Attenuated malaria sporozoite vaccine using a P. falciparum blood-stage auxotroph

使用恶性疟原虫血期营养缺陷型的减毒疟疾子孢子疫苗

• 批准号: 8607502
• 负责人: STEPHEN Lev HOFFMAN
• 金额: $ 30万
• 依托单位: SANARIA, INC.
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-02-01 至 2015-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8607502
• 关键词: 1-deoxy-2-pentulose; Adverse effects; Anabolism; Anopheles Genus; Antigens; Antimalarials; Area; Attenuated; Azithromycin; Biodistribution; Biological Assay; Biological Models; Bite; Blood; Chemoprophylaxis; Chloroquine; Clinical Trials; Culicidae; Cyclic GMP; Development; Diet; Disease; Dose; England; Enzymes; Epitopes; Erythrocytes; Excision; Exposure to; Falciparum Malaria; Female; Gene Targeting; Generations; Genes; Genome; Germany; Growth; Growth and Development function; Hepatocyte; Human; Immune system; Immunity; Immunization; In Vitro; Infection; Knock-out; Liver; Maintenance; Malaria; Malaria Vaccines; Mali; Maryland; Mediating; Methods; Midgut; Modeling; Monitor; Mus; Netherlands; Oocysts; Parasites; Pathway interactions; Patients; Pharmaceutical Preparations; Phase; Phase I Clinical Trials; Plasmodium falciparum; Process; Production; Proteins; Quality Control; Risk; Rodent; Rodent Model; Safety; Salivary Glands; Sporozoite vaccine; Sporozoites; Staging; Supplementation; System; Tanzania; Technology; Testing; Time; Tissue Model; Tissues; Toxicology; Vaccines; Validation; Zinc Fingers; asexual; base; controlled release; cost; disease transmission; drug metabolism; feeding; fosmidomycin; immunogenicity; improved; inorganic phosphate; isopentenyl pyrophosphate; isoprenoid; killings; mutant; nuclease; pre-clinical; prevent; protective efficacy; public health relevance; tool; transmission process; vaccine candidate; vaccine safety; volunteer; zinc finger nuclease

DESCRIPTION (provided by applicant): A highly effective vaccine that targets the asymptomatic sporozoite (SPZ) and liver stages of the most lethal human malaria parasite, Plasmodium falciparum (Pf), would be an ideal tool to prevent malaria infection, disease and transmission. After 3 decades of effort, the most effective subunit malaria vaccine only provides 50% protection against infection at 2 wks and 22% at 5 months after the last dose. PfSPZ are the only immunogens that induce sustained (at least 10-28 months), high level (>90%) protection against Pf. For decades the focus has been on irradiated (irr) PfSPZ. Recently, however, genetically attenuated SPZ disrupted in genes required for early/mid liver stage development have shown protection in mice. Furthermore, volunteers immunized by the bite of mosquitoes carrying fully infectious PfSPZ, and administered chloroquine to eliminate blood stage parasites, acquire protective immunity against liver stage parasites after exposure to 20 times fewer PfSPZ-infected mosquitoes than are required with mosquitoes carrying irrPfSPZ. This increased efficiency is likely due to increased abundance and diversity of parasite epitopes presented to the immune system during the mid/late liver stages. Clinical trials are underway or planned for PfSPZ-based vaccines, including PfSPZ administered with chloroquine. However, it would be ideal if the need for an antimalarial drug were eliminated. Furthermore, in a rodent malaria model the drug azithromcyin, which kills parasites at the late liver stage by disrupting apicoplast function, is more efficient than chloroquine in inducing protective immunity. We propose to eliminate the need for an antimalarial drug and improve the efficiency of induction of protective immunity by using a genetically attenuated strain of Pf lacking genes that are essential for development of only late liver and asexual blood stages. Using a newly developed, highly efficient method for genome editing based on customized zinc-finger nucleases, we will delete the Pf dxr and lspD genes that encode apicoplast enzymes deoxyxylulose 5-phosphate reductoisomerase (DXR) and methylerythritol phosphate cytidyltransferase (IspD), respectively. DXR catalyzes the 1st step in isoprenoid biosynthesis, followed by IspD, resulting in production of the essential metabolite isopentenyl diphosphate (IPP). DXR is essential for Pf blood stage growth. Inhibition of apicoplast development and isoprenoid biosynthesis can be reversed by supplementation with IPP allowing for generation of double knockout parasites. Knockout clones will be selected that generate acceptable numbers of gametocytes and PfSPZ when compared to wild type parasites, and Sanaria will produce purified, cryopreserved Pf?dxr+?lspD SPZ. Studies in hepatocytes and a new liver tissue model will enable our testing of the hypothesis that Pf?dxr+?lspD parasites have a profound, IPP-dependent, developmental arrest late during liver stage development when the apicoplast is most active. These studies will establish a vaccine candidate with optimal immunogenicity and a critical safety feature of being unable to sustain replication in erythrocytes if any parasites break through from the liver.

