To compare 6-month safety and efficacy outcomes of fractionated CO2 laser (laser) with topical clobetasol propionate (steroid) for treatment of symptomatic vulvar lichen sclerosus. We conducted a single-center randomized controlled trial that compared fractionated CO2 laser with steroid treatment for patients with biopsy-proven lichen sclerosus. Randomization was stratified by prior clobetasol propionate use. The primary outcome was mean change in Skindex-29 score at 6 months. A total sample size of 52 participants were recruited to detect a mean difference of 16 points on the Skindex-29 (SD±22) with 80% power, based on a one-sided two-sample t test with α=0.05, accounting for 10% attrition. Secondary outcomes included validated subjective and objective measures. Intention-to-treat, per protocol, and regression analysis based on prior steroid exposure were performed. From October 2015 to July 2018, 202 women were screened, 52 were randomized, and 51 completed a 6-month follow-up. No significant difference was found in baseline demographics, symptoms, and physician assessment scores. There was greater improvement in the Skindex-29 score in the laser arm at 6-months (10.9 point effect size, 95% CI 3.42-18.41; P=.007). Overall, 89% (23/27) of patients in the laser group rated symptoms as being "better or much better" compared with 62% (13/24) of patients in the steroid group, P=.07. More patients (81%, 21/27) were "satisfied or very satisfied" with laser treatment compared with steroid treatment (41%, 9/24); P=.01. After stratification for previous steroid use, the significant change of Skindex-29 score was only seen in the previously exposed group. There was one adverse event in each group: minor burning and blistering at the laser site and reactivation of genital herpes 1 week after starting steroid. Fractionated CO2 laser treatment showed significant improvement in subjective symptoms and objective measures compared with clobetasol propionate, without serious safety or adverse events at 6 months. ClinicalTrials.gov, NCT02573883.