Selective Inhibitors of the Hexose Transporter from African Trypansomes

非洲锥虫己糖转运蛋白的选择性抑制剂

• 批准号: 8607500
• 负责人: Scott M Landfear
• 金额: $ 19.25万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-02-01 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8607500
• 关键词: Affinity; African; African Trypanosomiasis; Amino Acids; Awareness; Biological Assay; Blood; Blood Circulation; Blood Glucose; Cattle; Cell membrane; Central Africa; Cessation of life; Citric Acid Cycle; Collaborations; Complement; Cytochalasins; Development; Disease; Drug Targeting; Drug effect disorder; Drug resistance; Economic Burden; Eflornithine; Environment; Enzymes; Epidemic; Exhibits; Future; Genes; Glucose; Glucose Transport Inhibition; Glucose Transporter; Glycolysis; Growth; Hexose Transporter; Hexoses; Human; Human Cell Line; In Vitro; Individual; Infection; Inhibitory Concentration 50; Laboratories; Leishmania mexicana; Lethal Dose 50; Libraries; Livestock; Measures; Messenger RNA; Metabolic; Modification; Molecular; One-Step dentin bonding system; Oxidative Phosphorylation; Parasites; Pathway interactions; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phloretin; Property; Proteins; Publications; Reagent; Relative (related person); SLC2A1 gene; Saint Jude Children' s Research Hospital; System; Testing; Therapeutic Index; Toxic effect; Toxicity Tests; Transgenes; Transgenic Organisms; Tropical Disease; Trypanosoma; Trypanosoma brucei brucei; analog; cell growth; cost; extracellular; glucose metabolism; glucose uptake; high throughput screening; improved; inhibitor/antagonist; killings; mouse model; mutant; nagana; novel; pandemic disease; permease; public health relevance; screening; uptake

DESCRIPTION (provided by applicant): Trypanosoma brucei causes the disease African trypanosomiasis that afflicts an estimated 50,000-70,000 individuals with occasional pandemics resulting in deaths of up to 800,000 individuals. The limited number of drugs for treatment of African trypanosomiasis suffer from major deficiencies, including high toxicity, high cost, and development of drug resistance. Hence, there is a widely recognized urgent need for discovery of novel anti-trypanosomal drugs. Trypanosomes are critically dependent upon uptake and metabolism of glucose in the bloodstream form (BF) that causes disease, and the hexose transporter THT1 that is expressed in BF parasites has been identified and experimentally validated as an essential permease and highly promising drug target in multiple publications. Despite awareness of the promise of targeting THT1, essentially nothing has been done regarding identifying compounds that selectively inhibit this parasite permease. The novelty of this project is that it will carry out a screen for compounds that selectively inhibit THT1 and do not inhibit mammalian hexose transporters. These compounds can ultimately be employed to develop improved anti- trypanosomal drugs. A cell growth assay has been established and fully validated for high-throughput screening (HTS) to identify selective inhibitors of THT1. This assay employs a glucose transporter null mutant of the related parasite Leishmania mexicana that has been complemented with either the gene encoding THT1 or the gene for a major human glucose transporter GLUT1. In Aim 1, ~600,000 drug-like compounds from the St. Jude Children's Research Hospital CBT library will be screened for ability to inhibit growth of the transgenic L. mexicana expressing THT1. Hits from this primary screen will then be secondarily screened to select those that do not inhibit growth of transgenic L. mexicana expressing GLUT1. This two-step screen will identify compounds that are likely selective inhibitors of THT1. In Aim 2, these hit compounds will be tested in uptake assays employing [3H]D-glucose to identify those that are high affinity selective inhibitors of glucose uptake by THT1 but not by GLUT1. Compounds will also be tested for ability to inhibit growth of T. brucei BFs in vitro and in an established mouse model of African trypanosomiasis. Toxicity tests against several human cell lines will be performed to assess the therapeutic index of hit compounds, and metabolic stability will also be determined. This project takes advantage of an established collaboration between the laboratory of the PI Dr. Scott Landfear, a molecular parasitologist who has studied parasite transporters extensively, and that of Dr. Kip Guy, who operates an internationally renowned compound screening and pharmaceutical chemistry facility at St. Jude and who has a strong track record of screening for anti-parasitic compounds.

描述（由申请人提供）：布氏锥虫引起非洲锥虫病，估计影响 50,000-70,000 人，偶尔发生大流行，导致多达 800,000 人死亡。用于治疗非洲锥虫病的药物数量有限，存在毒性大、成本高、产生耐药性等重大缺陷。因此，人们普遍认为迫切需要发现新型抗锥虫药物。锥虫严重依赖于引起疾病的血流形式 (BF) 葡萄糖的摄取和代谢，并且在 BF 寄生虫中表达的己糖转运蛋白 THT1 已在多个出版物中被鉴定和实验验证为重要的渗透酶和高度有前途的药物靶标。尽管人们意识到靶向 THT1 的前景，但在识别选择性抑制这种寄生虫渗透的化合物方面基本上没有采取任何措施。该项目的新颖之处在于，它将筛选选择性抑制THT1且不抑制哺乳动物己糖转运蛋白的化合物。这些化合物最终可用于开发改进的抗锥虫药物。细胞生长测定已建立并经过充分验证，可用于高通量筛选 (HTS)，以识别 THT1 的选择性抑制剂。该测定采用相关寄生虫墨西哥利什曼原虫的葡萄糖转运蛋白无效突变体，该突变体已与编码 THT1 的基因或主要人类葡萄糖转运蛋白 GLUT1 的基因互补。在目标 1 中，将从 St. Jude 儿童研究医院 CBT 库中筛选约 600,000 种药物样化合物，以确定其抑制表达 THT1 的转基因 L. mexicana 生长的能力。来自该初级筛选的命中然后将被二次筛选以选择那些不抑制表达GLUT1的转基因墨西哥乳杆菌生长的那些。这个两步筛选将鉴定可能是 THT1 选择性抑制剂的化合物。在目标 2 中，这些命中化合物将在使用 [3H]D-葡萄糖的摄取测定中进行测试，以鉴定那些是 THT1 而不是 GLUT1 葡萄糖摄取的高亲和力选择性抑制剂。还将测试化合物在体外和体内抑制 T. brucei BF 生长的能力。 已建立的非洲锥虫病小鼠模型。将针对几种人类细胞系进行毒性测试，以评估命中化合物的治疗指数，并确定代谢稳定性。该项目利用了 PI 博士 Scott Landfear 实验室和 Kip Guy 博士实验室之间的合作关系。Scott Landfear 博士是一名分子寄生虫学家，对寄生虫转运蛋白进行了广泛研究。Kip Guy 博士在圣裘德经营一家国际知名的化合物筛选和药物化学设施，在筛选抗寄生虫化合物方面拥有良好的记录。