Broad-range Inhibitors of Human Immunodeficiency Virus Entry

描述(申请人提供)：人类免疫缺陷病毒(HIV-1)是获得性免疫缺陷综合征(AIDS)的病原体。全球艾滋病毒-1大流行(约3500万人感染)每年有200-300万新感染者持续存在。改变这一大流行的进程需要预防艾滋病毒-1传播，其中大部分是通过性传播。在缺乏有效疫苗的情况下，迫切需要阻止艾滋病毒-1经性传播的方法。最近，研究表明，使用特定的抗逆转录病毒药物可以部分保护高危性伴免受艾滋病毒-1感染。然而，部分疗效、药物副作用和耐药病毒的出现限制了这些特殊药物作为预防措施的普遍适用性。新的广泛有效的抗病毒药物可以用作局部杀微生物剂，可以弥补这些不足。HIV-1包膜糖蛋白(Envs)介导病毒进入靶细胞，是此类预防性药物的诱人靶点。HIV-1env暴露在病毒表面，可被水溶性抑制剂访问，并且在每个病毒粒子上的数量很少。用作杀菌剂的环境病毒抑制剂不需要被宿主细胞系统吸收或吸收，从而限制了潜在的毒性。Env的保守元件介导受体结合、构象变化和膜融合，为抑制提供了几个潜在的靶点。虽然已经确定了一些HIV-1进入抑制剂，但耐药的HIV-1变种要么是自然存在的，要么是在治疗期间形成的。我们设计了一种筛选策略，以确定新的广泛抑制HIV-1进入的药物。在我们的筛选实验中，HIV-1env的功能导致表达env的细胞与表达病毒受体的细胞融合。这种细胞-细胞融合试验和特异性对照试验将同时用于NIH小分子文库的初步筛选。排名最高的化合物将通过二次筛查进行验证，评估重复性、剂量反应、广度和特异性。涉及带有多个HIV-1和其他逆转录病毒包膜糖蛋白的病毒的单轮复制的三次检测可以以适中的吞吐量进行，并将提供有关效力、广度和特异性的信息。用于评估作用机制的其他三级分析方法在 首席研究人员实验室，并将应用于选定的化合物。将对特定和广泛的抑制剂进行修改，以在保持广度的同时提高效力。这项研究中确定的化合物可能作为预防性杀微生物剂、作为已经感染的个人的治疗方法，以及作为了解HIV-1进入的复杂、多步骤过程的探针。

DESCRIPTION (provided by applicant): Human immunodeficiency virus (HIV-1) is the etiologic agent of acquired immunodeficiency syndrome (AIDS). The global HIV-1 pandemic (~35 million people infected) is sustained by 2-3 million new infections annually. Changing the course of this pandemic requires prevention of HIV-1 transmission, most of which occurs sexually. In the absence of an effective vaccine, modalities that block sexual HIV-1 transmission are desperately needed. Recently, the use of specific antiretroviral drugs has been investigated and shown to partially protect at-risk sex partners from HIV-1 infection. However, partial efficacy, drug side effects and the emergence of drug-resistant viruses limit the general applicability of these particular agents as prophylactic measures. New broadly active antiviral agents that can be used as topical microbicides could remedy these deficiencies. The HIV-1 envelope glycoproteins (Envs), which mediate virus entry into target cells, represent attractive targets for such prophylactic agents. The HIV-1 Envs are exposed on the viral surface, are accessible to water-soluble inhibitors and are present in low numbers on each virion. Env inhibitors used as microbicides need not be systemically absorbed or taken up by host cells, limiting potential toxicity. Conserved elements of Env mediate receptor binding, conformational changes, and membrane fusion, providing several potential targets for inhibition. Although some HIV-1 entry inhibitors have been identified, drug-resistant HIV-1 variants either exist naturally o develop during treatment. We have devised a screening strategy to identify new broad-range inhibitors of HIV-1 entry. In our screening assay, HIV-1 Env function leads to the fusion of Env-expressing cells with cells expressing the viral receptors. This cell-cell fusion assay and a specificity control assay will be used in parallel in a primary screen of the NIH small-molecule library. The highest-ranked compounds will be validated by a secondary screen that assesses reproducibility, dose-response, breadth and specificity. Tertiary assays involving single-round replication of viruses with multiple HIV-1 and other retroviral envelope glycoproteins can be performed at modest throughput and will provide information on potency, breadth and specificity. Additional tertiary assays to evaluate mechanism of action are well-established in the principal investigator's laboratory, and will be applied to selected compounds. Specific and broad-range inhibitors will be modified to improve potency while retaining breadth. The compounds identified in this study could potentially be useful as prophylactic microbicides, as treatments for already infected individuals, and as probes to understand the complex, multi-step process of HIV-1 entry.