Single-Cell Sequencing Exploration of Extrinsic Adaptive Resistance to PD-1 Inhibitors: Theory of Acupuncture-Associated Intervention in Advanced Non-Small Cell Lung Cancer

Vollständiger Abstract

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PD-1 and PD-L1 are respectively the receptor-ligand molecules on T cells and their target cells. The interaction between them can mediate immunosuppressive signals and induce immune escape of tumors. PD-1/PD-L1 inhibitors have revolutionized advanced non-small cell lung cancer (NSCLC) treatment, both primary resistance and acquired resistance persist due to the immunosuppressive tumor microenvironment and alternative immune checkpoints. Acupuncture shows immunomodulatory potential, but its synergy with PD-1 blockade remains unclear. To investigate the underlying mechanisms, we examined single-cell RNA sequencing profiles derived from NSCLC patients prior to and following PD-1 inhibitor treatment, utilizing the GSE207422 dataset. After quality control, 53,360 cells were retained. Cell types were annotated via unsupervised clustering, all cell types and T cell subsets were clustered, and T cells differentiation trajectories were reconstructed. Intercellular communication was assessed with CellChat, and a mechanistic framework for acupuncture’s synergistic effects was integrated from the literature. PD-1 inhibitor treatment remodeled the tumor microenvironment: T cells increased from 23.9% to 49.0%, while tumor cells decreased. CD8+ cytotoxic T cells expanded from 32.4% to 55.7% of T cells, whereas CD4+ naïve T cells declined from 67.1% to 42.7%. Pseudotime analysis indicated differentiation from CD4+ naïve toward CD8+ cytotoxic and CD8+ memory T cells. CellChat showed extensive T cell–myeloid interactions. Notably CD99-CD99 was highly expressed between T cells and tumor cells, suggesting an adaptive resistance mechanism. PD-1 inhibitors drive potent anti-tumor immunity in NSCLC, marked by CD8+ T cells expansion and tumor cells reduction, but tumor cells concurrently upregulate CD99-mediated compensatory suppression. We propose that acupuncture may synergize by enhancing T cell functional fitness, enabling T cells to resist CD99-PILRα inhibitory signaling, offering a promising strategy to overcome immunotherapy resistance.