Extracellular vesicles (EVs) can be functionalized to display specific protein receptors on their surface. However, surface-display technology typically labels only a small fraction of the EV population. Here, we show that the joint display of two different therapeutically relevant protein receptors on EVs can be optimized by systematically screening EV-loading protein moieties. We used cytokine-binding domains derived from tumour necrosis factor receptor 1 (TNFR1) and interleukin-6 signal transducer (IL-6ST), which can act as decoy receptors for the pro-inflammatory cytokines tumour necrosis factor alpha (TNF-α) and IL-6, respectively. We found that the genetic engineering of EV-producing cells to express oligomerized exosomal sorting domains and the N-terminal fragment of syntenin (a cytosolic adaptor of the single transmembrane domain protein syndecan) increased the display efficiency and inhibitory activity of TNFR1 and IL-6ST and facilitated their joint display on EVs. In mouse models of systemic inflammation, neuroinflammation and intestinal inflammation, EVs displaying the cytokine decoys ameliorated the disease phenotypes with higher efficacy as compared with clinically approved biopharmaceutical agents targeting the TNF-α and IL-6 pathways.

细胞外囊泡 (EV) 可以功能化以在其表面显示特定的蛋白质受体。然而，表面显示技术通常只标记一小部分电动汽车。在这里，我们表明可以通过系统地筛选 EV 加载蛋白部分来优化 EV 上两种不同治疗相关蛋白受体的联合展示。我们使用了源自肿瘤坏死因子受体 1 (TNFR1) 和白细胞介素 6 信号转导 (IL-6ST) 的细胞因子结合域，它们可以作为促炎细胞因子肿瘤坏死因子 α (TNF-α) 的诱饵受体和IL-6，分别。我们发现，表达寡聚化外泌体分选域和 syntenin（单跨膜域蛋白 syndecan 的胞质衔接子）N 端片段的 EV 产生细胞的基因工程提高了 TNFR1 和 IL-6ST 的展示效率和抑制活性并促进了他们在电动汽车上的联合展示。在全身性炎症、神经炎症和肠道炎症的小鼠模型中，与临床批准的靶向 TNF-α 和 IL-6 通路的生物药剂相比，显示细胞因子诱饵的 EV 改善了疾病表型，且疗效更高。