mdm2抑制剂协同mTOR抑制剂治疗肾癌的作用和机制研究

VHL/HIF-1α plays a key role in the development of renal cell carcinoma(RCC). mTOR inhibitor can block PI3-K/AKT/mTOR pathway and downregulate HIF-1α.So it was used to treat RCC in the past few years. However, the clinical effect was not very good. P53 was widely used to treat cancer and most RCC cell lines express wild p53. Up-regulation of p53 could be a possible way of treating RCC. MDM2 inhibitor can block the combination and ubiquitination of MDM2 on p53. Our previous studies showed mdm2 inhibitor can enhance sorafenib induced AIF nuclear translocation (programmed necrosis) in melanoma via p53 pathway. Some papers also proved mdm2 inhibitor can regulate HIF-1α. Therefore, we hypothesized that MDM2 inhibitor could enhance mTOR inhibitor induced programmed cell death on RCC via p53 pathway and PI3-K/AKT/mTOR pathway.We intent to testify this hypothesis by method of IHC, confocal andRNA interference. We will first observe the synergic effect of MDM2 inhibitor and mTOR inhibitor on renal cell cancinoma cell line A498; second the designed shRNA was applied to interfere the key molecule HIF-1α/p53 to elucidate the instinct mechanism; Finally, A498 xenograft was applied to observe the synergic effect of MDM2 inhibitor and mTOR inhibitor and elucidate the instinct mechanism. The study findings will help to provide new ideas for treatment on RCC.

肾细胞癌（RCC）发病率日益增高、对放化疗不敏感、进展快、死亡率高。目前观点认为VHL基因缺失/HIF-1α高表达在RCC发病进展中起着重要作用。mTOR抑制剂可以阻断PI3-K/AKT/mTOR途径而下调HIF-1α被用于治疗RCC，但效果仍不理想。p53作为抑癌基因被广泛用于治疗肿瘤，绝大多数RCC均表现为野生型p 53，使通过上调p 53治疗RCC的设想成为可能。MDM2抑制剂阻断MDM2结合并泛素化降解P53而上调P53，最近发现MDM2抑制剂可以直接或间接的影响HIF-1α。因此本课题设想运用免疫组化、confocal、RNA干扰等方法，首先在A498细胞株验证MDM2抑制剂联合mTOR抑制剂的治疗作用，然后对相关信号传导通路的分子进行RNA干扰，探讨作用机制，最后在A498肿瘤动物模型上验证并探索二者联用的疗效及作用机制。从而为进一步探索及治疗RCC提供新的治疗思路及理论依据。

背景: mTOR和MDM2信号通路在肿瘤激活过程非常频繁与MDM2经常放松管制在癌症发展和mTOR抑制剂或MDM2能独立增强癌细胞凋亡。然而，在肾细胞癌中mTOR和MDM2拮抗剂的反应(RCC)仍未知。.主要研究内容: 本研究使用A498细胞治疗MDM2抗Mi-319和/或mTOR抑制剂雷帕霉素。 方法为细胞凋亡和免疫印迹分析。.结果和结论: 我们发现MDM2抑制剂(Mi-319)主要依靠p53蛋白过表达诱导肾细胞癌凋亡，而mTOR抑制剂(雷帕霉素)通过上调HIF1α蛋白表达促进肾细胞癌细胞凋亡。重要的是，我们观察到在相对较低的浓度下对肾癌细胞凋亡的mi-319和雷帕霉素的组合强协同效应。p53和HIF1α下调拮抗剂引起的细胞凋亡差异的程度，其表达的变化响应的化学处理，双敲除p53和HIF1α显著阻碍了mi-319或雷帕霉素诱导的细胞凋亡，提示p53和MDM2 HIF1α参与或mTOR受体拮抗剂诱导的细胞凋亡。.总的来说，我们认为，p53和HIF1α激活可导致癌细胞的凋亡，结合MDM2拮抗剂和mTOR抑制剂可能在肾癌的治疗是有益的。