ELUCIDATING THE FUNCTION OF MATRIPTASE-ACTIVATED PDGF D IN BONE METASTATIC PROSTA

描述（由申请人提供）：癌细胞转移是一个复杂的多步骤过程，需要肿瘤和周围基质之间的相互作用。原发性肿瘤细胞必须能够增殖，脱离抑制性细胞-细胞连接复合物，迁移通过下面的基底膜和基质以进入附近的脉管系统，这些细胞最终在脉管系统中归巢并定植在继发性转移部位。前列腺癌是美国男性中诊断最多的癌症，仅在2009年就估计有192，000例，并且是同一组中癌症相关死亡的第二大原因。这种腺癌在90%的病例中转移到骨，导致骨微环境破坏，并引起病理性骨折和脊髓压迫，大大降低患者的生活质量。为了使转移性前列腺癌在骨中生长，它必须分泌不同的因子，为肿瘤细胞生长创造有利的环境。有丝分裂生长因子家族，血小板衍生生长因子（PDGF），已被证明支持成骨细胞和破骨细胞的增殖和活化。有趣的是，血小板衍生生长因子受体2（PDGFR 2）已被证明在前列腺癌的转移进展过程中上调。事实上，PDGFR 2抑制剂STI 571单独或与紫杉醇联合使用可显著降低体内前列腺癌细胞的骨生长。该受体被其配体PDGF B和PDGF D结合并激活。我们的实验室证明，PDGF D，而不是PDGF B，在前列腺癌进展过程中过表达，表明PDGF D可能在转移部位诱导PDGFR 2活化。PDGF D在上皮细胞中表达为无活性的二聚体，并被上皮特异性丝氨酸蛋白酶matriptase蛋白水解活化。释放的活性形式的PDGF D结合并激活周围基质细胞上的其同源受体。目的/假设：PDGF D和PDGFR 2在前列腺癌转移中均上调。此外，PDGF D的激活剂间质蛋白酶在前列腺癌骨转移期间与PDGF D共定位。因此，我们假设间质蛋白酶介导的PDGF D成熟可能激活肿瘤细胞衍生的生长因子，导致破骨细胞和成骨细胞的激活，从而导致异常的骨反应。具体目的：（1）阐明PDGF D和matriptase对破骨细胞信号转导和分化的影响。(2)分析PDGF D介导的成骨细胞发生中的信号级联反应。(3)评估PDGF D在体内成骨细胞生成和破骨细胞生成中的作用。研究设计：我们将利用低侵袭性和高侵袭性前列腺癌细胞系差异表达PDGF D或间质蛋白酶，以监测其对破骨细胞和成骨细胞发育的影响。此外，我们将采用重组蛋白质技术来剖析PDGF D在前列腺癌骨生长中的结构-功能，特别是破骨细胞和成骨细胞的生成。最后，我们将应用体内转移小鼠模型来检查PDGF D和间质蛋白酶在转移性前列腺癌中的功能。相关性：尽管早期发现提高了前列腺癌的生存率，但许多患者在诊断时仍表现出转移性疾病。我们相信，我们的研究结果将大大有助于了解PDGF-PDGFR轴在前列腺癌转移中的作用。与此同时，我们设想我们的突出贡献，导致新的和有针对性的治疗前列腺癌的发展，除了提高现有的治疗方式的疗效。 公共卫生相关性：前列腺癌倾向于扩散到骨骼，导致顽固性骨痛、骨折和脊髓压迫，从而降低患者的生活质量。我们发现PDGF D、其激活剂间质蛋白酶和受体PDGFR 2在骨转移性前列腺癌中上调。因此，我们假设在前列腺癌中，PDGF D可以通过其被间质蛋白酶激活而充当PDGFR 2激动剂以促进肿瘤进展，这将使用差异表达PDGF D或间质蛋白酶的前列腺癌细胞系来评估，以确定该生长因子及其同源受体在肿瘤进展中的作用。

DESCRIPTION (provided by applicant): Cancer cell metastasis is an intricate multistep process requiring interplay between the tumor and the surrounding stroma. The primary tumor cells must be able to proliferate, disengage from the inhibitory cell-cell junctional complex, migrate through the underlying basement membrane and matrix to gain access to nearby vasculature where these cells eventually home in and colonize secondary metastatic sites. Prostate cancer is the most diagnosed cancer in American men responsible for an estimated 192,000 cases in 2009 alone and it is the second leading cause of cancer-related death within the same group. This adenocarcinoma metastasizes to the bone in 90% of cases resulting in the disruption of the bone microenvironment and causing pathological bone fractures and spinal cord compressions drastically reducing the quality of life of patients. In order for metastatic prostate cancer to grow in the bone, it must secrete different factors creating a favorable environment for tumor cell growth. A family of mitogenic growth factors, platelet-derived growth factors (PDGF), have been shown to support osteoblast and osteoclast proliferation and activation. Interestingly, the platelet-derived growth factor receptor 2 (PDGFR2) has been demonstrated to be upregulated during the metastatic progression of prostate cancer. In fact the PDGFR2 inhibitor, STI571, alone or in combination with paclitaxel significantly reduced osseous growth of prostate