Targeting immunophilin- and MAPK-associated pathways to inhibit MERS-CoV replication and prevent lung injury

靶向亲免素和 MAPK 相关途径抑制 MERS-CoV 复制并预防肺损伤

• 批准号: 319808243
• 负责人: Professor Dr. Stephan Becker
• 金额: --
• 依托单位: Medizinische Klinik V, Klinik für Infektiologie und Krankenhaushygiene
• 依托单位国家: 德国
• 项目类别: Clinical Research Units
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/319808243?language=en
• 关键词: Targeting; immunophilin; MAPK; associated; pathways

The emergence of SARS (Severe Acute Respiratory Syndrome) in 2002 and MERS (Middle East Respiratory Syndrome) in 2012 revealed the potential of coronaviruses to induce severe pneumonia and lung injury with high fatality rates. Since novel highly pathogenic CoV are presumed to represent an emerging threat to humans in future, broad-range, readily-available therapeutics are of utmost importance. Using an established MERS-Coronavirus (MERS CoV) mouse infection model and primary human lung cells, we aim to characterize the antiviral mechanisms of compounds known to block MERS CoV-host interactions by inducing interferon lambda (IFNλ) and targeting immunophilin and downstream MAPK pathways in vitro and in vivo. The final goal is to establish readily-available treatment strategies for current and future emerging infections caused by highly pathogenic human coronaviruses. In the first funding period, we investigated the antiviral and lung injury-attenuating properties of Cyclosporin A (CsA) in MERS-CoV infection. We revealed that a CsA-induced, strong antiviral effect was largely mediated by induction of a pronounced type III interferon (IFNλ) response. Elevated IFNλ levels stimulated an IRF1 (interferon regulatory transcription factor 1)-dependent upregulation of interferon stimulated genes (ISG) and strongly attenuated viral growth in vitro, in human primary alveolar epithelial cells ex vivo, and, importantly, in our recently established MERS-CoV mouse model in vivo (Sauerhering et al., unpublished data). In addition, our data indicate that both immunophilin- and MAPK-dependent pathways convey the CsA-induced block of MERS-CoV replication. Interestingly, among various MAPK analyzed, only JNK inhibition led to a significant impairment of MERS-CoV particle release. In the upcoming funding period we aim to investigate i) how CsA-stimulated molecular signaling cascades lead to IFNλ induction and investigate whether MERS CoV-developed escape strategies counteract cellular defense mechanisms. We will further ii) investigate putative downstream effectors induced by CsA in MERS-CoV infected respiratory epithelial cells which were previously defined by RNA-Seq analyses. The identified differentially expressed genes will be evaluated for their suitability as targets for antiviral strategies. Moreover, we will elucidate iii) how JNK impairs MERS-CoV particle egress by characterizing its interplay with host cell factors including cyclophilin A to identify and evaluate novel molecular targets for their therapeutic value in our in vivo and ex vivo infection models. Finally, iv) we will investigate the therapeutic potential of treatment strategies targeting CsA-downstream molecules in MERS-CoV in vivo infection.The obtained data are expected to reveal immunophilin-targeting antiviral compounds for treatment of MERS-CoV, and potentially further respiratory virus infections in vivo.

2002年SARS(严重急性呼吸系统综合症)和2012年MERS(中东呼吸综合征)的出现揭示了冠状病毒可能导致严重肺炎和高死亡率的肺损伤。由于新的高致病性冠状病毒被推定为未来对人类的一种新的威胁，广泛的、容易获得的治疗方法是至关重要的。利用已建立的MERS冠状病毒小鼠感染模型和原代人肺细胞，我们的目标是在体外和体内研究通过诱导干扰素λ和靶向免疫亲和素及其下游的MAPK通路来阻断MERS冠状病毒与宿主相互作用的化合物的抗病毒机制。最终目标是为目前和未来由高致病性人类冠状病毒引起的新出现的感染建立现成的治疗策略。在第一个资助阶段，我们研究了环孢菌素A(CsA)在MERS冠状病毒感染中的抗病毒和减轻肺损伤的特性。我们发现，CsA诱导的强大的抗病毒作用在很大程度上是通过诱导明显的III型干扰素(干扰素λ)反应来调节的。干扰素λ水平升高可刺激依赖干扰素调节转录因子1的干扰素刺激基因上调，并强烈抑制体外培养的人肺泡上皮细胞中的病毒生长，更重要的是，在我们最近建立的体内中东呼吸综合征冠状病毒小鼠模型中(Sauerhering等人，未发表数据)。此外，我们的数据表明，免疫亲和素和MAPK依赖的途径都传递了CsA诱导的MERS-CoV复制的阻断。有趣的是，在分析的各种MAPK中，只有JNK抑制导致MERS-CoV颗粒释放显著障碍。在接下来的资金阶段，我们的目标是研究1)CsA刺激的分子信号级联如何导致干扰素λ的诱导，并研究MERS冠状病毒发展的逃逸策略是否抵消了细胞防御机制。我们将进一步研究CsA在MERS冠状病毒感染的呼吸道上皮细胞中可能诱导的下游效应，这是以前通过RNA-Seq分析定义的。识别出的差异表达基因将被评估它们是否适合作为抗病毒策略的靶点。此外，我们将通过表征JNK与包括亲环素A在内的宿主细胞因子的相互作用来阐明JNK如何损害MERS-CoV颗粒外泄，以确定和评估其在体内和体外感染模型中的治疗价值的新的分子靶点。最后，iv)我们将研究针对MERS冠状病毒CsA下游分子的治疗策略在体内感染的治疗潜力。所获得的数据有望揭示免疫亲和素靶向的抗病毒化合物用于治疗MERS冠状病毒，以及潜在的进一步体内呼吸道病毒感染。