Targeting the DNA damage response network as a therapeutical strategy in ovarian and pancreatic cancers

Abstract

High grade serous (HGS) epithelial ovarian cancer (EOC) and pancreatic ductal adenocarcinoma (PDAC) are among the poorest prognosis neoplasms; both are characterised by defects in DNA damage repair mechanisms, high levels of genomic instability, intra-tumoural heterogeneity and resistance, intrinsic or acquired, to conventional chemotherapy. The DNA damaging agents, gemcitabine and platinum are standard treatments for patients with PDAC and EOC, respectively. Here it is demonstrated that time of onset and levels of DNA double strand breaks (DSB), does not correlate with response to these drugs in PDAC and EOC cell lines. Instead, the duration of these lesions/time of repair differs between drug-sensitive and –resistant cell lines. The AKT pathway is central to cell survival and has been implicated in platinum resistance. Here, it is demonstrated that cisplatin and gemcitabine exposure induces an AKT-dependent, pro-survival, DNA damage response in clinically cisplatin-resistant but not -sensitive cells. AKT is phosphorylated specifically on Serine 473 by DNA-dependent protein kinase catalytic subunit (DNA-PKcs), which is involved in the early stages of DSB repair by the non-homologous end-joining (NHEJ) pathway. Inhibition of DNA-PKcs and/or AKT restores platinum sensitivity in a panel of clinically resistant HGS EOC and PDAC cell lines. This study also identifies that restoration of cisplatin sensitivity by DNA-PKcs inhibition is independent of its role in the NHEJ. Unexpectedly, inhibiting DNA-PKcs was shown to decrease both NHEJ and homologous recombination (HR) activities, and whilst studying the DNA-PKcs effect on EOC and PDAC cell lines, a novel regulatory interaction between DNA-PKcs and RAD51, which assists in the repair of DNA DSB via the HR pathway, was also identified. It is suggested here that the method by which DNA-PKcs is controlling RAD51 is via ubiquitin-mediated proteasomal degradation and this appears to be a cancer-specific feature. It was also identified that RAD51 can be ubiquitinated on three different lysine sites and all of these sites are required for the proteasomal-degradation of RAD51 following DNA-PKcs inhibition. Implications of this finding in the clinical setting could be substantial, as here it is shown that targeting DNA-PKcs downregulates both HR and NHEJ pathways, restoring chemotherapy response in EOC and PDAC cell lines.