MTA1介导的HER2/CD44信号通路在胃癌侵袭与转移过程中的作用及机制分析

Metastasis is one of main causes which result in the death of gastric cancer patients. Human epidermal growth factor receptor 2 (HER2) (neu/ErbB2) is overexpressed on many types of cancer cells, including gastric cancer cells. HER2 overexpression has been associated with metastasis and poor prognosis. However its detailed molecular mechanism is still not clear. We found that CD44 binds directly to HER2 with disulfide bond, which up-regulates the expression of metastasis-associated protein-1（MTA1） controlling miR-139/CXCR4. However, the detailed mechanism about HER2 and CD44 control miR-139/CXCR4 through regulating MTA1 expression is still unknown. This project plans to ensure that HER2 and CD44 induces deacetylation of histone H3 lysine 9, and suppresses transcription of microRNA-139 (miR-139) to inhibit expression of its target gene, C-X-C chemokine receptor type 4 (CXCR4) from cellular and animal levels, discuss the ubiquitination mechanism by which HER2 and CD44 control the MTA1 through cell and animal model levels, identify the important molecules accomplishing the ubiquitination mechanism with cellular cotransfection and functional identification of full clone of the promoter and analyze effects that this mechanism plays in the gastric cancer metastasis through function research in vivo and vitro. This project will provide theoretical values for the HER2 mediating molecular mechanism about gastric cancer metastasis and infer some targets for the molecular targeted therapy for gastric cancer.

转移是导致胃癌患者死亡的主要原因之一，HER2与CD44在此过程中发挥重要作用，我们在预实验中发现HER2与CD44共价连接，并有初步证据表明可能通过传递信号上调MTA1来控制miR-139/CXCR4，然而，HER2与CD44分子如何经MTA1实现对miR-139调控的具体分子机制尚属未知。本课题拟进一步从细胞和动物模型两个层次确证HER2与CD44对miR-139启动子区组蛋白H3第9位赖氨酸去乙酰化进行调控，进而下调miR-139表达水平，解除miR-139对其靶基因CXCR4的抑制作用，并探讨此信号通路中MTA1调控的泛素化机制，通过共转染、全长启动子区功能分析等研究手段鉴定实现此泛素化机制的重要分子，通过体内外功能学研究证实HER2和CD44介导的信号通路在胃癌转移中的作用。本研究对揭示HER2相关的胃癌转移分子机制具有一定的理论价值，对胃癌分子靶向治疗具有较强的提示意义。

胃癌是常见的恶性肿瘤，是消化系统最多发的肿瘤。胃癌早期并无典型表现，大多数患者在确诊时已属晚期并发生了多器官转移。转移是关系到胃癌患者生死攸关的瓶颈，胃癌一旦发生其他脏器转移则明显预后不良。因此，深入研究胃癌转移发生的分子机制，对于胃癌的靶向治疗有着重要的意义。大量细胞生物学实验证实，作为表皮生长因子受体家族的一员，HER2（neu, ErbB2）在肿瘤细胞中过度表达可抑制细胞的分化，促进细胞的增殖，而且HER2在肿瘤发生与转移中发挥重要的作用。我们发现HER2与CD44介导的对miR-139的下调有赖于miR-139启动子区域组蛋白去乙酰化水平的上调，HER2和CD44诱导的miR-139启动子区域组蛋白去乙酰化是由于MTA1表达上调并募集NuRD复合物引起的，且此机制是胃癌细胞独特的调控方式。在此基础上，我们观察到MTA1被泛素化途径所调控，同时XIAP能够调控MTA1，但这一调控是否经过泛素化途径实现尚待后续实验进一步证实。在完成原定工作计划的基础上，我们还进行了曲妥珠单抗耐药细胞株的鉴定，并且发现MKN7细胞中MTA1的表达水平不随曲妥珠单抗处理而下调，通过基因芯片证实了曲妥珠单抗敏感与耐药细胞株的差异表达谱分析；进一步的研究显示曲妥珠单抗敏感细胞株中HER2通过ERK通路调控XAF1，而曲妥珠单抗耐药细胞系中HER2通过AKT通路调控XAF1。我们在本实验中深入分析HER2、CD44通过调节MTA1泛素化过程、进而使miR-139启动子区发生去乙酰化来控制其靶分子CXCR4表达并促进胃癌转移过程的详尽分子机制，在此基础上借助体外模型阐明胃癌转移有关的HER2及相关分子精确调控网络，为未来进行体内研究提供前期的实验依据。