Vena Francesca, Jia Ruochen, Esfandiari Arman, Garcia-Gomez Juan J, Rodriguez-Justo Manuel, Ma Jianguo, Syed Sakeena, Crowley Lindsey, Elenbaas Brian, Goodstal Samantha, Hartley John A, Hochhauser Daniel
Cancer Research UK Drug-DNA Interactions Research Group, UCL Cancer Institute, Paul O'Gorman Building, University College London, London WC1E 6DD, UK.
Department of Research Pathology, UCL Cancer Institute, London EC1M6BQ, UK.
Oncotarget. 2018 Jan 22;9(14):11592-11603. doi: 10.18632/oncotarget.24294. eCollection 2018 Feb 20.
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 γH2AX and RAD51 immunofluorescence staining. 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 γH2AX 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.
针对DNA修复存在缺陷的肿瘤中的DNA损伤反应（DDR）是一种临床成功的策略。RAS/RAF/MEK/ERK信号通路在人类癌症中经常失调。在本研究中，我们探讨了MEK抑制对同源重组途径的影响，并探索了MEK抑制剂与DDR抑制剂及一种缺氧激活前药联合治疗的潜力。我们研究了MEK1/2的选择性变构抑制剂匹美替尼与聚（二磷酸腺苷[ADP] - 核糖）聚合酶（PARP）的小分子抑制剂奥拉帕利以及缺氧激活前药依沃福酰胺在卵巢癌和胰腺癌细胞系中的联合作用。通过Caspase 3/7检测评估细胞凋亡，通过免疫印迹检测蛋白质表达。用γH2AX和RAD51免疫荧光染色监测DNA损伤反应。在胰腺癌异种移植模型中评估匹美替尼与依沃福酰胺的抗肿瘤活性。我们发现，在缺氧条件下，匹美替尼治疗后BRCA2蛋白表达下调。这表现为RAD51焦点水平所示的同源重组修复减少。MEK抑制足以诱导γH2AX焦点形成，表明该途径的抑制会损害DNA修复。当与奥拉帕利或依沃福酰胺联合使用时，匹美替尼治疗增强了DNA损伤并增加了细胞凋亡。匹美替尼与依沃福酰胺的联合在BRCA野生型Mia - PaCa - 2异种移植模型中显示出增强的抗肿瘤活性，但在BRCA突变的BxPC3模型中则不然。我们的数据表明，靶向MEK抑制导致BRCA - 2功能正常的癌症中同源重组DNA损伤修复受损，并增加PARP抑制敏感性。
