Tumor-cell NY-ESO-1 expression predicts poor response to PD-1/PD-L1 blockade in NSCLC

Background: NY-ESO-1 is an immunogenic cancer-testis antigen and a therapeutic target for TCR-based strategies. Its significance as a baseline biomarker in non-small cell lung cancer (NSCLC) treated with PD-1/PD-L1 inhibitors is not well established. Because NY-ESO-1 has biological relevance only when expressed by tumor epithelial cells, we focused our analysis specifically on NY-ESO-1 expression within CK7⁺ tumor cells and evaluated its association with clinical outcomes in patients receiving immune checkpoint blockade. Methods: Baseline FFPE tumor samples from 195 patients with advanced NSCLC treated with PD-1/PD-L1 inhibitors were profiled using a validated multiplex immunofluorescence panel including CD8, CD68, CD103, CK7, HLA-ABC, cMAF, NY-ESO-1, and pSMAD3. Digital image analysis was performed using Vectra Polaris imaging, QuPath segmentation, and FlowJo-based phenotyping after signal normalization. Tumor-specific NY-ESO-1 expression was defined by CK7⁺/NYESO1⁺ co-localization. Associations with RECIST response, durable clinical benefit (DCB), progression-free survival (PFS), and overall survival (OS) were assessed using Wilcoxon tests, chi-square tests, and cutpoint-based survival modeling. CK7→CD8 and CK7→CD68 intercellular distances were also computed. Results: Higher CK7⁺/NY-ESO-1⁺ tumor cell levels were significantly associated with non-response to PD-1/PD-L1 blockade (p=0.011), and increased NY-ESO-1 expression intensity also associated with non-response (p=0.044). Tumor-restricted NY-ESO-1 was strongly predictive of shorter survival. Patients with high CK7⁺/NY-ESO-1⁺ tumor expression had a median PFS of 1.8 months, compared with 8.1 months in the low-expression group (p=7.2×10⁻⁵). Median OS was similarly reduced, with 11.0 months in the high-expression group versus 33.5 months in the low-expression group (p=3.9×10⁻³). High tumor NY-ESO-1 also correlated with substantially reduced durable clinical benefit (χ² p=0.013). Independently of NY-ESO-1, greater CK7→CD8 and CK7→CD68 distances predicted inferior PFS and OS, although no direct association between NY-ESO-1 levels and spatial distances was assessed. Conclusions: Tumor-cell NY-ESO-1 expression is a robust adverse biomarker in NSCLC treated with PD-1/PD-L1 inhibitors. High CK7⁺/NY-ESO-1⁺ tumor burden identifies patients with dramatically reduced PFS and OS, lower response rates, and poor likelihood of achieving durable clinical benefit. These findings support the role of tumor-restricted NY-ESO-1 as a clinically meaningful negative baseline predictor of immunotherapy outcome and may help refine patient selection for NY-ESO-1-targeted therapies.