The majority of therapies that target individual proteins rely on specific activity-modulating interactions with the target protein—for example, enzyme inhibition or ligand blocking. However, several major classes of therapeutically relevant proteins have unknown or inaccessible activity profiles and so cannot be targeted by such strategies. Protein-degradation platforms such as proteolysis-targeting chimaeras (PROTACs) 1 , 2 and others (for example, dTAGs 3 , Trim-Away 4 , chaperone-mediated autophagy targeting 5 and SNIPERs 6 ) have been developed for proteins that are typically difficult to target; however, these methods involve the manipulation of intracellular protein degradation machinery and are therefore fundamentally limited to proteins that contain cytosolic domains to which ligands can bind and recruit the requisite cellular components. Extracellular and membrane-associated proteins—the products of 40% of all protein-encoding genes 7 —are key agents in cancer, ageing-related diseases and autoimmune disorders 8 , and so a general strategy to selectively degrade these proteins has the potential to improve human health. Here we establish the targeted degradation of extracellular and membrane-associated proteins using conjugates that bind both a cell-surface lysosome-shuttling receptor and the extracellular domain of a target protein. These initial lysosome-targeting chimaeras, which we term LYTACs, consist of a small molecule or antibody fused to chemically synthesized glycopeptide ligands that are agonists of the cation-independent mannose-6-phosphate receptor (CI-M6PR). We use LYTACs to develop a CRISPR interference screen that reveals the biochemical pathway for CI-M6PR-mediated cargo internalization in cell lines, and uncover the exocyst complex as a previously unidentified—but essential—component of this pathway. We demonstrate the scope of this platform through the degradation of therapeutically relevant proteins, including apolipoprotein E4, epidermal growth factor receptor, CD71 and programmed death-ligand 1. Our results establish a modular strategy for directing secreted and membrane proteins for lysosomal degradation, with broad implications for biochemical research and for therapeutics. Lysosome-targeting chimaeras—in which a small molecule or antibody is connected to a glycopeptide ligand to form a conjugate that can bind a cell-surface lysosome-shuttling receptor and a protein target—are used to achieve the targeted degradation of extracellular and membrane proteins.

中文翻译：

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用于降解细胞外蛋白的溶酶体靶向嵌合体

大多数针对单个蛋白质的疗法依赖于与靶蛋白的特异性活性调节相互作用，例如酶抑制或配体阻断。然而，几类主要的治疗相关蛋白质具有未知或无法获得的活性谱，因此不能被这些策略靶向。蛋白质降解平台，例如蛋白水解靶向嵌合体 (PROTACs) 1、2 和其他（例如，dTAGs 3、Trim-Away 4、伴侣介导的自噬靶向 5 和 SNIPERs 6）已被开发用于通常难以处理的蛋白质目标; 然而，这些方法涉及对细胞内蛋白质降解机制的操作，因此从根本上仅限于含有细胞溶质结构域的蛋白质，配体可以结合和募集必要的细胞成分。细胞外和膜相关蛋白（40% 的蛋白质编码基因 7 的产物）是癌症、衰老相关疾病和自身免疫性疾病 8 的关键因子，因此选择性降解这些蛋白质的一般策略有可能改善人类健康。在这里，我们使用结合细胞表面溶酶体穿梭受体和靶蛋白的细胞外结构域的偶联物来确定细胞外和膜相关蛋白的靶向降解。这些最初的溶酶体靶向嵌合体，我们称之为 LYTAC，由与化学合成的糖肽配体融合的小分子或抗体组成，这些糖肽配体是阳离子非依赖性甘露糖 6-磷酸受体 (CI-M6PR) 的激动剂。我们使用 LYTAC 开发了一种 CRISPR 干扰筛选，该筛选揭示了 CI-M6PR 介导的细胞系货物内化的生化途径，并揭示了外囊复合物作为该途径的一个先前未识别但必不可少的组成部分。我们通过降解治疗相关蛋白（包括载脂蛋白 E4、表皮生长因子受体、CD71 和程序性死亡配体 1）证明了该平台的范围。我们的结果建立了一种模块化策略，用于指导分泌蛋白和膜蛋白进行溶酶体降解，具有广泛的对生化研究和治疗的影响。