Ack1: Activation and Consequences in Prostate Cancer

描述(申请人提供)：前列腺癌是最常见的男性恶性肿瘤，每年夺走30,000名男性的生命，因为它进展为一种激素不敏感的癌症，侵袭周围组织和骨骼。我们对前列腺癌细胞酪氨酸激酶信号的研究表明，AcK1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的癌基因，实际上可能对前列腺癌的进展到雄激素非依赖性有深远的影响。AcK1在晚期人类前列腺癌中被发现激活；此外，在免疫缺陷小鼠中，活性AcK1将低致瘤性、雄激素依赖的LNCaP细胞转化为快速生长的、侵袭性、雄激素非依赖性肿瘤。我们通过质谱学蛋白质测序确定AcK1是前列腺细胞中Mer受体酪氨酸激酶的下游靶点。稳定地转导活性AcK1的LNCaP细胞(与载体对照或死亡的AcK1相比)，在培养中表现出最低限度的增长，显著增加了软琼脂的生长，并如所述，显著加速了裸鼠的肿瘤形成，即使在肿瘤植入前去势也是如此。我们已经证明，AcK1直接使雄激素受体(AR)酪氨酸磷酸化，导致不依赖于配体的AR活性。对前列腺癌活检标本的检测发现，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)的AcK1和酪氨酸磷酸化的AR蛋白，而雄激素依赖性前列腺癌或良性前列腺癌中没有。我们提供的证据表明，活性AcK1至少有三个功能：i)部分通过AR依赖机制促进生长；ii)发出不依赖AR的生存信号；iii)刺激胶原基质和裸鼠模型的侵袭。我们已经建立了一种新的AcK1驱动的前列腺癌转基因小鼠模型，初步研究表明，活性AcK1的前列腺特异性表达诱导了从增生到前列腺上皮内瘤变到前列腺癌的进行性前列腺病理。我们的目标是：i)建立转基因小鼠模型，研究前列腺上皮中持续的AcK1活性的影响，从而能够对AcK1在前列腺肿瘤中的作用进行遗传学分析；ii)阐明AcK1活性改变的信号通路，这些信号通路有助于增加生长、存活、侵袭性和雄激素非依赖性。目的1是确定激活的AcK1转基因基因的前列腺特异性表达是否单独或与前列腺癌基因工程小鼠模型结合使用，加速小鼠肿瘤的发生、侵袭或转移。目的2是利用小鼠基因敲入模型，确定雄激素受体磷酸化在AcK1诱导的前列腺癌发生中的作用。目的3是研究体内的致瘤特性(转移、骨侵袭等)。研究AcK1促进前列腺癌生长的作用机制，并验证其在肿瘤生长中的作用。我们的数据建立了前列腺癌发生和进展的新机制，在人类前列腺癌标本中观察到活跃状态。我们提出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。公共卫生意义：尽管60多年来，雄激素剥夺疗法一直是转移性前列腺癌的主要治疗方法，但在没有有效治疗的情况下，不可避免地发展到晚期，这使得开发前列腺癌的新治疗方案成为当务之急。AcK1酪氨酸激酶似乎在前列腺癌的进展中起着关键作用。这项提议有可能产生关于AcK1如何促进前列腺癌进展的重要新信息，并可能导致通过用新药靶向AcK1酪氨酸激酶来治疗前列腺癌患者的新方法。

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.