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Arora S , Yan H , Cho I , Fan HY , Luo B , Gai X , Bodian DL , Vockley JG , Zhou Y , Handorf E , Egleston BL , Andrake M , Nicolas E , Serebriiskii I , Yen TJ , Hall MJ , Golemis EA , Enders GH
Genetic Variants That Presdispose to DNA Double-strand Breaks in Lymphocytes from a Subset of Patients With Familial Colorectal Carcinomas
Gastroenterology. 2015 Sep 4;149(7) :1872-1883 e9
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BACKROUND & AIMS: DNA structural lesions are prevalent in sporadic colorectal cancer, so we proposed that gene variants that predispose to DNA double-strand breaks (DSBs) would be found in patients with familial colorectal carcinomas of an undefined genetic basis (UFCRC). METHODS: We collected primary T cells from 25 patients with UFCRC and matched patients without colorectal cancer (controls) and assayed for DSBs. We performed exome sequence analyses of germline DNA from 20 patients with UFCRC and 5 undiagnosed patients with polyposis. The prevalence of identified variants in genes linked to DNA integrity was compared to that of individuals without a family history of cancer. The effects of representative variants found to be associated with UFCRC was confirmed in functional assays with HCT116 cells. RESULTS: Primary T cells from most patients with UFCRC had increased levels of the DSB marker gammaH2AX following treatment with DNA damaging agents, compared to T cells from controls (P<.001). Exome sequence analysis identified a mean 1.4 rare variants/patient that were predicted to disrupt functions of genes relevant to DSBs. Controls (from public databases) had a much lower frequency of variants in the same genes (P<.001). Knockdown of representative variant genes in HCT116 CRC cells increased gammaH2AX. Detailed analysis of immortalized patient-derived B cells, which contained variants in the Werner syndrome, RecQ helicase-like gene (WRN, encoding T705I), and excision repair cross-complementation group 6 (ERCC6, encoding N180Y), revealed reduced levels of these proteins and increased DSBs, compared to B cells from controls. This phenotype was rescued by exogenous expression of WRN or ERCC6. Direct analysis of the recombinant variant proteins confirmed defective enzymatic activities. CONCLUSIONS: These results provide evidence that defects in suppression of DSBs underlie some cases of UFCRC; these can be identified by assays of circulating lymphocytes. We specifically associated UFCRC with variants in WRN and ERCC6 that reduce capacity for repair of DNA DSBs. These observations could lead to a simple screening strategy for UFCRC, and provide insight into the pathogenic mechanisms of colorectal carcinogenesis.
Arora, Sanjeevani Yan, Hong Cho, Iltaeg Fan, Hua-Ying Luo, Biao Gai, Xiaowu Bodian, Dale L Vockley, Joseph G Zhou, Yan Handorf, Elizabeth Egleston, Brian L Andrake, Mark Nicolas, Emmanuelle Serebriiskii, Ilya Yen, Timothy J Hall, Michael J Golemis, Erica A Enders, Greg H R01 CA063366/CA/NCI NIH HHS/United States R01 GM084983/GM/NIGMS NIH HHS/United States Gastroenterology. 2015 Sep 4. pii: S0016-5085(15)01259-7. doi: 10.1053/j.gastro.2015.08.052.