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MEK inhibition leads to BRCA2 downregulation and sensitization to DNA damaging agents in pancreas and ovarian cancer models.

ABSTRACT: Targeting the DNA damage response (DDR) in tumors with defective DNA repair is a clinically successful strategy. The RAS/RAF/MEK/ERK signalling pathway is frequently deregulated in human cancers. In this study, we explored the effects of MEK inhibition on the homologous recombination pathway and explored the potential for combination therapy of MEK inhibitors with DDR inhibitors and a hypoxia-activated prodrug. We studied effects of combining pimasertib, a selective allosteric inhibitor of MEK1/2, with olaparib, a small molecule inhibitor of poly (adenosine diphosphate [ADP]-ribose) polymerases (PARP), and with the hypoxia-activated prodrug evofosfamide in ovarian and pancreatic cancer cell lines. Apoptosis was assessed by Caspase 3/7 assay and protein expression was detected by immunoblotting. DNA damage response was monitored with ?H2AX and RAD51 immunofluorescence staining. In vivo antitumor activity of pimasertib with evofosfamide were assessed in pancreatic cancer xenografts. We found that BRCA2 protein expression was downregulated following pimasertib treatment under hypoxic conditions. This translated into reduced homologous recombination repair demonstrated by levels of RAD51 foci. MEK inhibition was sufficient to induce formation of ?H2AX foci, suggesting that inhibition of this pathway would impair DNA repair. When combined with olaparib or evofosfamide, pimasertib treatment enhanced DNA damage and increased apoptosis. The combination of pimasertib with evofosfamide demonstrated increased anti-tumor activity in BRCA wild-type Mia-PaCa-2 xenograft model, but not in the BRCA mutated BxPC3 model. Our data suggest that targeted MEK inhibition leads to impaired homologous recombination DNA damage repair and increased PARP inhibition sensitivity in BRCA-2 proficient cancers.

SUBMITTER: Vena F

PROVIDER: S-EPMC5837749 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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MEK inhibition leads to BRCA2 downregulation and sensitization to DNA damaging agents in pancreas and ovarian cancer models.

Vena Francesca F Jia Ruochen R Esfandiari Arman A Garcia-Gomez Juan J JJ Rodriguez-Justo Manuel M Ma Jianguo J Syed Sakeena S Crowley Lindsey L Elenbaas Brian B Goodstal Samantha S Hartley John A JA Hochhauser Daniel D

Oncotarget 20180122 14

Targeting the DNA damage response (DDR) in tumors with defective DNA repair is a clinically successful strategy. The RAS/RAF/MEK/ERK signalling pathway is frequently deregulated in human cancers. In this study, we explored the effects of MEK inhibition on the homologous recombination pathway and explored the potential for combination therapy of MEK inhibitors with DDR inhibitors and a hypoxia-activated prodrug. We studied effects of combining pimasertib, a selective allosteric inhibitor of MEK1/ ...[more]

PMID: 29545922

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Project description:BackgroundAberrant lipogenicity and deregulated autophagy are common in most advanced human cancer and therapeutic strategies to exploit these pathways are currently under consideration. Group III Phospholipase A2 (sPLA2-III/PLA2G3), an atypical secretory PLA2, is recognized as a regulator of lipid metabolism associated with oncogenesis. Though recent studies reveal that high PLA2G3 expression significantly correlates with poor prognosis in several cancers, however, role of PLA2G3 in ovarian cancer (OC) pathogenesis is still undetermined.MethodsCRISPR-Cas9 and shRNA mediated knockout and knockdown of PLA2G3 in OC cells were used to evaluate lipid droplet (LD) biogenesis by confocal and Transmission electron microscopy analysis, and the cell viability and sensitization of the cells to platinum-mediated cytotoxicity by MTT assay. Regulation of primary ciliation by PLA2G3 downregulation both genetically and by metabolic inhibitor PFK-158 induced autophagy was assessed by immunofluorescence-based confocal analysis and immunoblot. Transient transfection with GFP-RFP-LC3B and confocal analysis was used to assess the autophagic flux in OC cells. PLA2G3 knockout OVCAR5 xenograft in combination with carboplatin on tumor growth and metastasis was assessed in vivo. Efficacy of PFK158 alone and with platinum drugs was determined in patient-derived primary ascites cultures expressing PLA2G3 by MTT assay and immunoblot analysis.ResultsDownregulation of PLA2G3 in OVCAR8 and 5 cells inhibited LD biogenesis, decreased growth and sensitized cells to platinum drug mediated cytotoxicity in vitro and in in vivo OVCAR5 xenograft. PLA2G3 knockdown in HeyA8MDR-resistant cells showed sensitivity to carboplatin treatment. We found that both PFK158 inhibitor-mediated and genetic downregulation of PLA2G3 resulted in increased number of percent ciliated cells and inhibited cancer progression. Mechanistically, we found that PFK158-induced autophagy targeted PLA2G3 to restore primary cilia in OC cells. Of clinical relevance, PFK158 also induces percent ciliated cells in human-derived primary ascites cells and reduces cell viability with sensitization to chemotherapy.ConclusionsTaken together, our study for the first time emphasizes the role of PLA2G3 in regulating the OC metastasis. This study further suggests the therapeutic potential of targeting phospholipases and/or restoration of PC for future OC treatment and the critical role of PLA2G3 in regulating ciliary function by coordinating interface between lipogenesis and metastasis.

Dataset Information

MEK inhibition leads to BRCA2 downregulation and sensitization to DNA damaging agents in pancreas and ovarian cancer models.

Publications

MEK inhibition leads to BRCA2 downregulation and sensitization to DNA damaging agents in pancreas and ovarian cancer models.

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