Matrix remodeling in tumor growth and metastasis in a murine breast cancer model

描述(申请人提供)：我的研究旨在了解成纤维细胞激活蛋白(FAP)的致癌作用，FAP是一种具有二肽基肽酶(DPP)和内肽酶/胶原酶活性的膜结合丝氨酸蛋白酶。FAP的独特之处在于，在90%的人类上皮源性癌症中，FAP在癌症相关成纤维细胞(CAF)中表达，而在正常或转化(恶性/肿瘤)的上皮细胞中不表达。重要的是，我们实验室(J.Tchou和E.Puri)的未发表工作表明，FAP可以作为一种强大的泛CAF标记物用于人类乳腺癌，它在选择性和特异性方面优于其他CAF标记物。CAF在细胞外基质重塑、血管生成和间质形成中发挥重要作用，并在肿瘤生长和转移中发挥重要作用。绝大多数关于FAP活性的报道描述了FAP表达与不良预后之间的相关性，这使得抑制FAP成为癌症治疗的一个有吸引力的潜在靶点。我们的实验室最近证明了FAP促进了突变K-ras驱动的肺癌和结肠癌和胰腺癌同基因移植模型的原生小鼠模型的肿瘤发生，并且FAP缺陷小鼠肿瘤生长的减少与紊乱的胶原蛋白的增加、异常的整合素信号以及间质形成和血管生成的减少有关。然而，FAP促进肿瘤发生的机制尚不明确，其在肿瘤转移中的作用尚未在任何肿瘤类型中被研究。我们的初步数据表明，FAP介导了胶原酶介导的切割产生的特征性3/4和1/4胶原蛋白片段的蛋白水解性加工(例如。基质金属蛋白酶-1)。这一提议的一个中心假设是，FAP通过其蛋白酶活性在ECM重塑中发挥关键作用，从而促进肿瘤的发生。具体地说，我假设FAP通过靶向胶原的降解和周转，调节肿瘤细胞增殖、间质形成、血管生成和转移的依赖于基质的信号。为了检验这些假说，我将比较MMTV-Neu驱动的FAP野生型小鼠、FAP缺失小鼠(表达荧光素酶取代FAP基因的敲入基因)和表达酶死亡的FAP突变体(FAPS624A)或保持DPP活性但缺乏内肽酶/胶原酶活性的FAP突变体(FAPA657S)的敲入小鼠的肿瘤发生和转移。因此，通过利用这些酶缺陷的FAP小鼠，本研究还将研究FAP在ECM重塑中的作用机制。这些新的FAP小鼠品系已经在实验室中生产出来，目前正在培育中，而MMTV-Neu小鼠则可以在商业上买到。好了！ 公共卫生相关性：乳腺癌是当今女性癌症死亡的第二大原因。癌症的代价是对被诊断为癌症的人、他们的家庭和整个社会造成经济和精神上的损失。2005年，美国乳腺癌造成的时间和经济生产力损失为120.96亿美元(NCI，2009/2010)，因此，迫切需要为这种毁灭性疾病的早期预测、治疗和根除确定新的分子靶点。本研究旨在通过研究乳腺肿瘤细胞与其微环境的关系，更好地了解肿瘤发生、发展和转移的原因和机制。这项拟议的研究将确定一种肿瘤基质细胞蛋白水解酶的促肿瘤活性和在自体乳腺癌模型中细胞外基质的重塑。拟议的研究结果将揭示这种蛋白水解酶调节肿瘤发展的新机制，从而将其作为癌症治疗的靶点。此外，我们的研究可能会带来一种早期乳腺癌诊断生物标记物，可以用于临床，以节省时间、资源，最重要的是，节省生命。好了！

DESCRIPTION (provided by applicant): My research aims to understand the tumorigenic role of fibroblast activation protein (FAP), a membrane-bound serine protease with dipeptidyl peptidase (DPP) and endopeptidase/collagenase activities. FAP is unique, in that it is expressed in cancer associated fibroblasts (CAFs) in 90% of all human epithelial derived cancers, but not in normal or transformed (malignant/neoplastic) epithelial cells. Importantly, unpublished work from our lab (J.Tchou and E. Puri) shows that FAP can be used as a robust pan-CAF marker in human breast cancer, it being superior to other CAF markers in its selectivity and specificity. CAFs play roles in ECM remodeling, angiogenesis and stromagenesis, and are important players in tumor growth and metastasis. A wide majority of reports on FAP activity describe correlations between FAP expression and poor prognosis, making FAP inhibition an appealing potential target for cancer therapeutics. Our lab recently demonstrated that FAP promotes tumorigenesis in an autochthonous mouse model of mutant K-ras- driven lung cancer and syngeneic transplant models of colon and pancreatic cancer and that the reduction in tumor growth in FAP-deficient mice was associated with enhanced accumulation of disorganized collagen, aberrant integrin signaling and reduced stromagenesis and angiogenesis. However, the mechanisms by which FAP promotes tumorigenesis are yet to be defined and its role in metastasis has not yet been investigated in any tumor type. Our preliminary data indicate that FAP mediates the proteolytic processing of the characteristic 3/4 and 1/4 fragments of collagen generated by collagenase mediated cleavage (eg. MMP-1). A central hypothesis of this proposal is that FAP promotes tumorigenesis through its protease activity by playing a critical role in ECM remodeling. Specifically, I hypothesize that by targeting collagen for degradation and turnover, FAP regulates matrix-dependent signaling of tumor cell proliferation, stromagenesis, angiogenesis and metastasis. To test these hypotheses I will compare tumorigenesis and metastasis of MMTV-Neu driven autochthonous mammary tumors in FAP wild-type mice, FAP-null mice (FAP-knock-ins expressing luciferase in place of the FAP gene) and knock-in mice expressing an enzymatic dead FAP mutant (FAPS624A) or a FAP mutant (FAPA657S) that retains DPP activity but lacks endopeptidase/collagenase activity. Therefore, by making use of these enzymatically defective FAP mice, this study will also examine the mechanism by which FAP acts on ECM remodeling. These novel FAP mouse strains have been produced in the lab and are currently breeding, while the MMTV-Neu mouse is commercially available. ! PUBLIC HEALTH RELEVANCE: Breast cancer is the second leading cause of cancer deaths in women today. The costs of cancer are a financial and mental toll on people diagnosed with cancer, their families, and society as a whole. In 2005 the burden of breast cancer in the U.S. was $12.096 billion in lost time and economic productivity (NCI, 2009/2010), therefore it is imperative to identify new molecular targets for early prognosis, treatment and eradication of this devastating disease. This research aims to better understand the causes and mechanisms underlying tumor initiation, progression and metastasis by studying the relationship between mammary tumor cells and their microenvironment. The research proposed will define the protumorigenic activity of a tumor stromal cell protease and extracellular matrix remodeling in an autochthonous breast cancer model. Results of the proposed studies will reveal novel mechanisms through which this protease regulates tumor development and therefore its value as a target for cancer therapeutics. Further, our studies may lead to an early breast cancer diagnostic biomarker that could be used in the clinic to save time, resources and most importantly, life. !