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Allosteric Pharmacologic Chaperones for alpha-1 Antitrypsin Mutants

α-1 抗胰蛋白酶突变体的变构药理学伴侣

基本信息

批准号：
10390150
负责人：
Adrian M. Guerrero
金额：
$ 3.38万
依托单位：
SCRIPPS RESEARCH INSTITUTE, THE
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-01 至 2024-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10390150
关键词：
Address Adult Affect Affinity Affinity Chromatography Alkynes Binding Binding Sites Biological Assay Biology Cell model Cells Chemicals Chemistry Chronic Chronic Obstructive Pulmonary Disease Clinical Clinical Medicine Clinical Pathology Competence Complex Crystallization Disease model Elastases Enzyme-Linked Immunosorbent Assay Exhibits Future Hepatocyte Homeostasis Hydrophobicity Impairment In Situ In Vitro Inherited Laboratories Libraries Ligand Binding Ligands Literature Liver Liver Failure Liver diseases Lung Lung diseases Mammalian Cell Maps Mass Spectrum Analysis Measures Methods Modeling Molecular Chaperones Morbidity - disease rate Mutation Mutation Analysis Pathology Patients Peptide Hydrolases Peptides Pharmaceutical Chemistry Pharmaceutical Preparations Pharmacology Phenotype Photoaffinity Labels Physicians Polymers Protease Inhibitor Proteins Proteome Proteomics Proxy Pulmonary Pathology Reporting Research Research Personnel Role Scientist Serum Site Structure Structure of parenchyma of lung Surface System Therapeutic Training Variant alpha 1-Antitrypsin alpha 1-Antitrypsin Deficiency assay development base career cellular pathology chronic liver disease crosslink cytotoxic cytotoxicity drug discovery early onset experience extracellular high throughput screening in silico in vivo innovation loss of function miniaturize monomer mortality multidisciplinary multiorgan injury mutant neutrophil novel peptidomimetics plasma protein Z polymerization pre-clinical preservation prevent protein misfolding screening small molecule tandem mass spectrometry trafficking

项目摘要

Project Summary/Abstract The major deficiency E342K mutant or “Z-variant” of the abundant serum antiprotease Alpha-1 antitrypsin (AAT) is responsible for the vast majority of morbidity and mortality associated with Alpha-1 antitrypsin deficiency (AATD), a leading cause of hereditary lung and liver disease affecting millions of patients globally. This missasembly-prone variant is known to be highly polymerogenic, owing to a widened -sheet A domain which predisposes the AAT-Z monomer to form cytotoxic loop-sheet oligomers. These toxic oligomers accumulate in producing hepatocytes leading to chronic liver disease, and build up extracellularly leading to both gain-of-toxic function in the lung with a concomitant loss of serum AAT-Z antiprotease activity which leads to proteolytic destruction of lung parenchyma by neutrophil proteases like elastase. Crystal structures of monomeric AAT-Z have been studied and previous mutational analyses have demonstrated the capacity for space-filling mutations within surface-accessible hydrophobic pockets on this protein to prevent polymerization without abrogating antiprotease activity. It has thus been hypothesized that small molecules could be discovered which act as pharmacological chaperones, preventing AAT-Z polymer formation while permitting native antiprotease activity of the stabilized monomer, serving to ameliorate the multiorgan injury (including lung pathology) associated with AATD. While stabilizing ligands for AAT-Z have previously been reported, these ligands universally fail to permit the native antiprotease activity of AAT-Z. In this training proposal, I propose to use Fully-Functionalized Fragment” (FFF) substructures along with photo-crosslinking, affinity chromatography and tandem mass-spectrometry in Aim 1 to identify the small molecule sites on the AAT-Z monomer accessible to binding by drug-like substructures. In Aim 2 I will develop a novel screening assay that I conceived of for identifying AAT-Z stabilizing ligands or pharmacologic chaperones that prevent RCL insertion while permitting antiprotease activity of ligand-bound AAT-Z. In Aim 3 I will employ a cell-based phenotypic assay already developed and validated by our collaborators in the Balch Lab to simultaneously evaluate the capacity for screening hits to restore functional monomeric AAT-Z secretion efficiency, while reducing intracellular oligomers in cultured hepatocyte and pulmonary cell models of AATD. This project will afford a multidisciplinary training experience with guidance from experts in the fields of protein misfolding biology, chemical proteomics, high-throughput assay development, as well as AATD patient treatment. Through the research proposed herein, I will develop a robust expertise in state-of-the-art methods for proteomic analysis, screening assay optimization, and applications of mammalian cell models of disease. These valuable discovery-oriented research competencies, together with my previous training background in pre-clinical medicine and medicinal chemistry will serve to enable a highly productive future career as a leading physician- scientist researcher in the challenging field of first-in-class drug discovery.

项目总结/文摘