Development of Improved HIV-1 Capsid Inhibitors

摘要 HIV-1衣壳蛋白(CA)是一种极具吸引力的新型抗逆转录病毒药物。 在病毒生物学中的重要和多方面的作用。Lenacapavir(LEN)是最近发现的一种一线药物， 具有皮摩尔效力的长效衣壳靶向抑制剂，抗HIV-1。2/3期临床试验 结果显示，皮下注射LEN，间隔6个月，可使HIV-1感染率高 有大量治疗经验的感染多重耐药病毒表型的患者的抑制。这些 研究结果表明，LEN是一种很有前途的药物，可以补充现有的抗逆转录病毒化合物来 治疗HIV-1感染者。然而，基于细胞培养的病毒突破性分析和临床试验 确定了许多对LEN具有实质性抗性的CA替换。具体来说，Q67H/N74D和 M66I替换是主要的耐药相关变异。因此，有必要发展 改进的LEN模拟，具有更高的抵抗力。我们已经在学术上建立了LEN的合成 通过利用模块化方法独立地制备三个独特的LEN亚基(A、B和C)来设置 商业上可以买到的试剂。这种模块化方法允许直接修改每个 亚基，然后可以以任何顺序组合以制备LEN及其类似物。因此，我们的药物化学 战略对模拟发展是非常有利的。我们互补的病毒学、生物化学和 结构生物学实验使我们能够描述LEN的多模式作用机制。 此外，我们最近还测定了CA六聚体的高分辨X射线晶体结构，其中包含 主要耐药相关基因Q67H/N74D和M66I突变。我们建议利用这些发现来 合理开发具有更高抵抗力的改良LEN类似物。在目标1中，我们的努力将集中在 对LEN亚基A和C的修饰以克服空间位阻和静电斥力 耐药的Q67H/N74D CA变异株。在目标2中，我们将使用我们最新的来自MiniFrags的有希望的发现 筛选研究，确定了与LEN附近的CA疏水口袋结合的片段。 我们将通过将这些片段连接到LEN来合成新的化学类型，以生成第二代 类似物针对主要的耐药M66I变种。目标1和目标2的新合成化合物 将通过使用抗病毒活性、细胞毒性、表面等离子体共振和X-射线扫描等逐步方法进行评估。 用射线结晶学鉴定和表征先导化合物(S)。综上所述，拟议的研究是 有望产生新的化学类型，提高对主要耐药CA的抗病毒活性 对双亲Len产生实质性抵抗的变种。

ABSTRACT The HIV-1 capsid protein (CA) is an attractive target for the development of novel antiretrovirals due to its essential and multifaceted roles in the virus biology. Lenacapavir (LEN) is a recently discovered first-in-class, long-acting capsid-targeting inhibitor with picomolar potency against HIV-1. Phase 2/3 clinical trials have revealed that subcutaneous administration of LEN with a six-month dosing interval enables high rates of HIV-1 suppression in heavily treatment-experienced patients infected with multi-drug resistant viral phenotypes. These findings have highlighted LEN as a promising agent that could complement current antiretroviral compounds to treat people living with HIV-1. However, cell culture-based viral breakthrough assays and clinical trials have identified a number of CA substitutions that confer substantial resistance to LEN. Specifically, Q67H/N74D and M66I substitutions emerged as major drug-resistance associated variants. Therefore, there is a need to develop improved LEN analogs with a higher barrier to resistance. We have established synthesis of LEN in academic setting by utilizing a modular approach of independently preparing three unique LEN subunits (A, B, and C) from commercially available reagents. Such modular approach allows for straightforward modifications of each subunit which can then be combined in any order to prepare LEN and its analogs. Thus, our medicinal chemistry strategy is highly advantageous for analog development. Our complementary virology, biochemistry and structural biology experiments have allowed us to characterize a multi-modal mechanism of action of LEN. Furthermore, we have recently determined high-resolution X-ray crystal structures of CA hexamers containing major drug-resistance associated Q67H/N74D and M66I changes. We propose to exploit these findings to rationally develop improved LEN analogs with a higher barrier to resistance. In Aim 1, our efforts will focus on modifications to LEN subunits A and C to overcome steric hindrance and electrostatic repulsions created by the drug resistant Q67H/N74D CA variant. In aim 2, we will use our recent promising findings from MiniFrags screening studies, which identified fragments that bind to the CA hydrophobic pocket in close vicinity to LEN. We will synthesize new chemotypes by connecting these fragments to LEN to generate second-generation analogs to target the major drug-resistant M66I variant. Newly synthesized compounds from both Aims 1 and 2 will be evaluated by a stepwise approach using antiviral activity, cytotoxicity, surface plasmon resonance and X- ray crystallography to identify and characterize the lead compound(s). Taken together, the proposed research is expected to generate novel chemotypes with improved antiviral activities against major drug resistant CA variants that confer substantial resistance to parental LEN.