Cannabis use is most likely to begin during adolescence or emerging adulthood; and such use during these developmental periods has been associated with some negative physical, mental, and cognitive health outcomes. Some of these long-term negative effects of this consumption during adolescence may be explained by epigenetic changes through a process called DNA methylation. However, little is known about the epigenetic impact of drug use during adolescence. The current study examined associations between DNA methylation and cannabis use during adolescence. Analyses focused on two genes, AHRR and NR2B because their methylation has been implicated in illicit drug use. Participants included twenty-seven adolescents separated into two groups, cannabis users and control. After identifying the subjects who would participate in the research, the general research framework was formed. The inclusion criteria were age between 13 and 18 years old. Those individuals diagnosed with cannabis dependence comprised the CUD (Cannabis Use Disorder) group, verified through a urine test for cannabis. Participants in the CONT group were healthy individuals, who did not have a history and/or criteria for diagnosing chemical dependency. DNA methylation values were measured with salivary DNA and averaged for each gene. Relationships between cannabis use and DNA methylation were tested using multivariate regressions, adjusting for sociodemographic data, and age. Adolescents with psychoactive substance use face a considerable risk of co-occurring mental health problems, something that may involve a more difficult life situation, social problems, as well as a worse result in complying with socio-educational measures.