DNA damage response (DDR) pathways are frequently dysregulated in ovarian and pancreatic cancers, and this can be associated with resistance to DNA-damaging chemotherapy. Targeting the DDR network is thus a promising strategy to enhance chemosensitivity in these malignancies. We have previously shown that inhibition of DNA-PKcs, a DDR protein involved in the non-homologous end joining pathway, which repairs DNA double strand breaks, reverses resistance to cisplatin in ovarian and pancreatic cancer cells. Here, we assess the potential of other DDR proteins as therapeutic targets for these poor prognosis cancers.
siRNA screening to assess the effect of 639 DDR-associated genes on cisplatin or gemcitabine response was carried out in PEO4 (ovarian) or Panc-1 (pancreatic) cell lines. Calculation of z-scores allowed identification of robust modulators of cell viability. Inhibition of VCP (valosin containing protein)/p97 was carried out using the pharmacological inhibitor NMS-873, and MG-132 was used to inhibit proteasome activity. Apoptotic induction was determined by measuring caspase-3/7 activity and protein expression was assessed using western blotting.
Our siRNA screen revealed a set of novel DDR proteins that can be targeted either alone or in combination with DNA damaging agents in ovarian and pancreatic cancers. Enrichment analysis of the top hits identified regulators of protein ubiquitination as the most enriched subset. Of these, VCP, a facilitator of ubiquitinated protein degradation, was validated as a therapeutic target in an expanded panel of ovarian and pancreatic cancer cell lines: treatment with NMS-873 increased apoptotic induction and sensitised resistant cell lines to cisplatin. Additionally, this inhibition resulted in reduced expression of DNA-PKcs and Rad51, a key protein in the homologous recombination DNA double strand break repair pathway. Reduction of Rad51 was prevented by MG-132 indicating that the regulation is at the proteasomal level.
Overall our data validates targeting DDR pathways as a therapeutic approach in ovarian and pancreatic cancers and demonstrates VCP/p97 as a potential target in these cancers.
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All authors have declared no conflicts of interest.