Melanoma Cell Surface Proteoglycans in Metastasis

黑色素瘤硫酸软骨素蛋白聚糖（MCSP）是一种高水平表达的细胞表面蛋白聚糖 在绝大多数人黑色素瘤上，并涉及促进肿瘤粘附、迁移和 入侵为了进一步表征MCSP在肿瘤进展中的作用，我们克隆并稳定表达了MCSP基因。 两种MCSP阴性黑素瘤细胞系中的MCSP核心蛋白。MCSP表达促进生长， 异种移植模型中黑色素瘤细胞的肿瘤形成，以及抗细胞外 MCSP结构域抑制体内肿瘤形成。MCSP的表达刺激整合素介导的信号 通过MCSP诱导的细胞铺展和粘着斑激酶的增加证明的转导 磷酸化此外，MCSP的表达刺激增强的酪氨酸磷酸化， ERK的磷酸化，这是锚定非依赖性生长所需的，而细胞质尾- 缺失的MCSP不能支持锚定非依赖性生长或增强ERK激活。成员 ERK/MAPK途径，包括BRAF（其在这些细胞中突变为V600 E活性形式）、pMEK和 pERK均与GST-MCSP胞质尾融合蛋白沉淀，而截短的融合蛋白 缺少MCSP尾的C-末端不能结合这些分子。这些结果表明，MCSP 作为ERK/MAPK通路成员的对接位点。我们假设MCSP作为一种新的 跨膜支架蛋白，有助于组装和有效激活关键信号通路， 促进黑色素瘤生长、存活和侵袭。目标#1将研究MCSP在肿瘤生长和维持中的作用 使用siRNA抑制人黑素瘤细胞中MCSP的表达。目标#2将重点关注成员如何 ERK/MAPK通路与MCSP的胞质结构域相连。目标#3将定义 MCSP的细胞外结构域是激活肿瘤生长所需的关键信号通路所必需的。 黑色素瘤是一种毁灭性的疾病，对目前的治疗几乎完全没有反应。以来 绝大多数黑色素瘤在原发性肿瘤和转移性病变中都表达MCSP，这表明 黑色素瘤细胞可以利用这种分子获得与MCSP阴性细胞相比的竞争优势， 恶性进展的多个阶段。这些研究的长期目标是确定 开发MCSP表达/功能的抑制剂作为治疗恶性黑素瘤的新疗法。

Melanoma chondroitin sulfate proteoglycan (MCSP) is a cell surface proteoglycan expressed at high levels on the vast majority of human melanomas and is implicated in promoting tumor adhesion, migration and invasion. To further characterize MCSP in tumor progression we have cloned and stably expressed the MCSP core protein in two MCSP-negative melanoma cell lines. MCSP expression enhanced growth and tumor formation of melanoma cells in xenograft models, and monoclonal antibodies against the extracellular domain of MCSP inhibited tumor formation in vivo. Expression of MCSP stimulates integrin-mediated signal transduction as evidenced by MCSP-induced increases in cell spreading and focal adhesion kinase phosphorylation. Furthermore, expression of MCSP stimulates enhanced tyrosine phosphorylation and the phosphorylation of ERK, which is required for anchorage-independent growth, while a cytoplasmic tail- deleted MCSP failed to support anchorage-independent growth or enhance ERK activation. Members of the ERK/MAPK pathway, including BRAF (which is mutated to the V600E active form in these cells), pMEK and pERK all precipitated with a GST-MCSP cytoplasmic tail fusion protein, while a truncated fusion protein lacking the c-terminal end of the MCSP tail failed to bind these molecules. These results indicate that MCSP acts as a docking site for ERK/MAPK pathway members. We hypothesize that MCSP functions as a novel transmembrane scaffold protein that helps assemble and efficiently activate key signaling pathways to promote melanoma growth, survival and invasion. Aim #1 will study MCSP in tumor growth and maintanence using siRNA to inhibit MCSP expression in human melanoma cells. Aim #2 will focus on how members of the ERK/MAPK pathway link to the cytoplasmic domain of MCSP. Aim #3 will define structural features of the extracellular domain of MCSP required for activation of key signalling pathways important for tumor growth. Melanoma is a devastating disesase that is almost completely nonresponsive to current therapies. Since the vast majority of melanomas express MCSP in both primary tumors and in metastatic lesions, this suggests that melanoma cells may use this molecule to attain a competitive advantage over MCSP-negative cells at multiple stages of malignant progression. The long term goal of these studies is to determine the potential to exploit inhibitors of MCSP expression/function as novel therapies in the treatment of malignant melanoma.