肿瘤来源的小细胞外囊泡 (sEV) 上的程序性死亡配体 1 (PD-L1) 通过与宿主免疫细胞上的 PD-1 受体相互作用来限制治疗效果。靶向 sEV PD-L1 的分泌已成为增强免疫治疗的一种有前景的策略。然而，小分子抑制剂的缺乏给临床转化带来了挑战。在本研究中，我们开发了一种靶标和表型双驱动高通量筛选（TAP-HTS）策略，该策略将虚拟筛选与基于纳流的实验验证相结合。我们确定布洛芬 (IBP) 是一种有效靶向 sEV PD-L1 分泌的新型抑制剂。 IBP 通过与 sEV 生物发生的关键调节剂、肝细胞生长因子调节的酪氨酸激酶底物 (HRS) 发生物理相互作用，破坏肿瘤细胞中 PD-L1 sEV 的生物发生和分泌。值得注意的是，IBP 的作用机制与其众所周知的靶标环加氧酶（COX1 或 COX2）不同。在黑色素瘤和口腔鳞状细胞癌 (OSCC) 小鼠模型中，IBP 的施用可刺激抗肿瘤免疫并增强抗 PD-1 疗法的疗效。为了解决潜在的副作用，我们进一步开发了一种用于局部应用的 IBP 凝胶，与抗 PD-1 治疗相结合时显示出显着的治疗效果。这种特定小抑制剂的发现为建立持久、系统性抗肿瘤免疫提供了一条有希望的途径。版权所有 © 2024。由 Elsevier Inc. 出版。

Programmed death-ligand 1 (PD-L1) on tumor-derived small extracellular vesicles (sEVs) limits therapeutic effectiveness by interacting with the PD-1 receptor on host immune cells. Targeting the secretion of sEV PD-L1 has emerged as a promising strategy to enhance immunotherapy. However, the lack of small-molecule inhibitors poses a challenge for clinical translation. In this study, we developed a target and phenotype dual-driven high-throughput screening (TAP-HTS) strategy that combined virtual screening with nanoflow-based experimental verification. We identified ibuprofen (IBP) as a novel inhibitor that effectively targeted sEV PD-L1 secretion. IBP disrupted the biogenesis and secretion of PD-L1+ sEVs in tumor cells by physically interacting with a critical regulator of sEV biogenesis, hepatocyte growth factor-regulated tyrosine kinase substrate (HRS). Notably, the mechanism of action of IBP is distinct from its commonly known targets, cyclooxygenases (COX1 or COX2). Administration of IBP stimulated antitumor immunity and enhanced the efficacy of anti-PD-1 therapy in melanoma and oral squamous cell carcinoma (OSCC) mouse models. To address potential adverse effects, we further developed an IBP gel for topical application, which demonstrated remarkable therapeutic efficacy when combined with anti-PD-1 treatment. The discovery of this specific small inhibitor provides a promising avenue for establishing durable, systemic antitumor immunity.Copyright © 2024. Published by Elsevier Inc.