Atrogin-1 inhibitors for Muscle Wasting

Atrogin-1 抑制剂治疗肌肉萎缩

• 批准号: 7107637
• 负责人: Michael R Mattern
• 金额: $ 24.91万
• 依托单位: PROGENRA, INC.
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-05-01 至 2008-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7107637
• 关键词: Saccharomyces cerevisiae; atrophy; calcineurin; drug screening /evaluation; enzyme activity; high throughput technology; muscle function; muscle proteins; p53 gene /protein; protease inhibitor; proteasome; sarcopenia; technology /technique development; ubiquitin; ubiquitin protein ligase

DESCRIPTION (provided by applicant): Muscle atrophy (wasting) is a serious clinical complication of diabetes and other chronic pathoses, leading to increased morbidity and reduced life expectancy. While symptoms have been addressed by a number of interventions, no successful therapy has been developed. Recently, various proteins whose expression is enhanced under conditions of atrophy-inducing starvation have been identified in rats. In particular, the expression of a set of novel genes called atrogins (atrophy-specific genes) increases significantly in skeletal muscles of fasting organisms and decreases rapidly when feeding is resumed. The product of one of these genes - atrogin-1 - is an F-box protein ubiquitin E3 ligase, a critical enzyme of the ubiquitin-proteasomal degradation pathway. This pathway is implicated in muscle atrophy, since proteasome inhibitors protect against muscle wasting in model systems. Selective attenuation of proteasomal activity associated with muscle wasting may be achievable using inhibitors of atrogin-1. In Phase 1, E3 ubiquitination activity of atrogin-1 ectopically expressed in yeast will be demonstrated against a known substrate, calcineurin. Then, a yeast-based assay for atrogin-1 will be developed and validated, in preparation for high throughput screening of compound and natural products collections to discover inhibitors of atrogin-1 in Phase 2. For the assay, the following will be cloned and expressed in yeast S. cerevisiae: the substrate of atrogin-1, fused to p53 linked to a beta-galactosidase reporter; atrogin-1 E3 ligase complex; and an E3 ligase suitable as a selectivity control. Results of pilot experiments to validate this modular assay demonstrate that human E3 ligases, p-53 activated reporter plasmid, and p53-fused ligase substrate ectopically expressed in yeast give a null signal, but produce a robust reporter signal when E3 ligase is left out, consistent with ubiquitination/degradation of fused p53 by E3. Successful completion of Phase 1 should result in a validated screening assay that can be utilized in Phase 2. In Phase 2, active principles will be isolated from the best extract leads, with the goal of identifying novel, potent, and selective inhibitors of atrogin-1 for development as adjuvant therapy of muscle wasting associated with diabetes.

描述（由申请人提供）：肌肉萎缩（消瘦）是糖尿病和其他慢性疾病的严重临床并发症，导致发病率增加和预期寿命缩短。虽然已经通过一些干预措施解决了症状，但尚未开发出成功的治疗方法。最近，已经在大鼠中鉴定了在萎缩诱导饥饿条件下表达增强的各种蛋白质。特别是，一组称为atrogins（萎缩特异性基因）的新基因的表达在禁食生物体的骨骼肌中显着增加，并在恢复进食时迅速减少。这些基因之一- atrogin-1 -的产物是F-box蛋白泛素E3连接酶，泛素-蛋白酶体降解途径的关键酶。该途径与肌肉萎缩有关，因为蛋白酶体抑制剂在模型系统中保护肌肉免于萎缩。使用atrogin-1的抑制剂可以实现与肌肉萎缩相关的蛋白酶体活性的选择性衰减。在第1阶段，E3泛素化活性atrogin-1异位表达在酵母中将证明对一个已知的底物，钙调磷酸酶。然后，将开发并验证基于酵母的atrogin-1测定法，为化合物和天然产物集合的高通量筛选做准备，以在第2阶段发现atrogin-1的抑制剂。对于该试验，将克隆以下基因并在酵母S.酿酒酵母：与连接β-半乳糖苷酶报告基因的p53融合的atrogin-1的底物; atrogin-1 E3连接酶复合物;和适合作为选择性对照的E3连接酶。验证该模块化测定的初步实验的结果表明，在酵母中异位表达的人E3连接酶、p-53活化的报告质粒和p53融合的连接酶底物产生无效信号，但当E3连接酶被排除时产生稳健的报告信号，这与E3对融合的p53的泛素化/降解一致。成功完成第1阶段应产生可用于第2阶段的经验证的筛选试验。在第二阶段，将从最好的提取物中分离出活性成分，目的是鉴定新型、有效和选择性的atrogin-1抑制剂，用于开发与糖尿病相关的肌肉萎缩的辅助治疗。

项目成果

Analysis of the biliary transcriptome in experimental biliary atresia.

实验性胆道闭锁的胆道转录组分析。

• DOI: 10.1016/j.gastro.2005.05.052
• 发表时间: 2005
• 期刊: Gastroenterology.
• 作者: Carvalho,Elisa;Liu,Cong;Shivakumar,Pranavkumar;Sabla,Gregg;Aronow,Bruce;Bezerra,JorgeA
• 通讯作者: Bezerra,JorgeA