ERG调节CRMP4异二聚体结合热休克蛋白HSP-90 启动或逆转CRPC的分子机制研究

Castration-resistant prostate cancer (CRPC) is the main cause for prostate cancer (PCa) metastasis, leading to the death of the patients. To date, the mechanism underlying these process remains obscure. In previous study, we discovered that long form of CRMP4 (LCRMP4) gene (isoform of CRMP4, forming heterodimer with CRMP4) could promote hormone sensitive PCa cell line ,LNCaP, to transform to the status of CRPC in two weeks whereas CRMP4 gene rather drive a CRPC cell line, C4-2, revert to the manner of hormone sensitiveness in two weeks. Moreover, we detected significant high level of LCRMP4 mRNA in all blood samples from metastatic CRPC patients compared with hormone naïve PCa patients. These findings, including that the specific amino acid sequence on N-terminal of LCRMP4 determines ERG-induced two antagonistic function based on the domains in CRMP4/LCRMP4, which turns on and turns off CRPC, provide us with an optimal model in vitro and in vivo to investigate the mechanism of CRPC through the interaction among ERG, CRMP4/LCRMP4, and HSP90, enhancing or inhibiting AKT-AR stability. In this study, Co-IP is used to analyze the combination affinity between CRMP4/LCRMP4 and HSP90. Then, another IP/WB technique is performed to check the stability of chaperone protein of HSP90 and its client protein, AKT and AR, leading to the activation of AR. Accordingly, the function of regional amino acid sequence of CRMP4/LCRMP4 is determined for the connection with AR amplification, which contributes to CRPC.

CRPC（去势抵抗性前列腺癌）的发生机制现仍不清楚。我们前期研究中发现，CRMP4异二聚体长链（LCRMP4）能使激素依赖性前列腺癌（PCa）细胞LNCaP短期内转变至激素非依赖；其短链（CRMP4）则使激素非依赖PCa细胞C4-2短期内转化为激素依赖。针对CRMP4和LCRMP4在N端氨基酸序列的不同和ERG对其调节后产生的相互拮抗作用，特别是LCRMP4在CRPC患者血液中高表达，我们提出ERG-CRMP4/LCRMP4-HSP90途径导致CRPC发生的新机制。本研究重点围绕ERG结合CRMP4/LCRMP4启动子后各自单体蛋白结构域改变，用Co-IP验证CRMP4/LCRMP4与PCa细胞的热休克蛋白HSP90结合,通过IP/WB检测AKT-AR复合物稳定性, 评估AR转录活性，阐明CRPC发生机制。并建立体外和体内模型证实上述设想，为临床治疗CRPC选择有效治疗靶点。

CRPC（去势抵抗性前列腺癌）的发生机制现仍不清楚。前期研究中发现CRMP4异二聚体长链（LCRMP4）能使激素依赖性前列腺癌（PCa）细胞LNCaP短期内转变至激素非依赖；其短链（CRMP4）能够使激素非依赖PCa细胞C4-2短期内转化为激素依赖。转录因子ERG能够分别通过Thr49与Thr13磷酸化位点结合LCRMP4与CRMP4，并且能够分别发挥正性/负性调控作用。本研究实验证明LCRMP4蛋白在前列腺癌细胞与动物模型中能够与HSP90蛋白促进AKT-AR蛋白结合稳定性并增加AR 核蛋白和PSA蛋白表达水平；而CRMP4与HSP90蛋白结合后能够发挥相反的作用；通过体外和体内模型证实ERG-CRMP4/LCRMP4-HSP90途径是前列腺癌发生CRPC的新机制；LCRMP4的N端82与84位氨基酸突变可逆转CRPC发生。本课题阐明CRPC的分子机制并为其临床治疗提供新靶点。本课题系列研究成果已经发表SCI论文3篇、核心内容已为转化医学重大成果提供实验数据。