Development of a live attenuated vaccine for visceral leishmaniasis

内脏利什曼病减毒活疫苗的开发

• 批准号: 9725441
• 负责人: Abhay R Satoskar
• 金额: $ 7.8万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-01 至 2022-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9725441
• 关键词: Adverse event; Animal Model; Animals; Antibiotics; Antigens; Attenuated; Attenuated Live Virus Vaccine; Bacteria; Biochemical; Biological Markers; Biological Products; CRISPR/Cas technology; Characteristics; Clinical; Clinical Trials; Cutaneous; Cutaneous Leishmaniasis; Cyclic GMP; Development; Diffuse; Disease; Dose; Environment; Etiology; FDA approved; Formulation; Foundations; Genes; Genetic; Genomics; Genotype; Goals; Hamsters; Human; Immune response; Immunity; Immunization; Immunocompromised Host; Immunologic Markers; Immunologics; India; Induced Hyperthermia; Infection; Interferon Type II; Interleukin-10; Intervention; Laboratories; Leishmania; Leishmania donovani; Leishmania major; Leishmaniasis; Lesion; Life; Manufactured Materials; MicroRNAs; Molecular; Monitor; Mucous Membrane; Mus; Mutation; Parasites; Patients; Peripheral Blood Mononuclear Cell; Phenotype; Pilot Projects; Preventive vaccine; Process; Production; Safety; Sand Flies; Site; Testing; Time; Trimethoprim-Sulfamethoxazole; Vaccination; Vaccines; Virulent; Virus; Visceral; Visceral Leishmaniasis; Zoonoses; experience; genome sequencing; global health; immunogenic; immunogenicity; industry partner; interest; manufacturing process; neglected tropical diseases; obligate intracellular parasite; parasite genome; pathogen; pre-clinical; preclinical study; response; skin lesion; vaccine candidate; vector; whole genome

Abstract Infections caused by the protozoan parasite Leishmania include cutaneous (CL), mucosal (ML), and visceral leishmaniasis (VL). The WHO classifies leishmaniasis as a neglected tropical disease with over 12 million current infections globally, and approximately 2 million new cases annually. Patients who recover from leishmaniasis develop protective immunity against reinfection, which altogether indicates that a vaccine is feasible. In the past, leishmanization, a process in which deliberate infections with a low dose of Leishmania major, etiologic agent of zoonotic cutaneous leishmaniasis (ZCL) causes a controlled skin lesion and provides > 90% protection against reinfection, was a common practice in ZCL-endemic regions. Under the current regulatory environment such practice is not acceptable due to possibility of complications including non-healing lesions. However, these studies suggest that live-attenuated parasites which don not cause a disease could be an effective vaccine for leishmaniasis. Genetically attenuated L. infantum and L. donovani including LdCen-/- have shown promise as a vaccine in animal models. However, using live-attenuated L. donovani as a vaccine in humans could raise safety concerns due to visceralizing potential of this Leishmania species. Attenuated dermatotrophic Leishmania that cross-protects against VL could be a safer vaccine against Leishmania because adverse events (e.g. development of a lesion as vaccination site) will be easy to monitor and can be treated using approved non-pharmacological interventions such as topical thermotherapy. Several clinical and preclinical animal studies have shown that an infection with dermatotrophic Leishmania such as L. major and L. tropica confers cross-protection against VL caused by L. donovani or L. infantum. Using CRISPR-Cas technology, we have generated antibiotic selection marker free centrin gene deficient L. major (LmCen-/-). Whole genome sequencing of LmCen-/- passed through mice multiple times has confirmed stable deletion of centrin gene without other mutations in the parasite genome. Our preliminary findings show that LmCen-/- are highly attenuated and fail to cause disease in immunocompromised animals. We have also found that immunization with LmCen-/- parasites induces disease protective Th1 response in hamsters and protects them against sand fly transmitted VL caused by L. donovani. Similarly, LmCen-/- immunization also protects against CL caused by L. major. These findings indicate that LmCen-/- is a promising vaccine for leishmaniasis. Our industry partner Gennova Biopharma has already established LmCen-/- production under cGMP conditions at their US-FDA approved facility. In this project, we propose to (Aim 1) perform phenotypic, genetic and biochemical characterization of GMP-LmCen-/-parasites and (Aim 2) determine whether GMP-LmCen-/- induce biomarkers of protection in peripheral blood mononuclear cells (PBMCs) from the patients with active VL, asymptomatics and those who have cured VL. The scientific promise of this project, if successful, could provide foundation for advancing LmCen-/- parasites as a vaccine against leishmaniasis in humans.

