Design, Synthesis, and Evaluation of <i>o</i>‑(Biphenyl-3-ylmethoxy)nitrophenyl Derivatives as PD-1/PD-L1 Inhibitors with Potent Anticancer Efficacy <i>In Vivo</i>

Two series of novel o-(biphenyl-3-ylmethoxy)­nitrophenyl compounds (A1–31 and B1–17) were designed as programmed cell death protein 1 (PD-1)/PD-ligand 1 (PD-L1) inhibitors. All compounds showed significant inhibitory activity with IC50 values ranging from 2.7 to 87.4 nM except compound A17, and compound B2 displayed the best activity. Further experiments showed that B2 bound to the PD-L1 protein without obvious toxicity in Lewis lung carcinoma (LLC) cells. Furthermore, B2 significantly promoted interferon-gamma secretion in a dose-dependent manner in vitro and in vivo. Especially, B2 exhibited potent in vivo anticancer efficacy in an LLC-bearing allograft mouse model at a low dose of 5 mg/kg, which was more active than BMS-1018 (tumor growth inhibition rate: 48.5% vs 17.8%). A panel of immunohistochemistry and flow cytometry assays demonstrated that B2 effectively counteracted PD-1-induced immunosuppression in the tumor microenvironment, thereby triggering antitumor immunity. These results indicate that B2 is a promising PD-1/PD-L1 inhibitor worthy of further development.两个系列的新型邻联苯-3-基甲氧基硝基苯基化合物（A1–31与B1–17）被设计为程序性死亡蛋白1（programmed cell death protein 1, PD-1）/PD-配体1（PD-L1）抑制剂。除化合物A17外，所有化合物均表现出显著的抑制活性，其半最大抑制浓度（IC50）范围为2.7~87.4 nM，其中化合物B2的活性最优。进一步实验表明，B2可与PD-L1蛋白结合，且在路易斯肺癌（LLC）细胞中无明显细胞毒性。此外，B2在体外与体内实验中均能以剂量依赖方式显著促进干扰素-γ的分泌。尤为突出的是，在5 mg/kg的低剂量下，B2在荷路易斯肺癌同种异体移植小鼠模型中展现出强效的体内抗肿瘤活性，其效果优于BMS-1018（二者肿瘤生长抑制率分别为48.5%与17.8%）。一系列免疫组织化学与流式细胞术实验证实，B2可有效拮抗PD-1介导的肿瘤微环境免疫抑制，从而激活抗肿瘤免疫应答。上述结果表明，B2是一款极具开发潜力的PD-1/PD-L1抑制剂，值得开展进一步的研发工作。