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Interference with Androgen Receptor and Its Ligands

干扰雄激素受体及其配体

基本信息

批准号：
8243673
负责人：
JAMES L MOHLER
金额：
$ 34.77万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-04-01 至 2015-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8243673
关键词：
Adrenal Glands American Androgen Metabolism Androgen Receptor Androgens CYP17A1 gene Castration Cell Death Cells Clinical Clinical Trials Complex Cytochromes Disease remission Drug Formulations Enzymes Freezing Gene Expression Growth Immunohistochemistry In Vitro Infection Laboratories Learning Ligands Malignant neoplasm of prostate Mass Spectrum Analysis Measures Medical Castration Messenger RNA Metabolism Metastatic Neoplasm to the Bone Mountain Lion Mutation Operative Surgical Procedures Oxidoreductase Pathway interactions Patients Pharmaceutical Preparations Pharmacologic Substance Pre-Clinical Model Prostate Proteins Radioimmunoassay Recurrence Research Personnel Source Stanolone Stress Suspension substance Suspensions Testing Testosterone Time Tissue Sample Tissues Transactivation Xenograft Model Xenograft procedure abiraterone cancer cell cell growth deprivation experience in vivo killings men novel novel strategies response small hairpin RNA small molecule

项目摘要

Project 1 has contributed to two paradigm changing discoveries. First, the androgen receptor (AR) pathway appears critical to the growth of castration-recurrent prostate cancer (CaP) in spite of castrate levels of circulating testicular androgens. Second, castration-recurrent CaP produces high tissue levels of testosterone (T) and dihydrotestosterone (DHT), the preferred AR ligand. Intracrine-produced testicular androgens present a target for novel therapies. Further advances may result from applying novel treatments to CaP coincidental with androgen deprivation therapy (ADT) when CaP cell death is at a maximum and surviving cells are under greatest stress. The central hypothesis of the proposed studies is that CaP can be cured or remission extended by a coordinated attack upon intracrine androgen metabolism and AR coincidental with elimination of circulating testicular androgens (medical or surgical castration). In order to test this hypothesis and allow the formulation of an appropriate clinical trial in advanced CaP, the response to ADT will be assessed using preclinical models in the immediate post-castration period. In general, cell or tissue sampling will be conducted just prior to castration and 12h and 1, 2, 4, 8, 16 and 30d after "castration." The effect of castration upon the androgen axis will be assessed on 4 levels in vitro and 6 levels in vivo. The 4 levels of assessment in vitro and in vivo will include 1) changes in androgen metabolism enzymes at the mRNA level using qRT-PCR and protein level using immunohistochemistry (IHC); 2) androgen levels using LC-MS; 3) androgen-regulated gene expression using IHC for PSA, Nkx3.1 and hK2; and 4) cell growth. Measures in vivo will include 5) time to progression and 6) survival. Androgen metabolism after ADT will be studied using androgen-sensitive CWR22 cell suspensions, the androgen-dependent CWR22 xenograft and a fresh surgical tissue xenograft model. Changes in androgen metabolism critical for survival after ADT will be targeted using shRNA or drug in vitro and lentiviral shRNA and/or drug in vivo (Aim 1). Testicular androgens formed by intracrine metabolism of adrenal androgens will be removed using 53-reductase or Sult2A1 delivered using lentiviral infection (Aim 2). Finally, AR will be removed using lentiviral AR shRNA constructs or AR degrading small molecules (Aim 3).

项目1促成了两个改变范式的发现。首先，雄激素受体（AR）途径似乎对去势复发性前列腺癌（CaP）的生长至关重要，尽管去势水平的循环睾丸雄激素。其次，去势复发性CaP产生高组织水平的睾酮（T）和双氢睾酮（DHT），优选的AR配体。内分泌产生的睾丸雄激素是新疗法的靶点。当CaP细胞死亡达到最大值且存活细胞处于最大压力下时，将新的治疗方法与雄激素剥夺疗法（ADT）同时应用于CaP可能会产生进一步的进展。提出的研究的中心假设是，通过对分泌内雄激素代谢和AR的协调攻击，同时消除循环睾丸雄激素（药物或手术去势），可以治愈CaP或延长缓解期。为了检验这一假设并允许在晚期CaP中制定适当的临床试验，将在去势后即刻使用临床前模型评估ADT的反应。一般来说，细胞或在去势前和去势后12小时、1、2、4、8、16和30天进行组织取样。“阉割对雄激素轴的影响将在体外4个水平和体内6个水平上进行评估。体外和体内评估的4个水平将包括1）使用qRT-PCR在mRNA水平上检测雄激素代谢酶的变化，使用免疫组织化学（MHC）在蛋白质水平上检测雄激素代谢酶的变化; 2）使用LC-MS检测雄激素水平; 3）使用免疫组化检测PSA、Nkx3.1和hK 2的雄激素调节基因表达;和4）细胞生长。体内测量将包括5）进展时间和6）存活。将使用雄激素敏感性CWR 22细胞悬液、雄激素依赖性CWR 22异种移植物和新鲜手术组织异种移植物模型研究ADT后的雄激素代谢。将使用shRNA或体外药物和慢病毒shRNA和/或体内药物靶向ADT后对生存至关重要的雄激素代谢变化（目的1）。肾上腺雄激素分泌内代谢形成的睾丸雄激素将使用53-还原酶或使用慢病毒感染递送的Slt 2A 1去除（Aim 2）。最后，将使用慢病毒AR shRNA构建体或AR降解小分子去除AR（目的3）。