食管鳞癌细胞通过miR-140-3p/NFYA/MDR1轴诱导化疗耐药的机制研究

多药耐药和异质性是导致食管癌治疗失败的重要原因。外泌体可介导细胞间物质交换和信号传导，miRNA是其重要组成部分，并与肿瘤耐药密切相关。课题组前期通过组学测序发现miR-140-3p在食管鳞癌耐药细胞系中高表达，细胞学实验提示miR-140-3p过表达促进耐药，抑制其表达会逆转耐药。NF-YA是miR-140-3p候选靶基因，可调控MDR1表达。同时发现miR-140-3p在耐药细胞外泌体中高度富集，应用耐药细胞上清培养敏感细胞可增加耐药性。我们推测，耐药细胞外泌体miR-140-3p可能通过自分泌或旁分泌靶向耐药及非耐药细胞NFYA进而上调MDR1表达，从而获得耐药。本项目拟通过细胞学实验、裸鼠荷瘤实验、临床标本检测对这一假设进行研究，探索外泌体miR-140-3p在ESCC临床化疗疗效预测中的作用。本课题对研究食管癌耐药机理、寻找新的预后和疗效预测标记物、实现个体化治疗具有重要意义。

Multidrug-resistance affects the effect of chemotherapy and leads a poor prognosis in ESCC patients. As mediators of intercellular communication, exosomes have been shown to be involved in many physiological and pathologic courses. Exosomes contain proteins, mRNAs, and microRNAs (miRNA), which could be transferred to target cells inducing epigenetic changes including drug-resistance. Our previous study showed miR-140-3p is increased in drug-resistant ESCC cell lines than in drug-sensitive cell lines via small-RNA sequence. And exogenous miR-140-3p increased the resistance of ESCC cells to chemotherapy, while inhibition of endogenous miR-140-3p leaded to reversal of drug resistance. Bioinformatics predicted NF-YA is a potential target of miR-140-3p and NF-YA can regulate the expression of MDR1 (multi drug resistance gene-1). Furtherly we found miR-140-3p was enriched in the exosomes of the drug-resistant ESCC cells, which supernatant increased the resistance of sensitive ESCC cells to chemotherapy. So we assumed that drug-resistant ESCC cells can release miR-140-3p into exosomes and transmit chemoresistance to the non-resistant cells via exosomes by targetting NFYA and then up-regulate the expression of MDR1. This proposal aims to validate the hypothesis and confirm the role of exosomes and miR-140-3p in drug-resistance of ESCC by MTT and zoopery, and further reveal it’s molecular mechanism. These results will facilitate the study of multidrug-resistant mechanism, clinical individual therapy, and precision medicine of ESCC in Chinese population.