Endocytosis and lysosomal trafficking of cell surface receptors can be triggered by endogenous ligands. Therapeutic approaches such as lysosome-targeting chimaeras^1,2 (LYTACs) and cytokine receptor-targeting chimeras³ (KineTACs) have used this to target specific proteins for degradation by fusing modified native ligands to target binding proteins. Although powerful, these approaches can be limited by competition with native ligands and requirements for chemical modification that limit genetic encodability and can complicate manufacturing, and, more generally, there may be no native ligands that stimulate endocytosis through a given receptor. Here we describe computational design approaches for endocytosis-triggering binding proteins (EndoTags) that overcome these challenges. We present EndoTags for insulin-like growth factor 2 receptor (IGF2R) and asialoglycoprotein receptor (ASGPR), sortilin and transferrin receptors, and show that fusing these tags to soluble or transmembrane target protein binders leads to lysosomal trafficking and target degradation. As these receptors have different tissue distributions, the different EndoTags could enable targeting of degradation to different tissues. EndoTag fusion to a PD-L1 antibody considerably increases efficacy in a mouse tumour model compared to antibody alone. The modularity and genetic encodability of EndoTags enables AND gate control for higher-specificity targeted degradation, and the localized secretion of degraders from engineered cells. By promoting endocytosis, EndoTag fusion increases signalling through an engineered ligand–receptor system by nearly 100-fold. EndoTags have considerable therapeutic potential as targeted degradation inducers, signalling activators for endocytosis-dependent pathways, and cellular uptake inducers for targeted antibody–drug and antibody–RNA conjugates.

细胞表面受体的内吞作用和溶酶体运输可由内源性配体触发。溶酶体靶向嵌合体^1,2 （LYTAC） 和细胞因子受体靶向嵌合体³ （KineTAC） 等治疗方法通过将修饰的天然配体融合到靶向结合蛋白来靶向特定蛋白进行降解。尽管功能强大，但这些方法可能会受到与天然配体的竞争和化学修饰要求的限制，这些限制了遗传可编码性，并使制造复杂化，并且更普遍地说，可能没有通过给定受体刺激内吞作用的天然配体。在这里，我们描述了克服这些挑战的内吞触发结合蛋白 （EndoTags） 的计算设计方法。我们提出了胰岛素样生长因子 2 受体 （IGF2R） 和去唾液酸糖蛋白受体 （ASGPR）、sortilin 和转铁蛋白受体的 EndoTags，并表明将这些标签与可溶性或跨膜靶蛋白结合物融合会导致溶酶体运输和靶标降解。由于这些受体具有不同的组织分布，因此不同的 EndoTag 可以靶向降解到不同的组织。与单独使用抗体相比，EndoTag 与 PD-L1 抗体的融合可显著提高小鼠肿瘤模型中的疗效。EndoTags 的模块化和遗传可编码性使 AND 门控制能够实现更高特异性的靶向降解，以及工程细胞中降解剂的局部分泌。通过促进内吞作用，EndoTag 融合通过工程配体-受体系统将信号转导增加近 100 倍。 EndoTags 具有相当大的治疗潜力，可作为靶向降解诱导剂、内吞依赖性途径的信号激活剂以及靶向抗体-药物和抗体-RNA 偶联物的细胞摄取诱导剂。