Vav3, a Rho GEF, Stimulates AR Activity and CaP Growth

描述（由申请人提供）： 前列腺腺癌从雄激素依赖性进展为雄激素非依赖性被定义为雄激素消融（前列腺癌的标准治疗）的临床复发，标志着这种疾病的不可治愈形式。大量研究支持雄激素受体（AR）在雄激素非依赖性疾病中的持续作用，尽管其机制知之甚少。雄激素非依赖性前列腺癌中AR作用的一种机制是通过AR共调节和调节蛋白的上调或活性增加。我们通过使用高密度微阵列对雄激素依赖性人前列腺癌细胞系LNCaP和雄激素非依赖性亚系LNCaP-R1进行基因谱分析，确定了与雄激素非依赖性进展相关的基因表达变化。最显著的变化之一是在雄激素非依赖性细胞中，Rho GT3鸟嘌呤核苷酸交换因子vav 3上调8.5倍。作为一个假定的原癌基因，vav 3在前列腺癌中是一个有趣的候选基因。RNA酶保护和免疫沉淀试验证实vav 3在雄激素非依赖性前列腺癌中升高。Vav 3增强了几种AR表达前列腺癌细胞系和不同启动子环境中的AR转录活性。此外，组成型活性vav 3突变体在体外增强前列腺癌细胞的生长，而显性负性转录变体vav 3.1抑制生长。我们的初步数据提示了以下假设，即vav 3上调或活性增加促进雄激素非依赖性的进展。我们将在此R21应用程序中解决以下问题：1。体内雄激素非依赖性前列腺癌中vav 3水平或活性是否被诱导？2. vav 3激活对于雄激素非依赖性肿瘤形成是必要的还是充分的？我们将使用三种不同的人类前列腺异种移植物（LNCaP、LAPC 4和LAPC 9）来模拟雄激素非依赖性的临床进展，以定义vav 3的作用。具体目标是：一。检查前列腺癌异种移植物LNCaP、LAPC 4和LAPC 9进展为雄激素非依赖性期间的vav 3水平和AR靶基因表达。二.通过检测表达组成型活性vav 3突变体的LNCaP细胞和表达显性阴性vav 3.1的LNCaP-R1细胞的雄激素非依赖性肿瘤形成，确定vav 3信号传导在雄激素非依赖性进展中的作用。

DESCRIPTION (provided by applicant): The progression of prostate adenocarcinoma from androgen dependence to androgen independence is defined as clinical relapse from androgen ablation, the standard therapy for prostate cancer, and marks the incurable form of this disease. A large body of research supports a continued role for the androgen receptor (AR) in androgen independent disease although the mechanisms are poorly understood. One mechanism for AR action in androgen independent prostate cancer is via the upregulation or increased activity of AR coregulatory and modulatory proteins. We identified gene expression changes associated with progression to androgen independence by gene profiling analyses of the androgen dependent human prostate cancer cell line, LNCaP, and an androgen independent subline, LNCaP-R1, using high density microarrays. One of the most striking changes was an 8.5-fold up-regulation of vav3, a Rho GTPase guanine nucleotide exchange factor, in the androgen independent cells. As a putative protooncogene, vav3 is an intriguing candidate in prostate cancer. RNAse protection and immunoprecipitation assays confirmed that vav3 is elevated in androgen independent prostate cancer. Vav3 enhances AR transcriptional activity in several AR-expressing prostate cancer cell lines and in different promoter contexts. Further, a constitutively active vav3 mutant enhances growth of prostate cancer cells in vitro and a dominant negative transcript variant, vav3.1, inhibits growth. Our preliminary data prompted the following hypothesis, that vav3 up-regulation or increased activity promotes progression to androgen independence. We will address the following questions in this R21 application: 1. Are vav3 levels or activity induced in androgen independent prostate cancer in vivo? 2. Is vav3 activation necessary or sufficient for androgen independent tumor formation? We will use three different human prostate xenografts (LNCaP, LAPC4 and LAPC 9) that mimic clinical progression to androgen independence to define the role of vav3. The specific aims are: I. Examine vav3 levels and AR target gene expression during progression of prostate cancer xenografts LNCaP, LAPC4 and LAPC9 to androgen independence. II. Determine the role of vav3 signaling in progression to androgen independence by examination of androgen independent tumor formation by LNCaP cells expressing a constitutively active vav3 mutant and LNCaP-R1 cells expressing the dominant negative vav3.1