Dynamic regulation of MT1-MMP at the tumor cell surface and malignancy

肿瘤细胞表面MT1-MMP的动态调控与恶性肿瘤

• 批准号: 7051208
• 负责人: Rafael A. Fridman
• 金额: $ 26.55万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-07-06 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7051208
• 关键词: enzyme activity; enzyme induction /repression; extracellular matrix proteins; metalloendopeptidases; neoplasm /cancer invasiveness; neoplastic cell; tissue /cell culture; tissue inhibitor of metalloproteinases; zymogens

DESCRIPTION (provided by applicant): Cancer progression depends on the action of proteolytic systems that facilitate the growth and invasion of tumor cells. The membrane type-1 matrix metalloproteinase (MT1-MMP) endows tumor cells with the ability to invade and grow within collagenous matrices and thus is a key protease in cancer progression. As a membrane-tethered protease, MT1-MMP is regulated by a dynamic interplay of regulatory mechanisms that collectively control the level of active enzyme on the tumor cell surface and in the pericellular space. The long term objective of this application is to unveil the mechanisms regulating MT1-MMP activity at the tumor cell surface and apply this knowledge towards the development of new approaches aimed at inhibiting MT1-MMP in cancer. Our previous effort has been focused on elucidating the structural features and biochemical processes that define the ability of MT1-MMP to undergo autocatalytic processing and ectodomain shedding on the cell membrane, two fundamental processes of enzyme regulation. Processing of active MT1-MMP yields an inactive membrane-tethered fragment of 44 kDa that maintains key enzyme domains but its function in MT1-MMP regulation is poorly understood. Ectodomain shedding of MT1-MMP yields a 50-kDa soluble form that is present in tumors and is a fully competent protease. However its contribution to tumor proteolysis is unknown. Herein, new evidence in vitro and in tumor xenografts shows that the membrane-tethered 44-kDa species, displays a dynamic interaction with active MT1-MMP and identifies this fragment as a complex regulator of enzyme function in tumor cells. The naturally shed ectodomain of MT1-MMP has been characterized and found to be a catalytically competent protease, sensitive to TIMP-2, which has the potential to expand the proteolytic repertoire of MT1-MMP from the confines of the cell membrane to the pericellular space and regulate the activity of membrane-anchored MT1-MMP. Collectively, these observations pose a new paradigm in the regulation of MT1-MMP activity and suggest the hypothesis that processed and soluble forms of MT1-MMP play critical roles in tumor malignancy. To test this hypothesis we propose to: (1) investigate the dynamic interplay between the processed and active forms of MT1-MMP in enzyme function, (2) define the structural basis for the effects of the 44-kDa species on MT1-MMP regulation, (3) investigate the role of the soluble MT1-MMP in the regulation of MT1-MMP activity and (4) investigate the role of processed and soluble forms of MT1-MMP in functional assays of tumor cell invasion and growth in vitro and in vivo. The results of this application will contribute to our understanding of MT1-MMP function in tumor cells and contribute to the collective effort aimed at inhibiting its activity in cancerous tissues.

描述（由申请人提供）：癌症进展取决于促进肿瘤细胞生长和侵袭的蛋白水解系统的作用。膜1型基质金属蛋白酶（MT1-MMP）赋予肿瘤细胞在胶原基质内侵袭和生长的能力，因此是癌症进展中的关键蛋白酶。作为一种膜系蛋白酶，MT1-MMP受多种调节机制的动态相互作用调节，这些调节机制共同控制肿瘤细胞表面和细胞周围空间的活性酶水平。这项应用的长期目标是揭示肿瘤细胞表面MT1-MMP活性的调节机制，并将这些知识应用于开发旨在抑制癌症MT1-MMP的新方法。我们之前的工作集中在阐明MT1-MMP的结构特征和生化过程，这些特征和生化过程定义了MT1-MMP进行自催化处理和细胞膜上的外胞结构域脱落的能力，这是酶调节的两个基本过程。活性MT1-MMP的加工产生一个44 kDa的无活性膜系链片段，维持关键的酶结构域，但其在MT1-MMP调节中的功能尚不清楚。MT1-MMP的外畴脱落产生一种50 kda的可溶性形式，存在于肿瘤中，是一种完全胜任的蛋白酶。然而，其对肿瘤蛋白水解的作用尚不清楚。在此，在体外和肿瘤异种移植物中的新证据表明，膜系结的44 kda物种与活性MT1-MMP表现出动态相互作用，并将该片段鉴定为肿瘤细胞中酶功能的复杂调节剂。MT1-MMP的自然脱落外结构域已被表征并发现是一种具有催化能力的蛋白酶，对TIMP-2敏感，它有可能将MT1-MMP的蛋白水解库从细胞膜范围扩展到细胞周围空间，并调节膜锚定MT1-MMP的活性。总的来说，这些观察结果提出了MT1-MMP活性调控的新范式，并提出了MT1-MMP加工和可溶性形式在肿瘤恶性肿瘤中发挥关键作用的假设。为了验证这一假设，我们提出：(1)研究MT1-MMP加工形式和活性形式在酶功能中的动态相互作用；(2)确定44-kDa物种对MT1-MMP调节作用的结构基础；(3)研究可溶性MT1-MMP在MT1-MMP活性调节中的作用；(4)研究MT1-MMP加工形式和可溶性形式在体外和体内肿瘤细胞侵袭和生长的功能分析中的作用。这一应用的结果将有助于我们理解MT1-MMP在肿瘤细胞中的功能，并有助于共同努力抑制其在癌组织中的活性。