Integrated genomic and epigenetic profiling reveals RAS pathway as a key driver of odontogenic tumors

Odontogenic tumors (OTs) are highly proliferative lesions with a low number of driving mutations, suggesting that concurrent alternative molecular mechanisms could support their extensive proliferation capacity. In this study, we analyzed 94 tissue samples from 79 patients with OTs and 15 healthy controls to explore their genetic and epigenetic alterations. Whole Exome Sequencing identified the BRAF V600E mutation in 75% of patients. A mutational hotspot analysis of six key genes (NRAS, EGFR, BRAF V600E, SMO L412, SMO W535, KRAS Q22K, PTCH1 R602*, and PTCH1 W129) revealed a high mutational rate, particularly in BRAF (91%), with 90% of BRAF V600E-positive patients being ameloblastomas. DNA methylation in the promoters of nine tumor-related genes (RB1, RASSF1A, BRCA1, BRCA2, MSH2, MLH1, MGMT, TIMP3, BRAF) was assessed in 67 OT patients and 15 controls. Five CpG sites showed significant hypermethylation (p<0.05; FDR q<0.05), notably in RASSF1A (cg50378469, cg50378539) and TIMP3 (cg33197381, cg33197394, cg33197400). Somatic hypermethylation of the full promoter was detected in RASSF1A (8 patients, mean methylation: 17.5%), BRCA1 (3 patients, mean methylation: 11.7%), and MLH1 (1 patient, mean methylation: 2%). Interestingly, 76.5% of BRAF V600E-positive patients had RASSF1A promoter hypermethylation. A strong correlation between BRAF V600E mutation and RASSF1A hypermethylation was detected. Our results might imply a synergistic effect of the BRAF V600E mutation and RASSF1A hypermethylation as determinants in the RAS pathway. These findings, together with observations from other studies, suggest that the RAS pathway is a key axis in OT biology.