Bioinformatics-based analysis reveals elevated CYTL1 as a potential therapeutic target for BRAF-mutated melanoma

Abstract

Introduction: Despite many recent emerging therapeutic modalities that have prolonged the survival of melanoma patients, the prognosis of melanoma remains discouraging, and further understanding of the mechanisms underlying melanoma progression is needed. Melanoma patients often have multiple genetic mutations, with BRAF mutations being the most common. In this study, public databases were exploited to explore a potential therapeutic target for BRAF-mutated melanoma. Methods: In this study, we analyzed differentially expressed genes (DEGs) in normal tissues and melanomas, Braf wild-type and Braf mutant melanomas using information from TCGA databases and the GEO database. Subsequently, we analyzed the differential expression of CYTL1 in various tumor tissues and its effect on melanoma prognosis, and resolved the mutation status of CYTL1 and its related signalling pathways. By knocking down CYTL1 in melanoma cells, the effects of CYTL1 on melanoma cell proliferation, migration and invasion were further examined by CCK8 assay, Transwell assay and cell migration assay. Results: 24 overlapping genes were identified by analyzing DEGs common to melanoma and normal tissue, BRAF-mutated and BRAF wild-type melanoma. Among them, CYTL1 was highly expressed in melanoma, especially in BRAF-mutated melanoma, and the high expression of CYTL1 was associated with epithelial-mesenchymal transition (EMT), cell cycle, and cellular response to UV. In melanoma patients, especially BRAF-mutated melanoma patients, clinical studies showed a positive correlation between increased CYTL1 expression and shorter overall survival (OS) and disease-free survival (DFS). In vitro experiments further confirmed that the knockdown of CYTL1 significantly inhibited the migration and invasive ability of melanoma cells. Conclusion: CYTL1 is a valuable prognostic biomarker and a potentially effective therapeutic target in melanoma, especially BRAF-mutated melanoma.

Keywords: BRAF mutations; CYTL1; cell migration and invasion; melanoma; molecular biomarker.

FIGURE 1

Screening of potential targets in melanoma cells with BRAF mutation. (A,B) Volcano plot of the differentially expressed genes in melanoma according to the TCGA dataset. (C) Venn of all overlapping DEGs. (D) Protein–protein interaction network of the overlapping DEGs. (E) Gene Ontology enrichment analyses of the overlapping DEGs.

FIGURE 2

High expression of CYTL1 is associated with a poor prognosis of melanoma. (A) Forest plot of the p-value, risk coefficient (HR) and univariate analysis of the overlapping DEGs in melanoma. (B) Kaplan-Meier survival curves of SKCM patients with high expression or low expression of the overlapping DEGs.

FIGURE 3

High expression of CYTL1 is associated with poor prognosis in BRAF mutant melanoma. (A). CYTL1 expression levels in different tumor tissues and adjacent normal tissues from TCGA and GTEx databases. The expression level of CYTL1 in GSE46517 (B) and GSE114445 (C,D). Heat map of the normalized coefficient of CYTL1 in Cox mode. Kaplan-Meier survival curves of OS (E) and DSS (F) in SKCM patients with high CYTL1 expression or low CYTL1 expression. Kaplan-Meier survival curves of OS (G) and DSS (H) in BRAF-mutant SKCM patients with high CYTL1 expression or low CYTL1 expression.

FIGURE 4

Genomic mutations in CYTL1 in melanoma. (A) Alteration frequency of CYTL1 in different data sets. (B) Mutation sites for CYTL1 gene mutations. (C) Type and frequency of CYTL1 gene mutations in SKCM.

FIGURE 5

Analysis of genes and pathways associated with CYTL1 in melanoma. (A) Volcano plot of the differentially expressed genes in melanoma according to the TCGA dataset. (B) The heatmap of the differential gene expression. (C–F) GO and KEGG signaling pathways enrichment analyses of the DEGs.

FIGURE 6

CYTL1-related signaling pathways in SKCM by GSEA software.

FIGURE 7

Correlation of CYTL1 expression with immune infiltration. (A–D) The correlation between CYTL1 expression and infiltration levels of immune cells. (E) Correlation analysis of CYTL1 expression and immune checkpoint-related genes in SKCM in the TCGA database. (F) Relationship between CYTL1 expression and SKCM tumor immune cell infiltration according to the TIMER database.

FIGURE 8

Knockdown of CYTL1 inhibits migration and invasion of BRAF mutant melanoma cells. (A) CYTL1 mRNA levels in different melanoma cells and HEM. **p ≤ 0.01, compared with HEM. (B) The protein levels of CYTL1 in different melanoma cells and HEM. (C) CYTL1 mRNA levels in A2058 cells transfected with si-CYTL1. **p ≤ 0.01, compared with siCon group. (D) The protein levels of CYTL1 in A2058 cells transfected with si-CYTL1. Cell viability (E), wound healing (F), cell migration (G) and cell invasion (H) of A2058 cells transfected with si-CYTL1.