MiRNA-20a-5p家族调控乳腺癌耐受紫杉醇的作用机制研究及其临床意义

Breast cancer is the most common malignancy in women. Drug Resistance is a major obstacle to the successful treatment of the patients with breast cancer. In addition to the P-gp-mediated multiple drug resistance, recent studies shown that a class of cell-derived RNA molecules (microRNA) might also be involved in the regulation of chemoresistance. To explore the possible roles and underlying mechanisms of micro-RNAs in drug resistance. We recently established two strains of breast cancer drug-resistant cell lines with paclitaxel, Bats-72, Bads-200 from Human myeloid breast tumor BCap37 cell. Subsequently, through miRNA array and miRNA library screening technology, we identified two miRNA molecules miRNA-20a-5p and miRNA-106a-5p. Further analyses demonstrated that these two microRNAs belong to the same family and the mechanisms of which in breast cancer chemoresistant progress have not been illustrated before. Preliminary results have demonstrated that the expressions of two miRNAs are highly corresponded with chemoresistant conditions in clinical breast cancer patients. The up-regulation of miRNA-20a-5p and miRNA-106a-5p can efficiently reduce the drug resistance to PTX in cytological experiments in vitro, and had synergy effect. However, whether they can reverse chemoresistance in vivo and their downstream target regions remains unknown. In this study, we will comprehensively utilize techniques of molecular biology, cytobiology, bioinformatics and laboratory animal science to illustrate the target region of two miRNAs and related signal pathway. The ability of miRNA-20a-5p and miRNA-106a-5p on reversing chemoresistance in vivo will be further studied for developing new strategies for targeted therapies in patients with chemoresistant breast cancer.

乳腺癌是女性最常见的恶性肿瘤，其耐药性的形成是患者化疗失败的主要原因之一。肿瘤耐药是一个很复杂的过程，目前要在临床逆转耐药还存在诸多困难。已有报道表明miRNA参与调控细胞耐药。我们前期从髓性乳腺癌细胞BCap37出发，构建了两株耐药细胞，并通过miRNA芯片和miRNA文库技术筛出两个在亲代细胞和耐药细胞中差异表达的miRNA分子：miRNA-20a-5p及miRNA-106a-5p，二者属于同一家族，目前还未见二者调控乳腺癌耐药的报道。我们初步的实验结果表明二者的表达变化与临床乳腺癌病人的化疗效果显著相关，细胞实验也表明上调其表达可有效降低细胞对紫杉醇的耐药性，且二者联合作用有协同效果，但其作用机制以及其能否在体内逆转耐药还不清楚。因此我们拟综合利用多种生物实验技术，阐明该两个miRNA分子的作用靶点和相关的信号通路，考察其在体内逆转耐药的效果，为提高肿瘤化疗的效果提供可能的新的靶点。

乳腺癌是最常见的女性恶性腺癌之一，其化疗耐药是癌症病人死亡的主要原因之一，耐药的分子机制尝未完全阐明。已有研究表明，miRNAs可通过转录后水平调控基因的表达，从而参与调控肿瘤细胞的耐药性。.本项目构建了两株紫杉醇耐药Bcap37细胞，筛出两个在亲代细胞和耐药细胞中差异表达的miRNA分子：miRNA-20a-5p及miRNA-20b-5p。统计分析发现，miRNA-20a-5p及miRNA-20b-5p在乳腺癌组织中相比正常乳腺组织表达下调，在复发病人中表达更低，可作为预测病人预后的独立风险因子。体外实验表明，两者可显著抑制多种乳腺癌细胞及宫颈癌细胞增殖与耐药。小鼠体内实验表明，miRNA-20a-5p可显著增强紫杉醇对耐药乳腺癌的抑制作用，降低了其耐药性。.研究发现TGFBR2、PDGFRA、CCND1、E2F1及MAPK1可能是miRNA-20a-5p在调控乳腺癌细胞耐药与增殖过程中的靶基因。进一步的研究表明，该miRNA是通过抑制MAPK1来抑制细胞的增殖与耐药的。一方面，该miRNA可通过抑制MAPK1来抑制MAPK/ERK信号通路，从而抑制细胞生长，促进细胞凋亡；另一方面，MAPK1被抑制后，耐药蛋白P-gp也被抑制，从而降低了细胞的耐药性。.miRNA-20a-5p表达失调的的原因则是该miRNA自身受到基因调控区组蛋白甲基化及MAPK/ERK通路下游转录因子c-Myc的调控。一方面，在耐药细胞中，组蛋白甲基化失调导致该miRNA表达下降；另一方面，作为MAPK/ERK信号通路的下游分子，c-Myc促进该miRNA家族的表达，形成一个反馈环路。因此，肿瘤的进展将最终取决于MAPK1和c-Myc逃避该miRNA抑制的能力。.本项目研究结果将有助于加深我们对乳腺癌耐药的理解，并为乳腺癌的治疗提供潜在靶点。