乳腺癌耐药分子标志物Transgelin 2耐药功能与位点分析研究

Chemotherapy plays a very important role in the comprehensive treatment of breast cancer. However, the occurrence of multidrug resistance (MDR) greatly hindered the efficacy of chemotherapy drugs. Therefore, screening and identification of breast cancer resistance markers to clarify its functions and signal modes in the regulation of drug resistance in breast cancer. Using structural biology information of proteins predicts the active sites of action in order to design specific targeted inhibitors, which is considered the current problems to be solved. The previous study had successfully established paclitaxel multidrug resistant breast cancer cell line, finding that transgelin 2 was a MDR target for breast cancer therapy. However, the active sites of transgelin 2 were not entirely clear, leading that its specific lead compounds were hardly discovered. This study aimed to further determine the functional mechanism of MDR regulated by the abnormal expression of transgelin 2 in breast cancer from vitro, vivo and clinical three levels using molecular biology analysis methods. Base on structural biology information, the binding mode of small molecule compounds with transgelin 2 will be studied using analytical chemistry methods, and combined with molecular docking to foretell active sites of transgelin 2, which is promising to lay the foundation of innovative breast cancer resistant associated lead compounds.

化疗在乳腺癌治疗中发挥着重要作用，然而，多药耐药的产生极大阻碍了化疗药物疗效。因此，筛选鉴定乳腺癌耐药标志物，阐明其在乳腺癌耐药中的功能，利用结构生物学信息，预测活性作用位点，以设计发现特异性的靶向抑制剂是目前亟待解决的问题。我们已成功建立人乳腺癌紫杉醇耐药细胞系，发现Transgelin 2是乳腺癌耐药分子靶标，但其在乳腺癌耐药中的生物学功能及调控模式尚未完全明确，Transgelin 2的活性作用位点未知，相应的特异性先导化合物难以发现。本项目是既往工作的深入，拟通过多种分析手段从体外、体内及临床三种水平探讨Transgelin 2异常表达调控乳腺癌多药耐药的功能机理；并利用结构生物学信息，应用分析化学等手段研究小分子化合物与Transgelin 2作用模式，联合分子对接方法分析Transgelin 2的活性作用位点，为创新性乳腺癌耐药相关先导化合物的发现及研究奠定基础。

化学治疗是乳腺癌综合治疗方案中不可或缺的重要环节，然而，由化疗引发的多药耐药极大限制了化疗药物的疗效。本课题组前期通过低浓度紫杉醇持续诱导法建立了乳腺癌紫杉醇耐药细胞株MCF-7/PTX，采用蛋白组学方法筛选得到了Transgelin-2，即肌动蛋白结合蛋白2，其在耐药细胞MCF-7/PTX中的表达比敏感细胞MCF-7/S中高15.48倍，是潜在的乳腺癌耐药靶标，但其在乳腺癌耐药中的生物学功能及调控模式有待进一步探索，此外，Transgelin-2的活性作用位点及其靶向抑制剂有待开发研究。.本研究证明我们建立的乳腺癌紫杉醇耐药细胞MCF-7/PTX具有多药耐药（MDR）的特性，且PI3K/Akt信号通路在MCF-7/PTX细胞中激活，而线粒体凋亡通路被抑制；临床水平上发现，与癌旁及正常组织相比，Transgelin-2在癌组织中显著高表达；生物信息学分析表明，Transgelin-2与临床病理参数显著相关，此外，我们还发现Transgelin-2对乳腺癌的诊断具有一定的参考价值；细胞水平证明，Transgelin-2通过激活PI3K/Akt通路，抑制线粒体凋亡通路介导乳腺癌紫杉醇耐药；动物水平发现，高表达Transgelin-2增强乳腺癌细胞致裸鼠成瘤能力。.通过细胞膜色谱筛选逆转乳腺癌紫杉醇耐药的天然化合物丹皮酚及丹酚酸A，从体外阐明其可通过下调MCF-7/PTX细胞中的耐药分子标志物Transgelin-2，使PI3K/Akt通路失活，进而促进下游线粒体凋亡通路活化，从而逆转乳腺癌紫杉醇耐药。.此外，通过分析Transgelin-2活性位点，采用SYBYL-X分子对接虚拟模拟Transgelin-2与数据库中化合物结合作用，筛选出靶向Transgelin-2的化合物TSG14，发现其可以靶向Transgelin-2抑制乳腺癌细胞的增殖，诱导其凋亡，抑制细胞的EMT过程及侵袭转移能力，有望成为靶向Transgelin-2来提高乳腺癌治疗疗效的小分子抑制剂。.作为潜在的乳腺癌治疗靶标，Transgelin-2为开发新的乳腺癌靶向治疗药物及临床乳腺癌的个体化治疗提供了坚实的理论基础和科学依据。