Identification of lymph node metastasis-related genes and patterns of immune infiltration in colon adenocarcinoma

Abstract

Backgrounds: Colon adenocarcinoma(COAD) is one of the most common tumors of the digestive tract. Lymph node metastasis (LNM) is a well-established prognostic factor for COAD. The mechanism of COAD lymph node metastasis in immunology remains unknown. The identification of LNM-related biomarkers of COAD could help in its treatment. Thus, the current study was aimed to identify key genes and construct a prognostic signature.

Methods: Gene expression and clinical data were obtained from The Cancer Genome Atlas (TCGA) database. Differentially expressed genes were calculated by using R software. GO functional and KEGG pathway enrichment analysis were processed. The CIBERSORT algorithm was used to assess immune cell infiltration. STRING database was used to screen key genes and constructed a protein-protein interaction network (PPI network). The LASSO-Cox regression analysis was performed based on the components of the PPI network. The correlation analysis between LNM-related signature and immune infiltrating cells was then investigated. TISIDB was used to explore the correlation between the abundance of immunomodulators and the expression of the inquired gene.

Results: In total, 394 differentially expressed genes were identified. After constructing and analyzing the PPI network, 180 genes were entered into the LASSO-Cox regression model, constructing a gene signature. Five genes(PMCH, LRP2, NAT1, NKAIN4, and CD1B) were identified as LNM-related genes of clinical value. Correlation analysis revealed that LRP2 and T follicular helper cells (R=0.34, P=0.0019) and NKAIN4 and T follicular helper cells (R=0.23, P=0.041) had significant correlations. Immunologic analysis revealed that LRP2 and NKAIN4 are potential coregulators of immune checkpoints in COAD.

Conclusion: In general, this study revealed the key genes related to lymph node metastasis and prognostic signature. Several potential mechanisms and therapeutic and prognostic targets of lymph node metastasis were also demonstrated in COAD.

Keywords: TCGA; bioinformatic analysis; colon adenocarcinoma; immune cell infiltration; lymph node metastasis.

Figure 1

The flow diagram of the present study. COAD, colon adenocarcinoma; DEGs, differentially expressed genes; TCGA, The Cancer Genome Atlas; PPI, ptotein-protein interaction;.

Figure 2

DEGs. Hierarchical clustering analysis of genes (A); Volcano plots were constructed using fold-change values and adjusted P (B); Red dots represent upregulated genes, and green dots represent downregulated genes. The GO analyses (C); KEGG pathway analyses (D); PPI network of differentially expressed genes (E). DEGs, differentially expressed genes; COAD, colon adenocarcinoma.

Figure 3

The PPI network signature. The PPI network signature was constructed based on the components of the PPI network associated with survival according to LASSO-Cox regression analysis (A, B). Kaplan-Meier survival curve (C), The results of ROC curve (D), nomogram (E), and calibration curve (F) analyses are shown. LASSO, least absolute shrinkage and selection operator; ROC, receiver operating characteristic; PPI, protein-protein interaction.

Figure 4

Immune cell infiltration. The composition of immune cells was assessed by the CIBERSORT algorithm in COAD (A, B). The proportions of T follicular helper cells and monocytes were different in N+ and N0 group tissues (C). The results of the correlation analysis of immune infiltrating cells are shown as a heatmap (D). CIBERSORT, cell-type identification by estimating relative subsets of RNA transcripts; COAD, colon adenocarcinoma.

Figure 5

Coexpression analysis. The results of the correlation analysis of LNM-related genes expression and immune infiltrating cell are shown as a heatmap (A). The correlations between LRP2 and T follicular helper cells, and NKAIN4 and T follicular helper cells (B, C); LNM,lymph node metastasis.

Figure 6

LNM-related genes with immune checkpoints in COAD. LNM-related genes and immune checkpoint alteration in COAD. Compact visualization of cases with multiple genetic alterations of LNM-related genes and immune checkpoints (origined from five studies) were individually shown by cBioPortal as indicated, including fusion, amplification, deep deletion, truncating mutation, and missense mutation (A). Mutual-exclusivity analysis between LNM-related genes and multiple-immune checkpoints in COAD. The altered relationship between LNM-related genes and each immune checkpoint, such as co-occurrence and mutual exclusivity, was presented as indicated (B). The detailed log2 odds ratio, P-value, Q-value, tendency, and significance were individually presented in each panel. Q-value < 0.05 was considered to be statistically significant (indicated as yes, and others as no); LNM, lymph node metastasis; COAD, colon adenocarcinoma.