Department of Pediatrics, Emory University School of Medicine, Atlanta, Georgia, United States of America.
PLoS One. 2011;6(5):e20069. doi: 10.1371/journal.pone.0020069. Epub 2011 May 16.
Therapeutic targeting of host cell factors required for virus replication rather than of pathogen components opens new perspectives to counteract virus infections. Anticipated advantages of this approach include a heightened barrier against the development of viral resistance and a broadened pathogen target spectrum. Myxoviruses are predominantly associated with acute disease and thus are particularly attractive for this approach since treatment time can be kept limited. To identify inhibitor candidates, we have analyzed hit compounds that emerged from a large-scale high-throughput screen for their ability to block replication of members of both the orthomyxovirus and paramyxovirus families. This has returned a compound class with broad anti-viral activity including potent inhibition of different influenza virus and paramyxovirus strains. After hit-to-lead chemistry, inhibitory concentrations are in the nanomolar range in the context of immortalized cell lines and human PBMCs. The compound shows high metabolic stability when exposed to human S-9 hepatocyte subcellular fractions. Antiviral activity is host-cell species specific and most pronounced in cells of higher mammalian origin, supporting a host-cell target. While the compound induces a temporary cell cycle arrest, host mRNA and protein biosynthesis are largely unaffected and treated cells maintain full metabolic activity. Viral replication is blocked at a post-entry step and resembles the inhibition profile of a known inhibitor of viral RNA-dependent RNA-polymerase (RdRp) activity. Direct assessment of RdRp activity in the presence of the reagent reveals strong inhibition both in the context of viral infection and in reporter-based minireplicon assays. In toto, we have identified a compound class with broad viral target range that blocks host factors required for viral RdRp activity. Viral adaptation attempts did not induce resistance after prolonged exposure, in contrast to rapid adaptation to a pathogen-directed inhibitor of RdRp activity.
针对宿主细胞因子而非病原体成分进行治疗性靶向，为对抗病毒感染开辟了新的前景。这种方法的预期优势包括对病毒耐药性发展的更高屏障和更广泛的病原体靶标谱。正粘病毒主要与急性疾病相关，因此特别适合这种方法，因为治疗时间可以保持有限。为了鉴定抑制剂候选物，我们分析了从大规模高通量筛选中出现的针对两种正粘病毒和副粘病毒家族成员复制的抑制化合物，以确定其抑制作用。这产生了一类具有广泛抗病毒活性的化合物，包括对不同流感病毒和副粘病毒株的有效抑制。在从命中化合物到先导化合物的化学过程中，在永生化细胞系和人 PBMC 中，抑制浓度在纳摩尔范围内。当暴露于人 S-9 肝细胞亚细胞部分时，该化合物表现出高代谢稳定性。抗病毒活性具有宿主细胞种属特异性，在高等哺乳动物来源的细胞中最为明显，支持宿主细胞靶标。虽然该化合物诱导暂时的细胞周期停滞，但宿主 mRNA 和蛋白质生物合成基本不受影响，并且处理后的细胞保持完全的代谢活性。病毒复制在进入后阶段被阻断，类似于已知的病毒 RNA 依赖性 RNA 聚合酶 (RdRp) 活性抑制剂的抑制谱。在存在试剂的情况下直接评估 RdRp 活性，在病毒感染和基于报告基因的小复制子测定中均显示出强烈的抑制作用。总的来说，我们已经鉴定出一类具有广泛病毒靶标范围的化合物，该类化合物可阻断病毒 RdRp 活性所需的宿主因子。与对 RdRp 活性的病原体导向抑制剂的快速适应相比，长时间暴露后病毒适应尝试并未诱导耐药性。
