USP20 as a key regulator of immunosuppression and a novel predictor of poor prognosis in lung adenocarcinoma

To systematically delineate the expression landscape, genetic alterations, prognostic value, and immunological relevance of ubiquitin-specific protease 20 (USP20) in lung adenocarcinoma (LUAD). By integrating bulk and single-cell transcriptomic analyses, we aimed to identify the cellular sources of USP20 and clarify its potential roles in remodeling the tumor immune microenvironment (TME), thereby evaluating its utility as a prognostic biomarker and candidate immunotherapeutic target. USP20 expression was profiled across The Cancer Genome Atlas (TCGA), Genotype-Tissue Expression (GTEx), and multiple Gene Expression Omnibus (GEO) cohorts. Prognostic significance was assessed using Kaplan-Meier analysis and Cox proportional hazards models. USP20 genetic alterations were interrogated via cBioPortal. Differentially expressed genes (DEGs) associated with USP20 were identified, followed by GO/KEGG enrichment and GSEA to infer biological functions. TME characteristics were evaluated using ESTIMATE and CIBERSORT, and associations with immune checkpoint molecules were examined. Consensus clustering was performed to define USP20-related molecular subtypes and compare their immune landscapes. Single-cell RNA-seq datasets (TISCH and GSE146100) were integrated to resolve USP20 expression across tumor and immune compartments. Finally, immunohistochemistry, multiplex immunofluorescence, flow cytometry, and in vitro/in vivo functional assays were conducted to validate the biological and immunoregulatory roles of USP20 in LUAD. USP20 was consistently upregulated in LUAD relative to normal lung tissues across independent cohorts. High USP20 expression was associated with inferior overall survival and remained an independent prognostic risk factor. Genetic profiling showed a low mutation rate of USP20, with copy number amplification as the predominant alteration. Functional analyses indicated that USP20-associated DEGs were enriched in immune-related programs, including T-cell activation, antigen processing and presentation, interferon signaling, and immune checkpoint regulation. High USP20 expression correlated with a distinct immune contexture, featuring reshaped immune cell composition, increased infiltration of immunosuppressive populations, and elevated expression of multiple immune checkpoint molecules. Consensus clustering further revealed USP20-related molecular subtypes with divergent immunosuppressive features and clinical outcomes. Single-cell analyses demonstrated heterogeneous and cell-type-specific USP20 expression across malignant and immune populations, supporting a context-dependent immunomodulatory role. Experimental validation confirmed that USP20 promotes LUAD cell proliferation and migration and is associated with immune dysregulation within the TME. USP20 is aberrantly overexpressed in LUAD and portends poor prognosis. USP20 may facilitate disease progression by shaping tumor-immune interactions, including immune infiltration patterns, T-cell activation/exhaustion states, and immune checkpoint signaling. USP20 and its related molecular subtypes may serve as prognostic biomarkers and potential indicators of immunotherapeutic responsiveness, offering insights for precision immunotherapy in LUAD.