Discovery and preclinical evaluation of Plasmodium falciparum and P. vivax coiled coil antigens for malaria vaccine development

用于疟疾疫苗开发的恶性疟原虫和间日疟原虫卷曲螺旋抗原的发现和临床前评估

• 批准号: 10079466
• 负责人: Socrates Herrera Valencia
• 金额: $ 38.26万
• 依托单位: CAUCASECO SCIENTIFIC RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-03 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10079466
• 关键词: Adjuvant; Animal Model; Animals; Antibodies; Antibody Response; Antibody titer measurement; Antibody-mediated protection; Antigens; Aotus primate; Area; Bioinformatics; Biological Assay; Blood; Cessation of life; Chemicals; Circular Dichroism; Circular Dichroism Spectroscopy; Clinical; Coiled-Coil Domain; Control Animal; Country; Data; Development; Enzyme-Linked Immunosorbent Assay; Epitopes; Erythrocytes; Evaluation; Genome; Goals; Growth; High Pressure Liquid Chromatography; Human; Immune response; Immune system; Immunity; Immunization; Immunize; Immunoglobulin G; Immunologics; In Vitro; Incidence; Individual; Infection; Malaria; Malaria Vaccines; Mass Spectrum Analysis; Methods; Monkeys; Mus; Parasitemia; Parasites; Peptide Synthesis; Peptide antibodies; Peptides; Phagocytosis; Plasmodium; Plasmodium falciparum; Plasmodium vivax; Preparation; Primates; Process; Protein Fragment; Proteins; Proteome; Public Health; Recombinants; Respiratory Burst; Risk; Rodent; Solid; Specificity; Sterility; Structure; Synthetic Antigens; Testing; Vaccines; aqueous; base; cost effective; cross reactivity; design; disorder risk; efficacy study; experience; experimental study; immunogenic; immunogenicity; improved; in silico; in vitro Assay; in vivo; inhibiting antibody; innovation; manufacturing facility; nonhuman primate; novel; novel vaccines; peptide vaccination; preclinical evaluation; protective efficacy; protein structure; screening; success; tool; vaccine candidate; vaccine development

PROJECT SUMMARY/ABSTRACT Despite the great global public health importance, malaria vaccine development has been slow with only a handful of candidates under development and the most advanced displaying modest efficacy. The current use of bioinformatics for genome/proteome screening, animal models, and more efficient manufacturing facilities for recombinant/synthetic antigens could significantly accelerate the development of novel vaccines. We propose to use these advances to identify and develop vaccine candidates targeting both, Plasmodium falciparum and P. vivax parasites, responsible for >99% of the malaria cases worldwide. Alpha-helical coiled-coil motifs, protein structures widely distributed in the Plasmodium proteomes known to spontaneously fold in aqueous solution, are able to induce antibodies that inhibit parasite growth in vitro. We previously in silico identified 220 proteins in both parasite proteomes, 170 of which were synthesized and tested. High antigenicity with human sera from endemic areas, and immunogenicity in mice led to selection of 145 fragments. Several of them displayed significant P. falciparum/P. vivax cross reactivity, and importantly in vitro parasite inhibition activity, which correlated with malaria clinical immunity. Furthermore, mice immunization with a P. falciparum poly-epitope construct (Pf-181) maintained individual epitopes specificity and parasite inhibitory activity. Our general hypothesis is that “Plasmodium poly-epitope constructs containing cross-reacting α-helical coiled-coil motifs induce protection in monkey against homologous and heterologous parasite challenges. The overall goal is to determine the vaccine potential of P. falciparum and P. vivax erythrocytic synthetic poly-epitope constructs containing α-helical coiled-coil motifs. This goal will be approached through the following specific aims: 1) Design, synthesis and characterization of P. falciparum and P. vivax poly-epitope constructs containing α-helical coiled-coil motifs; 2) Evaluation of the immunogenicity and protective efficacy of selected P. falciparum and P. vivax poly-epitope constructs against homologous and heterologous parasite challenges in Aotus monkeys; 3) In vitro characterization of α-helical coiled-coil fragments/constructs and immune responses elicited by animal immunization. Methods proposed are: a) F-moc peptide synthesis of poly-epitope constructs, followed by HPLC purification, mass spectrometry (MS) and circular dichroism (CD) analyses; b) Monkey immunization with selected poly-epitope constructs and evaluation of protective efficacy to homologous and heterologous challenges using P. falciparum (Santa Lucia) and P. vivax (Sal-I) parasite strains; c) ELISA and IFAT test of sera from immunized animals; d) antibody functional in vitro assays (GIA, ADRB,OPA).The innovation of this study is the highly efficient and cost-effective approach, using a comprehensive rational and rigorous epitope down- selection of hundreds of protein fragments from in silico epitope identification in Plasmodium proteomes for poly- epitope design and evaluation of their protective efficacy studies in monkeys. It will accelerate the development of malaria vaccines for human use.

