Abstract 2486: Tumor hypoxia induces DNA repair vulnerabilities through contextual loss of heterozygosity

Abstract

Abstract Introduction: Intratumoural hypoxia leads to decreased expression in DNA damage response (DDR) and repair pathways. Given the ‘two-hit’ model for loss of gene function, we hypothesize that hypoxia-mediated down-regulation of gene expression and function, coupled with an already inactive allele contributes to an exploitable contextual ‘loss-of- heterozygosity’ phenotype. Method: To interrogate this relationship, isogenic DLD-1 cells heterozygous and homozygous null for BRCA2, were placed under normoxic (21%) or hypoxic (0.2%) conditions for 72 hours. Hypoxia-mediated changes in DDR response and DNA repair were evaluated by cell proliferation, cell cycle analysis, western blots, qPCR, immunofluorescence, clonogenic assays and functional repair assays. Results: No differences in proliferation and cell survival were observed between oxic and hypoxic cells (72 hours - 0.2% O2). Under chronic hypoxic conditions, confirmed by the up-regulation of VEGF and HIF1a, mRNA and protein expression of key homologous recombination (HR) genes (BRCA1, BRCA2, RAD51) were down-regulated. Functionally, BRCA2-/- null cells proved unable to recruit Rad51 foci and were profoundly sensitivity to PARP inhibition. Conversely, heterozygote BRCA2+/- cells retained the ability to recruit Rad51 foci under both oxic and hypoxic conditions. However, exposure to chronic hypoxia resulted in a reduction of foci formation. Chronically hypoxic BRCA2+/- cells exhibited a 30-40% increase in sensitivity to PARP inhibition compared to their oxic counterparts. Preliminary data shows a similar synthetically lethal relationship in genetic (BRCA2-/- ) and tumor microenvironment induced (BRCA2+/-) HR deficient cells, when challenged with DNA damage response (DDR) kinase inhibitors ATRi and DNAPKi. Conclusions: Herein, we demonstrate a novel mechanism of contextual ‘loss-of heterozygosity’, which marries the tumour microenvironment and innate genetic alterations. The resultant increased sensitivity to DDR kinase inhibitors and PARPi highlights the therapeutic significance of this phenomenon. Citation Format: Osman Mahamud, Melvin Chua, Winnie Lo, Gaetano Zafarana, Robert Bristow. Tumor hypoxia induces DNA repair vulnerabilities through contextual loss of heterozygosity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2486. doi:10.1158/1538-7445.AM2017-2486