CRISPR-Cas9 screening identified novel subtypes of cutaneous melanoma based on essential cancer genes

Abstract

Our primary objective was to identify genes critical for cutaneous melanoma (CM) and related typing, based on essential genes, to generate novel insights for clinical management and immunotherapy of patients with CM. We analyzed RNA sequencing (RNA-seq) data from The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx), and sequencing data of 29 CM cell line from Cancer Cell Line Encyclopedia (CCLE) databases. Combined with DepMap database, 406 CM essential cancer genes were finally obtained. Based on the expression of essential genes in cancer, the patients included in TCGA and Gene Expression Omnibus (GEO) databases were divided into three different molecular subtypes (C1, C2, and C3) by the NMF algorithm. Data analysis from TCGA and GEO datasets revealed that subtype C3 had the poorest prognosis, while subtype C1 exhibited the best prognosis. Combined with the CIBERSORT, ESTIMATE and ssGSEA algorithm, patients with different molecular subtypes can be divided into two immune subtypes (hot and cold). We found that subtype C1 was characterized by hot tumors, in contrast to subtypes C2 and C3, which were characterized by cold tumors. Then, we used univariate Cox regression, LASSO, and multifactor Cox regression analysis to select risk genes and constructed a prognostic model based on eight genes: RABIF, CDCA8, FOXM1, SPRR2E, AIP, CAP1, CTSW, and IFITM3. All patients were divided into two risk subtypes (high and low ) according to the median of risk scores. We found that most hot tumor subtypes were found in the low-risk subtypes and most patients with this subtype survived for longer. Ultimately, we selected RABIF, which exhibits the highest risk coefficient, for histological and cytological verification. The results showed that RABIF was overexpressed in melanoma. Inhibition of RABIF expression could suppress the proliferation and invasion of melanoma cells and promote the apoptosis of melanoma cells. In conclusion, we used CRISPR-Cas9 screening to verify the association between molecular subtypes (C1, C2, and C3), immune subtypes (hot and cold), and risk subtypes (high and low) in patients with CM, particularly in distinguishing survival and prognosis. These findings can be used to guide clinical management and immunotherapy of patients with CM.

Keywords: CRISPR-Cas9; Clinical management; Cutaneous melanoma; Essential cancer genes; Immunohistochemistry; Molecular subtypes; Predictive model; RABIF; Risk subtypes; Tissue microarray.