抽象的 由原生动物寄生虫利什曼原虫引起的感染包括皮肤（CL）、粘膜（ML）和内脏感染 利什曼病（VL）。世界卫生组织将利什曼病列为一种被忽视的热带疾病，感染人数超过 1200 万 全球目前的感染情况，每年约有 200 万新病例。康复的患者 利什曼病会产生针对再次感染的保护性免疫力，这完全表明疫苗是有效的 可行的。过去，利什曼原虫化是一种故意感染低剂量利什曼原虫的过程 人畜共患皮肤利什曼病 (ZCL) 的主要病原体会导致受控制的皮肤病变，并提供 > 90% 的预防再感染率，是 ZCL 流行地区的常见做法。在目前的情况下 在监管环境下，这种做法是不可接受的，因为可能会出现不愈合等并发症 病变。然而，这些研究表明，不引起疾病的减毒活寄生虫可能是 一种有效的利什曼病疫苗。基因减毒的婴儿乳杆菌和多诺瓦尼乳杆菌，包括 LdCen-/- 在动物模型中显示出作为疫苗的前景。然而，使用减毒活杜氏乳杆菌作为疫苗 由于这种利什曼原虫具有内脏化的潜力，因此在人类中使用可能会引起安全问题。减弱的 交叉保护 VL 的皮肤营养性利什曼原虫可能是针对利什曼原虫的更安全的疫苗 因为不良事件（例如作为疫苗接种部位的病变的发展）将很容易监测并且可以 使用经批准的非药物干预措施（例如局部热疗）进行治疗。多项临床和 临床前动物研究表明，皮肤营养性利什曼原虫（如硕大利什曼原虫）的感染 热带乳杆菌可针对杜氏乳杆菌或婴儿乳杆菌引起的 VL 提供交叉保护。使用 CRISPR-Cas 技术，我们已经产生了无抗生素选择标记的中心蛋白基因缺陷的 L. Major (LmCen-/-)。 LmCen-/-经过小鼠多次全基因组测序证实稳定缺失 寄生虫基因组中没有其他突变的中心蛋白基因。我们的初步研究结果表明 LmCen-/- 是 高度减毒，不会在免疫功能低下的动物中引起疾病​​。我们还发现 LmCen-/- 寄生虫免疫可在仓鼠中诱导疾病保护性 Th1 反应并保护它们 对抗由杜氏乳杆菌引起的沙蝇传播的 VL。同样，LmCen-/- 免疫也可以预防 CL 由 L. Major 引起。这些发现表明 LmCen-/- 是一种有前途的利什曼病疫苗。我们的 行业合作伙伴 Gennova Biopharma 已在 cGMP 条件下建立了 LmCen-/- 生产， 他们的美国 FDA 批准的设施。在这个项目中，我们建议（目标 1）进行表型、遗传和 GMP-LmCen-/-寄生虫的生化特征，并（目标 2）确定 GMP-LmCen-/- 是否诱导 活动性 VL 患者外周血单核细胞 (PBMC) 的保护生物标志物， 无症状者和已治愈 VL 者。该项目的科学前景如果成功的话 为推进 LmCen-/- 寄生虫作为人类利什曼病疫苗奠定基础。