Zilurgisertib is an oral, activin receptor-like kinase 2 (ALK2) inhibitor being developed for the treatment of patients with fibrodysplasia ossificans progressiva. In vitro metabolism studies suggest zilurgisertib is primarily eliminated via cytochrome P450 (CYP) 3A4/5-mediated metabolism, with an additional contribution from renal excretion. A drug-drug interaction study was conducted to evaluate the impact of the strong CYP3A inhibitor itraconazole and the strong CYP3A4 inducer rifampin on the pharmacokinetics of zilurgisertib in healthy participants. Participants in Cohort 1 (n = 18) received zilurgisertib 100 mg on Days 1 and 10 and itraconazole 200 mg once daily on Days 6-13. Participants in Cohort 2 (n = 18) received zilurgisertib 100 mg on Days 1 and 13 and rifampin 600 mg once daily on Days 6-15. With the concomitant administration of itraconazole, the geometric mean maximum plasma drug concentration (C) and area under the concentration-time curve from time 0 to infinity (AUC) of zilurgisertib increased by 1.28- and 2.13-fold, respectively. With the concomitant administration of rifampin, there was a 2.27- and 4.76-fold reduction in the geometric mean C and AUC of zilurgisertib, respectively. No dose-limiting toxicities occurred in either cohort, and all treatment-emergent adverse events were Grade 1 in severity. Zilurgisertib dose adjustment may be necessary with concomitant administration of strong CYP3A4 inhibitors, and coadministration of zilurgisertib with strong CYP3A4 inducers is not recommended.