Division of Anti-tumor Pharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 201203, China.
J Biol Chem. 2012 Apr 6;287(15):12132-41. doi: 10.1074/jbc.M111.302299. Epub 2012 Feb 15.
CXCL12/CXCR4 plays an important role in metastasis of gastric carcinoma. Rapamycin has been reported to inhibit migration of gastric cancer cells. However, the role of mTOR pathway in CXCL12/CXCR4-mediated cell migration and the potential of drugs targeting PI3K/mTOR pathway remains unelucidated. We found that CXCL12 activated PI3K/Akt/mTOR pathway in MKN-45 cells. Stimulating CHO-K1 cells expressing pEGFP-C1-Grp1-PH fusion protein with CXCL12 resulted in generation of phosphatidylinositol (3,4,5)-triphosphate, which provided direct evidence of activating PI3K by CXCL12. Down-regulation of p110β by siRNA but not p110α blocked phosphorylation of Akt and S6K1 induced by CXCL12. Consistently, p110β-specific inhibitor blocked the CXCL12-activated PI3K/Akt/mTOR pathway. Moreover, CXCR4 immunoprecipitated by anti-p110β antibody increased after CXCL12 stimulation and G(i) protein inhibitor pertussis toxin abrogated CXCL12-induced activation of PI3K. Further studies demonstrated that inhibitors targeting the PI3K/mTOR pathway significantly blocked the chemotactic responses of MKN-45 cells triggered by CXCL12, which might be attributed primarily to inhibition of mTORC1 and related to prevention of F-actin reorganization as well as down-regulation of active RhoA, Rac1, and Cdc42. Furthermore, rapamycin inhibited the secretion of CXCL12 and the expression of CXCR4, which might form a positive feedback loop to further abolish upstream signaling leading to cell migration. Finally, we found cells expressing high levels of cxcl12 were sensitive to rapamycin in its activity inhibiting migration as well as proliferation. In summary, we found that the mTOR pathway played an important role in CXCL12/CXCR4-mediated cell migration and proposed that drugs targeting the mTOR pathway may be used for the therapy of metastatic gastric cancer expressing high levels of cxcl12.
趋化因子 CXCL12/CXCR4 在胃癌转移中发挥重要作用。雷帕霉素已被报道能抑制胃癌细胞的迁移。然而，mTOR 通路在 CXCL12/CXCR4 介导的细胞迁移中的作用以及针对 PI3K/mTOR 通路的药物的潜力仍不清楚。我们发现 CXCL12 激活了 MKN-45 细胞中的 PI3K/Akt/mTOR 通路。用 CXCL12 刺激表达 pEGFP-C1-Grp1-PH 融合蛋白的 CHO-K1 细胞，导致磷脂酰肌醇（3,4,5）-三磷酸的产生，这为 CXCL12 激活 PI3K 提供了直接证据。siRNA 下调 p110β 而不是 p110α 阻断了 CXCL12 诱导的 Akt 和 S6K1 的磷酸化。一致地，p110β 特异性抑制剂阻断了 CXCL12 激活的 PI3K/Akt/mTOR 通路。此外，CXCR4 在用 CXCL12 刺激后与抗 p110β 抗体免疫沉淀增加，G(i)蛋白抑制剂百日咳毒素消除了 CXCL12 诱导的 PI3K 激活。进一步的研究表明，针对 PI3K/mTOR 通路的抑制剂显著阻断了 MKN-45 细胞对 CXCL12 触发的趋化反应，这主要归因于 mTORC1 的抑制，并与阻止 F-肌动蛋白重组以及下调活性 RhoA、Rac1 和 Cdc42 有关。此外，雷帕霉素抑制了 CXCL12 的分泌和 CXCR4 的表达，这可能形成一个正反馈环，进一步消除导致细胞迁移的上游信号。最后，我们发现表达高水平 cxcl12 的细胞对雷帕霉素在抑制其迁移和增殖方面的活性敏感。总之，我们发现 mTOR 通路在 CXCL12/CXCR4 介导的细胞迁移中发挥重要作用，并提出针对 mTOR 通路的药物可能用于治疗表达高水平 cxcl12 的转移性胃癌。
