Identification and functional characterization of genes associated with lipopolysaccharide in lung adenocarcinoma.

BACKGROUND AND

objectivesLung cancer is one of the most common and deadliest cancers worldwide, with approximately 85% of cases classified as non-small cell lung cancer (NSCLC). Lung adenocarcinoma (LUAD) is the most prevalent subtype of NSCLC. Tumor-associated microbiota play an important role in the initiation, progression, and metastasis of tumors, but the underlying mechanisms remain unclear. Lipopolysaccharide (LPS), a major component of Gram-negative bacteria, is a key factor in inducing pathological responses. This study focuses on genes associated with lipopolysaccharide to explore the potential role of the microbiota in the pathogenesis of LUAD.

methodsGenes associated with lipopolysaccharide were identified using the Comparative Toxicogenomics Database (CTD). Gene expression data and clinical information for LUAD were obtained from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Univariate Cox regression analysis was performed to screen genes associated with lipopolysaccharide that were significantly associated with survival. These genes were further evaluated using Least Absolute Shrinkage and Selection Operator (LASSO) regression and multivariate Cox regression to construct a prognostic risk model. The model’s accuracy was validated using Kaplan-Meier (K-M) survival analysis, receiver operating characteristic (ROC) curves, and nomograms. We also examined the differences in immune cell infiltration, tumor mutational burden (TMB), and tumor immune dysfunction and exclusion (TIDE) between high- and low-risk groups.The expression of the key gene ITGB4 in LUAD tissues and cell lines was analyzed using qRT-PCR, WB, and IHC. The functional role of ITGB4 in LUAD cells was assessed through CCK-8, wound healing, Transwell assays, and flow cytometry. High-throughput sequencing was conducted on PC9 cells with ITGB4 knockdown to further investigate its mechanism of action. The role of ITGB4 was also validated in vivo using nude mouse xenograft models and the B6-KRASLSL−G12D orthotopic LUAD mouse model.

resultsFifty-five differentially expressed genes associated with lipopolysaccharide were initially identified, among which 25 demonstrated a significant association with LUAD patient prognosis. LASSO and multivariate Cox regression analyses ultimately identified four key genes that were used to construct a prognostic model for LUAD. The results from K-M survival analysis, ROC curves, and nomograms confirmed the model’s predictive accuracy. Analysis of the tumor immune microenvironment revealed significant differences in immune cell infiltration between high- and low-risk groups. Patients in the high-risk group exhibited higher TMB and TIDE scores than those in the low-risk group. Additionally, experimental validation demonstrated that ITGB4 was highly expressed in LUAD tissues and cell lines. Knockdown of ITGB4 inhibited LUAD cell proliferation, migration, and invasion, induced G0/G1 phase arrest, and promoted apoptosis. In vivo experiments further confirmed that ITGB4 knockdown significantly suppressed tumor growth.

conclusionMicrobiota may participate in the regulation of LUAD onset and progression through complex mechanisms. Genes associated with lipopolysaccharide provide a basis for constructing reliable prognostic models to predict outcomes in LUAD. Moreover, our study validated ITGB4 as a potential therapeutic target for LUAD at both cellular and animal levels, offering deeper insights into LUAD pathogenesis and valuable clues for therapeutic innovation.