Report on “Radiation Disaster Recovery Studies”
Course: Radiation Disaster Medicine
Name: Royba Ekaterina○Regarding “Radiation Disaster Recovery Studies”
Nominal coefficients of radiation risks were formulated by the International Commission of theRadiological Protection (ICRP). These standards were established uniformly to the public. However,numerous scientific publications have been postulated that individual difference of radiosensitivity
exists in human population [1,2]. A line of studies in the field of radiation biology suggests that theindividual difference is probably attributed to single nucleotide polymorphisms on DNA repair
genes.
Radiation sciences may take an advantage to investigate effects of mutations in genes that areencoding proteins involved in DNA damage response and repair [3]. For example, functionality of
the genes encoding molecular sensors and protein machines participating in reparation of damaged
DNA. Dampened DNA repair system causes chromosomal aberrations and cell death to induce acuteand/or chronic radiation syndromes. Hence, analysis of size effects of gene mutations is required forinvestigation of individual radiosensitivity.
While studying at Radiation Disaster Medicine course of Phoenix Leader Education Program,Hiroshima Initiative, I was engaged in the research project focused on development of strategy forquantitative assessment of individual radiosensitivity. Screening of individual-specific genealterations with established methodology can be used as reliable genetic biomarker to predict
radiation response of given person and precisely calculate its consequences for health.
○Title of Doctoral Thesis
Evaluation of ATM heterozygous mutations underlying individual differences in radiosensitivity
using genome editing in human cultured cells. Scientific reports 2017, in press.
DOI: 10.1038/s41598-017-06393-8○Summary of Doctoral Thesis
Introduction: Ataxia-telangiectasia is a rare autosomal-recessive disorder characterized byhyper-radiosensitivity, cancer predisposition, immunodeficiency and neurodegeneration. A-T iscaused by germline mutations in the ataxia-telangiectasia mutated (ATM) gene encoding ATM kinase,which modulates DNA damage response. A-T heterozygous carriers exist at a rate of approximately
1% in human populations and are clinically asymptomatic. Previous epidemiological studies
demonstrated that A-T carriers has several-fold increased risk of cancer in comparison with normalindividuals. It is therefore important to quantify the precise effect of ATM heterozygous mutations onradiosensitivity in the primary cells.
Purpose: Main purpose of my study was to clarify size effect of ATM gene variants on individual
differences in radiosensitivity using various biological assays. Since the radiosensitivity of humanprimary cells might be affected by confounding factors such as age, gender, smoking and the diverse
genetic backgrounds within human populations, it was necessary to generate a system for evaluating
genetic factors underlying individual differences in radiosensitivity in a human cultured cell line witha uniform genetic background. Since in my doctoral study the term “radiosensitivity” was attributed
to biological end-points reflecting number of DNA double-strand breaks that wereunrepaired/misrepaired even after an operation of DNA repair systems.
Methodology: In my doctoral study, I have established semi-automatic cytokinesis-blocked
micronucleus assay for evaluation of radiosensitivity of human adherent cell lines using Metafer
system. Since evaluation of size effects of mutations on DNA repair genes using CBMN assay might
be affected by genetic heterogeneity and confounding factors (such as age, gender, smoking, lifestyle, etc.) existing in human population, I generated human cell lines with uniform genetic
background (not affected by confounding factors) using CRISPR/Obligare genome-editing strategy.
Generated model cell lines were used for evaluation of impact of homozygous and heterozygous
mutations on ATM gene on individual radiosensitivity. Results were compared with chromosomal
aberration assay. The coefficients representing DNA repair capacity were extracted fromdose-response calibration curves using Cabas software.
Results: Cytokinesis-blocked micronucleus assay and chromosome aberration assay showed that theradiosensitivity of ATM+/− cell clones was significantly higher than that of ATM+/+ cells,suggesting that ATM gene variants are indeed involved in determining individual radiosensitivity.
Importantly, the differences in radiosensitivity among the same genotype clones were small, unlike
the individual differences in fibroblasts derived from A-T-affected family members. Targeted
introduction of heterozygous mutation in ATM gene had an effect of increasing cellularradiosensitivity by 2.6-fold, and suggested that they were indeed a genetic factor underlying
individual differences in radiosensitivity within human populations.
Conclusions: In my research, I have established an experimental flow combined with asemiautomatic CBMN assay and genome editing technology in a human cultured cell line as aunique system for evaluating genetic factors underlying individual radiosensitivity. Obtained results
suggests that the genome editing technique could be implemented into radiation biology in order toinvestigate size effects of mutations on DNA repair genes on individual radiosensitivity. In addition,
this methodology can be applied for estimation of radiation-induced cancer risk in humanpopulations.

○Other theses published in academic research journals
1. Miyamoto T, Hosoba K, Ochiai H, Royba E, Izumi H, Sakuma T, Yamamoto T, Dynlacht BD,
Matsuura S: The microtubule-depolymerizing activity of a mitotic Kinesin protein KIF2A drives
primary cilia disassembly coupled with cell proliferation. Cell Reports 10, 664-673, 2015.
2. Matsuura S, Royba E, Akutsu SN, Yanagihara H, Ochiai H, Kudo Y, Tashiro H, Miyamoto T.
Analysis of individual differences in radiosensitivity using genome editing. Ann ICRP 45,290-296, 2016
References
1. Foray N, Colin C, Bourguignon M. 100 Years of Individual Radiosensitivity: How We Have
Forgotten the Evidence. Radiology 264, 627-631, 2012
2. Foray N, Bourguignon M, Hamada N. Individual response to ionizing radiation. Mutat Res Mutat
Res 770, 369-386, 2016
3. Herskind C, Talbot CJ, Kerns SL et al. Radiogenomics: A systems biology approach tounderstanding genetic risk factors for radiotherapy toxicity? Cancer Lett 382, 95-109, 2016