Models of AT Deficiency Using Human Hepatocytes

使用人肝细胞的 AT 缺陷模型

• 批准号: 10630350
• 负责人: Ira J. Fox
• 金额: $ 60.47万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-24 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10630350
• 关键词: Adolescence; Adult; Age of Onset; Alleles; Applications Grants; Biology; CRISPR/Cas technology; Caenorhabditis elegans; Cells; Characteristics; Childhood; Chronic; Cirrhosis; Clinical; Custom; Degradation Pathway; Development; Disparate; Effectiveness; Endoplasmic Reticulum; Evaluation; Exhibits; Exons; Fibrinogen; Fibroblasts; Gene Modified; Genes; Genetic; Genetic Variation; Goals; Hepatic; Hepatocyte; Hepatotoxicity; Hereditary Disease; Heterozygote; Hong Kong; Human; Human Engineering; Immune; Impairment; In Vitro; Kinetics; Libraries; Liver; Liver Fibrosis; Liver diseases; Lung; Lung diseases; Modeling; Morphology; Mutation; Pathologic; Patients; Pharmaceutical Preparations; Pharmacotherapy; Phenotype; Physical shape; Polymers; Predisposition; Primary carcinoma of the liver cells; Property; Proteins; Rattus; Rodent; Role; Signal Pathway; Skin; Small Interfering RNA; System; Testing; Therapeutic; Transforming Growth Factor beta; Trypsin; Update; Validation; Variant; alpha 1-Antitrypsin Deficiency; clinical phenotype; differentiation protocol; disease phenotype; drug candidate; endoplasmic reticulum stress; genome editing; high throughput analysis; human model; human tissue; in vitro Model; in vivo; in vivo Model; induced pluripotent stem cell; infancy; liver injury; liver transplantation; middle age; mutant; novel therapeutics; programs; proteotoxicity; response; retrorsine; stem cell differentiation; stem cells

SUMMARY The proposed study aims to develop models of ATD using patient-derived cells and use these models to confirm previously identified modifiers of ATD and test potential drug therapies for ATD in human cells. We have shown that stem cell-derived hepatocyte-like cells (iHeps) generated by differentiating stem cells from patients with liver vs. lung manifestations of ATD exhibit differences in ultrastructural morphology and kinetics of mutant ATZ disposal. Based on these findings, we hypothesize that genetic variations other than the ATZ mutation determine the clinical phenotype of ATD. As a part of the previous cycle of this program project, we and our colleagues have used high throughput analysis in C. elegans using a siRNA library to identify several genes that modify ATZ accumulation. The proposed project will reveal the differences in accumulation of ATZ and its proteotoxicity in iHeps from various subsets of ATD patients delinieated by age of onset of liver disease, disparate hepatic phenotype, presence of co-existing pulmonary disease or hepatocellular carcinoma, and heterozygosity for the ATZ allele. We will also examine a) the extent to which in vitro characteristics of iHeps correspond to various ATD clinical subsets, b) the role of genetic modulators on ATD liver disease phenotypes, c) the response of iHeps to candidate drugs for the treatment of ATD, and d) whether the delay in ATZ disposal that characterizes iHeps from ATD patients with severe liver disease is due to aggregation-prone properties of ATZ. Finally, we will generate in vivo models of human ATD by repopulating the livers of retrorsine-treated immune deficient rats with primary human hepatocytes and iHeps from ATD patients and controls. The development of in vivo and in vitro models of ATD using patient-derived cells would greatly benefit the discovery and validation of target genes for novel therapies of ATD and evaluation of potential therapeutic drugs.

总结