Genome wide assessment of essential genes in Trypanosoma brucei

描述(由申请人提供)：布鲁氏锥虫是世卫组织的一类病原体(正在出现和未得到控制)，可引起人类非洲锥虫病(HAT，又称非洲睡眠病)，除非接受治疗，否则总是致命的。由于目前的治疗方法受到毒性、给药困难和传播耐药性的限制，迫切需要新的药物。在过去的20-30年里，通过各种假设驱动的研究计划，在布鲁氏毛滴虫的分子和细胞生物学方面积累了丰富的知识。虽然作为这些研究的副产品出现了几个有效的药物靶点，但还没有系统的全面努力来利用基因组项目的成果和布鲁氏毛滴虫遗传操作的最新进展来具体识别和评估药物靶点的潜力。此外，已投入大量资源建立药物发现计划，包括在世界各地的几个学术中心建立高通量筛选(HTS)设施。然而，尽管做出了这些努力，但既是必要的又可能是“可用药”的已验证药物靶标的名单很小，因此没有足够的高价值靶标来证明HTS或专门的药物发现努力的相当大的努力和资源是合理的。这项建议是我们总部设在美国的团队(Stuart和Phillips)与我们在英国的合作者(M.Fergusson博士和J.Mottram博士)协调努力的一部分，目的是在全基因组调查中进行RNAi(干扰RNA)分析，以寻找布氏支原体的基本和“可用药”靶标。世卫组织/TDR正在汇编一个全基因组数据库，其中列出了药物发现中靶标评估所需的关键数据，并将成为靶标选择的关键资源。虽然这项建议关注的是HAT，但布鲁氏毛滴虫的基因验证靶标可能会转化为克鲁兹毛滴虫和利什曼原虫物种，因此这些研究将为整个动植体群落提供资源。该项目产生的所有数据将提供给tdrTars.org馆长，以便在该网站上公开分发。这项为期三年的研究的目标是：1)在布鲁氏毛滴虫基因组中选择200-300个基因进行RNAi必要性分析；2)通过RNAi分析确定选定的基因是否对布鲁氏毛滴虫体内和体外的血型寄生虫的生长是必需的；3)为被证明是必需的基因开发高产的异源蛋白表达系统和合适的酶分析；以及4)组织数据和试剂以向布氏毛滴虫研究界提供分发。 公共卫生相关性：国家过敏和传染病研究所(NIAID)的目标是支持基础和应用研究，从而更好地了解传染病、免疫学和过敏性疾病，从而为这些疾病制定新的治疗或预防计划。这项建议的直接目标是确定新的药物靶点，用于开发针对NIAID投资组合中长期感兴趣的几种优先锥虫病原体的新治疗计划。

DESCRIPTION (provided by applicant): Trypanosoma brucei, a WHO category one pathogen (emerging and uncontrolled), causes human African trypanosomiasis (HAT, aka African Sleeping Sickness), which is invariably fatal unless treated. New drugs are badly needed since current therapy is limited by toxicity, difficulty of administration, and spreading drug resistance. A wealth of knowledge has accumulated over the last 20-30 years on the molecular and cellular biology of T. brucei through a variety of hypothesis driven research programs. While several validated drug targets have arisen as a spin-off of these studies, there has been no systematic comprehensive effort to exploit the output of the genome projects and the recent advances in genetic manipulation of T. brucei to specifically identify and assess potential of drug targets. In addition, significant resources have been invested in setting up drug discovery programs including the building of facilities for high throughput screening (HTS) in several academic centers around the world. However, despite these efforts the list of validated drug targets that are both essential and likely to be "druggable" is small, and thus there are not enough high value targets that justify the considerable effort and resource of a HTS, or a dedicated drug discovery effort. This proposal is part of a coordinated effort by our US-based group (Stuart and Phillips) and our collaborators in the UK (Drs. M. Fergusson and J. Mottram), to undertake RNAi (interfering RNA) analysis in a genome-wide survey for essential, and "druggable" targets in T. brucei. A genome-wide data-base that lists key data required for target assessment in drug discovery is being compiled by the WHO/TDR and will be a key resource for target selection. While this proposal focuses on HAT, genetically validated targets in T. brucei may translate to T. cruzi and Leishmania species, and thus these studies will provide a resource for the entire kinetoplastid community. All data generated from the project will be provided to the tdrtargets.org curators for public distribution on that web site. The goals of this three year study are to 1) Select 200 - 300 genes in the T. brucei genome for RNAi essentiality analysis based on an assessment of potential "druggability"; 2) to determine if the selected genes are essential for the growth of blood form T. brucei parasites in vitro and in vivo by RNAi analysis; 3) to develop high-yield heterologous protein expression systems and suitable enzyme assays for genes that are shown to be essential; and 4) to organize data and reagents to provide distribution to the T. brucei research community. PUBLIC HEALTH RELEVANCE: The goal of the National Institute of Allergy and Infectious Diseases (NIAID) is to support basic and applied research leading to a better understanding of infectious, immunologic, and allergic diseases with the goal of generating new treatments or prevention programs for these diseases. The direct goal of this proposal is to identify new drug targets for the development of novel treatment programs against several priority trypanosomal pathogens that have been of long standing interest in the NIAID portfolio.