Integrated multi-omics reveals glycolytic gene signatures of lung adenocarcinoma brain metastasis and the impact of Rac2 lactylation on immunosuppressive microenvironment.

PURPOSE

Tumor Metabolic Behavior modulates the immunosuppressive microenvironment through multiple pathways, thereby compromising anti-tumor immune responses. To date, there have been limited studies assessing the role of metabolic plasticity or immunometabolism in the tumor microenvironment (TME) during metastasis. Notably, emerging evidence suggests the presence of an immunosuppressive niche in brain metastases. This research aims to delineate distinct metabolic signatures in brain metastatic, investigate the impact of tumor-associated glycolysis on the development of brain metastases in lung adenocarcinoma, and characterize the lactylation regulation in this immunosuppressive microenvironment.

METHODS

The GSE131907 and GSE198291 datasets were retrieved for bioinformatic analysis. Combined with the results of proteomic and transcriptomic sequencing conducted on the lung adenocarcinoma brain metastasis model, differentially expressed signaling pathways were systematically identified through KEGG and GO functional annotations. A multimodal approach encompassing immunohistochemical (IHC) staining, immunofluorescence (IF) imaging, enzyme-linked immunosorbent assay (ELISA) quantification, and co-immunoprecipitation (Co-IP) assays was employed to experimentally validate the characteristics of the immunosuppressive microenvironment and the levels of tumor lactate/lactylation. Rescue experiments were performed by adding a lactylation-specific inhibitor (LDHi) or an H3K18la site-specific inhibitor. Finally, immunohistochemical staining was used to verify the expression level of H3K18la in clinical samples.

RESULTS

A total of 86,215 cells were extracted from the GSE131907 dataset, and the metabolic profiles of different cell types were analyzed. The results showed that glycolysis plays a dominant role in tumor cell metabolism. Further analysis revealed that early-stage primary lesions exhibit an inflammatory response signature, while advanced-stage primary lesions and brain metastatic lesions display an immunosuppressive signature. Elevated glycolytic flux showed a significant positive correlation with both the progression of brain metastasis and the immune evasion capacity of brain metastatic lesions. Pathological evaluation of tumor tissues from the LLC-BM (Lewis Lung Cancer Brain Metastasis) model confirmed its immunosuppressive characteristics. Additionally, obvious hypoxia was observed in the tumor tissues, accompanied by intratumoral vascular malformation and dysfunction. Significant lactate accumulation was present in the tumor microenvironment of LLC-BM tumors, and prominent lactylation modifications were detected in the tumor regions. In this model, Rac2 was identified as a potential core mediator of lactylation modification in macrophages, promoting the M2 polarization of macrophages. Meanwhile, CD40, TNFSF13 and CCL22 were identified as key immunoregulatory factors regulated by lactylation signaling. Notably, H3K18la was significantly highly expressed in lung cancer brain metastatic lesion samples.

CONCLUSIONS

The glycolytic pathway plays a critical role in the metabolic reprogramming of tumor cells during lung adenocarcinoma brain metastasis. Tumor glycolysis is closely associated with lung cancer progression, brain metastasis, and immune evasion. The Rac2 could be affected by lactylation, and then facilitate the formation of an immunosuppressive tumor microenvironment by induce the M2 polarization of macrophages.