长链非编码RNA-TROJAN通过调控NKRF促进腔面型乳腺癌对CDK4/6抑制剂耐药机制研究

Luminal type breast cancer patients occupy more than two-thirds of all breast cancer patients. Luminal type breast tumors have high degree of intratumor heterogeneity and cumulative annual rate of relapse. Although the potential improvements afforded by cyclin D-cyclin dependent kinase 4/6 (CDK4/6) kinase inhibitors, most patients ultimately lead to disease progression. Detailed researches of the resistance to CDK4/6 inhibitors mechanisms and screening the therapeutic targets are the basis of luminal breast cancer treatment. We screened out one long non-coding RNA, TROJAN, which was highly expressed in luminal breast cancer, correlated with poor survival, could approve the proliferation and the resistance to CDK4/6 inhibitor. Through RNA pull down as well as functional assays, TROJAN could bind to NKRF and enhance the oncogenic process through NKRF. The purpose of this study is to study the role of TROJAN regulate the resistance to CDK4/6 Inhibitors through NKRF. The methods include co-immunoprecipitation, next-generation sequencing, animal models and analysis of clinical data. We also expect to find the potential therapeutic targets in luminal breast cancer.

腔面（luminal）型乳腺癌占所有乳腺癌的2/3，其肿瘤内部异质性强、具有较高远期复发以及转移风险。尽管细胞周期蛋白依赖性激酶CDK4/6抑制剂可以改善患者预后，但多数患者最终仍产生耐药。阐明CDK4/6抑制剂耐药机制并寻找相关治疗靶点是改善luminal型乳腺癌患者预后的重要前提。申请者前期筛选出luminal型乳腺癌中高表达且与较差预后相关的长链非编码RNA-TROJAN。初步研究发现，TROJAN可促进细胞增殖以及CDK4/6抑制剂耐药；而TROJAN可与NKRF相互作用，共同调控肿瘤细胞增殖。我们提出假说：TROJAN与NKRF复合体可以通过推进细胞周期促进CDK4/6抑制剂耐药。在此基础上，本研究拟以“TROJAN-NKRF-CDK4/6抑制剂耐药”为研究线索，从细胞、分子、动物以及组织多个层面研究机制，为寻找luminal型乳腺癌相关治疗靶点提供理论和实验依据。

腔面型乳腺癌是一类雌/孕激受体阳性的乳腺癌，占所有乳腺癌的2/3。细胞周期信号通路激活是腔面型乳腺癌中驱动增殖的主要机制。尽管靶向细胞周期蛋白依赖性激酶CDK4/6的抑制剂可以改善患者预后，但多数患者最终仍产生耐药，导致预后不佳。阐明CDK4/6抑制剂耐药机制并寻找相关治疗靶点是改善腔面型乳腺癌患者预后的重要前提。申请者筛选出腔面型乳腺癌中高表达且与较差预后相关的长链非编码RNA-TROJAN。申请人通过一系列体外、体内、患者组织多个层面，论证TROJAN在腔面型乳腺癌中高表达、促进细胞增殖、且对CDK4/6抑制剂耐药；在分子机制层面，通过RNA pull down实验揭示了TROJAN可与NKRF相互作用。TROJAN通过NKRF转录抑制因子作用，共同调控细胞分子CDK2的表达，从而促进腔面型乳腺癌细胞增殖、CDK4/6抑制剂耐药。通过靶向TROJAN的反义寡核苷酸链（ASO），可以有效抑制TROJAN的表达，阻断其分子功能，抑制腔面型乳腺癌细胞增殖、并恢复对CDK4/6抑制剂敏感性。本项目已按计划完成预设的研究目标，明确TROJAN与NKRF调控腔面型乳腺癌CDK4/6抑制剂耐药的具体机制，开发了靶向TROJAN的药物ASO，完成临床前期探索，为CDK4/6耐药腔面型乳腺癌患者的治疗靶点开发提供了理论依据，同时发表高质量论著7篇。