Ack1: Activation and Consequences in Prostate Cancer

描述（由申请人提供）：前列腺癌是最常见的男性恶性肿瘤，每年夺去30，000名男性的生命，因为它进展为侵袭周围组织和骨的难治性癌症。我们对前列腺癌细胞酪氨酸激酶信号的研究表明，Ack 1是一种研究不足的细胞内酪氨酸激酶，以前从未被认为是前列腺癌癌基因，实际上可能对前列腺癌向雄激素非依赖性的进展有深远的影响。发现Ack 1在晚期人前列腺肿瘤中被激活;此外，活性Ack 1在免疫缺陷小鼠中将致瘤性差的雄激素依赖性LNCaP细胞转化为快速生长的侵袭性雄激素非依赖性肿瘤。我们通过质谱蛋白质测序确定Ack 1为前列腺细胞中Mer受体酪氨酸激酶的下游靶点。用活性Ack 1稳定转染的LNCaP细胞（与载体对照或激酶死亡的Ack 1相比）在培养物中表现出最低限度的生长增加，软琼脂生长显著增加，并且如所指出的，即使在肿瘤植入前去势时，也显著加速裸鼠的肿瘤发生。我们已经表明，Ack 1直接酪氨酸磷酸化雄激素受体（AR），导致配体非依赖性AR活性。对人前列腺癌活检标本的检查显示，约44%的雄激素非依赖性前列腺癌标本中存在酪氨酸磷酸化（自激活）Ack 1和酪氨酸磷酸化AR蛋白，但雄激素依赖性或良性前列腺中没有。我们提供了活性Ack 1至少具有三种功能的证据：i）生长刺激，推测部分通过AR依赖性机制，ii）发送不依赖于AR的存活信号，以及iii）刺激胶原基质和裸鼠模型中的侵袭。我们已经建立了一种新的转基因小鼠模型的Ack 1驱动的前列腺肿瘤和初步研究表明，前列腺特异性表达的活性Ack 1诱导进行性前列腺病理从增生前列腺上皮内瘤形成前列腺腺癌。我们的目标是：i）开发转基因小鼠模型，其中研究前列腺上皮中持续的Ack 1活性的影响，使得能够对Ack 1在前列腺瘤形成中的作用进行遗传分析，和ii）阐明由Ack 1活性改变的信号传导途径，其有助于增加生长、存活、侵袭性和雄激素依赖性。目的1是确定激活的Ack 1转基因的前列腺特异性表达是否加速小鼠中的肿瘤发生、侵袭或转移-单独或与前列腺肿瘤的基因工程小鼠模型组合。目的2是确定雄激素受体磷酸化在Ack 1驱动的前列腺癌发生中的作用，使用小鼠基因敲入模型。目的3是表征体内致瘤特性（转移、骨侵袭等）。Ack 1加速前列腺癌生长，定义其机制，并验证其参与肿瘤生长。我们的数据建立了一种新的前列腺肿瘤发展和进展的机制，在人类前列腺癌标本中观察到活跃状态。我们的工作将建立其作用和激活机制，从而为晚期前列腺癌的治疗提供新的靶点。公共卫生相关性：虽然雄激素剥夺疗法已成为转移性前列腺癌治疗的主要手段超过60年，但由于不可避免地进展至终末期且无有效治疗，因此开发前列腺癌的新治疗选择成为当务之急。Ack 1酪氨酸激酶似乎在前列腺癌进展中起关键作用。该提案有可能产生关于Ack 1如何促进前列腺癌进展的重要新信息，并可能通过靶向Ack 1酪氨酸激酶与新药来治疗前列腺癌患者。

DESCRIPTION (provided by applicant): Prostate cancer, the most common male malignancy, takes the lives of 30,000 men annually, as it progresses to a hormone-refractory cancer that invades surrounding tissue and bone. Our investigation into prostate cancer cell tyrosine kinase signaling revealed that Ack1, an under-studied, intracellular tyrosine kinase, never before suggested as a prostate cancer oncogene, may in fact have a profound effect on prostate cancer progression to androgen independence. Ack1 is found activated in advanced human prostate tumors; moreover, active Ack1 converts the poorly tumorigenic, androgen-dependent LNCaP cells into a rapidly- growing, invasive, androgen-independent tumor in immunodeficient mice. We identified Ack1 by mass spectroscopic protein sequencing as a downstream target of the Mer receptor tyrosine kinase in prostate cells. LNCaP cells, stably transfected with active Ack1 (as compared to vector control or kinase dead Ack1), demonstrated minimally increased growth in culture, substantially increased soft agar growth and, as noted, remarkably accelerated tumorigenesis in nude mice, even when castrated prior to tumor implantation. We have shown that Ack1 directly tyrosine phosphorylates the androgen receptor (AR), resulting in ligand-independent AR activity. Examination of human prostate cancer biopsy specimens revealed tyrosine phosphorylated (auto-activated) Ack1 and tyrosine phosphorylated AR protein in ~44 % of androgen- independent prostate cancer specimens, but not in androgen-dependent or benign prostate. We provide evidence that active Ack1 has at least three functions: i) growth stimulation, presumably in part through AR- dependent mechanism, ii) sending a survival signal that is not dependent on the AR, and iii) stimulating invasion in collagen matrix and in nude mouse models. We have created a novel transgenic mouse model of Ack1-driven prostate neoplasia and preliminary studies indicate that prostate-specific expression of active Ack1 induces progressive prostate pathology from hyperplasia to prostate intraepithelial neoplasia to adenocarcinoma of the prostate. Our objectives are to: i) develop transgenic mouse models in which to study the effects of persistent Ack1 activity in the prostate epithelium, enabling the genetic analysis of the role of Ack1 in prostate neoplasia and ii) elucidate the signaling pathways altered by Ack1 activity that contribute to increased growth, survival, invasiveness and androgen-independence. Aim 1 is to determine whether prostate-specific expression of an activated Ack1 transgene accelerates tumorigenesis, invasion or metastasis in the mouse - by itself or in combination with genetically engineered mouse models of prostate neoplasia. Aim 2 is to determine the role of androgen receptor phosphorylation in Ack1-driven prostate carcinogenesis using a mouse knock-in model. Aim 3 is to characterize the in vivo tumorigenic properties (metastasis, bony invasion, etc.) of Ack1 accelerated prostate cancer growth, define their mechanism, and validate their involvement in tumor growth. Our data establish a novel mechanism of prostate tumor development and progression that is observed in an active state in human prostate cancer specimens. Our proposed work will establish its mechanism of action and activation, and thus provide new targets for treatment of advanced prostate cancer. PUBLIC HEALTH RELEVANCE: Although androgen deprivation therapy has been the mainstay of treatment for metastatic prostate cancer for over 60 years, inevitable progression to the terminal stage with no effective treatment makes development of new therapeutic options for prostate cancer an urgent priority. Ack1 tyrosine kinase appears to play a critical role in prostate cancer progression. This proposal has the potential to yield significant new information about how Ack1 promotes prostate cancer progression and may lead to novel ways of treating prostate cancer patients by targeting Ack1 tyrosine kinase with new drugs.