NR5A2上调MDC1促进DNA损伤修复增强乳腺癌化疗耐药研究

Despite recent advances in the treatment of human breast cancer by chemotherapy, resistance to chemotherapeutic agents is still a major problem. However, the mechanisms of chemotherapy resistance have not been fully elucidated. Our preliminary experiments found that NR5A2 promoted chemoresistance in breast cancer. In addition, the interaction of NR5A2 and NCOA3 up-regulated the expression of MDC1, and attenuated DNA damage induced by chemotherapy drugs. This project will carry out the following work: 1) to verify the function of NR5A2 promotes breast cancer chemotherapy resistance by inducing MDC1 expression; 2) to clarify the mechanisms of NCOA3 interacts with NR5A2 and induces MDC1 expression, to verify the MDC1 gene promoter sites bound by NR5A2, to clarify the interaction domain between NR5A2 and NCOA3, to find some new factors which interacts with NR5A2 and regulate the transcription of MDC1, to clarify the mechanisms that the new factors interact with NR5A2 and regulate the expression of MDC1; 3) to investigate the homologous recombination repair and non homologous end of link repair mechanism by NR5A2 and MDC1, to find some new factors which interacts with MDC1 and promotes DNA repair, to clarify the mechanisms that the new factors interact with MDC1 and promote DNA repair. This study will provide a novel theory for the mechanisms of breast cancer chemotherapy resistance, and provide potential diagnostic markers and therapeutic targets for breast cancer.

化疗耐药是乳腺癌治疗中的难题，其机制仍未完全阐明。前期我们发现NR5A2促进了乳腺癌化疗耐药，初步表明NR5A2与NCOA3相互作用上调了MDC1的转录水平，并降低化疗药诱导的DNA损伤而介导耐药。本课题将开展的工作：1.完善NR5A2通过MDC1促进乳腺癌化疗耐药的功能验证；2.阐明NR5A2和NCOA3相互作用上调MDC1的机制：找出NR5A2结合在MDC1启动子的位点，阐明NR5A2与NCOA3互作结构域，挖掘与NR5A2互作调节MDC1转录的新因子，阐明新因子与NR5A2互作形式及调节MDC1表达机制；3.探讨NR5A2通过MDC1促进DNA损伤修复的机制，从同源重组修复和非同源末端连接修复机制探讨，搜寻与MDC1互作介导DNA修复的新因子，阐明新因子与MDC1互作形式与机制。本课题将为乳腺癌化疗耐药机制提供新的理论，为耐药乳腺癌提供潜在的诊断标志物及药物治疗靶点。

化疗耐药是乳腺癌复发的重要原因，耐药癌细胞DNA损伤修复能力的增强是导致耐药的重要机制之一。然而目前促进癌细胞DNA损伤修复能力的基因报道的较少。因而挖掘相关癌基因，并针对这些靶点开发靶向抑制剂药物对克服乳腺癌化疗耐药有着十分重要的意义。本项目中发现NR5A2促进了乳腺癌化疗耐药，并且证明了NR5A2与NCOA3相互作用上调了MDC1的转录水平，并降低化疗药诱导的DNA损伤而介导耐药。具体研究发现如下：1.通过细胞实验、动物成瘤实验以及临床病人标本分析，确切的证实了NR5A2通过MDC1促进乳腺癌化疗耐药；2.临床病理组织研究发现NR5A2和NCOA3在化疗后的乳腺癌复发的病人组织中表达显著增强，并且与病人的预后不良正相关。3.阐明NR5A2通过其LBD区与NCOA3相互作用同时募集AP1，从而形成转录复合物去上调MDC1的表达。4.发现了NR5A2结合在MDC1启动子区的DNA基序。5.定位了影响NR5A2转录活性的氨基酸位点S243和S238，并证明NR5A2的转录活性受该位点磷酸化调节。6.发现ATM磷酸化MDC1介导了NR5A2促进的化疗耐药，抑制ATM活性能够显著减少NR5A2对乳腺癌化疗耐药的增强作用。7.发现ATM抑制剂联合NR5A2抑制剂有显著的协同杀伤化疗耐药乳腺癌细胞的作用。8.探讨NR5A2通过MDC1促进DNA损伤修复的机制，从同源重组修复和非同源末端连接修复机制探讨，结果表明MDC1促进的化疗耐药与以上两种修复机制都有相关性。总之本课题的完成为乳腺癌化疗耐药机制提供新的理论，为耐药乳腺癌提供潜在的诊断标志物及药物治疗靶点。