Evaluating the Functional Impact of Genetic Diversity on Malaria Vaccine Candidates

摘要 由恶性疟原虫引起的疟疾仍然是全球导致这两种疾病死亡的主要原因之一 儿童和孕妇。最近全球疟疾死亡人数的减少停滞不前 开发一种高效疫苗至关重要。开发有效疫苗的主要挑战 是恶性疟原虫抗原的广泛多样性。虽然遗传多样性在生物多样性中发挥着重要作用 免疫逃避，是发展自然和疫苗诱导的保护性免疫的障碍， 在评估疟疾疫苗候选时，它没有得到足够的重视。这项提案将使用基因组 下一代疟疾疫苗候选疫苗的认证方法。反向疫苗接种是一种 识别疫苗的潜在抗原，该疫苗以生物体的基因组序列开始，并使用 这些信息可以识别可能成为合适候选疫苗的表位和抗原。自.以来 恶性疟原虫基因组序列公布，仅有四种新的潜在候选疫苗 已经进入临床开发，包括PfRh5。拟议研究的主要目标是使用一种 利用直接来自感染患者的寄生虫基因组数据的反向疫苗学方法来识别和 从功能上询问这些抗原多样性的重要性。对于这些当前和新奇的候选人来说， 包括PfRh5和结合伙伴，我们将通过以下方式测试遗传多样性对免疫中和的作用 使用高效的CRISPR-Cas9基因组编辑创建转基因寄生虫。这些寄生虫系将会是 用于在第二阶段临床试验之前评估特定变种在免疫逃避中的作用。我们将使用免疫球蛋白 来自疟疾免疫个体，在长期纵向队列中密切跟踪，以及来自 通过生长抑制试验评估疟疾寄生虫复制抑制程度的疫苗试验， 中性粒细胞呼吸爆发，裂殖子吞噬细胞。这种方法需要凝聚力 基因组测序技术，以确定潜在的候选抗原和自然产生的多样性， 跟踪感染和免疫进化的特征良好的人类纵向队列，标准化 作为免疫的体外相关性的检测，疫苗设计的基于结构的方法，以及强大的 与流行国家的科学家和机构都有联系。我们的研究团队具有独特的优势 结合这些关键要求，调查寄生虫多样性对发展的影响 加强保护性免疫和疫苗效力，这是加速发现疟疾疫苗的一个重要因素。这 方法填补了疟疾疫苗开发领域的一个关键需求，因为它将遗传多样性带到了 候选抗原走在疫苗候选验证和认证的前沿。这项研究适用于 发现新的候选疫苗组合的特殊前景，这些组合将提供广泛的中和 包括在全球有效疫苗中的抗体，这种疫苗可以绕过寄生虫的自然策略 逃避免疫系统。

ABSTRACT Malaria caused by Plasmodium falciparum remains one of the leading causes of death globally of both children and pregnant women. The recent global stall in the reduction of malaria deaths has made the development of a highly effective vaccine essential. A major challenge to developing an efficacious vaccine is the extensive diversity of Plasmodium falciparum antigens. While genetic diversity plays a major role in immune evasion and is a barrier to the development of both natural and vaccine-induced protective immunity, it has been underprioritized in the evaluation of malaria vaccine candidates. This proposal will use genomic approaches to credential next generation malaria vaccine candidates. Reverse vaccinology is a method of identifying potential antigens for a vaccine that starts with the genomic sequence of an organism and uses that information to identify epitopes and antigens that might make suitable vaccine candidates. Since the genome sequence of Plasmodium falciparum was published, only four new potential candidate vaccines have entered clinical development, including PfRh5. The main objective of the proposed study is to use a reverse-vaccinology approach utilizing parasite genomic data directly from infected patients to identify and functionally interrogate the importance of diversity in these antigens. For these current and novel candidates, including PfRh5 and binding partners, we will test the role of genetic diversity on immune neutralization by creating transgenic parasites by using efficient CRISPR-Cas9 genome editing. These parasite lines will be used to assess the role of specific variants in immune evasion prior to Phase 2 clinical trials. We will use IgG from malaria-immune individuals, followed closely in long-term longitudinal cohorts, and IgG from subjects in vaccine trials to assess the degree of inhibition of replication of malaria parasites by growth inhibition assays, neutrophil respiratory burst, and opsonophagocytosis of merozoites. This approach requires the cohesion of genomic sequencing technologies to identify potential candidate antigens and naturally occurring diversity, well-characterized human longitudinal cohorts to follow evolution of infection and immunity, standardized assays to serve as in vitro correlates of immunity, structure-based approaches for vaccine design, and strong ties to both scientists and institutions in endemic countries. Our research team is uniquely positioned to combine these critical requirements to investigate the implications of parasite diversity on the development of protective immunity and vaccine efficacy, an essential factor to accelerate malaria vaccine discovery. This approach fills a critical need in the malaria vaccine development field in that it brings genetic diversity in candidate antigens to the forefront of vaccine candidate validation and credentialing. This study holds exceptional promise to discover new vaccine candidate combinations that will provide broadly neutralizing antibodies for inclusion in a globally effective vaccine, one that circumvents the parasite’s natural strategy to evade the immune system.