The HIV-1 Nef accessory factor enhances the viral life cycle in vivo, promotes immune escape of HIV-infected cells, and represents an attractive antiretroviral drug target. However, Nef lacks enzymatic activity and an active site, complicating traditional occupancy-based drug development. Here we describe the development of proteolysis targeting chimeras (PROTACs) for the targeted degradation of Nef. Nef-binding compounds, based on an existing hydroxypyrazole core, were coupled to ligands for ubiquitin E3 ligases via flexible linkers. The resulting bivalent PROTACs induced formation of a ternary complex between Nef and the cereblon E3 ubiquitin ligase thalidomide-binding domain in vitro and triggered Nef degradation in a T cell expression system. Nef-directed PROTACs efficiently rescued Nef-mediated MHC-I and CD4 downregulation in T cells and suppressed HIV-1 replication in donor PBMCs. Targeted degradation is anticipated to reverse all HIV-1 Nef functions and may help restore adaptive immune responses against HIV-1 reservoir cells in vivo.

HIV-1 Nef辅助因子可增强体内病毒生命周期，促进HIV感染细胞的免疫逃逸，是一个有吸引力的抗逆转录病毒药物靶点。然而，Nef 缺乏酶活性和活性位点，使传统的基于占据的药物开发变得复杂。在这里，我们描述了用于靶向降解 Nef 的蛋白水解靶向嵌合体 (PROTAC) 的开发。 Nef 结合化合物基于现有的羟基吡唑核心，通过柔性接头与泛素 E3 连接酶的配体偶联。由此产生的二价 PROTAC在体外诱导 Nef 和 cereblon E3 泛素连接酶沙利度胺结合域之间形成三元复合物，并引发 T 细胞表达系统中的 Nef 降解。 Nef 定向的 PROTAC 有效地挽救了 T 细胞中 Nef 介导的 MHC-I 和 CD4 下调，并抑制了供体 PBMC 中的 HIV-1 复制。靶向降解有望逆转所有 HIV-1 Nef 功能，并可能有助于恢复体内针对 HIV-1 储存细胞的适应性免疫反应。