Loss of CLIC3 in CD8+ cytotoxic T lymphocytes is associated with immune dysfunction and poor prognosis in lung adenocarcinoma

Background Lung adenocarcinoma (LUAD) is the most common histological subtype of lung cancer, with a dismal prognosis partly attributed to immune evasion mediated by T cell dysfunction within the tumor microenvironment. However, the molecular determinants governing CD8 + cytotoxic T lymphocyte functional competence remain incompletely understood. Chloride intracellular channel 3 (CLIC3) has been implicated in cancer progression, but its role in T cell immunity within the LUAD TME is unknown. Methods We performed single-cell RNA sequencing on tumor tissues from nine LUAD patients spanning three pathological stages (adenocarcinoma in situ [AIS], minimally invasive adenocarcinoma [MIA], and invasive adenocarcinoma [IAC]), integrated with spatial transcriptomics, TCGA-LUAD bulk RNA-seq analysis, and flow cytometric validation in an independent cohort of 32 LUAD patients. Functional module scoring and pseudotime trajectory analysis were applied to characterize CLIC3-associated CTL functional states. Results CLIC3 was predominantly expressed in CTLs and showed a stage-associated decline from AIS to MIA to IAC. CLIC3 + CTLs exhibited significantly higher module scores for cytotoxicity, cytoskeleton remodeling, interferon signaling, and chemotaxis compared to CLIC3- CTLs. Pseudotime trajectory analysis indicated that CLIC3 + cells were positioned at one end of the inferred transcriptional continuum, corresponding to a functionally active state, while CLIC3 - cells were positioned at the other, corresponding to a dysfunctional state, accompanied by a decline in CD247, GZMB, PRF1, and TNF along the trajectory. High CLIC3 expression in TCGA-LUAD was associated with significantly longer overall survival. Flow cytometry showed a stage-associated decrease in CLIC3 expression on CD8 + T cells with increasing tumor invasiveness and poor differentiation. Conclusions CLIC3 serves as a novel marker of CTL functional competence in LUAD, and its stage-associated downregulation reflects T cell dysfunction during tumor progression. CLIC3 expression in CD8 + T cells may represent a predictive biomarker for immunotherapeutic response and a potential target for restoring T cell function in LUAD.