MEK inhibitor GSK1120212-mediated radiosensitization of pancreatic cancer cells involves inhibition of DNA double-strand break repair pathways
MEK inhibitor GSK1120212-mediated radiosensitization of pancreatic cancer cells involves inhibition of DNA double-strand break repair pathways
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DOI：
10.1080/15384101.2015.1104437
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发表时间：
2015-12-02
期刊：
影响因子：
4.3
通讯作者：
Williams, Terence M.
中科院分区：
文献类型：
--
作者：
Estrada-Bernal, Adriana;Chatterjee, Moumita;Williams, Terence M.
目的：超过90%的胰腺癌PC表达致癌突变KRAS，其组成性激活Raf-MEK-MAPK途径，从而赋予对放射和化学疗法的抗性。MEK抑制剂在最近的临床前和临床研究中显示出有希望的抗肿瘤反应，并且目前正在临床试验中与辐射组合进行测试。在这里，我们评估了一种新的MEK 1/2抑制剂GSK 1120212的放射增敏潜力（GSK 212或曲美替尼），并评估MEK 1/2抑制是否改变多种PC细胞系中的DNA修复机制。通过克隆形成试验、彗星试验、核灶形成试验等方法评价放射增敏作用和DNA双链断裂（DSB）修复作用（H2 AX、DNA-PK、53 BP 1、BRCA 1和RAD 51），并通过同源重组（HR）和非同源末端连接（NHEJ）的功能性GFP-报告基因测定。结果：GSK 212可阻断ERK 1/2活性，并对多种KRAS突变PC细胞株具有放射增敏作用。需要用GSK 212延长预处理24- 48小时以观察显著的放射增敏作用。GSK 212处理导致彗星试验的DNA损伤延迟消退和持久的H2 AX核灶。GSK 212治疗还导致辐射后BRCA 1、RAD 51、DNA-PK和53 BP 1核灶外观和分辨率的改变。使用功能性报告基因，GSK 212引起HR和NHEJ修复活性的抑制。结论：GSK 212通过抑制DSB修复途径的主要通路，对KRAS诱导的PC细胞具有放射增敏作用。这些数据为MEK 1/2抑制和放射治疗PC的组合提供了支持。
Purpose: Over 90% of pancreatic adenocarcinoma PC express oncogenic mutant KRAS that constitutively activates the Raf-MEK-MAPK pathway conferring resistance to both radiation and chemotherapy. MEK inhibitors have shown promising anti-tumor responses in recent preclinical and clinical studies, and are currently being tested in combination with radiation in clinical trials. Here, we have evaluated the radiosensitizing potential of a novel MEK1/2 inhibitor GSK1120212 (GSK212,or trametinib) and evaluated whether MEK1/2 inhibition alters DNA repair mechanisms in multiple PC cell lines.Methods: Radiosensitization and DNA double-strand break (DSB) repair were evaluated by clonogenic assays, comet assay, nuclear foci formation (H2AX, DNA-PK, 53BP1, BRCA1, and RAD51), and by functional GFP-reporter assays for homologous recombination (HR) and non-homologous end-joining (NHEJ). Expression and activation of DNA repair proteins were measured by immunoblotting.Results: GSK212 blocked ERK1/2 activity and radiosensitized multiple KRAS mutant PC cell lines. Prolonged pre-treatment with GSK212 for 24-48hours was required to observe significant radiosensitization. GSK212 treatment resulted in delayed resolution of DNA damage by comet assays and persistent H2AX nuclear foci. GSK212 treatment also resulted in altered BRCA1, RAD51, DNA-PK, and 53BP1 nuclear foci appearance and resolution after radiation. Using functional reporters, GSK212 caused repression of both HR and NHEJ repair activity. Moreover, GSK212 suppressed the expression and activation of a number of DSB repair pathway intermediates including BRCA1, DNA-PK, RAD51, RRM2, and Chk-1.Conclusion: GSK212 confers radiosensitization to KRAS-driven PC cells by suppressing major DNA-DSB repair pathways. These data provide support for the combination of MEK1/2 inhibition and radiation in the treatment of PC.