first study to demonstrate the implementation of both the cell surviving fraction and the DNA rejoining kinetics with the estimated initial DNA damage, in a realistic cell geometrical model simulated by full track structure MC simulations at DNA level and for various LET. These simulation and model make the link between mechanistic physical/chemical damage processes and these two specific biological endpoints.

1. Introduction Ionising radiation induced DNA damage and subsequent biological responses depend on the radiation’s track structure and its energy loss distribution pattern. To investigate the underlying biological mechanisms involved in such complex system, there is need of predicting biological response by integrated Monte Carlo (MC) simulations across physics, chemistry and biology. Typically, modeling of survival fraction and/or kinetics of repair proteins attracts interests of researchers in the fields of radiobiology including medical physics. Hence, in this work, we have developed an application using the open source Geant4‐DNA toolkit to propose a realistic “fully integrated” MC simulation to calculate both early DNA damage and subsequent biological responses with time. 2. Materials and Methods We had previously developed an application allowing simulations of radiation induced early DNA damage on a naked cell nucleus model and its sub-components model (shown in top panels of Fig.1) [1]. In the new version presented in this work [2], we have developed three additional important features: (1) modeling of a realistic cell geometry including cytoplasm and the materials surrounding irradiated cells (shown in bottom panels of Fig.1), (2) inclusion of a biological repair model, (3) refinement of DNA damage parameters for direct damage and indirect damage scoring. The simulation results are validated with experimental data in terms of Double Strand Break (DSB) yields for human fibroblast cell as well as yield of accumulated γ-H2AX. In addition, the yields of indirect DSBs are compatible with the experimental scavengeable damage fraction. 3. Results and Discussion The simulated quantities are showing good agreements with the reference experimental quantities. For instance, as shown in Fig.2, the simulation application also demonstrates agreement with experimental data of γ-H2AX yields for gamma ray irradiation. 4. Conclusion As a new important step for radiobiological simulation using Geant4-DNA, the first fully integrated simulation chain has been developed across the physical, chemical and biological stages of cellular radiation action in a single application. The application is validated with the reference experimental results., 第121回 日本医学物理学会学術大会

Molecular targeted cancer therapy is a promising treatment strategy. Considering the central role of the epidermal growth factor receptor in cell proliferation and survival, there are indications that targeted agents like tyrosine kinase inhibitors, i. e., erlotinib, may enhance the antitumor treatment by radiation. The aim of this study is to analyze the inactivation effects of g-rays and to test the radiosensitizing potential of erlotinib on human lung adenocarcinoma cells in vitro. Irradiations were performed with doses ranging from 1 Gy to 8 Gy. In order to increase the radiosensitivity of CRL-5876 lung adenocarcinoma cells, the cells were treated with a clinically relevant concentration of 2 µM erlotinib. The effects of single and combined treatments were monitored using clonogenic survival, cell viability and proliferation assays at different time points. For the detection and visualization of the phosphorylated histone H2AX (γ-H2AX), an important biological marker of DNA double-strand break formation, fluorescence immunocytochemistry, was performed. The response to the treatment was monitored at four time points: 30 min, 2, 6, and 24 h. Irradiations with g-rays resulted in significant cell inactivation regarding all analyzed biological endpoints. Combined treatments revealed consistent cell inactivation. Moreover, compared to g-rays alone, elevated levels of g-H2AX foci were observed after pretreatment with erlotinib, indicating radiosensitization through impaired DNA repair. [Projekat Ministarstva nauke Republike Srbije, br. 173046 i br. 171019]

In order to contribute to the understanding of mechanisms by which regulatory proteins recognize genetic information stored in DNA, analyses of their interaction with specific nucleotides are usually performed. In this study, the electrophoretic mobility shift assay (EMSA) was applied to analyze the interaction of nuclear proteins from the liver of rats of different age i.e., young (3-month-old), middle- aged (12-month-old) and aged (24-month-old), with radioactively labelled synthetic oligonucleotide analogues, corresponding to GRE. The levels of GRE binding activity were assessed by quantitative densitometric scanning of the autoradiograms. The results showed statistically significant decreasing values of up to 78% and 49% in middle aged and old animals, respectively, compared to young animals (p < 0.05). The specificity of the nuclear proteins-GRE interaction was demonstrated by competition experiments with unlabelled GRE. In a supershift assay, using the antibody BuGR2, it was shown that the GR proteins present in nuclear extracts have a high affinity for the GRE probe. The stabilities of the protein-DNA complexes were analysed and it was concluded that they changed during ageing. .