Mutant TP53 Differentially Rewires FOXP3-Mediated Immune Checkpoint Regulation in a Cancer Lineage-Dependent Manner: A Comprehensive Pan-Cancer Computational Analysis

Background: TP53 is the most frequently mutated gene in human malignancy, yet its role in rewiring tumor-intrinsic immune checkpoint signaling via transcription factor intermediaries remains poorly characterized. FOXP3, canonically recognized as the master regulator of regulatory T cells (Tregs), is increasingly documented in tumor epithelial cells where it may directly regulate immune checkpoint gene expression. I hypothesized that mutant TP53 (mutp53) interferes with FOXP3-mediated immune checkpoint regulatory circuits in a cancer-cell-intrinsic manner shaped by histological lineage. Methods: I performed a multi-level computational analysis integrating RNA-sequencing (STAR-Counts), somatic mutation (MuTect2), and clinical data from 517 TCGA-LUAD primary tumours (246 mutp53 vs 271 TP53 wild-type). Differential expression was assessed using DESeq2 with apeglm shrinkage. Spearman correlation analysis evaluated FOXP3-checkpoint coupling stratified by TP53 status. Pan-cancer validation encompassed seven additional TCGA cohorts (total n=4,205 across 8 cohorts). Immune deconvolution using ssGSEA with Treg-adjusted partial correlation addressed tumor-intrinsic confounding. FOXP3 expression in tumor epithelial cells was confirmed using Cancer Cell Line Encyclopedia (CCLE/DepMap) data and published LUAD single-cell RNA sequencing (Kim et al. 2020; GSE131907; n=44,011 cells). Four independent GEO microarray datasets provided external validation (GSE68465 n=462, GSE72094 n=442, GSE30219 n=307, GSE31210 n=246; total n=1,457). Results: mutp53 significantly upregulated CD274/PD-L1 in LUAD (log2FC=0.53, padj<0.0001; 365 total DEGs). Pan-cancer Spearman correlation analysis revealed a striking lineage-specific pattern: mutp53 reduced FOXP3-checkpoint coupling in 67% of adenocarcinoma cohort-gene pair comparisons (10/15), while paradoxically enhancing it in all squamous (HNSC), urothelial (BLCA), and hepatocellular (LIHC) carcinoma cohorts (0/9 pairs supported). FOXP3-CD274 delta-rho=+0.083, FOXP3-CTLA4 delta-rho=+0.054 in LUAD. Immune deconvolution confirmed persistence of FOXP3-checkpoint associations after Treg adjustment. CCLE data confirmed FOXP3 expression in lung adenocarcinoma cell lines absent immune cells. scRNA-seq data revealed 46.9% of EPCAM+ tumor epithelial cells express FOXP3 detectably. FOXP3 proxy validation against direct TP53 annotation achieved 100% agreement in GSE68465 (n=462). GEO validation corroborated discovery findings. Survival analysis showed numerically inferior outcomes in the mutp53/FOXP3-high subgroup, consistent with the hypothesis direction. Conclusions: mutp53 differentially rewires FOXP3-mediated immune checkpoint regulation in a cancer lineage-dependent manner, disrupting the FOXP3-checkpoint axis in adenocarcinomas while enhancing it in non-adenocarcinoma histologies. Multi-level evidence from TCGA, CCLE, scRNA-seq, and four independent GEO cohorts supports a tumor-cell-intrinsic mechanism, providing a novel framework for differential checkpoint immunotherapy responses across cancer types.