描述（由申请人提供）：一种针对无症状的孢子岩（SPZ）和肝脏阶段的高效疫苗，最致命的人类疟原虫寄生虫，恶性疟原虫（PF），将是预防疟疾感染，疾病和传播的理想工具。经过30年的努力，最有效的亚基疟疾疫苗仅在最后剂量后5个月时提供50％的感染保护感染，22％。 PFSPZ是唯一诱导持续（至少10-28个月），高水平（> 90％）保护PF的免疫原子。几十年来，重点一直在辐照（IRR）PFSPZ上。然而，最近，在早期/中期肝脏阶段发育所需的基因中破坏了遗传减弱的SPZ已显示出对小鼠的保护。此外，志愿者通过携带完全具有感染性PFSPZ的蚊子的叮咬来免疫，并管理氯喹，以消除血液阶段的寄生虫，在暴露于20倍的PFSPSZ感染蚊子后，对肝脏寄生虫的保护性免疫，比蚊子携带的蚊子更少。这种提高的效率可能是由于肝脏中/晚期阶段中呈现给免疫系统的寄生虫表位的丰度和多样性所致。临床试验正在进行或计划用于基于PFSPZ的疫苗，包括用氯喹施用的PFSPZ。但是，如果需要消除对抗疟药的需求，那将是理想的选择。此外，在啮齿动物疟疾模型中，药物azithromcyin在肝脏阶段杀死寄生虫，通过破坏蛋白质成体功能，在诱导保护性免疫方面更有效。我们建议消除对抗疟疾药物的需求，并通过使用缺乏遗传性的PF缺乏基因的遗传菌株来提高保护性免疫的效率，而PF缺乏基因，这对于仅肝脏和无性血液阶段的发展至关重要。使用基于定制的锌指成核的基因组编辑的新开发，高效的方法，我们将删除编码Apicoplast酶Deoxyxylose 5-磷酸还原酶的PF DXR和LSPD基因，并删除PF DXR和LSPD基因，并删除PF DXR和LSPD基因，并删除pficoplast酶，并删除ph磷酸盐还原酶（DXR）和甲基磷酸二磷酸二磷酸二氨基酯酶（Isplantyltrantertrantspasse）。 DXR催化了类异丙生素生物合成的第一步，其次是ISPD，导致产生必需的代谢物异戊烯基二磷酸（IPP）。 DXR对于PF血液阶段的生长至关重要。通过补充IPP允许产生双基因敲除寄生虫的抑制作用抑制蛋白酶发育和类异型生物合成。与野生型寄生虫相比，将选择敲除克隆，以产生可接受的配子细胞和PFSPZ，而Sanaria将产生纯净的，冷冻保存的PF？DXR+？LSPD SPZ。在肝细胞和新的肝组织模型中进行的研究将使我们能够测试Pf？dxr+？lspd寄生虫在肝脏阶段发育后期具有深刻的，IPP依赖性的发育停滞。这些研究将建立具有最佳免疫原性的疫苗候选者，并且如果有任何寄生虫从肝脏中断出来，则无法维持红细胞复制的关键安全特征。