Exploring the role of disulfidptosis‑related signatures in immune microenvironment, prognosis and therapeutic strategies of cholangiocarcinoma

Cholangiocarcinoma (CCA) is an aggressive malignancy with poor prognosis and a limited number of treatments is available. Disulfidptosis, a newly identified form of cell death triggered by disulfide bond accumulation during glucose deprivation, may influence cancer progression but its role in CCA is poorly understood. The present study investigated disulfidptosis‑related genes (DRGs) and their impact on CCA prognosis and immune modulation. Differential expression analysis of 100 DRGs using RNA sequencing data from The Cancer Genome Atlas and EMBL‑EBI identified 74 dysregulated genes in CCA. Unsupervised clustering stratified patients with CCA into two distinct subtypes (Subs): i) SubA; and ii) SubB. A four‑gene prognostic signature was developed using least absolute shrinkage and selection operator regression and validated via Kaplan‑Meier survival analysis and receiver operating characteristic curves. Immune infiltration and tumor microenvironment were evaluated using Cell‑type Identification by Estimating Relative Subsets of RNA Transcripts, Estimation of Stromal and Immune cells in Malignant Tumor tissues using Expression data and single‑sample Gene Set Enrichment Analysis. Functional assays, including small interfering RNA knockdown of <i>CD109</i> and <i>EFNB2</i> in CCA cell lines were used to investigate proliferation, migration, invasion and F‑actin staining. Results showed SubB, associated with higher disulfidptosis activity, had worse prognosis, increased immune cell infiltration and elevated immune checkpoint gene expression. The four‑gene signature effectively stratified patients into risk groups. Knockdown of <i>CD109</i> and <i>EFNB2</i> significantly suppressed CCA cell proliferation, migration and invasion while it promoted disulfidptosis under glucose deprivation. The present study established an association between DRGs and CCA prognosis/immune dynamics, provided a robust four‑gene prognostic signature, and identified <i>CD109</i> and <i>EFNB2</i> as potential therapeutic targets, positioning disulfidptosis as a promising focus for precision medicine in CCA.