A General Molecular Strategy for Attenuation of Human Pathogenic Arenaviruses

人类致病性沙粒病毒减毒的通用分子策略

• 批准号: 9217579
• 负责人: Juan C. de la Torre
• 金额: $ 24.06万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-02-05 至 2019-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9217579
• 关键词: Adverse effects; Africa; Animal Model; Antigens; Area; Arenaviridae; Arenavirus; Attenuated; Attenuated Live Virus Vaccine; Cavia; Cell Line; Cells; Clinical; Clinical Trials; Competence; Consensus Sequence; Cultured Cells; Development; Dideoxy Chain Termination DNA Sequencing; Disease; Engineering; Exhibits; Future; Genetic; Genome; Goals; Growth; Heterophile Antigens; Human; Immune response; Immunity; Immunize; Individual; Infection; Interferons; Knowledge; Label; Lassa Fever; Lassa virus; Lujo virus; Lymphocytic choriomeningitis virus; Mediating; Modeling; Molecular; Morbidity - disease rate; Mus; Mutation; Old World Arenaviruses; Pathogenicity; Phenotype; Population; Production; Property; Protocols documentation; Public Health; Reassortant Viruses; Recombinants; Resources; Ribavirin; Serial Passage; South Africa; Symptoms; Testing; Travel; Vaccines; Viral; Viral Hemorrhagic Fevers; Virulence; Virus; Virus Diseases; attenuation; base; biodefense; clinically significant; combat; design; epidemiology study; genetic approach; in vivo; interest; metropolitan; mortality; mouse model; neglect; novel; novel strategies; pathogen; public health relevance; pyrosequencing; response; reverse genetics

 DESCRIPTION (provided by applicant): Several arenaviruses cause hemorrhagic fever (HF) disease in humans and pose a serious public health problem in their endemic regions. Thus, the Old World arenavirus (OWA) Lassa (LASV) infects several hundred thousand individuals yearly in West Africa resulting in a high number of Lassa fever (LF) cases associated with high morbidity and mortality. Moreover, evidence indicates that the worldwide-distributed OWA LCMV is a neglected clinically important human pathogen. In addition, several arenaviruses including LASV and LCMV pose a credible biodefense threat. Existing anti-arenaviral therapy is limited to an off-label use of ribavirin that is only partially effective, and there are no FDA-licensed, or currently in clinical trials, arenavirus vaccines. However, the MOPV/LASV reassortant (ML29) is a candidate live-attenuated vaccine (L-AttV) for LASV that has shown promising results in animal models. Nevertheless, as with many other traditional L-AttV, the mechanism of ML29 attenuation remains unknown, which raises concerns about the phenotypic stability of ML29 in response to additional mutations. The central goal of this application is to test the hypothesis that we can convert a pathogenic OWA into an attenuated form with features of L-AttV via replacement of its L IGR by a genetically defined synthetic S-like IGR (Ssyn). Our hypothesis is supported by our recent following findings: 1) We could rescued rLCMV(IGR/S-S) with the same S-IGR in both S and L genome segments, and found that it was highly attenuated in vivo. 2) Mice immunized with rLCMV(IGR/S-S) were protected against a lethal challenge with wild type (WT) LCMV. 3) We have obtained evidence that a high degree of sequence plasticity within the S IGR is compatible with virus viability, which supports the feasibility of engineering recombinant arenaviruses with a synthetic S IGR (Ssyn) in the L segment as a general molecular strategy for arenavirus attenuation. To test our hypothesis we propose to complete the following specific aims: Aim 1. Characterize rLCMV (IGR/S-Ssyn): We have rescued rLCMV(IGR/S-Ssyn) where a synthetic S IGR (Ssyn) substituted for the L-IGR. We will characterize rLCMV(IGR/S-Ssyn) in cultured cells and using the mouse model of LCMV infection test the hypothesis that rLCMV(IGR/S-Syn) displays key features for L-AttV. Aim 2: Generate and characterize rLASV(IGR/S-Ssyn): We will generate rLASV(IGR/S-Ssyn) and test the hypothesis that it is attenuated in vivo but able to induce protective immunity against a letha challenge with WT LASV in a well-established guinea pig model of LASV infection. Aim 3. Examine the stability of rLCMV(IGR/S-Ssyn): We will test the hypothesis that rLCMV(IGR/S-Ssyn) is genetically stable during multiplication under different growth conditions. The successful completion of this application will uncover a general molecular strategy for arenavirus attenuation that can facilitate a novel strategy to develop L-AttV to combat human pathogenic arenaviruses.