cancer cells in vivo. This receptor is bound and activated by its ligands, PDGF B and PDGF D. Our lab demonstrated that PDGF D, and not PDGF B, is overexpressed during prostate cancer progression suggesting that PDGF D may induce PDGFR2 activation at the metastatic site. PDGF D is expressed in epithelial cells as an inactive dimer and is proteolytically activated by the epithelial specific serine protease, matriptase. The released active form of PDGF D binds to and activates its cognate receptor on surrounding stromal cells. Objective/Hypothesis: Both PDGF D and PDGFR2 are upregulated within prostate cancer metastasis. Furthermore, the activator of PDGF D, matriptase, co-localizes with PDGF D during prostate cancer bone metastasis. Therefore, we hypothesize that matriptase mediated maturation of PDGF D may activate the tumor cell derived growth factor resulting in the activation of osteoclasts and osteoblasts leading to an aberrant osseous response. Specific Aims: (1) Elucidate the effects of PDGF D and matriptase on osteoclast signaling and differentiation. (2) Dissect the PDGF D mediated signaling cascade in osteoblastogenesis. (3) Assess the role of PDGF D in osteoblastogenesis and osteoclastogenesis in vivo. Study Design: We will utilize minimally aggressive and highly invasive prostate cancer cell lines differentially expressing PDGF D or matriptase to monitor their influence on osteoclast and osteoblast development. Furthermore, we will employ recombinant protein technology to dissect the structure-function of PDGF D in prostate cancer bone growth, specifically looking at osteoclast- and osteoblastogenesis. Finally, we will apply in vivo metastasis mouse models to examine the function of PDGF D and matriptase in metastatic prostate cancer. Relevance: Although early detection has improved prostate cancer survival rate, many patients still exhibit metastatic disease at diagnosis. We believe our findings will contribute greatly toward understanding the role of the PDGF-PDGFR axis in prostate cancer metastasis. Concomitantly, we envision our salient contributions leading to the development of novel and targeted therapeutics for prostate cancer in addition to improving the efficacy of existing treatment modalities. PUBLIC HEALTH RELEVANCE: Prostate cancer has a predilection to spread to the bone leading to intractable bone pain, bone fractures, and spinal cord compressions which decrease patient quality of life. We found that PDGF D, its activator matriptase and receptor PDGFR2 are upregulated in bone metastatic prostate cancer. Hence we hypothesized that in prostate cancer, PDGF D may serve as the PDGFR2 agonist through its activation by matriptase to promote tumor progression which will be assessed using prostate cancer cell lines differentially expressing PDGF D or matriptase to determine the role of this growth factor and its cognate receptor in tumor progression.