项目概要/摘要 尽管对全球公共卫生具有重要意义，但疟疾疫苗的开发进展缓慢，仅有 少数候选药物正在开发中，最先进的候选药物显示出有限的功效。目前使用情况 用于基因组/蛋白质组筛选、动物模型和更高效的生产设施的生物信息学 重组/合成抗原可以显着加速新型疫苗的开发。我们建议 利用这些进展来识别和开发针对恶性疟原虫和疟原虫的候选疫苗。 间日疟原虫，造成全球 99% 以上的疟疾病例。 α-螺旋卷曲螺旋基序，蛋白质 广泛分布于已知在水溶液中自发折叠的疟原虫蛋白质组中的结构， 能够在体外诱导抑制寄生虫生长的抗体。我们之前在计算机上鉴定了 220 种蛋白质 在这两种寄生虫蛋白质组中，合成并测试了其中 170 个。与人血清具有高抗原性 流行地区和小鼠的免疫原性导致选择了 145 个片段。其中有几张展示了 显着的恶性疟原虫/P.间日疟原虫交叉反应性，以及重要的体外寄生虫抑制活性， 与疟疾临床免疫力相关。此外，用恶性疟原虫多表位对小鼠进行免疫接种 构建体（Pf-181）保持了个体表位特异性和寄生虫抑制活性。我们的将军 假设是“含有交叉反应 α 螺旋卷曲螺旋基序的疟原虫多表位构建体 诱导猴子免受同源和异源寄生虫挑战的保护。总体目标是 确定恶性疟原虫和间日疟原虫红细胞合成多表位构建体的疫苗潜力 含有α-螺旋卷曲螺旋图案。该目标将通过以下具体目标来实现：1) 含有 α 螺旋的恶性疟原虫和间日疟原虫多表位构建体的设计、合成和表征 盘绕线圈图案； 2) 所选恶性疟原虫和恶性疟原虫的免疫原性和保护功效评价 间日疟原虫多表位构建体对抗Aotus猴中的同源和异源寄生虫挑战； 3） α-螺旋卷曲螺旋片段/结构的体外表征和动物引起的免疫反应 免疫接种。提出的方法是：a) 多表位构建体的 F-moc 肽合成，然后进行 HPLC 纯化、质谱 (MS) 和圆二色性 (CD) 分析； b) 猴子免疫接种 选择的多表位构建体以及对同源和异源的保护功效的评估 使用恶性疟原虫 (Santa Lucia) 和间日疟原虫 (Sal-I) 寄生虫菌株的挑战； c) 血清的ELISA和IFAT检测 来自免疫动物； d) 抗体功能体外测定（GIA、ADRB、OPA）。本研究的创新点 是高效且具有成本效益的方法，使用全面合理且严格的表位向下- 从疟原虫蛋白质组中的计算机表位鉴定中选择数百个蛋白质片段，用于多聚 表位设计及其在猴子中的保护功效研究的评估。将会加速发展 供人类使用的疟疾疫苗。