Engineering high affinity integrin binding proteins

工程化高亲和力整合素结合蛋白

• 批准号: 6892795
• 负责人: JENNIFER R COCHRAN
• 金额: $ 15.95万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-05-10 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6892795
• 关键词: angiogenesis; binding proteins; biotechnology; carcinogenesis inhibitor; cell adhesion; chemical registry /resource; chemical stability; crosslink; cyclic peptides; drug design /synthesis /production; flow cytometry; high performance liquid chromatography; immunologic substance resource /registry /referral center; integrins; mass spectrometry; matrix assisted laser desorption ionization; metastasis; protein binding; protein engineering; protein folding; protein structure function; receptor coupling; tissue support frame; vascular endothelium; yeast two hybrid system

DESCRIPTION (provided by applicant): The candidate has a multidisciplinary background in chemistry, biochemistry and engineering. These fields will be combined to develop stable, high affinity integrin binding proteins for mediating cell adhesion and angiogenesis in cancer. The candidate will spend a year in a mentored position in the laboratories of William F. DeGrade and Joel S. Bennett at the University of Pennsylvania School of Medicine. During this period, the applicant will develop skills in peptide and protein design, and organic synthesis while in the DeGrade lab. Experience in integrin receptor function and biological assays will be gained in the Bennett lab. This period will be important in establishing the groundwork for the unmentored portion of the award. The remainder of the award will be spent in an independent, preferrably tenure-track, academic position, where the applicant will build upon this additional expertise to develop biomolecules for cancer therapeutics and tumor imaging applications. The University of Pennsylvania School of Medicine has a strong commitment to research in health and human disease, and several of its faculty are conducting integrin and/or cancer related research. The mentors chosen by the applicant have well-established, productive collaborations in studying the structure and function of integrin proteins. In addition, Prof. DeGrade has much expertise in development of integrin-specific mimics through rational design and combinatorial library panning in collaboration with DuPont. Integrin-specific peptide mimics and antibodies have shown much therapeutic promise in the inhibition of angiogenesis and metastases in tumors, but can benefit greatly from stability and affinity maturation. The mentored period of the award will involve incorporation of integrin-specific peptide motifs into constrained molecular scaffolds to increase stability and binding affinity. These scaffolds will provide the framework for a portion of the unmentored period of the award, where integrin binding proteins will be engineered to even higher affinity using directed molecular evolution by yeast surface display. In addition, the mentored period of the award will focus on the design and synthesis of a series of novel peptide-based crosslinking reagents. These compounds will be used for creating multivalent integrin-specific peptides and proteins for high avidity binding and enhanced antagonism. In the unmentored period of the application, a nonimmune human library of scFv antibody fragments displayed on the surface of yeast will be used to isolate a panel of integrin-specific binders. These antibodies will be characterized for their ability to inhibit RGD-mediated binding and cell adhesion. Promising antibodies will be engineered to ultra high affinity and stability using yeast surface display to develop potent integrin antagonists.

描述（由申请人提供）：候选人具有化学，生物化学和工程学的多学科背景。这些领域将结合起来，开发稳定的，高亲和力的整合素结合蛋白介导的细胞粘附和血管生成的癌症。候选人将在威廉F. Degrade和Joel S.宾夕法尼亚大学医学院的班尼特说。在此期间，申请人将发展肽和蛋白质设计以及有机合成方面的技能。在整合素受体功能和生物测定的经验将获得在班尼特实验室。这段时间对于为该奖项的无人指导部分奠定基础至关重要。该奖项的其余部分将用于一个独立的，最好是终身制的学术职位，申请人将在此基础上开发用于癌症治疗和肿瘤成像应用的生物分子。宾夕法尼亚大学医学院对健康和人类疾病的研究有着坚定的承诺，其几位教师正在进行整合素和/或癌症相关的研究。申请人选择的导师在研究整合素蛋白的结构和功能方面有着良好的、富有成效的合作。此外，DeGrade教授还与杜邦公司合作，通过合理设计和组合文库淘选开发整合素特异性模拟物。 整合素特异性肽模拟物和抗体在抑制肿瘤中的血管生成和转移方面已经显示出很大的治疗前景，但是可以从稳定性和亲和力成熟中大大受益。该奖项的指导期间将涉及整合素特异性肽基序纳入约束分子支架，以增加稳定性和结合亲和力。这些支架将为该奖项的一部分未指导期提供框架，其中整合素结合蛋白将通过酵母表面展示使用定向分子进化工程化至甚至更高的亲和力。此外，该奖项的指导期将专注于一系列新型肽基交联试剂的设计和合成。这些化合物将用于产生多价整联蛋白特异性肽和蛋白质，用于高亲合力结合和增强的拮抗作用。在未指导的申请期间，将使用酵母表面展示的scFv抗体片段的非免疫人类文库来分离一组整合素特异性结合剂。这些抗体将表征其抑制RGD介导的结合和细胞粘附的能力。有前途的抗体将工程超高亲和力和稳定性使用酵母表面展示开发有效的整合素拮抗剂。