Protein Synthesis Inhibitors as anti-T. Cruzi agents

蛋白质合成抑制剂作为抗 T。

• 批准号: 10043125
• 负责人: BERTAL H. AKTAS
• 金额: $ 22.38万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-19 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10043125
• 关键词: Acute; Adopted; Amino Acids; Antibiotics; Antineoplastic Agents; Antiparasitic Agents; Area; Bacteria; Bacterial Proteins; Biological; Biological Assay; Biological Process; Biology; Cells; Chagas Disease; Chemical Agents; Chemicals; Chronic; Communities; Cycloheximide; Cytostatics; Development; Diagnosis; Dimethyl Sulfoxide; Drug Targeting; Elongation Factor; Eukaryota; Fostering; Gene Expression; Genetic Translation; Goals; Health; Human; In Vitro; Incubated; Industry; Infection; Label; Laboratories; Lead; Leishmania; Libraries; Life Cycle Stages; Luciferases; Mammalian Cell; Mammals; Messenger RNA; Methionine; Molecular Weight; Natural regeneration; Oral; Oryctolagus cuniculus; Parasites; Patients; Peer Review; Peptide Initiation Factors; Pharmaceutical Preparations; Pharmacotherapy; Positioning Attribute; Preparation; Property; Protein Biosynthesis; Protein Synthesis Inhibitors; Proteins; Protozoa; Publications; Renilla Luciferases; Reporter; Reporter Genes; Reporting; Reproducibility; Research; Reticulocytes; Ribosomal Proteins; Ribosomal RNA; Ribosomes; Structure; Structure-Activity Relationship; Symptoms; System; Testing; Time; Toxic effect; Translation Initiation; Translations; Trypanosoma; Trypanosoma cruzi; Tumor stage; Urea; Validation; assay development; base; cancer therapy; chronic infection; compound 30; cost; counterscreen; cytotoxic; cytotoxicity; design; drug development; experience; high throughput screening; inhibitor/antagonist; lead optimization; lead series; miniaturize; novel; pathogen; screening; small molecule libraries; translation assay; water solubility

Project Abstract Trypanosoma cruzi is the causative agent of the Chagas’ disease. Despite the devastating effects of chronic T. cruzi infections on human health, there are no satisfactory options for their treatment. The goal of this proposal is to develop effective, affordable, and safe therapies against chronic T. cruzi infections. Such therapies must target a biological process essential for T. cruzi viability without disrupting it in humans. This concept has successfully been used for the development of antibiotics, many of which target bacterial protein synthesis with minimal toxicity to humans. T. cruzi protein synthesis differs substantially from that of humans at the levels of mRNA structure, translation initiation and elongation factors, ribosomal RNA and proteins and ribosome structure. These differences render every stage of T. Cruzi protein synthesis potentially suitable for the development of T,Cruzi specific protein synthesis inhibitors for the treatment of chronic infections. We therefore propose to optimize and adapt a T. cruzi in vitro translation assay developed in our laboratory to high throughput screening (Specific Aim 1) and implement an HTS campaign to identify agents that specifically inhibit T. cruzi protein synthesis and selectively kill these parasites (Specific Aim 2). PI has developed and implemented several high throughput screening (HTS) assays, obtained three first-in- class compounds and optimized these by evaluating focused libraries and structure-activity-relationships studies. These probes have been widely adapted by the scientific community and fostered research that resulted in more than 100 peer-reviewed publications by PI, his collaborators and others. Two of these lead series have been licensed to industry for further development into drugs for cancer therapy. In this application we propose to utilize our expertise in biology of translation and HTS assay development and implementation to identify the specific/selective inhibitors T.Cruzi protein synthesis. In addition to primary screening assay we will employ a GFP reporter T. cruzi in vitro translation assay to eliminate false positives and a counter dual luciferase mammalian in vitro translation assay to eliminate hit compounds not specific for T. cruzi, i.e. those that also inhibit mammalian protein synthesis. We will test specific inhibitors of T. cruzi protein synthesis for anti-parasitic activity and selective toxicity to parasite vs. host cells. While what we propose here is a pilot screen, we possess all the expertise and facilities needed to conduct a large scale screen and/or hit-to-lead optimization for the hits we will identify. Effort towards the hit-to-lead optimization would require that one or more of our specific/selective T.cruzi protein synthesis inhibitor compounds will display toxicity towards T.cruzi but not the mammalian cells and will be chemically accessible for design of focused libraries and structure-activity-relationship studies. Identification of the direct target of hit compounds is another area of future research that will elucidate biological differences between translation machineries of human and T. cruzi, and all well within the PI’s capabilities.

项目摘要 尽管克氏锥虫具有毁灭性的影响，但它仍然是恰加斯病的病原体。 克鲁兹感染对人类健康的影响，目前尚无令人满意的治疗方案。该提案的目标是。 开发针对慢性克氏锥虫感染的有效、负担得起且安全的疗法是必须的。 目标是克氏锥虫生存所必需的生物过程，而不破坏它在人类中的作用。 已成功用于抗生素的开发，其中许多抗生素以细菌蛋白质合成为目标 对人类的毒性最小。克氏锥虫的蛋白质合成水平与人类有很大不同。 mRNA 结构、翻译起始和延伸因子、核糖体 RNA 和蛋白质以及核糖体 这些差异使得 T. Cruzi 蛋白质合成的每个阶段都可能适合 因此，我们开发了 T,Cruzi 特异性蛋白质合成抑制剂来治疗慢性感染。 建议优化和调整我们实验室开发的 T. cruzi 体外翻译测定，以达到高 通量筛选（具体目标 1）并实施 HTS 活动来识别专门针对 抑制克氏锥虫蛋白质合成并选择性杀死这些寄生虫（具体目标 2）。 PI 开发并实施了多项高通量筛选 (HTS) 检测方法，获得了三项首创 类化合物并通过评估重点库和结构-活性关系来优化它们 这些探针已被科学界广泛采用，并促进了以下研究： PI、他的合作者和其中两位领导者发表了 100 多篇经过同行评审的出版物。 系列已授权给工业界以进一步开发成癌症治疗药物。 在此应用中，我们建议利用我们在翻译生物学和 HTS 检测开发方面的专业知识， 实施以鉴定除初级蛋白质合成之外的特异性/选择性抑制剂 T.Cruzi。 筛选试验 我们将采用 GFP 报告基因 T. cruzi 体外翻译试验来消除假阳性 以及反双荧光素酶哺乳动物体外翻译测定，以消除非特异性的命中化合物 克氏锥虫，即那些也抑制哺乳动物蛋白质合成的抑制剂。我们将测试克氏锥虫的特定抑制剂。 蛋白质合成具有抗寄生虫活性和对寄生虫与宿主细胞的选择性毒性。 这是一个试点屏幕，我们拥有进行大规模测试所需的所有专业知识和设施 对我们将确定的点击进行筛选和/或点击到引导优化 努力实现点击到引导优化。 需要我们的一种或多种特异性/选择性 T.cruzi 蛋白质合成抑制剂化合物 对 T.cruzi 表现出毒性，但对哺乳动物细胞没有毒性，并且可以通过化学方法设计 重点库和结构-活性-关系研究的确定是命中化合物的直接目标。 未来研究的另一个领域将阐明翻译机器之间的生物学差异 人类和克氏锥虫，并且都在 PI 的能力范围内。