Single-cycle Infectious Candid#1 to Study Hemorrhagic Fever New World Arenavirus

单周期传染性坦诚

• 批准号: 8385029
• 负责人: Luis Martinez-Sobrido
• 金额: $ 7.68万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8385029
• 关键词: Adverse effects; Antiviral Agents; Arenavirus; Arenavirus Infections; Attenuated Vaccines; Biological; Biological Assay; Biology; Bioterrorism; Cell Line; Cells; Detection; Development; Disease; Drug Delivery Systems; Family; Generations; Genetic; Genome; Glycoproteins; Goals; Green Fluorescent Proteins; Human; Infection; Junin virus; Label; Laboratories; Lassa virus; Lead; Licensing; Life; Life Cycle Stages; Lymphocytic choriomeningitis virus; Mediating; Morbidity - disease rate; Old World Arenaviruses; Plasmids; Prevention; Property; Public Health; RNA; Reagent; Recombinants; Reporter; Reporter Genes; Research; Research Personnel; Ribavirin; Screening procedure; System; Tacaribe Complex Viruses; Technology; Therapeutic; Time; Vaccines; Viral; Viral Hemorrhagic Fevers; Virus; Virus Inhibitors; Virus Replication; Work; base; biodefense; biosafety level 2 facility; drug discovery; high throughput screening; inhibitor/antagonist; innovation; member; mortality; neutralizing antibody; nucleoside analog; pathogen; positional cloning; prophylactic; receptor; stable cell line; vaccine candidate; vaccine development; vector; viral detection; virus identification

DESCRIPTION (provided by applicant): Arenaviruses include several causative agents of Hemorrhagic Fever (HF) disease in humans that are associated with high morbidity and significant mortality. Moreover, weaponized forms of arenaviruses pose a serious threat as agents of bioterrorism. Public health concerns posed by arenaviruses are aggravated by the lack of licensed vaccines and by current anti-arenavirus therapy limited to the off-label use of ribavirin, which is only partially effective and often associated with severe side effects. Limitations in the study of HF arena-viruses include the manipulation of live forms of these agents under BioSafety Level (BSL) 4 laboratories and the requirement of secondary assays for detection of the virus. Development of valid single-cycle infectious surrogates that allow the study of HF arenavirus under less-strict BSL2 facilities and that express a reporter gene for easy viral detection will facilitate the identification of prophylactic and therapeutic strategies using safe, sensitive and specific screening assays compatible with High Throughput Screening (HTS) technologies. Furthermore, they will also represent promising vaccine approaches that circumvent concerns about potential out-of-control replication of attenuated vaccine strains that might lead to disease. Recently, we have described, for the first time, the generation of a single-cycle infectious, reporter-expressing, Old World Lymphocytic Choriomeningitis Virus (LCMV) where we replaced the viral glycoprotein (GP) with a reporter green fluorescent protein (GFP), rLCMV?GP/GFP. Infectious virus was achieved via genetic trans- complementation with cell lines constitutively expressing LCMV GP. This system allowed us to study multiple aspects of the virus and to develop screening assays for detection and quantification of neutralizing antibodies and identification of viral inhibitors. We were also able to extend our work to Lassa virus (LASV), an HF member of the Old World arenavirus, by generating LASV GP-expressing cell lines and LASV GP-pseudotyped rLCMV?GP/GFP. Since Old World and New World arena-viruses differ in their reservoirs, cellular receptor use for viral entry, and RNA genome composition, the development of valid surrogates to study New World arenavirus is imperative to provide researchers a way to study all aspects of the virus biology, to identify antivirals, and to develop vaccines against New World arenaviruses under less strict biosafety laboratories. In this application we propose to expand our recently described technology to the study of New World arenavirus by generating a single-cycle infectious, reporter-expressing, attenuated vaccine strain Candid#1 (rCan?GP/GFP). PUBLIC HEALTH RELEVANCE: Arenaviruses are important viral pathogens that cause common and in some case severe infections in humans. Among them, Hemorrhagic Fever (HF) arenaviruses represent a serious public health problem as well as a threat as agents of bioterrorism. No licensed anti-arenavirus vaccines are available, and current anti-arenavirus therapy is limited to the use of ribavirin, which is only partially effective and often associated with severe side effects. Our goal is to generate a valid surrogate system that facilitates the study of HF New World members in the family under less-strict biosafety conditions by generating a single-cycle infectious, reporter-expressing, attenuated vaccine strain Candid#1.

描述（由申请方提供）：沙粒病毒包括与高发病率和显著死亡率相关的人类出血热（HF）疾病的几种病原体。此外，沙粒病毒的武器化形式作为生物恐怖主义制剂构成严重威胁。沙粒病毒引起的公共卫生问题因缺乏许可的疫苗和目前的抗沙粒病毒治疗仅限于利巴韦林的标签外使用而加剧，利巴韦林仅部分有效且通常与严重副作用相关。HF沙粒病毒研究的局限性包括在生物安全等级（BSL）4实验室下操作这些病原体的活形式，以及检测病毒的二级检测要求。开发有效的单周期感染性替代物，允许在不太严格的BSL 2设施下研究HF沙粒病毒，并表达报告基因以便于病毒检测，这将有助于使用与高通量筛选（HTS）技术兼容的安全、灵敏和特异性筛选试验来确定预防和治疗策略。此外，它们还将代表有希望的疫苗方法，避免对可能导致疾病的减毒疫苗株的潜在失控复制的担忧。最近，我们已经描述了，第一次，一代的单周期传染性，表达，旧世界淋巴细胞脉络丛脑膜炎病毒（LCMV），我们取代了病毒糖蛋白（GP）与记者绿色荧光蛋白（GFP），rLCMV？GP/GFP。通过与组成型表达LCMV GP的细胞系的遗传反式互补获得感染性病毒。该系统使我们能够研究病毒的多个方面，并开发用于检测和定量中和抗体以及鉴定病毒抑制剂的筛选测定法。我们还能够扩大我们的工作，拉沙病毒（LASV），HF成员的旧世界沙粒病毒，通过产生LASV GP表达细胞系和LASV GP假型rLCMV？GP/GFP。由于旧世界和新世界沙粒病毒在其储存库、用于病毒进入的细胞受体和RNA基因组组成方面不同，因此开发有效的替代物来研究新世界沙粒病毒对于为研究人员提供一种研究病毒生物学的各个方面、鉴定抗病毒药物和 在不太严格的生物安全实验室下开发针对新世界沙粒病毒的疫苗。在本申请中，我们建议将我们最近描述的技术扩展到新世界沙粒病毒的研究，通过产生单循环感染性的、表达病毒的减毒疫苗株RCAND#1（rCan？GP/GFP）。 公共卫生相关性：沙粒病毒是重要的病毒病原体，可引起人类常见和某些情况下的严重感染。其中，出血热（HF）沙粒病毒是一个严重的公共卫生问题，也是生物恐怖主义的威胁。没有获得许可的抗沙粒病毒疫苗可用，目前的抗沙粒病毒治疗仅限于使用利巴韦林，其仅部分有效且通常与严重副作用相关。我们的目标是产生一个有效的替代系统，通过产生一个单周期感染性的，表达抗原的，减毒疫苗株p53 d#1，促进在不太严格的生物安全条件下研究家族中的HF新世界成员。