The implications of TMSB4X in TIM3 hypermethylation and CD8⁺ T cell exhaustion in diffuse large B-cell lymphoma

Epigenetic modifications of immune checkpoints (IC) are crucial for immune checkpoint blockade (ICB) efficacy. TIM3 accelerates CD8⁺ T cell exhaustion in diffuse large B-cell lymphoma (DLBCL), yet its epigenetic regulation remains unclear. We analyzed CD8⁺ T cell localization and TIM3 expression using single-cell RNA sequencing (scRNA-seq) and immunofluorescence. DNA methylation analysis evaluated the epigenetic status of typical IC genes across TIM3 expression groups. Integrating genetic mutation, DNA methylation, scRNA-seq, and bulk RNA sequencing data identified a hub gene for TIM3 regulation. The biological and clinical significance of this hub gene, including impacts on immune activity, tumor microenvironment composition, clinicopathological features, prognosis, ICB response, and potential therapeutic options, was comprehensively investigated using scRNA-seq, bulk RNA sequencing data, and immunohistochemistry. Differentially methylated positions were identified in key exhaustion signature genes (TIM3, PD1, CTLA4, LAG3, STAT1) between TIM3-high and TIM3-low groups. Hypermethylation of TIM3's 3'UTR region correlated with transcriptional upregulation, suggesting positive regulation through TMSB4X loss, frequently altered by somatic mutation. Decreased TMSB4X expression is associated with epigenetic dysregulation, impaired immune function, aggressive clinicopathological features, poor prognosis, and inferior chemotherapy response. Patients with low TMSB4X may benefit from ICB therapy and certain epigenetic-targeting agents. TMSB4X links mutation and epigenetics in TIM3-induced CD8⁺ T cell exhaustion, providing a prognostic biomarker and potential therapeutic strategies for DLBCL.