 描述（由适用提供）：几种体育症病毒会导致人类出血热（HF）疾病，并在其内在地区构成严重的公共卫生问题。这就是旧世界体育症病毒（OWA）LASSA（LASV）每年在西非感染数十万个人，导致大量的LASSA热（LF）病例与高发病率和死亡率相关。此外，证据表明，全球分布的OWA LCMV是人类病原体。此外，包括LASV和LCMV在内的几个体育症病毒构成了可靠的生物形式威胁。现有的抗动脉粥样硬病疗法仅限于仅部分有效的利巴韦林的标签外使用，并且没有FDA许可，或者目前正在临床试验中，Arenavirus疫苗。但是，MOPV/LASV REASORTANTANT（ML29）是LASV的候选疫苗（L-ATTV），在动物模型中显示出有望的结果。然而，与许多其他传统的L-ATTV一样，ML29衰减的机制仍然未知，这引起了人们对ML29对其他突变的表型稳定性的担忧。该应用程序的核心目的是测试我们可以通过遗传定义的合成S-like IGR（SSYN）将致病性OWA转换为具有L-ATTV特征的衰减形式的假设。我们最近的以下发现得到了我们的假设：1）我们可以在S和L基因组段中对RLCMV（IGR/S-S）进行相同的S-gr，并发现它在体内受到了高度减弱。 2）保护用RLCMV（IGR/S-S）免疫的小鼠受到对野生型（WT）LCMV的致命挑战的保护。 3）我们获得了证据表明，SIGR内的高度序列可塑性与病毒活力兼容，该病毒的可行性支持了L段中具有合成的SSSYN（SSYN）的工程重组体系毒病毒作为一般分子策略的可行性。为了检验我们的假设，我们建议完成以下特定目的：AIM 1。表征RLCMV（IGR/S-SSYN）：我们已经恢复了RLCMV（IGR/S-SSYN），其中合成S IGR（SSYN）代替了L-gr-gr。我们将在培养细胞中表征RLCMV（IGR/S-SYN），并使用LCMV感染的小鼠模型检验RLCMV（IGR/S-SYN）显示L-ATTV的关键特征。 AIM 2：生成和表征RLASV（IGR/S-SSYN）：我们将生成RLASV（IGR/S-SSYN），并检验了以下假说：它在体内会减弱，但可以在lasv感染的几内亚猪模型中诱导WT LASV诱导保护性免疫，以防止letha挑战。 AIM 3。检查RLCMV（IGR/S-SSYN）的稳定性：我们将测试以下假设：在不同生长条件下乘积期间，RLCMV（IGR/S-SYN）在繁殖过程中通常是稳定的。该应用程序的成功完成将揭示用于轨道病毒衰减的一般分子策略，该策略可以促进开发L-ATTV来打击人类致病性体育疾病病毒的新型策略。

项目成果

A Lassa Virus Live-Attenuated Vaccine Candidate Based on Rearrangement of the Intergenic Region

• DOI: 10.1128/mbio.00186-20
• 发表时间: 2020-03-01
• 期刊: MBIO
• 影响因子: 6.4
• 作者: Cai, Yingyun;Iwasaki, Masaharu;de la Torre, Juan Carlos
• 通讯作者: de la Torre, Juan Carlos

Molecular Engineering of a Mammarenavirus with Unbreachable Attenuation.

具有不可破坏的减毒能力的乳腺病毒的分子工程。

• DOI: 10.1128/jvi.01385-22
• 发表时间: 2023
• 期刊: Journal of virology
• 影响因子: 5.4
• 作者: Sakabe,Saori;Cubitt,Beatrice;Martinez-Sobrido,Luis;delaTorre,JuanC
• 通讯作者: delaTorre,JuanC