Toward Selective Androgen Deprivation by Targeting Androgen Activation of SRF

项目摘要/摘要 尽管雄激素受体(AR)在前列腺癌(CAP)进展中起着核心作用，但对特定的 AR控制推动帽子进展的细胞过程的分子机制在很大程度上仍然存在 难以捉摸。缺乏这种知识是一个重要的问题，因为如果没有这种知识，新的治疗方法的发展 针对致命性表型背后的AR依赖事件的方法是不太可能的。长的- 学期目标是了解AR控制临床相关基因表达的机制(S)。 CAP细胞可以被操控用于治疗干预。这里的目标是确定治疗方法 破坏AR反应的PKN1-SRF相互作用的可能性。中心假设是以PKN1为靶点 将防止由AR反应的SRF作用介导的CAP进展。这一假设是基于 申请人实验室产生的初步工作。拟议研究的理由是，一旦它被 了解CAP中SRF操作的AR规则如何发生，选择与以下相关的AR操作的目标 疾病的发展将是可能的。通过追求3个具体目标来检验中心假设：1)确定 PKN1在AR反应的SRF转录复合体中的作用；2)确定PKN1‘S激酶的底物 在AR依赖的SRF转录复合体中的作用；以及3)确定靶向AR-1的治疗潜力。 响应的PKN1-SRF相互作用。目标1将确定遗传或药物失活的影响 PKN1对靶基因AR依赖的SRF复合体的组成和功能的影响 RNA-Seq、ChIP-Seq和ATAC-Seq检测。在目标2中，最先进的基于生物素的邻近连接分析 以及表达野生型或激酶死亡的PKN1或其中PKN1的作用是 药物抑制将定义依赖于PKN1的SRF的磷酸化，SRF转录 复合体及其染色质环境。目标3将确定抑制PKN1-SRF的治疗潜力 临床相关的PDX与患者新鲜的头盖骨组织体外移植的相互作用。临床上 将使用基因表达和邻近连接来寻找对这种治疗反应的适用生物标记物 化验。这项拟议的研究具有创新性，因为它关注的是一种完全不同的针对AR的方法 CAP中的行动：解开并靶向AR下游的AR依赖信号通路，该通路控制 与CAP进展相关的基因的表达。这一贡献之所以意义重大，是因为它是第一个 在一系列研究中迈出一步，预计将导致开发新的治疗模式， 靶向AR介导的致死帽表型下的基因表达。关于…… 预期结果，拟议的研究将确定PKN1引入 导致侵袭性上限的AR-PKN1-SRF信号级联中的治疗脆弱性 表型。这些结果将产生重要的积极影响，因为它们将从根本上推动 了解AR在CAP中的作用，特别是为CAP治疗提供新的靶点。

Project summary/Abstract Despite the central role of androgen receptor (AR) in prostate cancer (CaP) progression, insights into the specific molecular mechanisms by which AR controls the cellular processes that drive CaP progression remain largely elusive. Lack of this knowledge is an important problem, because without it, development of novel therapeutic approaches that target specifically AR-dependent events underlying the lethal phenotype is unlikely. The long- term goal is to understand how the mechanism(s) by which AR controls clinically relevant gene expression in CaP cells can be manipulated for therapeutic intervention. The objective here is to determine the therapeutic potential of disrupting AR-responsive PKN1-SRF interactions. The central hypothesis is that targeting PKN1 will prevent CaP progression mediated by AR-responsive SRF action. This hypothesis is formulated based on preliminary work produced in the applicant's laboratory. The rationale for the proposed studies is that, once it is understood how AR regulation of SRF action in CaP occurs, select targeting of AR action that is relevant to disease progression will be possible. The central hypothesis is tested by pursuing 3 specific aims: 1) determine the role of PKN1 in the AR-responsive SRF transcriptional complex; 2) define the substrate for PKN1's kinase action in the AR-dependent SRF transcriptional complex; and 3) determine therapeutic potential of targeting AR- responsive PKN1-SRF interactions. Aim 1 will determine the impact of genetic or pharmacological inactivation of PKN1 on the composition and function of AR-dependent SRF complex at target genes using a combination of RNA-Seq, ChIP-Seq and ATAC-Seq assays. In Aim 2, state-of-the-art biotin-based proximity ligation assays and mass spectrometry in CaP models that express wild-type or kinase-dead PKN1 or in which PKN1 action is pharmacologically inhibited will define PKN1-dependent phosphorylation of SRF, the SRF transcriptional complex and its chromatin environment. Aim 3 will determine the therapeutic potential of inhibiting PKN1-SRF interactions in clinically relevant PDXs and fresh ex vivo explants of CaP tissue obtained from patients. Clinically applicable biomarkers of response to such treatment will be sought using gene expression and proximity ligation assays. The proposed research is innovative because it focuses on an entirely different approach to target AR action in CaP: unraveling and targeting an AR-dependent signaling pathway downstream of AR that controls the expression of genes that are relevant for CaP progression. This contribution is significant because is it the first step in a continuum of research that is expected to lead to the development of novel treatment modalities that target specifically AR-mediated gene expression that underlies the lethal CaP phenotype. With respect to expected outcomes, the proposed studies will identify the molecular mechanisms by which PKN1 introduces a therapeutic vulnerability in the AR-PKN1-SRF signaling cascade that gives rise to the aggressive CaP phenotype. These results will have an important positive impact because they will fundamentally advance knowledge about AR action in CaP, in general, and provide new targets for CaP therapy, specifically.