PI3K/AKT/mTOR通路对上皮细胞间质化的调控在胃癌化疗耐药中的作用及其机制研究

Poor prognosis of gastric cancer is mainly related to treatments resistance. And acquisition of EMT is correlated with enhanced invasiveness and treatments resistance. Emerging evidences showed that EMT phenotypes were related to intrinsic and acquired chemoresistance, and mesenchymal-subtype gastric cancer cells were sensitive to the inhibitors of PI3K/AKT/mTOR signaling. Previous studies showed that enhanced EMT phenotypes and decreased sensitivity of chemotherapy drugs were found in OXA resistant gastric cancer cell line, and that the patients with over-expression of AKT and mTOR exhibited resistant to adjuvant chemotherapy after gastrectomy. It suggests that gastric cancer chemoresistance should be associated with EMT via activation of the PI3K/AKT/mTOR pathway. In this study, we will investigate the association of EMT, chemoresistance and PI3K/AKT/mTOR pathway from cellular experiments, animal models and clinical study. In vitro and vivo, we will observe the changes of EMT phenotypes, biological characteristic and sensitivities of drugs when the activities of PI3K/AKT/mTOR pathway were inhibited by inhibitors, or the expressions of mTOR were regulated by mTORshRNA or pcDNA3.0-mTOR. Moreover, clinical study was performed to further verify the relation among EMT phenotypes, activities of PI3K/AKT/mTOR pathway, responses to chemotherapy and prognosis. It will contribute to understanding heterogeneity of gastric cancer, improving efficacy of chemotherapy, prolonging survival, and reversing chemoresistance.

化疗耐药是胃癌预后差的主要原因。EMT与肿瘤侵袭性增强及治疗抵抗密切相关。研究发现EMT参与肿瘤先天性及获得性耐药，且间充质亚型胃癌对PI3K/AKT/mTOR通路抑制剂敏感。前期研究发现OXA耐药的胃癌细胞发生EMT并对其他药物敏感性下降；且AKT和mTOR高表达与术后辅助化疗耐药相关。提示胃癌化疗耐药可能与EMT和PI3K/AmTOR通路相关。本研究将从细胞实验、动物实验和临床研究三个层面展开：体内外实验中，探讨EMT表型、细胞侵袭力与化疗药物敏感性的关系；并利用PI3K/mTOR抑制剂抑制通路活性、利用不同的mTOR表达载体上调或下调其表达，观察胃癌EMT表型、侵袭力及化疗药物敏感性变化；结合临床病例，分析高侵袭性及难治性胃癌EMT表型、PI3K/ mTOR通路状态及其与化疗疗效、预后的关系，加深对胃癌异质性及生物学特征的认识，为提高胃癌化疗疗效、逆转耐药及改善预后提供新思路。

化疗耐药是胃癌预后差的主要原因。上皮细胞间质化（EMT）被认为是实体肿瘤细胞浸润和转移的关键环节而备受关注。近年研究发现EMT与肿瘤侵袭性增强及治疗抵抗密切相关，并参与肿瘤先天性及获得性耐药，且间充质亚型胃癌对PI3K/AKT/mTOR通路抑制剂敏感。项目组前期研究发现OXA耐药的胃癌细胞发生EMT并对其他药物敏感性下降；且AKT和mTOR高表达与术后辅助化疗耐药相关。提示胃癌化疗耐药可能与EMT和PI3K/AmTOR通路相关。项目组从细胞实验、 动物实验和临床研究三个层面开展相关研究。研究发现：1)奥沙利铂（OXA）获得性耐药的SGC7901/OXA细胞株较亲本细胞株SGC7901的EMT标志物表达升高，肿瘤细胞迁移及侵袭能力增加，对OXA、紫杉醇的敏感性降低；证实耐药细胞株具有EMT表型；2) ZEB1 shRNA和PI3K/AKT/mTOR 通路抑制剂可使耐药细胞株上皮标志物表达增加，而间充质标志物表达减少，且化疗敏感性增加，提示PI3K/AKT/mTOR 通路参与调控 EMT 介导的胃癌化疗耐药的机制；3) 亲代细胞和耐药细胞株的全转录组测序分析显示，差异基因2570/26848（9.6%），其中1161个上调，1409个下调，除PI3K/mTOR通路外，MAPK, TGF-beta，Norch通路可能也参与了胃癌耐药的发生；4) 通过铂类耐药胃癌的原代肿瘤组织移植模型（PDX）发现，依维莫司+5FU组较5FU组移植肿瘤缩小明显，联合治疗组间充质标志物和pmTOR表达均低于单药5FU组；证实mTOR抑制剂可能通过调控EMT而逆转耐药；5) 胃癌患者的组织标本的免疫组化实验发现，mTOR高表达与高BMI，以及E-cadherin低表达相关，且均与较差的无疾病生存时间有关，进一步证实EMT表型，PI3K/mTOR通路的活化与高侵袭性和化疗耐药相关。上述发现为阐明胃癌的进展和耐药机制提供了证据，为为今后开展逆转耐药的临床研究提供理论基础，为临床实践提供新的治疗思路。