Harnessing Transcriptomics to Identify and Test Novel Hookworm Vaccine Targets

利用转录组学来识别和测试新型钩虫疫苗靶标

基本信息

批准号：
9063524
负责人：
RAFFI V AROIAN
金额：
$ 24.36万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-05-05 至 2017-10-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9063524
关键词：
Adult Agriculture Albendazole Ancylostoma (genus)Anthelmintics Binding Binding Proteins Bioinformatics Biology Blood Candidate Disease Gene Cathepsins B Complementary DNA Data Digestion Disease Experimental Models Family Gene Targeting Genes Genomics Goat Haemonchus Hamsters Health Homologous Gene Hookworms Human Humoral Immunities Immune system Immunity Immunocompetent Immunocompromised Host Individual Infection Injection of therapeutic agent Integral Membrane Protein Intestinal parasite Intestines Larva Legal patent Life Livestock Mammals Membrane Mesocricetus auratus Methods Multi-Drug Resistance Necator americanus Nematoda Old World Hookworm Organism Parasite Control Parasites Peptide Hydrolases Pharmaceutical Preparations Pharmacotherapy Pichia Population Protease Inhibitor Proteins RNA Reporting Reverse Transcriptase Polymerase Chain Reaction Sheep Staging T cell response T-Lymphocyte Testing Tissues Translating Vaccinated Vaccination Vaccines Yeasts comparative genomics cost feeding gene product genome sequencing genome-wide killings member next generation novel novel vaccines parasite genome preference protein expression response success transcriptome transcriptome sequencing transcriptomics vaccine candidate vaccine development vaccine trial vaccinology

项目摘要

DESCRIPTION (provided by applicant): We will develop vaccines against hookworms using RNA-seq and comparative genomics to identify targets essential for infection. Hookworms such as Ancylostoma duodenale and Necator americanus infect over 400 million human beings, stunting and impoverishing them. Existing drugs are only partially effective, and no reliably effective vaccines against hookworms exist. The zoonotic hookworm Ancylostoma ceylanicum easily infects both humans and other mammals (such as golden hamsters). It is therefore a useful experimental model in which to test possible treatments for hookworm disease. Our first hypothesis is that by using transcriptional analysis of genes upregulated during infection by both A. ceylanicum and the related parasite Haemonchus contortus, we have identified a novel set of secreted proteases and protease inhibitors that are essential for parasite survival and that are promising vaccine targets. Our second hypothesis is that we can use RNA-seq and tissue-specific expression to identify a deeper set of vaccine targets, encoded by genes whose products are required for hookworms to negate the host immune system and that are accessible via vaccination. We have recently generated and analyzed an A. ceylanicum genomic sequence of 313 Mb containing ~27,000 genes, along with RNA-seq data during an A. ceylanicum infection from third-stage larvae to fully mature adults. This has already provided us with our top priority protease and protease inhibitor vaccine candidates. Our first aim will be to prioritize these candidates by tissue-specific expression, synthesize them in the yeast Pichia pastoris, and then test them in mixtures as vaccines against A. ceylanicum. Although previous efforts at vaccination with crude mixtures of nematode proteins from killed or weakened parasites have given promising results, vaccinations with single proteins from individual parasite genes have generally failed. We suspect that multiple gene groups expressed during infection by a single parasite genome can be collectively expressed and coinjected to elicit effective immunity. T cell responses to successful vaccine formulations will be characterized. Our second aim will be to identify immunoresponsive intestinal hookworm proteins as further vaccine candidates. We propose to perform additional RNA-seq on dissected intestines and non-intestinal tissues, from A. ceylanicum infecting golden hamsters with either normal or suppressed immune systems. We will use bioinformatics to look for genes encoding secreted or membrane-bound proteins, which, during infection, are either interacting with the host's immune system and/or robustly translated in the parasite's intestines. These are good candidates for proteins used by the parasite for neutralization of the immune system. We will further prioritize candidate secreted and membrane-bound gene products by their conservation in the human hookworm Necator americanus and in H. contortus. Vaccination will begin with the top one or two candidates, to be expanded upon in a subsequent R01.

描述（由申请人提供）：我们将使用 RNA 测序和比较基因组学来开发针对钩虫的疫苗，以确定十二指肠钩虫和美洲钩虫等感染所需的目标，现有药物使他们发育迟缓和贫困。仅部分有效，并且没有可靠有效的针对钩虫的疫苗。因此，它是一个有用的实验模型，可用于测试钩虫病的可能治疗方法，即通过对锡兰线虫和金仓鼠感染期间上调的基因进行转录分析。相关的寄生虫捻转血矛线虫，我们已经确定了一组新的分泌蛋白酶和蛋白酶抑制剂，它们对于寄生虫的生存至关重要，并且是有希望的疫苗靶标。我们的第二个假设是我们可以使用 RNA-seq。 313 Mb 包含约 27,000 个基因，以及从第三阶段幼虫到完全成熟成虫的锡兰弧菌感染过程中的 RNA 测序数据，这已经为我们提供了我们的最佳结果。我们的首要目标是通过组织特异性表达对这些候选疫苗进行优先排序，在毕赤酵母中合成它们，然后将它们作为针对锡兰弧菌的疫苗进行测试。来自被杀死或弱化的寄生虫的线虫蛋白的粗制混合物已经给出了有希望的结果，用来自单个寄生虫基因的单一蛋白质进行疫苗接种通常会失败，我们怀疑在单个寄生虫基因组感染期间表达的多个基因组可以共同表达和。我们的第二个目标是确定免疫反应性肠钩虫蛋白作为进一步的候选疫苗，以诱导 T 细胞对解剖的肠道和非肠道组织的有效免疫反应。我们将使用生物信息学来寻找编码分泌蛋白或膜结合蛋白的基因，这些蛋白在感染过程中会被感染。与宿主的免疫系统相互作用和/或在寄生虫的肠道中强有力地翻译，这些是寄生虫用于中和免疫系统的蛋白质的良好候选者，我们将通过它们的保守性进一步优先考虑候选分泌和膜结合的基因产物。人类钩虫美洲钩虫和捻转钩虫的疫苗接种将从前一两个候选者开始，并在随后的 R01 中进行扩展。