Tumour-Infiltrating Lymphocyte Subsets, PD-L1 Expression, HPV/p16 Status, and Neutrophil-to-Lymphocyte Ratio in Oral Squamous Cell Carcinoma: A Longitudinal Stratified Study of Non-Recurrent and Recurrent Disease

<title>Abstract</title> <p>Oral squamous cell carcinoma (OSCC) imposes a disproportionate epidemiological burden in India, yet lacks validated predictive biomarkers for immune-based therapies. We characterised the tumour immune microenvironment (TME) by evaluating tumour-infiltrating lymphocyte (TIL) subsets (CD3+, CD4+, CD8+, FOXP3+ and CD57+), PD-L1 expression, HPV/p16 status and the systemic peripheral neutrophil-to-lymphocyte ratio (NLR) in resection specimens from 299 patients (stratified by disease outcome: non-recurrent, n=106; recurrent, n=193) treated at two tertiary oncology centres in Bangalore and Cuttack, India, between 2020 and 2023. TIL subsets were assessed by immunohistochemistry; PD-L1 was scored using the Dako 22C3 assay with a cut-off of …., and p16 was interpreted per College of American Pathologists guidelines. FOXP3+ regulatory T cells showed the broadest association with adverse histopathology, including lymphovascular invasion (LVI; P &lt; 0.001), worst pattern of invasion (WPOI; P = 0.038), and lymph node metastasis (P = 0.003). PD-L1 expression correlated positively with all five TIL subsets (P &lt; 0.001 for each), defining an “inflamed but suppressed” TME. p16-positive tumours (4.0%, associated with recurrent patients) showed markedly reduced FOXP3+ infiltration (P &lt; 0.001). In multivariate regression, only LVI and WPOI were associated with elevated pre-treatment log-NLR. The immune microenvironment was statistically indistinguishable between non-recurrent and recurrent tumours across all immune parameters. This integrated multi-biomarker framework — combining intratumoral TIL subset profiling, PD-L1 scoring, HPV/p16 status, and systemic NLR — demonstrates practical utility for immunotherapy patient stratification in South Indian OSCC. Notably, in our study, this utility operates independently of recurrence prediction, suggesting immunotherapy eligibility is a distinct biological dimension from recurrence risk</p>