MED12 mutation activates Notch signaling to enhance cancer stemness and suppress CD8+ T cell cytotoxicity in lung cancer

Abstract

BACKGROUND

Mediator complex subunit 12 (MED12) somatic mutations occur frequently in various tumors and are involved in tumor progression.

AIM

To evaluate the role and mechanism in lung cancer.

METHODS

MED12-associated genes were subjected to enrichment analysis. MED12-mutant NCI-H322 cells and MED12-non-mutant PC9 cells were used to evaluate the mutation’s effects on stemness, employing sphere formation, chromatin immunoprecipitation quantitative polymerase chain reaction, and flow cytometry. The association between MED12 and the tumor immune microenvironment was analyzed by integrating bulk transcriptomic and single-cell transcriptomic data. Chemotaxis and cytotoxic functions of CD8+ T cells were assessed using a co-culture model of tumor cells and CD8+ T cells.

RESULTS

Stem cell maintenance and Notch pathway regulation were identified as the main enriched categories for MED12-associated genes by bioinformatics analysis. Compared with non-mutant cells, MED12 mutation was associated with enhanced the sphere-forming ability, increased the ALDH1-positive cells and elevated Notch pathway activity. Treatment with the Notch inhibitor γ-secretase inhibitor (DAPT) partially reversed the observed effect of MED12 mutation on maintaining cancer stemness. Integration of bulk and single-cell transcriptomics revealed that MED12 expression is inversely associated with CD8+ T cell infiltration, with its differentially expressed genes being enriched in CD8+ T cell pathways, pointing to a potential immunomodulatory role. MED12-mutant tumor cells exhibited significantly higher surface programmed death ligand-1 expression than non-mutant cells, which was reversed by DAPT. In co-culture experiments using these cell lines, CD8+ T cells co-cultured with MED12-mutant lung cancer cells showed inhibited chemotaxis and reduced release of interferon-γ, tumor necrosis factor-α, perforin, and granzyme-B, while exhibiting significantly increasing programmed cell death 1. Furthermore, the killing capacity of CD8+ T cells against MED12-mutant lung cancer cells was attenuated. However, DAPT treatment partially reversed the MED12 mutation-induced suppression of CD8+ T cell function.

CONCLUSION

This study reveals the key role of the MED12 mutation in lung cancer progression, providing a new theoretical basis for lung cancer immunotherapy.

Keywords: Lung cancer; MED12 mutation; Stemness; CD8+ T; Notch signaling

Core Tip: Mediator complex subunit 12 mutation in lung cancer cells activates Notch signaling, which concurrently promotes cancer stemness - characterized by enhanced sphere formation and increased ALDH1+ cells - and induces immune evasion via upregulation of surface programmed death ligand-1 on tumor cells. This leads to impaired CD8+ T cell chemotaxis, reduced release of effector molecules (interferon-γ, tumor necrosis factor-α, perforin, granzyme-B), and increased programmed cell death 1 expression. Notch inhibitor DAPT reverses these pro-tumor effects.