Mechanism of promiscuous AR activation in PTEN-mutated prostate cancer

PTEN突变前列腺癌中AR混杂激活的机制

• 批准号: 8853827
• 负责人: Haojie Huang
• 金额: $ 32.72万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-06-01 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8853827
• 关键词: Androgen Receptor; Androgens; Animals; Binding; Biological; Castration; Cell Nucleus; Cells; Clinical Research; Cultured Cells; Data; Development; Disease; Disease Progression; Disease Resistance; EP300 gene; Exclusion; Genes; Genetic Transcription; Growth; In Vitro; Interleukin-6; Knock-out; Ligands; Light; Malignant neoplasm of prostate; Mediating; Metastatic Prostate Cancer; Modeling; Molecular; Mus; Mutate; Nuclear; Nuclear Receptors; PTEN gene; Pathway interactions; Patients; Phosphorylation; Play; Prostate; Prostatic; Prostatic Neoplasms; RNA; Receptor Activation; Regulation; Relapse; Residual state; Resistance; Role; STAT3 gene; Signal Pathway; Signal Transduction; Testing; Transactivation; Transcription Coactivator; Transcriptional Activation; Tumor Suppressor Genes; Xenograft Model; castration resistant prostate cancer; deprivation; design; in vivo; mutant; prostate cancer cell; protein complex; research study; response; standard care; targeted treatment; tumor

DESCRIPTION (provided by applicant): The androgen receptor (AR) belongs to the superfamily of nuclear receptors that become active in gene transactivation in response to their ligands. Because the AR plays a central role in growth and survival of prostate cancer (PCa) cells, androgen deprivation therapy (ADT) has been a standard treatment for advanced/metastatic PCa. Unfortunately, a majority of tumors invariably relapse and become a disease called androgen-deprivation independent or castration-resistant prostate cancer (CRPC), from which most patients eventually die. Increasing evidence obtained from both clinical and experimental studies indicates that the AR remains active in the absence or presence of residual (castration) levels of androgens and plays a pivotal role in the development of CRPC. These findings have elevated the need to understand the molecular mechanisms by which the AR becomes promiscuously activated following ADT. Our preliminary data demonstrate that loss of the PTEN tumor suppressor gene increases AR activity in both the presence of low levels of androgens and their complete absence. This effect of PTEN is mediated primarily by its downstream nuclear factor FOXO1. Silencing of FOXO1 in PTEN-positive cells not only increases AR basal activity, but also sensitizes the AR to activation by very low concentrations of androgens and nonandrogenic factors such as interleukin-6 (IL-6). In contrast, forced expression of FOXO1 in PTEN-null PCa cells inhibits both androgen-dependent and -independent activation of the AR. Mechanistically, FOXO1 directly binds to AR's transcription activation unit 5 (TAU5) domain, a region known to interact with a number of functionally important transcription coactivators. The AR inhibitory function of FOXO1 requires its nuclear localization but not its transcriptional activity. Most importantly, expression of the transcription-deficient but AR-inhibitory mutant of FOXO1 inhibits growth of CRPC cells. STAT3 is important for IL-6-induced activation of the AR through its interaction with AR. We demonstrate that forced expression of the nuclear form of FOXO1 diminishes activation of the AR by IL-6 and the STAT3-AR interaction. Moreover, our in vitro studies show that the increase of AR activity with PTEN loss and FOXO1 inactivation is completely abolished by treatment of cells with small inference RNA of p300, an important coactivator of the AR. Conditional knockout of p300 in the mouse prostate shows that deletion of p300 inhibits AR activity in vivo. The unifying hypothesis of this application is that in prostatic cells with a functional PTEN, FOXO1 localizes primarily in the nucleus where it forms a protein complex with the AR that limits the association of the AR with coactivators and thereby prohibits aberrant AR activation. In PTEN-mutated PCa cells, however, highly activated Akt leads to phosphorylation and nuclear exclusion of FOXO1, which results in unopposed access of the AR to the coactivators such as p300 and STAT3 that promotes promiscuous AR activation in the presence of castration levels of androgens or nonandrogenic factors (e.g. IL-6) and castration-resistant disease progression. We propose three specific aims to test this hypothesis: (1) to define the precise mechanisms that mediate the inhibitory effect of the PTEN/FOXO1 signaling axis on androgen- independent activation of the AR and castration-resistant growth of PCa cells using xenograft models; (2) to determine the importance of p300 in androgen-independent and yet AR-dependent progression of prostate tumors in the PTEN-knockout model; and (3) to elucidate the molecular mechanism and biological importance of the PTEN/FOXO1 pathway in regulation of IL-6-induced androgen-independent activation of the AR in PCa cells. Information obtained from this study will not only shed new light on the molecular mechanism of aberrant AR activation in CRPC, but also define new molecules and pathways for targeted therapy of this fatal disease.

