PROJECT SUMMARY/ABSTRACT Alcohol Use Disorder (AUD) is a chronic relapsing illness associated with high rates of relapse, and thus, there is great need to develop and evaluate novel treatments to decrease relapse and improve alcohol use outcomes in AUD. We previously conducted a novel dose finding human laboratory, safety and pilot efficacy study to assess whether the neuroactive steroid (NAS) precursor pregnenolone (PREG) that influences GABAergic functioning may normalize alcohol-related stress disruption and improve alcohol use outcomes in AUD. Pilot data showed that PREG at 300mg/day reduced stress- and cue- induced alcohol craving, anxiety and normalized chronic alcohol-related disruption in stress biology and also reduced alcohol drinks/day (AvgD), percent drinking days and heavy drinking days (%DD and %HDD) compared to placebo (PBO) in an 8- week clinical study. On the basis of these findings, this project proposes a 12-week double blind, randomized Phase II clinical trial to evaluate the safety and efficacy of PREG treatment (300 mg/day) versus PBO in 150 AUD men and women. The following specific aims will be addressed: Aim #1: To establish the safety and tolerability of PREG (300mg/day) vs. PBO in men and women with AUD over the 12-week treatment period and at the 1-month follow up. Aim #2: To test the efficacy of PREG vs. PBO on the primary alcohol use outcome of PSNHDD and secondary drinking outcomes of HDD%, DD% and AvgD during the trial. Aim #3: To assess the effects of PREG vs. PBO on other secondary stress-related outcomes of alcohol craving, anxiety, depression and patient-related functioning during the trial. Aim #4: To assess the effects of PREG vs. PBO on PREG and other NAS levels and examine their relationship to primary and secondary alcohol use and related outcomes. Exploratory Aim 1: To assess enduring short-term treatment effect of PREG vs. PBO on primary and other secondary outcomes at a 1-month post-treatment follow-up. Exploratory Aim 2: To explore whether pre-treatment patient characteristics (sex, trauma history, AUD severity and co-occurring psychiatric disorders) influence PREG effects on primary and secondary outcomes. It is well known that chronic alcohol use downregulates GABA which plays a significant role in the stress pathophysiology of AUD and also in loss of control drinking. The proposed study is based on our novel preliminary findings and will test our innovative approach of boosting endogenous neuroactive steroid levels to increase their function and thereby improve AUD outcomes. If successful, the proposed research will establish PREG and neuroactive steroids as key targets in the treatment of AUD, and also provide data on neuroactive steroid levels and whether they may serve as biomarkers in AUD treatment.