Spatial heterogeneity of PD-L1 expression influence its assessment in esophageal squamous cell carcinoma

Immune checkpoint inhibitors are a promising treatment for esophageal squamous cell carcinoma (ESCC). However, the predictive value of programmed death-ligand 1 (PD-L1) expression, the most common biomarker for immunotherapy, remains controversial, particularly in the neoadjuvant setting. We hypothesized that the spatial heterogeneous of PD-L1 expression within tumors might render limited biopsy samples unrepresentative of the bulk tumor. In this study, we assessed the spatial heterogeneity in PD-L1 expression within ESCC by sampling four distinct regions using endoscopic biopsy forceps and the largest longitudinal sections on complete resected tumor from treatment-naïve patients. Our findings demonstrated the insufficiency of using limited biopsy tumor tissue to accurately determine the combined positive score (CPS) within the tumor. Notably, spatial heterogeneity was reduced when tumor’s CPS was sufficiently high. Multi-region sampling assessment revealed that the maximum CPS derived from three regions provided a more accurate approximation of the bulk tumor's PD-L1 status. Additionally, the densities of CD8⁺/CD4⁺ T cells were positively correlated with CPS. These findings emphasize the clinical need for standardized and modified biopsy assessment strategies to improve the accuracy of PD-L1 evaluation, thereby guiding therapeutic decision-making in ESCC.