PROJECT SUMMARY/ABSTRACT Sexual minority (SM, i.e., people who are not heterosexual) and gender minority (GM, i.e., people who have a gender that is discordant from the sex they were assigned at birth, as opposed to cisgender people who have a gender that is concordant with the sex they were assigned at birth) people (collectively abbreviated as SGM) are at greater risk for health disparities including very high rates of substance use. Some subgroups of GM people evidence greater risk for cannabis use. The primary explanation for the higher rates of cannabis use among SGM people is minority stress (for example mistreatment related to one’s SGM status), but there may also be hormonal contributors to cannabis use. Epigenetic modifications (e.g., DNA methylation) are one way to track molecular modifications in response to cannabis use and may serve as biomarkers for cannabis use. Understanding the epigenetics of cannabis use may help us to develop better ways to identify and treat substance use disorders and to understand the downstream health outcomes of SGM people. This study will leverage The PRIDE Study, a national longitudinal cohort study of SGM people to identify changes in trajectories of cannabis use among SGM people over 9 years of annual data collection (N>14,000 aged 18+), examine differences in these trajectories based on hormonal exposures and examine how minority stress changes these relationships (Aim 1). This study will then identify the relationships between cannabis use trajectories and DNA methylation among SGM people assigned female at birth (N=600), identify differences in the relationships between cannabis use and DNA methylation based on hormonal exposures, and identify which of these relationships between DNA methylation and cannabis use persist even after accounting for minority stress (Aim 2). This study will help us to understand longitudinal trajectories of cannabis use among SGM people to inform targeted prevention and intervention development. This study will also inform the development of biomarkers for cannabis use to improve substance use treatment and prevention.