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 克鲁齐锥虫是查加斯病的病因。尽管慢性T。 克鲁兹（Cruzi）关于人类健康的感染，他们的治疗没有满意的选择。该提议的目标 是为了开发有效，负担得起和安全的疗法，以抵抗慢性T. cruzi感染。这种疗法必须 针对克鲁氏菌生存能力至关重要的生物学过程，而不会在人类中破坏它。这个概念有 成功地用于开发抗生素，其中许多靶向细菌蛋白合成 对人类的毒性最小。 T. cruzi蛋白合成与人类在 mRNA结构，翻译起始和伸长因子，核糖体RNA和蛋白质以及核糖体 结构。这些差异使克鲁兹蛋白质合成的每个阶段都可能适合 T，Cruzi特异性蛋白质合成抑制剂的开发用于治疗慢性感染。因此，我们 提议优化和适应在我们实验室中开发的体外翻译测定法至高的提案 吞吐量筛选（特定目标1）并实施HTS活动，以确定特定的代理 抑制克鲁齐蛋白质的合成并有选择地杀死这些寄生虫（特定目标2）。 PI已开发并实施了几个高通量筛选（HTS）测定法，获得了三个首次 类化合物并通过评估集中的库和结构活动相关性来优化它们 研究。这些问题已被科学界广泛适应，并培养了研究 PI，他的合作者和其他人进行了100多个同行评审的出版物。其中两个领先 系列已获得工业许可，以进一步发展为癌症治疗的药物。 在此应用中，我们建议利用我们在翻译和HTS分析开发生物学领域的专业知识以及 实施以识别特异性/选择性抑制剂T.cruzi蛋白合成。除了主要 筛选测定法我们将在体外翻译测定法中使用GFP记者T. Cruzi 和反双重荧光素酶哺乳动物哺乳动物在体外翻译测定法，以消除非特定的命中化合物 T. Cruzi，即那些也抑制哺乳动物蛋白质合成的T.。我们将测试克鲁齐的特定抑制剂 蛋白质合成抗寄生活性和对寄生虫与宿主细胞的选择性毒性。而我们 这里的建议是一个试点屏幕，我们拥有大规模进行所需的所有专业知识和设施 屏幕和/或对我们将确定的命中的命中优化。努力进行命中率优化 将要求我们的一个或多个我们的特定/选择性t.cruzi蛋白质合成抑制剂化合物将 对T.Cruzi显示毒性，但不显示哺乳动物细胞，可以在化学上访问 集中的图书馆和结构活动关系研究。识别命中化合物的直接目标是 未来研究的另一个领域将阐明翻译机之间的生物学差异 Human and T. Cruzi，以及Pi的功能中都很好。