Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, Michigan 48109-5637, USA.
Clin Cancer Res. 2013 Aug 15;19(16):4422-32. doi: 10.1158/1078-0432.CCR-13-0788. Epub 2013 Jun 18.
To identify targets whose inhibition may enhance the efficacy of chemoradiation in pancreatic cancer and thus improve survival, we conducted an siRNA library screen in pancreatic cancer cells. We investigated PPP2R1A, a scaffolding subunit of protein phosphatase 2A (PP2A) as a lead radiosensitizing target.
We determined the effect of PP2A inhibition by genetic (PPP2R1A siRNA) and pharmacologic (LB100, a small molecule entering phase I clinical trials) approaches on radiosensitization of Panc-1 and MiaPaCa-2 pancreatic cancer cells both in vitro and in vivo.
PPP2R1A depletion by siRNA radiosensitized Panc-1 and MiaPaCa-2 cells, with radiation enhancement ratios of 1.4 (P < 0.05). Likewise, LB100 produced similar radiosensitization in pancreatic cancer cells, but minimal radiosensitization in normal small intestinal cells. Mechanistically, PPP2R1A siRNA or LB100 caused aberrant CDK1 activation, likely resulting from accumulation of the active forms of PLK1 (pPLK1 T210) and CDC25C (pCDC25C T130). Furthermore, LB100 inhibited radiation-induced Rad51 focus formation and homologous recombination repair (HRR), ultimately leading to persistent radiation-induced DNA damage, as reflected by γ-H2AX expression. Finally, we identified CDC25C as a key PP2A substrate involved in LB100-mediated radiosensitization as depletion of CDC25C partially reversed LB100-mediated radiosensitization. In a mouse xenograft model of human pancreatic cancer, LB100 produced significant radiosensitization with minimal weight loss.
Collectively, our data show that PP2A inhibition radiosensitizes pancreatic cancer both in vitro and in vivo via activation of CDC25C/CDK1 and inhibition of HRR, and provide proof-of-concept evidence that PP2A is a promising target for the improvement of local therapy in pancreatic cancer.
为了寻找可能增强胰腺癌放化疗疗效从而提高生存率的靶点，我们对胰腺癌细胞进行了 siRNA 文库筛选。我们将蛋白磷酸酶 2A（PP2A）的支架亚基 PPP2R1A 作为潜在的放射增敏靶点进行了研究。
我们通过基因（PPP2R1A siRNA）和药理学（小分子 LB100，正在进行 I 期临床试验）方法来抑制 PP2A，以确定其对体外和体内 Panc-1 和 MiaPaCa-2 胰腺癌细胞的放射增敏作用。
siRNA 敲低 PPP2R1A 可使 Panc-1 和 MiaPaCa-2 细胞增敏，放射增敏比为 1.4（P < 0.05）。同样，LB100 也使胰腺癌细胞产生类似的放射增敏作用，但对正常小肠细胞的放射增敏作用很小。从机制上讲，PPP2R1A siRNA 或 LB100 导致 CDK1 异常激活，可能是由于 PLK1（pPLK1 T210）和 CDC25C（pCDC25C T130）的活性形式积累所致。此外，LB100 抑制了辐射诱导的 Rad51 焦点形成和同源重组修复（HRR），最终导致持续的辐射诱导的 DNA 损伤，如 γ-H2AX 表达所示。最后，我们鉴定出 CDC25C 是 LB100 介导的放射增敏作用中的关键 PP2A 底物，因为 CDC25C 的耗竭部分逆转了 LB100 介导的放射增敏作用。在人胰腺癌细胞的小鼠异种移植模型中，LB100 产生了显著的放射增敏作用，同时体重减轻最小。
总的来说，我们的数据表明，PP2A 抑制通过激活 CDC25C/CDK1 和抑制 HRR，在体外和体内均可增强胰腺癌细胞的放射敏感性，并提供了 PP2A 是改善胰腺癌局部治疗的有前途的靶点的概念验证证据。
