Gene Set Enrichment Analysis (GSEA) and single-GSEA (sGSEA)

GSEA was used to assess the data on genomic expression levels. Relative to the median expression of the hub genes, the 515 lung cancer samples from the RNA-seq data were divided into two groups, high-expression and low-expression samples. These two GSEA groups were used to identify the potential functions of the hub genes with the c5.all.v5.2.symbols.gmt annotations being selected as the reference gene sets. Nominal differences with p < 0.05, false discovery rate (FDR) < 0.05, and enrichment score (ES) > 0.6 were defined as the cutoff standards.

The only gene related to the gene sets from the MSigDB [25] that was identified in the study to correlate with metastasis and prognosis (RN7SL494P) was used to determine whether the sets showed statistical differences between the low-expression and high-expression categories. The analysis was performed using the java-dependent GSEA 3.0 software package [26].