Abstract Opioid use disorder (OUD) is rare (2%) but associated with extraordinarily high morbidity and mortality. Opioids are the primary contributors to the soaring numbers of U.S. drug overdose deaths, with increasing co-use of psychostimulants exacerbating the burden. Alongside this opioid crisis, are increasing rates of cannabis use, a drug that is now legal for recreational use in 24 states. Early cannabis use onset (eCan) has been documented in cross-national, cross-sectional, and longitudinal studies to increase the likelihood of using and misusing both cocaine/psychostimulants and opioids. Shared genetic vulnerability has been suggested as a mechanism undergirding both eCan and OUD, although it has not been estimated. However, even within discordant twin pairs, twins who endorse eCan more commonly report stimulant misuse and OUD, when compared to their co- twins (each at 4 times increased odds), suggesting that genetic factors alone do not explain this association. Also, rodents pretreated with delta-9 tetrahydrocannabinol during adolescence show variable behavioral and epigenetic responses to opioids and cocaine, suggesting that eCan may modify sensitization to these drugs. Yet, due to the rarity of OUD in samples that typically assess eCan and the absence of eCan measurement in large biobanks from which OUD data are drawn, a detailed examination of the association between eCan and opioid involvement is pending. In this R21 data analysis proposal in response to NOT-DA-22-003, using data from multiple deeply phenotyped samples, predominantly those ascertained for substance use disorders (N~138,000), we examine the association between eCan and opioid involvement. In aim 1, we estimate the association between early cannabis use (eCan) and proximal (e.g., initial subjective responses, indexing sensitization) and distal aspects of opioid involvement (e.g., OUD criterion count), and whether associations are modified by co-occurring stimulant (non-prescription, including cocaine) misuse. In aim 2, we test whether eCan and OUD (including overdose in the presence of psychostimulants) share an architecture of common genetic influences and parse evidence for non-genetic, putatively causal influences of eCan on opioid involvement using both genomic causality and data from pairs of twins and siblings discordant for eCan. In aim 3, we estimate the independent role of Adverse Childhood Experiences (ACEs) on eCan and opioid involvement and its interplay with genetic susceptibility in worsening the likelihood of OUD in those who start using cannabis at an earlier age. Understanding the relationship between eCan and specific aspects of opioid involvement provides insights into whether future changes in cannabis use might exacerbate OUD, and establishes targets for experimental and mechanistic studies. If eCan modifies opioid sensitization and increases likelihood of OUD, or is correlated with opioid overdose death, then heightened vigilance is urgently ne...