One of the risks associated with radiotherapy for cancer treatment is the potential induction of a second cancer in the normal tissues surrounding the treatment volume. Some individuals are predisposed to both spontaneous and radiation-induced cancer, often due to inherited defects in DNA repair genes. The Peter MacCallum Cancer Centre Late Effects Clinic follows a cohort of patients who were treated with radiotherapy for a first cancer, and who went on to develop a second independent cancer years later. We hypothesise that this population might be selected for individuals who are more sensitive to radiation-induced cancer than parallel control patients who were treated with radiotherapy but have not developed a second cancer (1). To test this hypothesis, we have conducted a functional assay of cellular radiosensitivity in lymphocytes from these patients and matched controls, examining the induction and repair of γ-H2AX foci in response to ex vivo radiation. The foci were counted automatically from confocal microscopy images using the custom-built software JQuantPro. The repair kinetics and dose-responses suggest that there are individuals with signs of radiosensitivity (slower repair rate and/or increased levels of residual damage) but that these are not different between those with or without a second cancer. Next, we have conducted whole-exome sequencing on germline DNA from the second cancer group and matched first-cancer-only controls to determine whether we can detect genomic determinants of radiosensitivity. We will present the results of the functional and genomics assays, along with dosimetry data which has been retrospectively calculated to determine the estimated doses to the tissues at risk of second cancer. These data will contribute to efforts to minimise radiotherapy-induced second cancer risk, particularly in those individuals who are at inherently greater risk.