Engineering selective inhibition of metalloproteinases by tissue inhibitors of metalloproteinases (R01 GM132100 RESUB - *TIMPs)

通过金属蛋白酶组织抑制剂对金属蛋白酶进行工程选择性抑制（R01 GM132100 RESUB - *TIMP）

• 批准号: 10319170
• 负责人: Evette S Radisky
• 金额: $ 31.3万
• 依托单位: MAYO CLINIC JACKSONVILLE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10319170
• 关键词: Address; Affinity; Arthritis; Autoimmune Diseases; Binding; Biological Assay; Biological Models; Biology; C-terminal; Cardiovascular Diseases; Catalytic Domain; Complement; Complex; Computer Analysis; Computer Models; Crotalus adamanteus proteinase II; Crystallization; Data; Development; Directed Molecular Evolution; Discrimination; Disease; Disease model; Diversity Library; Dose-Limiting; Drug Targeting; Endopeptidases; Engineering; Enzymes; Epitopes; Family; Foundations; Free Energy; Generations; Goals; Individual; Inflammatory; Inhibition of Matrix Metalloproteinases Pathway; Kinetics; Length; Libraries; Malignant Neoplasms; Maps; Matrix Metalloproteinase Inhibitor; Matrix Metalloproteinases; Mediating; Metalloproteases; Methodology; Methods; Modeling; Molecular; Molecular Probes; Morphogenesis; Musculoskeletal; Mutagenesis; Mutation; N-terminal; Names; Outcome; Pathologic; Pathology; Peptide Hydrolases; Pharmaceutical Preparations; Physiological; Play; Process; Progression-Free Survivals; Protease Inhibitor; Protein Dynamics; Protein Engineering; Proteins; Publishing; Reagent; Roentgen Rays; Role; Scaffolding Protein; Shapes; Specificity; Structural Models; Structure; Surface; Therapeutic; Tissue Engineering; Tissue Inhibitor of Metalloproteinase-1; Tissue Inhibitor of Metalloproteinases; Tissues; Toxic effect; Variant; Work; X-Ray Crystallography; Yeasts; Zinc; base; design; early phase clinical trial; inhibitor; molecular dynamics; molecular recognition; next generation; novel; protein complex; protein folding; scaffold; screening; synergism; therapeutic protein; therapeutic target; therapeutically effective; tool

PROJECT SUMMARY/ABSTRACT Matrix metalloproteinases (MMPs) have long been regarded as promising therapeutic targets, but clinical trials of early-generation MMP inhibitors in arthritis and cancer proved disappointing. Broad-spectrum MMP inhibitors produced serious dose-limiting musculoskeletal toxicity and failed to extend progression-free survival in cancer trials, partly due to the inability of the inhibitors to distinguish among different MMPs. This was a critical problem, because some MMPs serve a primarily protective function, and it is now clear that indiscriminate inhibition of all MMPs inevitably leads to poorer outcomes. Based on our published and preliminary data, we hypothesize that tissue inhibitors of metalloproteinases (TIMPs), endogenous regulators of the MMP family, can be engineered into highly selective MMP inhibitors free of undesired off-target activities, to produce probes and therapeutics targeting individual MMPs with exquisite selectivity. In this application, we will (a) optimize novel methodology for directed evolution of selective binders to discriminate within the large families of related MMPs and adamalysin proteases, (b) uncover mechanisms of molecular recognition that govern MMP-TIMP binding specificity, and (c) develop a toolbox of engineered TIMPs that selectively target individual MMPs with highly enhanced specificity. To accomplish these goals, we will use state-of-the-art directed evolution approaches to engineer the TIMP-1 scaffold for fine discrimination between closely similar MMPs, reengineering N- and C-terminal TIMP domain epitopes and exploiting cooperativity between domains. Additionally, we will integrate X-ray crystallographic and computational approaches to elucidate the protein structural and dynamic features that govern affinity and selectivity of TIMP/MMP complexes. This project will thus elucidate fundamental principles of molecular recognition governing TIMP/MMP selectivity, and will produce designer TIMPs targeting single MMPs with highly enhanced specificity, with potential for development as useful molecular probes and as protein therapeutics for the many diseases driven by MMP dysregulation.

项目总结/文摘