Title: Quantifying the Potential Genotoxic Effects of Ionising Radiation on BRCA1/2 Mutant Cells
Funder: Department of Health and Social Care

Abstract: Background There is evidence that exposure to ionising radiation (IR) from CT scans can cause mutagenic/carcinogenic DNA double strand breaks (DSBs) and increase the risk of cancer. Recent studies have also suggested that this effect is potentiated by the presence of iodinated contrast reagents. Certain population sub-groups have defective DNA damage responses (e.g. due to hereditary/sporadic mutations in DNA damage repair genes) and are thus likely to be more susceptible to genotoxicity (and mutagenicity/cancer) after CT scans. To test this, the proposed study will use mutant BRCA1/BRCA2 breast cancer (defective in DNA DSB repair) as a model disease state. Cancer diagnosis/treatment in such patients can involve the use of multiple CT scans, with the potential adverse effects on genomic stability currently unknown. Aims 1) Investigate whether BRCA1/BRCA2-mutant breast cancer cells have elevated levels of DNA DSBs or micronuclei after therapeutically relevant doses of IR (compared to normal/non-cancer cells). 2) Investigate whether contrast reagents enhance IR-induced DNA DSBs or micronuclei. Research Plan and Methods of Investigation A panel of normal, BRCA1 mutant and BRCA2 mutant lymphoblastoid cells from breast cancer patients will be obtained from the Coriell Biorepository. Eight of each variant will be obtained. Cell lines will be exposed to a therapeutically relevant dose of IR and the levels of DSBs quantified over time by immunofluorescence detection of &gamma;-H2AX (histone H2AX phospho-Ser139 foci). To investigate chromosome instability, micronuclei will be quantified in the panel of cells after single and multiple exposures (as a model for the repeat CT scans often used during breast cancer diagnosis/treatment) to therapeutically-relevant doses of IR (+/- contrast reagent). Experiments will be performed in both proliferating and non-proliferating cells (likely to exhibit different responses/susceptibility to IR). Research Team Consistent with the nature of the research, this project will be led by a scientist (Dr Paul Jowsey) and a scientist/clinician (Professor Peter Blain), with relevant expertise/experience. As part of recent government funding, Dr Jowsey and Professor Blain have published several papers defining the toxic molecular mechanism of the radiomimetic chemical sulphur mustard (causes DNA DSBs) and how this relates to adverse health effects after exposure to this agent. The techniques used are directly applicable to those in the proposed research. A postdoctoral scientist with relevant skills will be appointed to perform the experimental work. As part of current NIHR funding, Dr Jowsey leads the Newcastle-based laboratory research investigating DNA damage markers in patients undergoing CT scan procedures in local hospitals. In addition, research success will be aided by an existing collaboration with colleagues at PHE CRCE, who have extensive experience in radiation biology/protection and policy. Potential Impact 1) Increased understanding of the potential genotoxic effects of CT scans and contrast reagents. 2) Inform policy makers/clinicians on the safe/appropriate use of CT scans in the diagnosis/treatment of BRCA1/BRCA2 mutant breast cancer. This could include modifying CT scanning procedures, or introducing non-ionising radiation imaging techniques (e.g. MRI), to limit potential adverse effects to patients health. 3) Ensure patients are not exposed to avoidable or unnecessary levels of DNA damage from CT scans (e.g. as part of cancer treatment or screening programmess). On the contrary, if CT scans do not increase levels of DNA damage in BRCA1/BRCA2 mutant cells, then patients can be given this information, thus alleviating the natural concerns regarding exposure to radiation in the general public. 4) Extend studies to individuals with other gene mutations/cancer predisposition conditions. 5) Identify an appropriate biomarker of genotoxicity that could be used to estimate the individual-specific risk of significant DNA damage prior to a CT scan procedure (e.g. measurement of specific biomarker in patient s blood after ex vivo exposure to IR).

Lay abstract: CT scans are used frequently in the diagnosis and treatment of injury and disease. CT scans involve the exposure of patients to X-ray radiation to generate a detailed image of internal organs/tissues. Whilst the form of radiation used is the same as for conventional X-rays, doses from CT scans are significantly higher, thus potential adverse effects need to be fully assessed. X-rays can damage our genetic material (DNA), possibly leading to cancer. Consistent with this, research has demonstrated increased levels of DNA damage in patient blood cells and increased risk of certain cancers after CT scan procedures. Importantly, certain population sub-groups have problems repairing DNA damage due to specific gene mutations and we suggest that these individuals will be more susceptible to the potential adverse effects of CT scans. For example, mutations in two genes called BRCA1 and BRCA2 are associated with defective DNA repair and cause breast cancer. CT scans are frequently used in the diagnosis/treatment of breast cancer and we want to find out whether patients with mutations in BRCA1 or BRCA2 have increased levels of DNA damage after CT scans, which could increase the risk of further gene mutations and cancer progression. To investigate this, cells from breast cancer patients with BRCA1/BRCA2 mutations will be exposed to relevant doses of radiation and levels of DNA damage measured. The project will be led by a combination of scientists and clinicians and will complement our ongoing research looking at various markers of DNA damage in patients undergoing CT scans in Newcastle hospitals. In addition, research success will be aided by collaboration with colleagues at Public Health England Centre for Radiation, Chemical and Environmental Hazards (PHE CRCE). The project plan and findings will be discussed in regular PPI meetings that are already ongoing as part of our existing research on CT scan toxicity. Data will be described in detailed reports to the Department of Health and inform policy on the safe and appropriate use of CT scan procedures. Finally, findings will be disseminated to the wider clinical/scientific community by presentation at relevant conferences and publications in peer-reviewed journals.

Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: CANCER AND NEOPLASMS