Future pandemic influenza necessitates the development of new drugs against the current circulating, amantadine and rimantadine drugs resistant, influenza A M2 S31N viruses. The possibility of an antigenic shift to M2 S31 necessitates ranking the biological activities of amantadine variants. Several amantadine variants have been tested by different laboratories, but various M2 wild type influenza A strains have been used with different sensitivity against amantadine and the unambiguous comparison between potencies is not straightforward. Here, we compared the anti-influenza activities of 57 synthetic amantadine variants against influenza A WSN/33 viruses with amantadine-sensitive M2 WT, with a range of over three digits providing a reference set of potencies for structure-activity relationships, and amantadine-resistant M2 S31N proteins (and observed no potent compounds). 17 compounds were selected and tested against M2 L26F, V27A, A30T, G34E viruses. We tested few reference compounds using electrophysiology and explored point mutations which both showed that M2 is the target of potent antiviral potency against the M2 WT, L26F, V27A viruses. Major findings are: (a) Several amantadine variants from Kolocouris group block only M2 WT and M2 L26F-mediated proton current and the corresponding viruses replication. (b) A compound from Vazquez’s group is a triple blocker of M2 WT, L26F, V27A channels and viruses replication. (c) A compound from Vazquez’s group blocks only M2 L26 channel and virus replication. (d) Several compounds from Kolocouris group have potent activity against several influenza A M2 WT and three M2 S31N viruses, eg. the pandemic A/H1N1/California/07/2009 (H1N1pdm09) or A/H1N1/PuertoRico/08/1934 without blocking M2 S31N. The compounds and their cocktails while not to be more toxic than amantadine might be useful for re-purposing of amantadine class of drugs in the case (i) of the prevalence of M2 L26F and or M2 V27A strains (ii) of an antigenic shift of the virus to M2 WT and (iii) because they inhibited a broad panel of M2 WT and M2 S31N viruses including the H1N1pdm09). (d) We showed that the mechanism of antiviral activity against A/California/07/2009 or A/PR/08/1934 and possibly also M2 WT viruses compared to WSN/33 viruses is not due to inhibition of an early stage of virus infection or a late stage of M2 channel function during endocytosis or inhibition of HA binding to host cells or a different pH for HA fusion or a lysosomotropic effect.

中文翻译：

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金刚烷胺变体对多种甲型流感病毒的活性

未来的大流行性流感需要开发针对当前流行的、对金刚烷胺和金刚乙胺耐药的甲型流感 M2 S31N 病毒的新药。抗原转变为 M2 S31 的可能性需要对金刚烷胺变体的生物活性进行排序。不同的实验室已经测试了几种金刚烷胺变体，但已使用各种 M2 野生型 A 型流感病毒株对金刚烷胺具有不同的敏感性，并且效力之间的明确比较并不简单。在这里，我们比较了 57 种合成金刚烷胺变体对具有金刚烷胺敏感 M2 WT 的甲型流感 WSN/33 病毒的抗流感活性，范围超过三位数，为结构-活性关系提供了一组参考效力，和金刚烷胺抗性 M2 S31N 蛋白（未观察到有效化合物）。选择了 17 种化合物，并针对 M2 L26F、V27A、A30T、G34E 病毒进行了测试。我们使用电生理学测试了少数参考化合物并探索了点突变，这两者都表明 M2 是针对 M2 WT、L26F、V27A 病毒的有效抗病毒效力的目标。主要发现是：（a）来自 Kolocouris 组的几个金刚烷胺变体仅阻断 M2 WT 和 M2 L26F 介导的质子电流和相应的病毒复制。(b) Vazquez 小组的一种化合物是 M2 WT、L26F、V27A 通道和病毒复制的三重阻滞剂。(c) Vazquez 小组的一种化合物仅阻止 M2 L26 通道和病毒复制。(d) 来自 Kolocouris 组的几种化合物对几种甲型流感 M2 WT 和三种 M2 S31N 病毒具有强效活性，例如。大流行 A/H1N1/California/07/2009 (H1N1pdm09) 或 A/H1N1/PuertoRico/08/1934 而不阻塞 M2 S31N。在 (i) M2 L26F 和/或 M2 V27A 毒株流行 (ii)病毒对 M2 WT 和 (iii) 因为它们抑制了广泛的 M2 WT 和 M2 S31N 病毒，包括 H1N1pdm09）。(d) 我们表明，与 WSN/33 病毒相比，针对 A/California/07/2009 或 A/PR/08/1934 以及可能还有 M2 WT 病毒的抗病毒活性机制不是由于抑制病毒的早期阶段感染或内吞过程中 M2 通道功能的后期阶段或抑制 HA 与宿主细胞的结合或不同的 pH 值以促进 HA 融合或溶酶体作用。