ALK磷酸化TOPK，促进ALK阳性肺癌发生的作用机制研究

The direct upstream and downstream signaling pathways of ALK are still unclear. Further exploration of its molecular mechanism of tumorigenesis is of great benefit in solving the problem of targeted drug therapy for ALK-positive lung cancer. We initially found that: 1) ALK directly phosphorylated tumor-associated protein kinase TOPK at Y74; 2) TOPK was highly expressed in ALK-positive lung cancer patients; 3) ALK was interacted with TOPK; 4) The phosphorylation of TOPK by ALK promoted cell proliferation and colony formation of lung cancer. Thus, we hypothesize that: The phosphorylation of TOPK by ALK promotes the tumorigenesis of ALK-positive lung cancer, and the following study would be performed to: 1) confirm that ALK interacts with TOPK, phosphorylates and activates TOPK at the Y74 site in vitro and in vivo; 2) identify that the phosphorylation of TOPK by ALK promotes tumorigenesis of lung cancer ex vivo and in vivo; 3) explore the substrates of TOPK and downstream signaling pathways by SILAC Quantitative Phosphorylation Proteomics, and confirm the analysis results in cell lines; 4) collect the clinical tissue samples, detect the expression level of TOPK and p-TOPK(Y74) and analyze their correlation with the prognosis of patients. This study will reveal the ALK-TOPK signaling pathway in ALK-positive lung cancer and may provide a new strategy for its molecular targeted therapy.

ALK直接上下游信号通路尚不清楚，进一步探索其导致肿瘤发生的分子机制对解决ALK阳性肺癌靶向药物治疗问题大有裨益。我们初步发现：1）ALK直接在Y74位点磷酸化肿瘤相关蛋白激酶TOPK；2）ALK阳性肺癌患者TOPK的表达高；3）ALK与TOPK相互作用；4）ALK磷酸化TOPK促进肺癌细胞的增殖和克隆形成。由此，我们提出假说“ALK磷酸化TOPK促进ALK阳性肺癌发生”，拟进行以下研究证实并探讨机制：1）体内外证实ALK与TOPK相互作用，且ALK在Y74位点磷酸化激活TOPK；2）细胞模型和动物模型证实ALK磷酸化TOPK促进肺癌的发生；3）SILAC标记技术探讨ALK-TOPK调控的下游信号分子及通路；4）扩大临床样本量，检测分析TOPK及p-TOPK（Y74）的表达与ALK阳性肺癌预后的相关性。这一研究将阐明ALK-TOPK信号通路促进肺癌发生的机制，为其分子靶向治疗提供新的策略。

TOPK是潜在的肿瘤治疗靶标，然而其在ALK阳性非小细胞肺癌中的作用尚未报道。本项目研究发现TOPK在ALK阳性非小细胞肺癌患者中高表达；体外激酶实验筛选到ALK是TOPK直接上游激酶，同时我们在体内外证实ALK可在Y74位点磷酸化TOPK，增强TOPK的稳定性促进肺癌细胞发生；本研究还证实抑制TOPK增强肺癌细胞对ALK靶向药艾乐替尼的敏感性，并且联用艾乐替尼和HI-032（TOPK抑制剂）可抑制ALK阳性非小细胞肺癌细胞的生长，促进其凋亡。本研究揭示了新的ALK-TOPK信号通路，联用二者抑制剂将为增强ALK阳性非小细胞肺癌的靶向治疗敏感性提供新的线索。