REGULATION OF ECM PROTEIN BIOSYNTHESIS IN NORMAL, MALIGNANT AND METASTATIC CELLS

已知转移是由金属蛋白酶增加引起的 癌细胞的产生/活动导致破坏 细胞外基质（ECM）和基底膜，作为屏障 转移细胞通过循环系统进出， 淋巴系统很少有研究来确定 结缔组织蛋白（CTP）在肿瘤发生过程中的合成状态 进展这项研究的目的是确定其他 调节事件发生在癌细胞中，其进一步促进了细胞的增殖。 转移的过程。具体而言，本研究将探讨 ECM蛋白合成的下调可能是一个不可或缺的因素， 基底膜结构完整性丧失的组成部分， ECM，从而促进转移。我们之前的研究表明 核心蛋白聚糖、纤维连接蛋白、骨连接蛋白和I型胶原的表达改变 当比较乳腺结肠癌中的转录谱时， 和皮肤（正常、良性、原发癌和转移性）细胞系， 临床样本。此外，限制性片段长度多态性 （RFLP）分析揭示了突变改变的基因（核心蛋白聚糖， 纤连蛋白和骨连蛋白）在两种转移性细胞系（黑素瘤和 乳房）。本建议的具体目标是：（1）进行系统的 正常、良性、原发癌和转移细胞系的分析 和临床样品，用于ECM合成减少的进一步相关性 和增加的转移潜力，（2）发展一个转移概况， ECM产生减少和金属蛋白酶活性增加， 临床来源的组织，（3）确定结构和功能 显示RFLP多态性的核心蛋白聚糖和骨粘连蛋白的结构域 通过单链构象多态性（SSCP）分析，RT-PCR 克隆和序列分析以及（4）确定 选择的ECM基因和它们的结合蛋白，它们参与了 观察到的变化规律。完成上述具体目标应 提供了一种确定原发性肿瘤转移能力的方法， 原位肿瘤

It is known that metastasis results from increased metalloprotease production/activity by cancerous cells resulting in the breaching of the extracellular matrix (ECM) and basement membranes, which serve as barriers to ingress and egress of metastatic cells through the circulatory and lymphatic systems. Few studies have been done to ascertain the regulatory and synthetic status of the connective tissue proteins (CTP) during tumor progression. The goal of this research is to determine what other regulatory events occur in cancerous cells which further facilitate the process of metastasis. Specifically, this research will explore the possibility that down-regulation of ECM protein synthesis is an integral component in the loss of structural integrity of the basement membrane and ECM, thus facilitating metastasis. Our previous studies demonstrated altered expression of decorin, fibronectin, osteonectin and type I collagen when transcriptional profiles were compared among breast colon and skin (normal, benign, primary cancerous and metastatic) cell lines and clinical samples. Additionally, restriction fragment length polymorphism (RFLP) analyses have revealed mutationally altered genes (decorin, fibronectin and osteonectin) in two metastatic cell lines (melanoma and breast). The specific aims of this proposal are: (1) perform a systematic analysis of normal, benign, primary cancerous and metastatic cell lines and clinical samples for further correlations of decreased ECM synthesis and increased metastatic potential, (2) develop a metastatic profile of decreased ECM production and increased metalloprotease activity in clinical derived tissues, (3) determine the structural and functional domains of decorin and osteonectin which demonstrate RFLP polymorphisms via single strand conformational polymorphism (SSCP) analysis, RT-PCR cloning and sequence analysis and (4) determine regulatory regions of selected ECM genes and their binding proteins which are involved in the observed altered regulation. Completion of the above specific aims should provide a means of determining the metastatic capability of a primary tumor in situ.