四跨膜蛋白TSPAN9介导酸性微环境下的自噬效应促进胃癌5-FU耐药的机制研究

Resistance to 5-fluorouracil (5-FU) of gastric cancer, is the main reason of treatment failure. Studies on mechanisms of resistance to 5-FU mostly focused on cancer cells rather than their microenvironment. We analyzed the biopsy samples and found that resistant tumor had lower intercellular pH than that of sensitive ones, whilst tetraspanins TSPAN9 overexpressed. We then simulated in vivo acidic microenvironment and established 5-FU-resistant gastric cancer cell line named SGC-7901/R, which characterized by TSPAN9 overexpression and high level of autophagy. After silencing of TSPAN9 by RNAi in SGC-7901/R, intercellular acidity and intracellular autophagy level were both declined, followed with reversed-chemoresistance to 5-FU of the cell. These results indicated that TSPAN9 may promote 5-FU drug-resistance of gastric cancer by regulating autophagy in tumor acidic microenvironment. The present project intends to use real-time PCR, western blot, RNA interfering and laser confocal microscope etc. to investigate the cross-actions among acidic microenvironment, autophagy pathway and 5-FU resistance of gastric cancer, and define the mechanisms of tetraspanins TSPAN9 in promoting drug-resistance to 5-FU of gastric cancer through regulating autophagy in tumor acidic microenvironment. This study may provide new thoughts in finding effective targets on chemoresistance reverse and personalized treatment.

胃癌5-FU类药物化疗常因耐药而导致治疗失败。相关机制研究多集中于耐药肿瘤细胞，而对其微环境较少涉及。课题组对活检标本检测显示，耐药患者胃癌组织的微环境pH比敏感者低，四跨膜蛋白TSPAN9过量表达。随后模拟体内酸性微环境建立的5-FU耐药胃癌细胞株SGC-7901/R，其TSPAN9表达上调且自噬水平增加；RNAi沉默SGC-7901/R 中TSPAN9的表达后，细胞外微环境pH升高，胞内自噬水平减少，5-FU耐药被逆转，提示TSPAN9可能通过调控酸性微环境下的自噬而导致胃癌5-FU耐药。课题组拟通过Real-time PCR、Western blot、RNA干扰和激光共聚焦显微镜等方法，分析TSPAN9与酸性微环境、自噬通路间的关系及在胃癌5-FU耐药中的交互作用，明确"TSPAN9介导酸性微环境下的自噬促进胃癌5-FU耐药"的机制，为寻找有效的逆转耐药靶点和个体化治疗提供新思路。

胃癌是最常见的消化道肿瘤之一，并且仍然是世界上第五大常见肿瘤，也是癌症相关死亡的第二大重要原因。Tetraspanins是一种进化上保守的四跨膜蛋白超家族，具有四个跨膜区段，一个小的细胞外区域和一个大的细胞外环。我们发现TSPAN9可以影响胃癌细胞的侵袭和迁移，而EMILIN1可以协同促进TSPAN9的抗癌作用。因此，在检测TSPAN9的抗癌作用以及挖掘其信号通路的过程中，我们的研究发现，在胃癌细胞中，通过过表达和敲低TSPAN9来抑制FAK-Ras-Erk1/2途径，从而影响了胃癌细胞的侵袭和迁移。在细胞体外实验中，EMILIN1通过促进TSPAN9的表达，协同调节了胃癌细胞的侵袭和转移。该研究为胃癌的靶向治疗提供了理论和实验依据。