Small Molecule Probes to Correct AGT Mistargeting in Primary Hyperoxaluria 1

用于纠正原发性高草酸尿症中 AGT 误定位的小分子探针 1

基本信息

批准号：
8913596
负责人：
Carla M Koehler
金额：
$ 23.1万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-08-21 至 2018-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8913596
关键词：
Alanine-glyoxylate aminotransferase Alleles Area Attenuated Back Biogenesis C-terminal Calcium Oxalate Cell Line Cell model Cells Chemicals Chinese Hamster Ovary Cell Chloride Ion Chlorides Code Collaborations Collection Cultured Cells Cytosol Data Defect Development Dialysis patients Disease Enzymes Event FDA approved Fanconi Syndrome Fibroblasts Future Genes Genetic Polymorphism Genetic Screening Glutamates Glycine Glyoxylates Goals Human Kidney Kidney Calculi Kidney Diseases Lead Left Letters Life Expectancy Location Lysine Mitochondria Mitochondrial Proteins Modeling Molecular Mutation N-terminal Neuropathy Oxalates Pathway interactions Patients Peptide Hydrolases Point Mutation Positioning Attribute Primary Hyperoxaluria Process Protein Import Protein translocation Proteins Public Health Publications Research Supplementation Symptoms Testing Therapeutic Tissues Uremia Vitamin B6 Yeasts base cofactor disease diagnosis disease-causing mutation exome sequencing expectation fatty acid oxidation genome wide association study glyoxylate in vivo induced pluripotent stem cell insight interest liver transplantation mitochondrial processing peptidase mutant peroxisome protein activation protein transport public health relevance reconstitution small molecule success targeted sequencing tool trafficking yeast genetics

项目摘要

DESCRIPTION (provided by applicant): Primary hyperoxaluria type I (PH1) and renal Fanconi's syndrome are two rare kidney diseases that share a common theme in which a peroxisomal protein is mistargeted to mitochondria. As additional kidney diseases are diagnosed with genome-wide association studies, this theme in which proteins are mistargeted to mitochondria is likely to become more common, because mitochondrial targeting sequences are degenerate. PH1 is an autosomal recessive disease caused by mutations in the gene coding for alanine-glyoxylate aminotransferase (AGT/AGXT, protein abbrev. AGT). PH1 is marked by an inability to efficiently metabolize glyoxylate, leading to the accumulation of calcium oxalate in various bodily tissues, especially the kidney. In addition to uremia, which can be transiently treated by dialysis, patients have other complications such as neuropathy from excess oxalates. Administration of pyridoxine (B6), a cofactor of AGT, has alleviated some symptoms in a subset of patients; however, adequate treatments are lacking and the disease is typically terminal. Whereas some mutations in AGT result in protein activation, a subset of mutations (one-third of patients have an allele with 2 point mutations, P11L and G170R) results in mistargeting of functional AGT from peroxisomes, where it is active in humans (omnivores), to mitochondria, which is the normal location in carnivores. Similarly, renal Fanconi's syndrome is caused by mistargeting of the peroxisomal bifunctional enzyme (PBE, coded by EHHADH) to mitochondria. PBE subsequently blocks mitochondrial fatty acid oxidation, resulting in an autosomal dominant disease. Our hypothesis is that small molecules that attenuate mitochondrial protein import are significant for dissecting the molecular mechanisms in AGT/PBE trafficking and, long-term, as a therapeutic strategy to retarget the protein from mitochondria back to peroxisomes. We have completed a high throughput in vivo screen in yeast to find attenuators of mitochondrial protein translocation. Strong preliminary data supports that these small molecule probes attenuate import in both yeast and mammalian mitochondria. Moreover, small molecule candidates have been identified that indeed retarget AGT from mitochondria back to peroxisomes and these probes are well tolerated by cells. Thus, the small molecules partially inhibit mitochondrial protein translocation at a level that is not toxic to cels. The specific aims of this proposal are: (1) to elucidate the specific trafficking pathway of AGT in cells; (2) to characterize the mechanism by which the small molecules retarget AGT from mitochondria to peroxisomes and to determine if the small molecules may be beneficial for patients with different mutations in AGT; and (3) to study the trafficking pathway of PBE mistargeting to mitochondria. This study is relevant to public health because of the potential development of new strategies to understand and treat kidney diseases.

描述（由适用提供）：原发性高黄油I型（pH1）和肾芬科综合征是两种罕见的肾脏疾病，它们具有共同的主题，其中过氧化物体蛋白被对线粒体错误定位。由于其他肾脏疾病被诊断为全基因组关联研究，因此将蛋白质定位于线粒体的主题可能会变得更加普遍，因为线粒体靶向序列是退化的。 PH1是由编码丙氨酸 - 甘氧基氨基转移酶的基因突变引起的常染色体隐性疾病（AGT/AGXT，蛋白质adbrev。Agt）。 PH1的特征是无法有效代谢乙二基，导致草酸钙在各种身体组织，尤其是肾脏中的积累。除了可以通过透析瞬时治疗的尿素外，患者还具有其他并发症，例如过量草酸盐的神经病。 AGT的辅助因子吡ido醇（B6）的给药缓解了一部分患者的某些症状。但是，缺乏足够的治疗方法，该疾病通常是终末。尽管AGT的某些突变导致蛋白质激活，但突变的子集（三分之一的患者具有2点突变，P11L和G170R）会导致过氧化物体的功能AGT失误，该功能AGT来自人体（Omnivores）（Omnivores）（Omnivores），在Carnivores中是正常位置。同样，肾芬科综合征是由过氧化物体双功能酶（PBE，由Ehhadh编码）与线粒体编码的错误定位引起的。 PBE随后阻断线粒体脂肪酸氧化，导致常染色体显性疾病。我们的假设是，减弱线粒体蛋白进口的小分子对于剖析AGT/PBE运输中的分子机制是重要的，并且长期作为一种治疗策略，是将蛋白从线粒体重新定位的治疗策略。我们已经在酵母中完成了高吞吐量的体内筛选，以找到线粒体蛋白易位的衰减器。强大的初步数据支持这些小分子问题减弱了在酵母和哺乳动物线粒体中的进口。此外，已经确定了小分子的候选物，确实，实际上从线粒体重新定位了AGT回到过氧化物体，并且这些问题受细胞的耐受性耐受性。这是小分子部分抑制线粒体蛋白的易位，在对碳无毒的水平上。该提案的具体目的是：（1）阐明AGT的特定贩运途径细胞；（2）表征小分子从线粒体重新定位到过氧化物体的机制，并确定小分子是否可能对AGT突变不同的患者有益；（3）研究PBE错误定位到线粒体的贩运途径。这项研究与公共卫生有关，因为新策略的潜在发展和治疗肾脏疾病。