Oral administration of vismodegib for basal cell carcinoma treatment is limited by significant class-specific systemic side-effects. We investigated the approach of combining ablative fractional laser (AFL)-assisted drug delivery with an extended-release microemulsion formulation of vismodegib to provide efficient cutaneous delivery in vivo. The developed formulation consisted of an oil-in-water (o/w) microemulsion stabilized by tween-80. Pig skin was exposed to AFL followed by topical application of vismodegib microemulsion for 4 hours. At 4h, 2d, 5d and 9d we evaluated vismodegib biodistribution in superficial, mid and deep dermis as well as plasma (n=189 measurements) and assessed local skin reactions. Sustained topical delivery of vismodegib was detected in all depths of AFL-exposed skin over the course of the study with peak concentrations found at 5d and 9d. The highest vismodegib concentrations reached 1,409.7 μmol/L in superficial dermis and 62.3 μmol/L in deep dermis, exceeding steady-state plasma concentrations previously reported for oral administration of vismodegib (5.5-56.0 μmol/L). AFL increased vismodegib uptake up to 16.6-fold compared to intact skin. Only mild local skin responses to vismodegib were observed and no vismodegib was detected in plasma. We demonstrate sustained topical delivery of vismodegib in vivo at high concentrations with favorable skin tolerability, suggesting a safer future vismodegib treatment.