Scanning the Cryptosporidium proteome for vaccine antigens

隐孢子虫病是低收入和中等收入国家（LMIC）婴儿腹泻的十大原因， 营养不良和神经认知发育受损，导致每年大量的发病率、死亡率和损失。 残疾调整生命年隐孢子虫病的疫苗是不可用的，但将是有吸引力的， 在感染负担高的中低收入国家预防儿童隐孢子虫病。研究由博士。 William a.弗吉尼亚大学的佩特里和卡罗尔·吉尔克里斯特博士发现了IgA与保护作用的相关性。 针对隐孢子虫感染，其可用于疫苗的开发。但 保护性IgA抗体靶向的隐孢子虫蛋白仍有待发现。Antigen Discovery，Inc (ADI)已经开发了一种中试规模的隐孢子虫蛋白质微阵列， 扩增并用于筛选针对隐孢子虫蛋白质组的抗体应答。蛋白质组规模 隐孢子虫病研究以前从未有过抗体免疫分析平台 这项技术有能力迅速推进我们对保护性免疫的理解， 针对隐孢子虫蛋白的反应。一个泛蛋白质组隐孢子虫微阵列将被开发， 在两个母婴出生队列中测量特异性抗隐孢子虫IgA水平，并随访数年， 腹泻临床终点的详细表征，包括许多疾病归因检测 引起腹泻的病原体，包括隐孢子虫属。本研究的目的是鉴定孕妇血清中IgA的含量， 婴儿摄入的母乳与随后的隐孢子虫感染风险降低相关， 腹泻阵列结果和最有希望的疫苗候选物将通过生产蛋白质来验证。 包括翻译后修饰和正确三级结构的真核表达系统， 将用于开发ELISA和Western blot检测方法，用于检测母乳样本。的成功 这项研究将为开发预防婴儿隐孢子虫病的母体疫苗提供机会 在生命最初几个月的脆弱时期。我们假设在乳腺癌中发现的保护性抗体 当由婴儿粘膜免疫应答产生时，牛奶也可以是保护性的，因此呈现出 为儿童疫苗的持续保护提供了发展的机会。保护性抗体也可以是 作为治疗性抗体开发，可用于治疗免疫功能低下患者的持续感染， 患者，如艾滋病患者。我们期望鉴定240 - 400种免疫反应性隐孢子虫蛋白， 其中至少有3种疾病的母亲母乳中IgA水平升高， 率或保护免受腹泻，以及粪便IgA从婴儿的适应性免疫。这份资助申请 解决了儿童隐孢子虫病的重大问题，为疫苗奠定了基础， 与保护相关的特异性粘膜IgA反应的研究。

Cryptosporidiosis is a top ten cause of infant diarrhea in low and middle-income countries (LMICs), as well as malnutrition and impaired neurocognitive development, leading to substantial yearly morbidity, mortality and lost disability-adjusted life years. A vaccine for cryptosporidiosis is not available, but would be appealing for prevention of cryptosporidiosis in children in LMICs where the burden of infection is high. Studies done by Dr. William A. Petri and Dr. Carol Gilchrist at University of Virginia have uncovered IgA correlates of protection against Cryptosporidium infection, which may be expoited for development of a vaccine. However, the cryptosporidial proteins targeted by protective IgA antibodies remain to be discovered. Antigen Discovery, Inc (ADI) of Irvine, California has developed a pilot scale Cryptosporidium protein microarray, which can be expanded and used to screen antibody responses against the cryptosporidial proteome. A proteome-scale platform for antibody immune-profiling has never before been available to the cryptosporidiosis research community, and this technology has the power to rapidly advance our understanding of the protective immune response directed to cryptosporidial proteins. A pan-proteome Cryptosporidium microarray will be developed to measure specific anti-cryptosporidial IgA levels in two mother-infant birth cohorts with years of follow-up and detailed characterization of clinical endpoints for diarrhea, including disease attributable detection of many diarrhea-causing pathogens, including Cryptosporidium spp. The aim of the study is to identify the IgA in maternal breast milk ingested by infants that correlates with reduced risk of subsequent Cryptosporidium infection and diarrhea. The array results and most promising vaccine candidates will be validated by producing the proteins in a eukaryotic expression system to include post-translational modifications and correct tertiary structure, which will be used to develop ELISA and Western blot assays for testing breast milk samples. The success of this study will present an opportunity for development of maternal vaccines to prevent cryptosporidiosis in infants during the vulnerable period of the first months of life. We postulate that protective antibodies identified in breast milk may also be protective when produced by the infant mucosal immune response, thus presenting the opportunity for development of a pediatric vaccine for continued protection. Protective antibodies may also be developed as therapeutic antibodies that can be given to treat persistent infection in immunocompromised patients, such as those with HIV/AIDS. We expect to identify 240-400 immunoreactive Cryptosporidium proteins, at least 3 of which will have increased IgA levels in the breast milk of mothers with infants that had lower incidence rates or protection from diarrhea, as well as fecal IgA from the infants’ adaptive immunity. This grant application addresses the significant problem of cryptosporidiosis in children by laying the foundation for a vaccine through the study of specific mucosal IgA responses associated with protection.