STIM1在胃癌增殖和多药耐药中的功能研究

Multi-drug resistance is a major cause of gastric cancer (GC) chemotherapy failure, and its mechanisms are still unclear. As the ubiquitous intracellular signal transduction composition, Ca2+ plays an important role in the development and progression of tumors. In nonexcitable cells, especially the tumor cells, store-operated Ca2+ entry (SOCE) is the predominant Ca2+ entry mechanism. It has now been confirmed that STIM1 is one of the most important compositions of SOCE. Our previous studies indicate that STIM1 is significantly downregulated in GC cells and drug-resistant GC cells, and plays a inhibiting role in tumor cell proliferation and drug resistance. We also found that there may be multiple regulatory relationships between chemotherapy,miR-185 and STIM1 which may be the cause of the dysregulated STIM1. In this project we will use RIP and gene reporter systerm to further test and verify the multiple regulation of STIM1. Moreover, we will use calcium imaging, truncation /mutation, stimulant/ inhibitor and other experimental method to study the role and its exact mechanism of SOCE and Ca2+ in the GC proliferation and drug resistance inhibited by STIM1. And clear is there any non-SOCE functionality mode of STIM1. Together, the current project will provide a novel theory and empirical basis for the treatment of gastric cancer.

多药耐药是胃癌化疗失败的主要原因，其根本分子机制仍不清楚。钙离子作为细胞内最普遍的信号转导成分在肿瘤发生发展过程中起重要作用。钙库操纵的钙内流(SOCE)是非兴奋细胞特别是肿瘤细胞最主要的钙离子进入机制，现已证实STIM1是触发SOCE最关键的功能分子之一。我们前期的研究证实STIM1在胃癌细胞及胃癌耐药细胞中表达降低，可发挥抑制增殖及调节多药耐药的作用，并发现化疗/miR-185 /STIM1之间可能存在的多重调控关系是STIM1表达失调的原因。本项目将通过RIP、荧光报告基因等实验进一步研究验证STIM1的多重调控机制。通过钙离子成像技术、截短/突变、兴奋/抑制实验等深入探讨SOCE和钙离子在STIM1调控增殖耐药过程中所发挥的作用及其具体机制，并明确STIM1是否存在非SOCE的功能模式，为干预胃癌增殖耐药提供新的理论依据和实验基础。

多药耐药是胃癌化疗失败的主要原因，其根本分子机制仍不清楚。钙离子作为细胞内最普遍的信号转导成分在肿瘤发生发展过程中起重要作用。钙库操纵的钙内流(SOCE)是非兴奋细胞特别是肿瘤细胞最主要的钙离子进入机制，现已证实STIM1是触发SOCE最关键的功能分子之一。我们前期的研究证实STIM1在胃癌细胞及胃癌耐药细胞中表达降低，可发挥抑制增殖及调节多药耐药的作用，本项目通过利用免疫组化、qRT-PCR、Western blot技术、STIM1过表达及 siRNA 慢病毒载体的构建、荧光报告基因、细胞内钙离子检测、WST-1增殖实验、平板克隆形成实验、裸鼠皮下成瘤实验、WST-1实验、流式细胞术凋亡检测、裸鼠体内药敏实验等技术证实，在胃癌细胞系及胃癌耐药细胞中，化疗/miR-185 /STIM1 多重机制参与了STIM1 的表达调控。明确了STIM1/ORAI1/SOCE 在胃癌增殖、耐药中发挥重要调节作用，并发现其作用机制与内质网应激（未折叠蛋白反应（UPR））及线粒体功能相关，此外，我们还证实 STIM1 可能通过不依赖于SOCE 的其他途径，对胃癌细胞增殖及耐药发挥作用。因此，此研究拓展了我们对胃癌生长及多药耐药机制的认识，STIM1相关调控通路有希望成为胃癌治疗、干预的新靶点。