E3泛素连接酶RFFL是一种高表达在癌症中的凋亡抑制剂，它的沉默抑制了肿瘤细胞的生长并增强对化疗的敏感性。RFFL还参与外周蛋白质质量控制，去除了功能性细胞表面ΔF508-CFTR通道并减少了囊性纤维化（CF）的药物治疗的功效。尽管RFFL抑制剂在肿瘤和CF的治疗潜力，但是它们仍然没有被发现。在这里，化学阵列筛选发现α-生育酚琥酸酯(αTOS)是RFFL的配体。NMR分析揭示αTOS直接结合到RFFL的底物结合区域，而不影响E3酶活性。因此，αTOS抑制了RFFL-底物之间的相互作用，ΔF508-CFTR泛素化并从上皮细胞的细胞膜上消失，从而增加了功能性CFTR通道。在我们测试的α-生育酚（αTOL）类似物中，只有αTOS抑制了RFFL-底物之间的相互作用并增加了细胞表面的ΔF508-CFTR，并且这种效应取决于RFFL的表达。类似地，αTOS的独特促凋亡效应也依赖于RFFL的表达。因此，与其他αTOL类似物不同，αTOS作为一种RFFL蛋白与蛋白相互作用的抑制剂可能解释了它在αTOL类似物中的独特生物学性质。此外，αTOS可能作为一种CFTR稳定剂，这是一类延长细胞表面ΔF508-CFTR寿命的新型药物。版权所有 © 2023 Elsevier Inc.

The E3 ubiquitin ligase RFFL is an apoptotic inhibitor highly expressed in cancers and its knockdown suppresses cancer cell growth and sensitizes to chemotherapy. RFFL also participates in peripheral protein quality control which removes the functional cell surface ΔF508-CFTR channel and reduces the efficacy of pharmaceutical therapy for cystic fibrosis (CF). Although RFFL inhibitors have therapeutic potential for both cancer and CF, they remain undiscovered. Here, a chemical array screening has identified α-tocopherol succinate (αTOS) as an RFFL ligand. NMR analysis revealed that αTOS directly binds to RFFL's substrate-binding region without affecting the E3 enzymatic activity. Consequently, αTOS inhibits the RFFL-substrate interaction, ΔF508-CFTR ubiquitination and elimination from the plasma membrane of epithelial cells, resulting in the increased functional CFTR channel. Among the α-tocopherol (αTOL) analogs we tested, only αTOS inhibited the RFFL-substrate interaction and increased the cell surface ΔF508-CFTR, depending on RFFL expression. Similarly, the unique proapoptotic effect of αTOS was dependent on RFFL expression. Thus, unlike other αTOL analogs, αTOS acts as an RFFL protein-protein interaction inhibitor which may explain its unique biological properties among αTOL analogs. Moreover, αTOS may act as a CFTR stabilizer, a novel class of drugs that extend cell surface ΔF508-CFTR lifetime.Copyright © 2023. Published by Elsevier Inc.