De novo engineering of protein agonists and antagonists

蛋白质激动剂和拮抗剂的从头工程

基本信息

批准号：
8570272
负责人：
Casim Sarkar
金额：
$ 19.95万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-07-01 至 2013-07-02
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8570272
关键词：
Active Sites Affinity Agonist Amino Acid Motifs Ankyrin Repeat Ankyrins Antibodies Binding Buffers C-terminal Cell Line Cell Surface Receptors Cell membrane Cell surface Cells Cysteine Disease Drug Formulations Drug usage Engineering Epithelial Evolution Extracellular Domain Family Generations Genes Goals HIV Hematological Disease Immune System Diseases Individual Interleukin 12 Receptor Beta Interleukin-12 Lead Libraries Ligands Malignant Neoplasms Mediating Medical Membrane Methodology Methods Modeling Patient Care Peptide Hydrolases Physiology Production Property Protein Engineering Proteins Randomized Ribosomes Role Serine Protease Signal Transduction Site Technology Terminal Repeat Sequences Testing Therapeutic Therapeutic antibodies Tumor-Derived cancer therapy crosslink cytokine design directed evolution disulfide bond improved inhibitor/antagonist insight interest interleukin-12 receptor matriptase mimetics mutant novel novel therapeutics pressure public health relevance receptor mediated endocytosis research study scaffold small molecule tumor

项目摘要

DESCRIPTION (provided by applicant): In addition to their vital roles in normal physiology, recombinantly produced cytokines and antibodies constitute a large, critical family of drugs used in the treatments for cancer, HIV, and a host of hematological and immune disorders. However, these molecules are often difficult to produce in mass quantities, suffer from poor folding and aggregation due to the presence of disulfide bonds, and are relatively unstable as therapeutic formulations. Recently, alternative scaffolds with superior biophysical properties to cytokines and antibodies have been used in directed evolution experiments to engineer high-affinity binders to targets of interest. However, cytokine and antibody therapeutics can serve as either agonists or antagonists, and current directed evolution methodologies only select for target binding, not consequent agonism or antagonism. Here, we propose to exploit the modular fold of an alternative scaffold - the designed ankyrin repeat protein - to develop novel directed evolution approaches for isolating new agonists and antagonists. The goals of this R21 proposal are to: 1) engineer anykrin agonists that are mimetics of interleukin-12, a potent anti-tumor cytokine, and 2) create site-specific ankyrin inhibitors of matriptase, a cell-surface protease tha is implicated in a variety of epithelial-derived tumors. More generally, these methods will not only enable the production of new therapeutics, but will also provide unique insights into the basic requirements for specific, high-affinity protein recognition.

描述（由申请人提供）：除了其在正常生理学中的重要作用外，重组产生的细胞因子和抗体还构成了用于癌症，HIV治疗，HIV和许多血液学和免疫疾病的大型药物家族。但是，由于存在二硫键，这些分子通常很难以质量量产生，折叠和聚集较差，并且作为治疗制剂相对不稳定。最近，在定向的进化实验中使用了具有优质生物物理特性的替代支架，用于对目标靶标进行高亲和力粘合剂的设计。但是，细胞因子和抗体疗法可以用作激动剂或拮抗剂，而当前的定向进化方法仅选择目标结合，而不是随之而不是激动剂或拮抗作用。在这里，我们建议利用替代支架的模块化褶皱 - 设计的弯蛋白重复蛋白 - 开发出新的定向进化方法，以隔离新的激动剂和拮抗剂。该R21提案的目标是：1）工程师Anykrin激动剂，它们是白介素12的模拟物，有效的抗肿瘤细胞因子和2）创建位点特异性的Matriptase的特异性弯曲蛋白抑制剂，一种细胞表面蛋白酶THA的tha含义在各种上皮层虫的the剂中。更普遍的是，这些方法不仅可以产生新的治疗剂，而且还将为特定高亲和力蛋白质识别的基本要求提供独特的见解。