描述（由申请人提供）：雄激素受体（AR）属于核受体超家族，其在响应其配体的基因反式激活中变得活跃。由于 AR 在前列腺癌 (PCa) 细胞的生长和存活中发挥着核心作用，因此雄激素剥夺疗法 (ADT) 已成为晚期/转移性 PCa 的标准治疗方法。不幸的是，大多数肿瘤总是会复发并成为一种称为雄激素剥夺无关型或去势抵抗性前列腺癌（CRPC）的疾病，大多数患者最终死于该病。从临床和实验研究中获得的越来越多的证据表明，无论是否存在残留（去势）雄激素水平，AR 仍保持活跃，并在 CRPC 的发展中发挥关键作用。这些发现提高了了解 ADT 后 AR 被混杂激活的分子机制的必要性。我们的初步数据表明，无论是在雄激素水平较低还是完全不存在的情况下，PTEN抑癌基因的缺失都会增加AR活性。 PTEN 的这种作用主要由其下游核因子 FOXO1 介导。 PTEN 阳性细胞中 FOXO1 的沉默不仅会增加 AR 的基础活性，还会使 AR 对极低浓度的雄激素和非雄激素因子（如白细胞介素 6 (IL-6)）的激活敏感。相反，在 PTEN 缺失的 PCa 细胞中强制表达 FOXO1 可抑制雄激素依赖性和非依赖性 AR 激活。从机制上讲，FOXO1 直接结合 AR 的转录激活单元 5 (TAU5) 结构域，该区域已知与许多功能上重要的转录共激活因子相互作用。 FOXO1 的 AR 抑制功能需要其核定位，但不需要其转录活性。最重要的是，转录缺陷但 AR 抑制的 FOXO1 突变体的表达会抑制 CRPC 细胞的生长。 STAT3 通过与 AR 相互作用，对于 IL-6 诱导的 AR 激活非常重要。我们证明 FOXO1 核形式的强制表达会减少 IL-6 和 STAT3-AR 相互作用对 AR 的激活。此外，我们的体外研究表明，用p300（AR的重要共激活剂）的小推理RNA处理细胞可以完全消除随PTEN丢失和FOXO1失活而增加的AR活性。小鼠前列腺中 p300 的条件性敲除表明 p300 的缺失会抑制体内 AR 活性。本申请的统一假设是，在具有功能性 PTEN 的前列腺细胞中，FOXO1 主要定位于细胞核，在细胞核中它与 AR 形成蛋白质复合物，限制 AR 与共激活剂的关联，从而阻止异常的 AR 激活。然而，在 PTEN 突变的 PCa 细胞中，高度激活的 Akt 会导致 FOXO1 磷酸化和核排斥，从而导致 AR 不受阻碍地接触共激活因子，例如 p300 和 STAT3，从而在雄激素去势水平存在的情况下促进混杂的 AR 激活或非雄激素因素（例如 IL-6）和去势抵抗性疾病进展。我们提出了三个具体目标来检验这一假设：（1）利用异种移植模型，确定介导 PTEN/FOXO1 信号轴对雄激素非依赖性 AR 激活和 PCa 细胞去势抵抗生长的抑制作用的精确机制； (2) 在 PTEN 敲除模型中确定 p300 在雄激素非依赖性和 AR 依赖性前列腺肿瘤进展中的重要性； (3) 阐明 PTEN/FOXO1 通路在调节 PCa 细胞中 IL-6 诱导的雄激素非依赖性 AR 激活中的分子机制和生物学重要性。从这项研究中获得的信息不仅将为CRPC中异常AR激活的分子机制提供新的线索，而且还将定义针对这种致命疾病的靶向治疗的新分子和途径。