Prediction of Target Genes and Pathways Associated With Cetuximab Insensitivity in Colorectal Cancer

Abstract

Background: Cetuximab has been regularly added to the treatments for metastatic colorectal cancer worldwide. However, due to its therapeutic insensitivity and underlying mechanisms being largely unknown, the clinical implementation of cetuximab in colorectal cancer remains limited.

Methods: The gene expression profile GSE56386 was retrieved from the Gene Expression Omnibus database. Differentially expressed genes were identified between cetuximab-responsive patients and nonresponders, annotated by gene ontology, Kyoto Encyclopedia of Genes and Genomes pathway analysis, and further analyzed by protein-protein interaction networks. The integrative prognostic analysis was based on The Cancer Genome Atlas and PrognoScan.

Results: 1350 differentially expressed genes were identified with 298 upregulated and 1052 downregulated. Epidermis development, the cornified envelope, calcium ion binding, and amoebiasis were enriched in upregulated genes while digestion, the apical part of the cell, the 3',5'-cyclic-adenosine monophosphate phosphodiesterase activity and pancreatic secretion were found enriched in downregulated genes. The top 10 hub genes were identified, including epithermal growth factor, G-protein subunit β 5, G-protein subunit γ 4, fibroblast growth factor 2, B-cell lymphoma protein 2, acetyl-coenzyme A carboxylase β, KIT proto-oncogene receptor tyrosine kinase, adenylate cyclase 4, neuropeptide Y, and neurotensin. The hub genes exhibited distinct correlations in cetuximab-treated and untreated genomic profiles (GSE56386, GSE5851 and GSE82236). The highest correlation was found between B-cell lymphoma protein 2 and acetyl-coenzyme A carboxylase β in GSE56386. The mRNA expression of hub genes was further validated in the genomic profile GSE65021. Furthermore, B-cell lymphoma protein 2 and acetyl-coenzyme A carboxylase β also exhibited highest degrees among the hub genes correlation networks based on The Cancer Genome Atlas. Both B-cell lymphoma and acetyl-coenzyme A carboxylase β were not independent prognostic factors for colorectal cancer in univariate and multivariate Cox analysis. However, integrative survival analysis indicated that B-cell lymphoma protein 2 was associated with favorable prognosis (hazard ratio = 0.62, 95% confidence interval, 0.30-0.95, P = .024).

Discussion: This in silico analysis provided a feasible and reliable strategy for systematic exploration of insightful target genes, pathways and mechanisms underlying the cetuximab insensitivity in colorectal cancer. B-cell lymphoma protein 2 was associated with favorable prognosis.

Keywords: KEGG pathway; cetuximab; colorectal cancer; differentially expressed genes; gene ontology; protein–protein interaction.

Figure 1.

Gene ontology and KEGG analysis of differentially expressed genes associated with cetuximab insensitivity of CRC. (A) Biological function of gene ontology in upregulated/downregulated groups; (B) cellular component of gene ontology in upregulated/downregulated groups; (C) molecular function of gene ontology in upregulated/downregulated groups; (D) the KEGG pathway analysis results of differentially expressed genes in upregulated/downregulated groups. BP indicates biological function; CC, cellular component; CRC, colorectal cancer; MF, molecular function; KEGG, Kyoto Encyclopedia of Genes and Genomes.

Figure 2.

Protein–protein interaction network of the DEGs. Red nodes stand for upregulated genes while the blue nodes represent downregulated genes, with the lines representing interactions between each gene. DEGs indicates differentially expressed genes.

Figure 3.

The top 3 modules extracted from the protein–protein interaction network: (A) module 1, (B) the KEGG pathway analysis of module 1, (C) module 2, (D) the KEGG pathway analysis of module 2, (E) module 3, and (F) the KEGG pathway analysis of module 3. KEGG indicates Kyoto Encyclopedia of Genes and Genomes.

Figure 4.

The illustration of cetuximab treatment during the time courses with 3 profiles (GSE56386, GSE5851, and GSE82236) associated with the insensitivity of cetuximab in CRC and the pairwise correlation of the hub genes expression. The red circle indicated negative correlation, the blue indicated positive correlation. The values of correlation coefficients were represented by the color bar aside. Color intensity and the circle size were proportional to the correlation coefficients. CRC indicates colorectal cancer.

Figure 5.

Integrative analysis of BCL2 and ACACB in TCGA. (A) The correlations of hub genes based on gene expressions in TCGA; the degree was in proportion to the red color; the correlation values were illustrated by the thickness of the connecting lines. (B) The mRNA expression of BCL2 in tumor versus normal tissues (red: tumor; blue: normal); (C) the mRNA expression of BCL2 in tumor versus normal tissues (red: tumor; blue: normal); (D) the stage distribution of the mRNA expression of BCL2; and (E) the stage distribution of the mRNA expression of ACACB. BCL2 indicates B-cell lymphoma protein 2; ACACB, acetyl-coenzyme A carboxylase β; TCGA, The Cancer Genome Atlas; mRNA, messenger RNA.

Figure 6.

The integrative analysis of the prognostic values of BCL2 and ACACB in multiple gene expression profiles. (A) Integrative prognostic analysis of BCL2 and (B) integrative prognostic analysis of ACACB. BCL2 indicates B-cell lymphoma protein 2; ACACB, acetyl-coenzyme A carboxylase β.