Rap1Gap and Tumor Progression

描述（由申请人提供）：恶性进展中最关键的步骤之一是肿瘤细胞获得转移能力。本提案中待检验的假设是Rap 1GAP的下调有助于甲状腺肿瘤的进展。这一观点是基于数据显示Rap 1GAP在正常人甲状腺细胞中高度表达，并且其表达在大多数浸润性甲状腺乳头状癌中显著降低。在人甲状腺癌细胞系中的研究揭示了Rap 1GAP的丢失与上皮结构的丢失之间的显著相关性。Rap 1GAP缺陷型肿瘤细胞缺乏E-钙粘蛋白，并获得波形蛋白的表达，表明上皮细胞向间充质细胞转化。与保留Rap 1GAP的肿瘤细胞相比，这些细胞表现出增强的迁移和侵袭特性。将Rap 1GAP恢复到Rap 1GAP缺陷细胞抑制细胞迁移、侵袭和非贴壁依赖性增殖。沉默Rap 1GAP表达的甲状腺癌细胞，保留上皮形态导致细胞分散，增强细胞聚集体的解离，和失调的E-钙粘蛋白，提示细胞/细胞粘附的缺陷。由于细胞/细胞连接的破坏与严重的病理后果有关，我们认为Rap 1GAP的下调通过减弱细胞/细胞粘附而有助于甲状腺肿瘤的发病。体外研究将探索Rap 1GAP表达降低减弱细胞/细胞粘附的分子机制，并评估消除Rap 1GAP是否赋予肿瘤细胞改变的迁移特性和侵袭潜力。由于TSH对Rap 1GAP的调节在人甲状腺肿瘤细胞系中丢失，但可能保留在原发性甲状腺肿瘤中，因此将在分化的大鼠甲状腺细胞中进行补充研究。这些研究将在三维环境中的细胞中进行，这些条件更接近地再现人体组织的基质顺应性，并且细胞/细胞接触最大化。体外研究与人类肿瘤标本中的研究相辅相成，这些研究将确定Rap 1GAP表达降低的甲状腺肿瘤亚型，并探讨Rap 1GAP缺失的临床意义。总之，该提案提出了一个高度一致的计划，以研究Rap 1GAP缺失对人类甲状腺肿瘤进展的贡献和临床意义。

DESCRIPTION (provided by applicant): One of the most critical steps in the progression to malignancy is the acquisition by tumor cells of the ability to metastasize. The hypothesis to be tested in this proposal is that downregulation of Rap1GAP contributes to thyroid tumor progression. This notion is based on data showing that Rap1GAP is highly expressed in normal human thyroid cells, and that its expression is dramatically reduced in the majority of invasive papillary thyroid carcinomas. Studies in human thyroid carcinoma cell lines revealed a striking correlation between loss of Rap1GAP and loss of epithelial structure. Rap1GAP-deficient tumor cells lacked E-cadherin and acquired the expression of vimentin, indicative of epithelial-to-mesenchymal transition. These cells exhibited enhanced migratory and invasive properties compared to tumor cells that retained Rap1GAP. Restoring Rap1GAP to Rap1GAP-deficient cells inhibited cell migration, invasion and anchorage-independent proliferation. Silencing Rap1GAP expression in thyroid carcinoma cells that retain an epithelial morphology caused the cells to disperse, enhanced the dissociation of cell aggregates, and dysregulated E-cadherin, suggestive of defects in cell/cell adhesion. As disruption of cell/cell junctions is associated with serious pathological consequences, we propose that downregulation of Rap1GAP contributes to the pathogenesis of thyroid tumors by attenuating cell/cell adhesion. In vitro studies will explore the molecular mechanism through which decreased Rap1GAP expression attenuates cell/cell adhesion and assess whether eliminating Rap1GAP endows tumor cells with altered migratory properties and invasive potential. As TSH regulation of Rap1GAP is lost from the human thyroid tumor cell lines, but is likely to be retained in primary thyroid tumors, complementary studies will be performed in differentiated rat thyroid cells. These studies will be conducted in cells in three-dimensional environments, conditions that more closely reproduce the matrix compliance of human tissues and where cell/cell contacts are maximized. The in vitro studies are complemented with studies in human tumor specimens that will identify the subtypes of thyroid tumors in which Rap1GAP expression is decreased and probe the clinical significance of loss of Rap1GAP. In summary, this proposal presents a highly cohesive plan to investigate the contribution and clinical significance of Rap1GAP depletion to the progression of human thyroid tumors.