核受体LRH-1在前列腺癌干细胞干性维持中的作用及分子机制研究

Currently, Castration-resistant prostate cancer (CRPC) is an incurable disease and represents a major clinical hurdle. Although the exact mechanisms underlying the progression to hormone-independent stage are still incompletely understood, emerging evidence has shown that prostate cancer stem cells (CSCs) endowed with self-renewal capacity and differentiation potential (stemness) may be involved in prostate tumor initiation, recurrence and progression to castration-resistant stage. Our previous results showed LRH-1 exhibited up-regulated expression pattern in enriched prostate cancer stem cells and CRPC xenograft model. Functional studies revealed that LRH-1 could promote stem-like properties in prostate cancer cells, as its over-expression enhanced the self-renewal capacity and conferred resistance to anti-androgen drugs (bicalutamide), chemo-therapeutic agents (paclitaxel) and androgen deprivation condition (charcoal-stripped serum) in prostate cancer cells, whereas its knockdown or inactivation triggered self-renewal arrest in vitro and suppressed tumourigenicity in vivo. Moreover, we demonstrated that OCT4, a vital non-cell surface marker for CSCs, could be transactivated by LRH-1. Based on these intriguing results, we hypothesize that LRH-1 may play a tumour-promoting role in the malignant growth of prostate cancer through its maintenance of CSCs via its transactivation of the OCT4 gene. This proposal will achieve the following goals: (1) to determine the functional role of LRH-1 in regulating the stemness of prostate cancer stem cells by “loss-of-function” and “gain-of-function” study or GFP-tracking system, (2) to explore and identify the mechanism underlying the LRH-1-mediated maintenance of stemness in prostate cancer stem cells by bioinformatics analysis, luciferase reporter assay, ChIP assay and rescue experiments. (3) To assess the therapeutic significance and application of targeting LRH-1 in CRPC treatment via inhibition of LRH-1 activity in vivo by a LRH-1-specific inverse agonist or knockdown of LRH-1 expression in vivo by siRNA. We expect through this study, it could help to provide a better understanding on the mechanisms responsible for the enhanced stemness in CRPC progression and resistance to hormone- or chemo-therapy in CRPC patients, and also to appraise the potential value of LRH-1 as a novel therapeutic target in CRPC treatment.

去势抵抗性前列腺癌（CRPC）是前列腺癌治疗的难点。既往研究证实：前列腺癌干细胞在CRPC发生和转化过程中发挥重要作用，但其分子机制尚不明确。本课题前期研究结果提示：LRH-1在具有干性的前列腺癌细胞中高表达；干预LRH-1表达可影响前列腺癌干细胞的功能；同时，LRH-1可调控肿瘤干细胞关键因子OCT4的表达。我们推测：LRH-1可能通过诱导OCT4的表达参与前列腺癌干细胞干性的维持，进而促进CRPC的发生发展。本项目拟：（1）通过干预LRH-1的表达或活性进一步阐明LRH-1调控前列腺癌干细胞干性的功能；（2）验证LRH-1通过OCT4调控前列腺癌干细胞干性的机制；（3）通过干预CRPC移植瘤内或携瘤小鼠体内LRH-1的表达或活性，评估LRH-1作为CRPC治疗靶点的临床转化价值。本研究成果将首次阐明LRH-1维持前列腺癌干细胞干性的作用和机制，为CRPC治疗新靶标的开发提供科学依据。

前列腺癌是欧美等发达国家男性最常见的恶性肿瘤之一。随着人口老龄化和饮食结构西方化，我国前列腺癌的发病率也逐年上升。早期前列腺癌多属于雄激素依赖性肿瘤，主动监测，根治性手术和放疗是局限性前列腺癌的主要治疗手段，然而，大约1/3的患者会发生复发或转移，需要接受内分泌治疗。但是，绝大多数患者都会在接受内分泌治疗后2-3年内进展为雄激素非依赖性前列腺癌，临床上称为去势抵抗性前列腺癌 (CRPC)。CRPC预后很差，死亡率高，是临床上亟待攻克的难题。CRPC的发生机制尚不明确。既往研究发现：肿瘤干细胞在CRPC发生和转化过程中发挥重要作用。肿瘤干细胞是存在于肿瘤组织中，数量极少 (<1%)，具有自我更新、多向分化潜能和高致瘤性的细胞亚群，其对放疗、化疗及内分泌治疗等常规治疗手段不敏感，常被视为肿瘤发生、进展、复发和转移的源动力。 本项目从临床、动物、细胞和分子水平深入剖析核受体LRH-1在CRPC发生发展中的功能和机制。我们的研究结果发现：核受体LRH-1在CRPC细胞系、动物模型和临床组织中高表达，与患者预后负相关；此外，LRH-1在具有干性的前列腺癌细胞系和临床标本中高表达。功能研究显示： 敲低LRH-1表达或反向激动剂抑制LRH-1活性可抑制前列腺癌干细胞的干性，而LRH-1过表达可增强前列腺癌细胞的干性。机制研究表明：LRH-1可直接调控干细胞关键因子OCT4的表达参与前列腺癌干细胞干性的维持；而雄激素受体AR可负调控LRH-1的表达。最后，我们发现：LRH-1特异性反向激动剂ML-180 可抑制CRPC移植瘤的体内生长。因此，我们得到如下结论：LRH-1作为治疗靶标，不仅可以降低前列腺癌细胞内雄激素的合成进而清除AR阳性的主群细胞 （之前已发表文章， Cancer Research 2018），还能抑制肿瘤干细胞的干性从而杀伤AR阴性的前列腺癌干细胞 (本课题)，进而在最大程度上消灭肿瘤细胞，预防复发和进展，为前列腺癌，尤其CRPC的靶向治疗提供新的临床策略和科学依据。 在本项目的资助下，项目负责人以第一作者或通讯作者发表SCI发表论文3篇，培养硕士研究生两名。