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3万名男性的生命，因为它进展为一种激素难治性癌症，侵犯周围组织和骨骼。我们对前列腺癌细胞酪氨酸激酶信号传导的研究表明，Ack1，一种未被充分研究的细胞内酪氨酸激酶，从未被认为是前列腺癌的致癌基因，实际上可能对前列腺癌进展到雄激素依赖性有深远的影响。Ack1在晚期人类前列腺肿瘤中被发现激活；此外，在免疫缺陷小鼠中，活性Ack1将低致瘤性、雄激素依赖性的LNCaP细胞转化为快速生长、侵袭性、雄激素非依赖性的肿瘤。我们通过质谱蛋白测序鉴定Ack1是前列腺细胞中Mer受体酪氨酸激酶的下游靶点。LNCaP细胞稳定地转染了活性Ack1（与载体对照或激酶死亡Ack1相比），在培养中表现出最低程度的生长增加，显著增加了软琼脂生长，并且如前所述，在裸鼠中显著加速了肿瘤发生，即使在肿瘤植入前阉割。我们已经证明Ack1直接酪氨酸磷酸化雄激素受体（AR），导致与配体无关的AR活性。人类前列腺癌活检标本的检查显示，在约44%的雄激素非依赖性前列腺癌标本中，酪氨酸磷酸化（自激活）Ack1和酪氨酸磷酸化AR蛋白，但在雄激素依赖性或良性前列腺中没有。我们提供的证据表明，活性Ack1至少具有三种功能：i)刺激生长，可能部分通过AR依赖机制，ii)发送不依赖于AR的生存信号，以及iii)刺激胶原基质和裸鼠模型中的入侵。我们已经建立了一种新的转基因小鼠Ack1驱动的前列腺肿瘤模型，初步研究表明，活性Ack1的前列腺特异性表达可诱导从前列腺增生到前列腺上皮内瘤变到前列腺腺癌的进行性前列腺病理。我们的目标是：1)建立转基因小鼠模型，研究Ack1在前列腺上皮中持续活性的影响，从而能够对Ack1在前列腺肿瘤中的作用进行遗传分析；2)阐明由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、侵袭性和雄激素依赖性。目的1是确定激活Ack1基因的前列腺特异性表达是否会加速小鼠前列腺肿瘤的发生、侵袭或转移——单独或与基因工程小鼠前列腺肿瘤模型联合。目的2是通过小鼠敲入模型确定雄激素受体磷酸化在ack1驱动的前列腺癌发生中的作用。目的3：表征Ack1加速前列腺癌生长的体内致瘤特性（转移、骨侵袭等），明确其机制，验证其参与肿瘤生长。我们的数据建立了前列腺肿瘤发展和进展的新机制，在人类前列腺癌标本中观察到活跃状态。我们的工作将建立其作用机制和激活机制，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：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，前列腺癌不可避免地会发展到终末期，这使得开发新的前列腺癌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关键作用。这一建议有可能产生关于Ack1如何促进前列腺癌进展的重要新信息，并可能导致用新药靶向Ack1酪氨酸激酶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.