Capsid-Targeting Small Molecules Blocking HIV through Novel Mechanism of Action

通过新颖的作用机制靶向衣壳的小分子阻断 HIV

• 批准号: 8802860
• 负责人: Stefan G Sarafianos
• 金额: $ 22.61万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-02-06 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8802860
• 关键词: Affect; Anti-Retroviral Agents; Antiviral Agents; Binding; Biological; Biological Assay; C-terminal; Capsid; Capsid Proteins; Cell Culture Techniques; Cell Line; Cells; Clinic; Clinical; Complement; Complex; Cysteine; DNA; Data; Drug resistance; Engineering; Event; Future; HIV; HIV Infections; HIV-1; Health; Highly Active Antiretroviral Therapy; Housing; Individual; Infection; Integration Host Factors; Laboratories; Lead; Length; Life Cycle Stages; Link; Morbidity - disease rate; Morphology; Multi-Drug Resistance; Mutation; Nuclear; Patients; Peripheral Blood Lymphocyte; Pharmaceutical Preparations; Play; Polyadenylation; Process; Relative (related person); Resistance; Reverse Transcription; Role; Series; Site; Staging; Structure; Testing; Therapeutic; Time; Transmission Electron Microscopy; Tube; Vial device; Viral; Viral Drug Resistance; Viral Load result; Viral Proteins; Virus; base; crosslink; cytotoxic; cytotoxicity; design; drug discovery; inhibitor/antagonist; insight; knock-down; macrophage; monocyte; mortality; novel; research study; screening; small molecule; small molecule libraries; viral DNA

DESCRIPTION: Highly Active Antiretroviral Therapies (HAART) have been successful at controlling viral load in HIV-infected patients and have significantly decreased the morbidity and mortality associated with HIV-1 infection. However, drug resistant HIV-1 strains arise and are transmitted between individuals, reducing the efficacy of currently available antivirals. Thus, identification of novel antiretrovirals (ARTs) that act by new mechanisms and have no cross-resistance with existing therapeutics is required. HIV-1 capsid (CA) is a viral protein essential for early and late events of the replication cycle and so far has been an untapped target, thus providing excellent opportunities for the discovery of novel ARTs that act by novel mechanisms of action. We screened an in-house chemical library of compounds and identified18E8, a small molecule that interferes with multimerization of HIV-1 CA. We demonstrated in cell-based assays with fully-infectious HIV that 18E8 showed broad antiretroviral activity (EC50 as low as ~1?M) against multiple laboratory strains and several multi-drug resistant clinical isolates. In additional preliminary experiments we showed that 18E8 exerts its antiretroviral activity by binding to HIV-1 CA. Six additional hits from this screening have not yet been characterized. In order to gain insight into the mode of action of 18E8 and to determine the specific step of the viral replication cycle that it affects, we performed time-of-drug-addition experiments that define how long the addition of 18E8 could be postponed before losing its antiviral activity in cell culture. Surprisingly, our preliminary data suggest that 18E8 targets an early step in the HIV replication cycle, after reverse transcription. 18E8 did not appear to affect late stages of the vial life cycle. Based on these data, we hypothesize that the unique mechanism of action of 18E8 is due to its binding in a manner different than other CA-targeting compounds that affect other steps of the virus life cycle. We will explore this hypothesis and use the same approaches to evaluate and characterize additional compound hits in the following two aims: Specific Aim 1. Functional and mechanistic characterization of 18E8 and other compounds Specific Aim 2. Crystallographic characterization of CA interactions with 18E8 and other compounds. The deliverables of the proposed studies include at least one novel CA-targeting antiviral with a new mechanism of action, its biological and structural mechanism of action that could guide the design of novel antivirals.

描述：高效抗逆转录病毒疗法 (HAART) 已成功控制 HIV 感染患者的病毒载量，并显着降低了与 HIV-1 感染相关的发病率和死亡率。然而，耐药性 HIV-1 毒株的出现并在个体之间传播，降低了现有抗病毒药物的功效。因此，需要鉴定出通过新机制发挥作用且与现有疗法不存在交叉耐药性的新型抗逆转录病毒药物（ART）。 HIV-1衣壳（CA）是一种病毒蛋白，对于复制周期的早期和晚期事件至关重要，迄今为止一直是一个尚未开发的靶标，从而为发现通过新作用机制发挥作用的新型ART提供了绝佳的机会。我们筛选了内部化合物化学库并鉴定了 18E8，这是一种干扰 HIV-1 CA 多聚化的小分子。我们在完全感染性 HIV 的细胞检测中证明，18E8 对多种实验室菌株和几种多重耐药临床分离株表现出广泛的抗逆转录病毒活性（EC50 低至约 1?M）。在其他初步实验中，我们表明 18E8 通过与 HIV-1 CA 结合发挥其抗逆转录病毒活性。本次筛选的另外 6 个命中尚未确定特征。 为了深入了解 18E8 的作用模式并确定其影响的病毒复制周期的具体步骤，我们进行了药物添加时间实验，定义了 在细胞培养中失去其抗病毒活性之前，18E8 的添加可以推迟多长时间。令人惊讶的是，我们的初步数据表明，18E8 靶向逆转录后 HIV 复制周期的早期步骤。 18E8 似乎不会影响小瓶生命周期的后期阶段。基于这些数据，我们假设 18E8 的独特作用机制是由于其结合方式不同于影响病毒生命周期其他步骤的其他 CA 靶向化合物。我们将探索这一假设，并使用相同的方法来评估和表征以下两个目标中的其他化合物： 具体目标 1. 18E8 和其他化合物的功能和机制表征 具体目标 2. CA 与 18E8 和其他化合物相互作用的晶体学表征。拟议研究的成果包括至少一种具有新作用机制的新型 CA 靶向抗病毒药物，其生物学和结构作用机制可以指导新型抗病毒药物的设计。