Defining conserved epitopes on polymorphic malaria antigens

项目摘要 据估计，疟疾每年仍造成429 000人死亡，2.12亿人感染，其中绝大多数是疟疾患者。 在年轻的孩子中。生活在流行地区的个体逐渐产生免疫反应， 对抗有症状的疾病自然获得的免疫针对寄生虫生命周期的血液阶段， 很大程度上取决于抗体的反应性。在年轻人中通过接种疫苗引发类似的免疫反应， 没有免疫力的儿童将大大降低疟疾发病率和死亡率。各种寄生虫抗原具有 与预防临床疾病有关。这些抗原大多数在裂殖子中表达 阶段，并参与红细胞的侵入。然而，基于这些血液阶段的疫苗试验 抗原迄今尚未成功。导致血液阶段疫苗失败的一个因素是 不同寄生虫菌株之间广泛的遗传多样性，导致菌株特异性，但不是菌株， 超越抗体反应。对自然获得免疫力的研究表明，受保护的个体 具有针对保守表位的抗体。这些保守表位的鉴定可以作为 疟疾血液阶段疫苗的结构设计蓝图。我们假设， 疟疾依赖于对裂殖子抗原具有跨株反应性的高亲和力IgG， 这些抗体可用于定义裂殖子疫苗候选物上的保守表位。测试 根据这一假设，我们将比较针对 生活在托罗罗的免疫成人、半免疫儿童和非免疫儿童中的裂殖子抗原， 乌干达是疟疾传播强度极高的地区， 用于表位作图的反应性抗体。在特异性目标1中，我们将分离裂殖子抗原特异性记忆B 细胞，测试这些抗体在体外抑制各种疟原虫株的能力， 并绘制跨菌株反应性抗体的表位以确定裂殖子抗原上的保守表位。在 具体目标2，我们将对记忆B细胞受体库进行深度测序，以揭示终身影响 疟疾暴露对体液免疫应答的影响，分析抗裂殖子血清抗体谱 并确定优势血清抗体克隆型的跨株抑制活性和表位。 总的来说，这些实验的结果将提供有关抗体类型的详细信息。 血液阶段的疫苗应该理想地引发，并将使新的免疫原的设计，以引发应变- 超越了疟疾血液阶段的抗体。此外，它将提供对重叠的深入了解， 记忆B细胞和血清学隔室之间的差异，以及驱动记忆的选择性机制。 B细胞分化为浆细胞和记忆B细胞。

PROJECT SUMMARY Malaria is still responsible for an estimated 429,000 deaths and 212 million infections per year, the large majority among young children. Individuals living in endemic regions gradually develop an immune response that protects against symptomatic disease. Naturally acquired immunity targets the blood stage of the parasite’s life cycle and is largely dependent on antibody reactivity. Elicitation of a similar immune response by vaccination in young, non-immune children would dramatically reduce malaria morbidity and mortality. Various parasite antigens have been associated with protection against clinical disease. Most of these antigens are expressed in the merozoite stage and are involved in invasion of the red blood cell. However, vaccine trials based on these blood-stage antigens have thus far been unsuccessful. One factor contributing to the failure of blood stage vaccines is the extensive genetic diversity between different parasite strains that has resulted in strain-specific, but not strain- transcending, antibody responses. Studies of naturally acquired immunity suggest that protected individuals harbor antibodies against conserved epitopes. The identification of such conserved epitopes could serve as a blueprint for structure-based design of a malaria blood stage vaccine. We hypothesize that immunity against malaria is dependent on high affinity IgGs with cross-strain reactivity against merozoite antigens and that these antibodies can be used to define conserved epitopes on merozoite vaccine candidates. To test this hypothesis, we will compare the molecular and functional characteristics of antibodies directed against merozoite antigens among immune adults, semi-immune children and non-immune children living in Tororo, Uganda, a region with extremely high malaria transmission intensity, and use the most potent and broadly reactive antibodies for epitope mapping. In Specific Aim 1, we will isolate merozoite antigen-specific memory B cells from these individuals, test the ability of these antibodies to inhibit various malaria parasite strains in vitro, and map the epitopes of cross-strain reactive antibodies to define conserved epitopes on merozoite antigens. In Specific Aim 2, we will deep-sequence the memory B cell receptor repertoire to uncover the impact of life-long malaria exposure on the humoral immune response, analyze the repertoire of anti-merozoite serum antibodies and determine the cross-strain inhibitory activity and epitopes of dominant serum antibody clonotypes. Collectively, the results from these experiments will provide detailed information about the types of antibodies that a blood-stage vaccine should ideally elicit and will enable the design of novel immunogens to elicit strain- transcending antibodies against the malaria blood stage. In addition, it will provide insight into the overlap and differences between the memory B cell and serological compartments, and the selective mechanisms that drive B cell differentiation into plasma cells and memory B cells.