Tirbanibulin is a novel tubulin polymerization and Src kinase signaling inhibitor. This study was designed to fully characterize tirbanibulin pharmacokinetics (PK) when applied topically under maximal use conditions. This was an open-label, parallel-group PK safety study of tirbanibulin ointment 1% applied to 25 cm2 of the face or balding scalp in adults with actinic keratosis (AK). Eligible subjects self-applied tirbanibulin once-daily for 5 days. PK sampling occurred on days 1, 3 and 4 at 0 hour (before dosing), and on day 5 at prespecified time points up to 24 hours after application. Safety assessments included adverse events and local skin reactions were evaluated up to day 29. Eighteen subjects (face or scalp, n = 9 each) completed the study. Subjects were White (100%), of mean [range] age 66.4 [43-83] years, predominantly men (83.3%) with Fitzpatrick skin type I to III (94.4%); baseline AK lesion count, mean [range] 8.2 [6‒14]. All subjects had quantifiable but low plasma concentrations of tirbanibulin. On day 5, overall mean (standard deviation) maximum concentration (Cmax) was 0.26 (0.23) ng/mL (or 0.60 nM), median time to maximum concentration was 6.91 hours, and mean (standard deviation) area under the plasma concentration–time curve from time 0 to 24 hours was 4.09 (3.15) ng ∙ h/mL. Four subjects experienced a total of 5 treatment-emergent adverse events that resolved. Mild to moderate erythema, flaking, or scaling in the treatment area peaked around day 8 before resolving or returning to baseline by day 29. In conclusion, under maximal use conditions, tirbanibulin ointment 1% for 5 days in the treatment of AK on the face or scalp was well tolerated and resulted in low systemic exposure with subnanomolar plasma concentrations.