The combination of PD-1 blockade with neoadjuvant chemotherapy (NAC) has achieved unprecedented clinical success in non-small cell lung cancer (NSCLC) compared to NAC alone, but the underlying mechanisms by which PD-1 blockade augments the effects of chemotherapy remain incompletely elucidated. Single-cell RNA sequencing was performed on CD45⁺ immune cells isolated from surgically resected fresh tumors of seven NSCLC patients receiving NAC or neoadjuvant pembrolizumab and chemotherapy (NAPC). Multiplex fluorescent immunohistochemistry was performed on FFPE tissues before and after NAC or NAPC from 65 resectable NSCLC patients, and results were validated with GEO dataset. NAC resulted in an increase only of CD20⁺ B cells, whereas NAPC increased the infiltration of CD20⁺ B cells, CD4⁺ T cells, CD4⁺CD127⁺ T cells, CD8⁺ T cells, CD8⁺CD127⁺ and CD8⁺KLRG1⁺ T cells. Synergistic increase in B and T cells promotes favorable therapeutic response after NAPC. Spatial distribution analysis discovered that CD8⁺ T cells and their CD127⁺ and KLRG1⁺ subsets were in closer proximity to CD4⁺ T/CD20⁺ B cells in NAPC versus NAC. GEO dataset validated that B-cell, CD4, memory, and effector CD8 signatures correlated with therapeutic responses and clinical outcomes. The addition of PD-1 blockade to NAC promoted anti-tumor immunity through T and B cells recruitment in the tumor microenvironment and induced tumor-infiltrating CD8⁺ T cells skewed toward CD127⁺ and KLRG1⁺ phenotypes, which may be assisted by CD4⁺ T cells and B cells. Our comprehensive study identified key immune cell subsets exerting anti-tumor responses during PD-1 blockade therapy and that may be therapeutically targeted to improve upon existing immunotherapies for NSCLC.

与单独使用 NAC 相比，PD-1 阻断联合新辅助化疗 (NAC) 在非小细胞肺癌 (NSCLC) 中取得了前所未有的临床成功，但 PD-1 阻断增强化疗效果的潜在机制仍不完全阐明。^{对从接受 NAC 或新辅助 pembrolizumab 和化疗 (NAPC) 的 7 名 NSCLC 患者手术切除的新鲜肿瘤中分离出的 CD45 +}免疫细胞进行单细胞 RNA 测序。对来自 65 名可切除 NSCLC 患者的 NAC 或 NAPC 前后的 FFPE 组织进行多重荧光免疫组织化学，并使用 GEO 数据集验证结果。NAC 仅导致 CD20 ^{+ B 细胞的增加，而 NAPC 增加了 CD20 +}的浸润B 细胞、CD4 ⁺ T 细胞、CD4 ⁺ CD127 ⁺ T 细胞、CD8 ⁺ T 细胞、CD8 ⁺ CD127 ⁺和 CD8 ⁺ KLRG1 ⁺ T 细胞。B 细胞和 T 细胞的协同增加促进了 NAPC 后良好的治疗反应。空间分布分析发现，CD8 ⁺ T 细胞及其 CD127 ⁺和 KLRG1 ⁺亚群更接近于 CD4 ⁺ T/CD20 ⁺NAPC 与 NAC 中的 B 细胞。GEO 数据集验证了 B 细胞、CD4、记忆和效应 CD8 特征与治疗反应和临床结果相关。在 NAC 中加入 PD-1 阻断剂可通过肿瘤微环境中的 T 细胞和 B 细胞募集促进抗肿瘤免疫，并诱导肿瘤浸润性 CD8 ⁺ T 细胞偏向 CD127 ⁺和 KLRG1 ⁺表型，这可能由 CD4 ⁺ T辅助细胞和 B 细胞。我们的综合研究确定了在 PD-1 阻断治疗期间发挥抗肿瘤反应的关键免疫细胞亚群，并且可能在治疗上有针对性地改善现有的 NSCLC 免疫疗法。