DNMT3A调控DDIT3影响胃癌细胞顺铂化疗抵抗的作用机制研究

Drug resistance often causes recurrence, metastasis and ineffectiveness of chemotherapy in patient with tumours. However, the underlying mechanism remains largely unclear. DNA methyltransferase 3A (DNMT3A) plays an important roles in gastric cancer (GC) progression, its high expression is positively correlated with tumor cell TNM stage, lymph node metastasis and poorer clinical outcomes in patients. In our preliminary results, we found that DNMT3A was highly up-regulated in cisplatin-resistance GC cells. Overexpression of DNMT3A significantly decreased the chemosensitivity of GC cells in response to cisplatin, accompanied with the changes of cellular function. Furthermore, cell apoptosis-induced factor DDIT3 was identified as a downstream target of DNMT3A. Mechanistic investigations suggested that DNMT3A silences DDIT3 in a manner that is both hypermethylation dependent and DNA methylation independent; methylation-independent silencing occurs via cooperation with HDAC1. The above results implied that DNMT3A is implicated in cisplatin-resistance of GC, and regulate DDIT3 in a variety of ways. Thus, we focus on the precise roles of DNMT3A-mediated cisplatin-resistance via regulation of DDIT3 in the levels of cell model, animal model and clinical samples. This study will provide not only an experimental basis for the mechanism of DNMT3A underlying drug resistance, but also a therapeutic target for the epigenetic intervention for GC.

细胞耐药是引起肿瘤侵袭转移和术后复发的重要原因，其分子机制有待进一步揭示。DNMT3A在肿瘤发生发展中起重要作用。我们以往研究发现，胃癌组织细胞中DNMT3A的异常高表达不仅与胃癌临床分期等显著相关，且能指示胃癌预后不良。顺铂耐药胃癌细胞中DNMT3A高表达，上调DNMT3A可明显降低胃癌细胞的顺铂敏感性。耐药相关靶基因DDIT3的表达受DNMT3A的DNA甲基化作用及DNMT3A结合HDAC1的调控。以上结果提示，DNMT3A可能通过调控DDIT3的表达促进胃癌细胞顺铂化疗抵抗。本项目在原有工作基础上，对DNMT3A影响胃癌耐药进行研究。拟从细胞模型、动物实验、临床样本等多个层面，通过多种实验方法分析DNMT3A在胃癌耐药中的可能作用，探究DNMT3A调控DDIT3表达参与胃癌细胞顺铂耐药的具体机制。通过此研究为理解胃癌耐药的分子机制提供实验基础，也为胃癌表观遗传策略干预治疗提供新靶点。

顺铂耐药是引起肿瘤侵袭转移和术后复发的重要原因之一，其分子机制有待研究完善。DNA甲基转移酶3A(DNMT3A)在胃癌发生发展中起重要调节作用。以往研究发现，胃癌组织细胞中DNMT3A的异常高表达与多项胃癌临床病理信息显著相关，且能指示胃癌预后不良，但其参与胃癌细胞耐药过程的作用和分子机制尚不明确。在本项目中，我们通过体内、外实验发现，DNMT3A在胃癌顺铂耐药细胞中高表达，其表达升高能够降低胃癌细胞对顺铂作用的敏感性，抑制细胞凋亡，促进增殖，诱导上皮间质转化，促进转移等；反之，当在耐药细胞中降低DNMT3A表达后能够减弱其所引起的恶性表型。进一步的分子机制研究发现，凋亡促进因子DDIT3作为DNMT3A重要的靶基因，受DNMT3A的直接结合进而DNA甲基化修饰而抑制转录。重要的是，这一过程需要组蛋白去乙酰化酶HDAC1协同，并且由转录因子c-Myc介导完成。在临床组织样本分析中，我们发现DDIT3在胃癌组织中低表达且其启动子甲基化水平较癌旁组织升高。DNMT3A高表达和DDIT3低表达患者组的生存时间短，且二者可以作为独立预后分子。综上结果表明，DNMT3A在胃癌顺铂耐药中发挥重要作用，其通过多种调控方式影响下游基因表达介导恶性表型。本研究为理解胃癌耐药的分子机制提供实验基础，也为胃癌表观遗传策略干预治疗提供新靶点。