Proteolytic matriptase-prostasin axis in breast cancer

描述（由申请人提供）：间质蛋白酶是一种细胞表面锚定的丝氨酸蛋白酶，其首先在人乳腺癌细胞系中鉴定，随后涉及乳腺癌病理学的许多方面。间质蛋白酶在人乳腺癌癌细胞中上调，其表达增加已被证明与患者预后不良相关。目前，尚不清楚间质蛋白酶是否在乳腺癌发生中起因果作用或主要促成癌症生长和进展的继发事件，并且其作为治疗靶点的潜力仍未得到测试。间质蛋白酶功能丧失小鼠中的围产期致死性迄今为止已经排除了对乳腺中间质蛋白酶消融作用的分析;然而，我们的提议提出了绕过该限制的新技术。在我们提出的研究中，将使用平行和互补的体外和体内“功能丧失”技术来进行间质蛋白酶在乳腺癌中的作用的功能和机制表征。我们以前的研究已经确定GPI锚定的丝氨酸蛋白酶prostastin作为一个生理底物和下游效应的matriptase蛋白水解活性，我们将专注于这个蛋白水解轴的潜在贡献乳腺癌生物学。我们认为这两种蛋白酶与癌症进展之间存在显著相关性，因为我们已经证明，间质蛋白酶和前列腺素在体内细胞与细胞粘附中起关键作用，并且任一蛋白质水平的变化可以通过破坏上皮紧密连接的形成和完整性来扰乱这些粘附。 待测试的假设是间质蛋白酶通过激活前列腺素酶原发挥关键功能，并且间质蛋白酶/前列腺素蛋白水解途径对于乳腺癌发生是关键的，特别是通过细胞-细胞粘附的丧失，从而促进体内致癌作用。 为了验证这一假设，我们制定了两个具体目标。在第一个目标中，将使用间质蛋白酶和前列腺素功能丧失的新型遗传小鼠模型来确定间质蛋白酶/前列腺素蛋白水解轴在乳腺癌起始和进展中的重要性。在第二个目标中，将研究前列腺素在乳腺癌细胞间粘附中的作用，重点是其在紧密连接功能和完整性中的作用。 将最先进的小鼠遗传学与基于2D和3D细胞培养的测定相结合，涵盖了研究人类癌症疾病的创新策略，并可能为乳腺癌的诊断和治疗提供新的途径。

DESCRIPTION (provided by applicant): Matriptase is a cell-surface anchored serine protease that was first identified in human breast cancer cell lines and has subsequently been implicated in many aspects of breast cancer pathology. Matriptase is up-regulated in human breast carcinoma cancer cells, and its increased expression has been shown to correlate with poor patient outcome. Presently, it is unknown if matriptase plays a causal role in breast carcinogenesis or contributes primarily to secondary events of cancer growth and progression, and its potential as a therapeutic target remains untested. Perinatal lethality in matriptase loss-of-function mice has thus far precluded analysis of the effect of matriptase ablation in the mammary gland; however our proposal presents novel techniques to bypass this limitation. In our proposed study both functional and mechanistic characterization of matriptase's role in breast cancer will be performed using parallel and complimentary in vitro and in vivo "loss-of-function" techniques. Our previous research has identified the GPI-anchored serine protease prostastin as a physiological substrate and downstream effector of matriptase proteolytic activity, and we will focus on this proteolytic axis for its potential contribution to breast cance biology. We believe there to be a significant correlation between these two proteases and cancer progression, as we have demonstrated that both matriptase and prostasin play critical roles in cell to cell adhesions in vivo, and changes in levels of either protein can perturb these adhesions via disruption of epithelial tight junction formation and integrity. The hypothesis to be tested is that matriptase exerts critical functions through activation of the prostasin zymogen and that the matriptase/prostasin proteolytic pathway is critical for breast oncogenesis, specifically through the loss of cell-cell adhesions, and thereby the promotion of carcinogenesis in vivo. To test this hypothesis, we formulated two specific aims. In the first aim the significance of the matriptase/prostasin proteolytic axis in breast cancer initiation and progression will be determined using both matriptase and prostasin loss-of-function novel genetic mouse models. In the second aim the role of prostasin in cell-cell adhesion in breast cancer will be studied with emphasis on its role in tight junction function and integrity. The combination of state-of-the art mouse genetics with 2D and 3D cell culture based assays encompasses an innovative strategy for studying human cancerous disease, and may offer new avenues for diagnosis and therapy of breast cancer.