Targeting AR and AR-Variants in Castration-Resistant Prostate Cancer

项目摘要/摘要 目前的AR抑制剂(苯扎鲁胺和阿比特龙)抑制雄激素向 二氢睾酮(DHT)，阻断雄激素与雄激素受体(AR)或AR信号的结合。虽然 这些治疗最初有效，但最终由于AR-剪接变异体(AR-SVS)等因素而失败 AR对雄激素以外的刺激有反应。控制发病和治疗的有效途径 抗去势前列腺癌(CRPC)的研究是开发抑制AR表达的小分子 或者促进它的退化。如果没有AR，则无论是否或以何种方式，都不会有AR信号 CRPC中存在许多雄激素或AR信号的其他刺激。我们的初步研究表明 天然产生的尿素素A(UroA)抑制AR信号转导，选择性抑制肿瘤细胞生长 AR阳性(AR：IC50~35M)而AR阴性(AR-：IC50~70µM)CRPC的体外和体内实验 模型组(50 mg/kg/体重)。由于需要更高浓度的尿素来抑制CRPC的生长， 我们的目标是设计和开发有效的UROA类似物，在较低的浓度下选择性地抑制AR激活 浓度(NM)，口服生物利用度，并抑制CRPC的生长。后续的发展 一系列UROA类似物和结构-活性关系(SAR)研究导致鉴定出两个 新型先导化合物ASR-600和ASR-603(美国临时62/941588)表现更好 在抑制CRPC细胞AR信号转导方面，NM比Uroa(ASR-600：40倍和ASR-603：&gt；12倍)有效 台词。分子研究表明，这两种ASR通过泛素化抑制AR和AR-SVS的表达 以及胞质内的降解。分子动力学(MD)系统模拟研究、生物物理分析 基于核磁共振和热位移的分析表明，这些小分子 结合AR的N-末端结构域(NTD)，阻断AR和AR-SV的激活。在生理上 可达到的浓度，ASR-600抑制AR和AR-SV(C4-2B和22Rv1)的肿瘤生长 异种移植物。根据我们的初步结果，我们假设，合理设计的ASR将有效地 通过直接靶向AR-NTD抑制和/或促进AR和AR-SV表达的降解 抑制CRPC的生长。我们将通过以下具体目标来检验这一假设。目标 1.探讨天冬氨酸受体对AR CRPC细胞的作用机制。目的2.确定体内信号转导 AR和AR-SVS的表达机制及ASRs剂量优化。目标3.确定治疗方法 ASR在原位和患者来源的异种移植(PDX)模型中抑制CRPC生长的有效性。这个 拟议的研究将阐明ASR的作用机制，并有助于更好地理解 这些化合物对CRPC的化疗性能。

Project Summary/Abstract Current AR inhibitors (enzalutamide and abiraterone) inhibit either the conversion of androgen to dihydrotestosterone (DHT), block androgen binding to androgen receptor (AR), or AR signaling. Although initially effective, these treatments eventually fail because of factors such as, AR-splice variants (AR-SVs) and AR responding to stimuli other than androgen. An effective way to control the emergence and treatment of castration-resistant prostate cancer (CRPC) is to develop small molecules that either inhibit AR expression or promote its degradation. If there is no AR, there would be no AR signaling, regardless of whether or how much androgen or other stimuli of AR signaling are present in CRPC. Our preliminary studies demonstrated that naturally occurring Urolithin A (UroA) inhibited the AR signaling and selectively suppressed the growth of AR-positive (AR+: IC50 ~35 M) but not AR-negative (AR-: IC50 ~70 µM) CRPC in both in vitro and in vivo models (50 mg/kg/body weight). As higher concentrations of UroA are required to suppress CRPC growth, we aimed to design and develop potent UroA analogs that could selectively inhibit AR activation at lower concentrations (nM), are orally bioavailable and suppress the growth of CRPC. The subsequent development of a series of UroA analogs and structure-activity relationship (SAR) studies led to the identification of two novel lead compounds ASR-600 and ASR-603 (US Provisional 62/941588) which demonstrated better efficacy (nM) than UroA (ASR-600: >40-fold and ASR-603: >12-fold) in inhibiting AR signaling in CRPC cell lines. Molecular studies suggest these two ASRs inhibit both AR and AR-SVs expression via ubiquitination and degradation within the cytosol. Molecular dynamic (MD) system simulation studies, biophysical analysis based on nuclear magnetic resonance (NMR), and thermal shift assays showed that these small molecules bind to the N-terminal domain (NTD) of AR and block the activation of both AR and AR-SV. At physiologically achievable concentrations, ASR-600 abrogated AR+ and AR-SV (C4-2B and 22Rv1) tumor growth in xenografts. Based on our preliminary results, we hypothesized that ‘Rationally designed ASRs will effectively inhibit and/or promote the degradation of both AR and AR-SV expression by directly targeting the AR-NTD and inhibit the growth of CRPC. We will test this hypothesis with the following specific aims. Aim 1. Investigate the mechanism of action of ASRs on AR+ CRPC cells. Aim 2. Determine in vivo signaling mechanism of AR and AR-SVs expression and optimize the dose of ASRs. Aim 3. Determine the therapeutic efficacy of the ASRs to inhibit CRPC growth in orthotopic and patient-derived xenograft (PDX) models. The proposed studies will elucidate the mechanism of action of the ASRs and lead to a better understanding of the chemotherapeutic properties of these compounds against CRPC.