Antiviral Research Unit, IPC 424, Pfizer PGRD, Discovery Biology, Sandwich Laboratories, Sandwich, Kent CT13 9NJ, United Kingdom.
J Virol. 2010 Jul;84(14):7288-99. doi: 10.1128/JVI.00535-10. Epub 2010 Apr 28.
A low-molecular-weight human immunodeficiency virus type 1 (HIV-1) inhibitor, PF-68742 (molecular weight, 573), has been identified in a high-throughput screen for compounds that block HIV-1 envelope glycoprotein (Env)-mediated fusion. The compound is shown to be potent against R5 and X4 isolates in both cell-cell fusion and antiviral assays (50% effective concentrations of approximately 0.1 to 1 muM). Postfusion and HIV-1 pseudotyping control experiments confirm that PF-68742 is an entry inhibitor with Env as the specific target for antiviral action. PF-68742 was not able to block binding of monomeric gp120 to soluble CD4 or the binding of gp120:CD4 complexes to cell-associated CCR5, thus distinguishing PF-68742 from described gp120 antagonists and coreceptor binders. Escape variants of HIV-1(NL4-3) were selected, and all resistant viruses were found to contain a common G514R (HxB2 numbering) mutation in Env, located proximal to the furin cleavage site in the fusion peptide of gp41. When introduced into wild-type NL4-3 gp41, G514R conferred resistance to PF-68742. Resistance via G514R is shown to be associated with enhancement of virion infectivity by PF-68742 that may result from altered properties of inhibitor-bound Env, rather than from a loss of compound binding. Wild-type viruses and those with substitutions in the disulfide loop (DSL) region of gp41 were also examined for PF-68742 sensitivity. Here, complete resistance to PF-68742 was found to occur through changes outside of position 514, including in the gp41 DSL region. The results highlight PF-68742 as a starting point for novel therapies against HIV-1 and provide new insights into models of Env-mediated fusion.
一种低分子量的人类免疫缺陷病毒 1 型（HIV-1）抑制剂 PF-68742（分子量 573）已在高通量筛选可阻断 HIV-1 包膜糖蛋白（Env）介导融合的化合物中被鉴定出来。该化合物在细胞-细胞融合和抗病毒测定中对 R5 和 X4 分离株均表现出强大的抑制作用（50%有效浓度约为 0.1 至 1 μM）。融合后和 HIV-1 假型控制实验证实，PF-68742 是一种进入抑制剂，Env 是抗病毒作用的特定靶标。PF-68742 不能阻断单体 gp120 与可溶性 CD4 的结合，也不能阻断 gp120:CD4 复合物与细胞相关 CCR5 的结合，因此 PF-68742 有别于已描述的 gp120 拮抗剂和核心受体结合剂。选择了 HIV-1（NL4-3）的逃逸变异体，所有耐药病毒均发现含有 Env 中靠近 gp41 融合肽中弗林裂解位点的共同 G514R（HxB2 编号）突变。当将 G514R 引入野生型 NL4-3 gp41 中时，G514R 赋予对 PF-68742 的耐药性。通过 G514R 产生的耐药性与 PF-68742 增强病毒感染力有关，这可能是由于结合抑制剂的 Env 的特性发生改变，而不是由于化合物结合的丧失。还研究了带有 gp41 二硫键环（DSL）区域取代的野生型病毒和那些病毒对 PF-68742 的敏感性。在这里，发现完全耐药性是通过 514 位以外的位置发生变化而产生的，包括 gp41 DSL 区域。这些结果强调了 PF-68742 作为针对 HIV-1 的新型治疗方法的起点，并提供了有关 Env 介导融合模型的新见解。
