靶向mTORC2抑制骨肉瘤及其分子机制研究

The mammalian target of rapamycin (mTOR) is a serine/threonine kinase at the nexus between oncogenic phosphoinositide 3-kinase (PI3K)/Akt signaling and critical downstream pathways that drive cancer cell growth, survival and resistance to therapeutic agents. mTOR complex 1 and 2 (mTORC1/2) are the functional units of mTOR. Because of their feedback loop can activate the Akt phosphorylation in Ser 473 site, rapamycin and its analogues always show poor performances of anti-tumor effect. As a result, increasing interests have been showed recently in the characterism of the second generation inhibitor targeting mTORC1/2. However, to date no publication has well established the role of mTORC2 alone in the process of oncogenesis, nor the effects of its targeted treatment in osteosarcoma. Our prelimary results showed that inhibition of mTORC2 but not mTORC1, could significantly reduce the migration and induce apoptosis in osteosarcoma. Consequently, our study aims to investigate the role of mTORC2 in tumorigenesis and progression of osteosarcoma. Furthermore, the effects of targeting mTORC1 and mTORC2 signaling on migration, apoptosis of osteosarcoma cell and tumor growth and metastasis will be compared using osteosarcom cellular and xneograft animal models.The study will provide theoretical and experimental evidence for mTORC2-based targeted therapy in treatment of osteosarcoma.

哺乳动物雷帕霉素靶蛋白(mTOR)是促进细胞生长增殖的关键分子并在多数肿瘤中过度激活。mTOR以复合物(mTORC1/2)形式存在并发挥功能。由于抑制mTORC1可负反馈激活Akt，雷帕霉素类似物靶向mTORC1治疗多数肿瘤包括骨肉瘤效果不佳。最近，靶向mTORC1/2的第二代抑制剂成为mTOR靶向治疗的研究热点，但mTORC2在骨肉瘤发生进展中的作用以及靶向mTORC2治疗骨肉瘤尚未见报道。课题组在前期研究中发现抑制mTORC2而不是mTORC1可明显抑制骨肉瘤细胞迁移并诱导其发生凋亡。本项目拟通过骨肉瘤病例及标本，研究mTORC2成员表达及活性与骨肉瘤发生进展的关系；利用细胞与动物模型，比较靶向抑制mTORC2与mTORC1对骨肉瘤细胞迁移与凋亡，以及骨肉瘤生长与转移的影响。从分子、细胞与整体水平探讨mTORC2在骨肉瘤发生发展中的作用，为靶向mTORC2治疗骨肉瘤提供理论与实验依据。

人类许多恶性疾病通常发生哺乳动物雷帕霉素靶(mTOR)信号失调,这使它成为治疗癌症的一个潜在的靶点。但是,特异性抑制哺乳动物雷帕霉素靶复合物2(mTORC2)对骨肉瘤的效果尚未报道。本研究用三种骨肉瘤细胞系(MG63 / U2OS / Saos-2)。用mTOR抑制剂PP242和靶向siRNA抑制mTORC2。通过划痕实验和Transwell实验评估抑制迁移效果。单独用mTORC2抑制剂或联合顺铂处理骨肉瘤细胞,PI染色评估细胞凋亡;评估PARP和caspase 7表达水平。激酶抑制剂或敲除rictor靶向抑制mTORC2促进顺铂诱导凋亡,但雷帕霉素或raptor敲除抑制mTORC1并促进顺铂诱导凋亡。此外,抑制mTORC2而不是抑制mTORC1有效阻止骨肉瘤细胞迁移。这些结果表明,抑制mTORC2比mTORC1抑制剂治疗骨肉瘤更有优势。此研究为mTORC1/2抑制剂或mTORC1/2抑制剂联合顺铂治疗骨肉瘤提供了一个有利的实验依据。