There is a lack of effective treatments to overcome resistance to EGFR-TKIs in EGFR mutant tumors. A deeper understanding of resistance mechanisms can provide insights into reducing or eliminating resistance, and can potentially deliver targeted treatment measures to overcome resistance. Here, we identified that the dynamic changes of the tumor immune environment were important extrinsic factors driving tumor resistance to EGFR-TKIs in EGFR mutant cell lines and syngeneic tumor-bearing mice. Our results demonstrate that the acquired resistance to EGFR-TKIs is accompanied by aberrant expression of PD-L2, leading a dynamic shift from an initially favorable tumor immune environment to an immunosuppressive phenotype. PD-L2 expression significantly affected EGFR mutant cell apoptosis that depended on the proportion and function of CD8⁺ T cells in the tumor immune environment. Combined with single-cell sequencing and experimental results, we demonstrated that PD-L2 specifically inhibited the proliferation of CD8⁺ T cells and the secretion of granzyme B and perforin, leading to reduced apoptosis mediated by CD8⁺ T cells and enhanced immune escape of tumor cells, which drives EGFR-TKIs resistance. Importantly, we have identified a potent natural small-molecule inhibitor of PD-L2, zinc undecylenate. In vitro, it selectively and potently blocks the PD-L2/PD-1 interaction. In vivo, it abolishes the suppressive effect of the PD-L2-overexpressing tumor immune microenvironment by blocking PD-L2/PD-1 signaling. Moreover, the combination of zinc undecylenate and EGFR-TKIs can synergistically reverse tumor resistance, which is dependent on CD8⁺ T cells mediating apoptosis. Our study uncovers the PD-L2/PD-1 signaling pathway as a driving factor to mediate EGFR-TKIs resistance, and identifies a new naturally-derived agent to reverse EGFR-TKIs resistance.

缺乏有效的治疗方法来克服 EGFR 突变肿瘤对 EGFR-TKI 的耐药性。对耐药机制的更深入了解可以为减少或消除耐药性提供见解，并有可能提供有针对性的治疗措施来克服耐药性。在这里，我们发现肿瘤免疫环境的动态变化是驱动EGFR突变细胞系和同基因荷瘤小鼠肿瘤对EGFR-TKI产生耐药性的重要外在因素。我们的结果表明，对 EGFR-TKI 的获得性耐药伴随着 PD-L2 的异常表达，导致从最初有利的肿瘤免疫环境动态转变为免疫抑制表型。 PD-L2表达显着影响EGFR突变细胞凋亡，其依赖于肿瘤免疫环境中CD8 ⁺ T细胞的比例和功能。结合单细胞测序和实验结果，我们证明PD-​​L2特异性抑制CD8 ⁺ T细胞的增殖以及颗粒酶B和穿孔素的分泌，从而减少CD8 ⁺ T 细胞和增强的肿瘤细胞免疫逃逸，从而驱动 EGFR-TKI 耐药。重要的是，我们已经鉴定出一种有效的天然 PD-L2 小分子抑制剂，十一烯酸锌。在体外，它选择性地有效阻断 PD-L2/PD-1 相互作用。在体内，它通过阻断 PD-L2/PD-1 信号传导来消除 PD-L2 过表达的肿瘤免疫微环境的抑制作用。此外，十一烯酸锌与EGFR-TKIs联合可以协同逆转肿瘤耐药，而肿瘤耐药依赖于CD8 ⁺ T细胞介导的细胞凋亡。 我们的研究揭示了 PD-L2/PD-1 信号通路作为介导 EGFR-TKI 耐药的驱动因素，并确定了一种新的天然衍生药物来逆转 EGFR-TKI 耐药。