An open-label pharmacokinetics (PK) clinical trial was conducted to comparatively assess the PK and explore the pharmacodynamics (PD) of miltefosine in children and adults with cutaneous leishmaniasis (CL) in Colombia. Sixty patients, 30 children aged 2 to 12 years and 30 adults aged 18 to 60 years, were enrolled. Participants received miltefosine (Impavido) at a nominal dose of 2.5 mg/kg/day for 28 days. Miltefosine concentrations were measured in plasma and peripheral blood mononuclear cells by liquid chromatography-tandem mass spectrometry of samples obtained during treatment and up to 6 months following completion of treatment, when therapeutic outcome was determined. Fifty-two patients were cured, 5 pediatric patients failed treatment, and 3 participants were lost to follow-up. Leishmania (Viannia) panamensis predominated among the strains isolated (42⁄46; 91%). Noncompartmental analysis demonstrated that plasma and intracellular miltefosine concentrations were, overall, lower in children than in adults. Exposure to miltefosine, estimated by area under the concentration-time curve and maximum concentration, was significantly lower in children in both the central and intracellular compartments (P < 0.01). Leishmania persistence was detected in 43% of study participants at the end of treatment and in 27% at 90 days after initiation of treatment. Clinical response was not dependent on parasite elimination. In vitro miltefosine susceptibility was similar for Leishmania strains from adults and children. Our results document PK differences for miltefosine in children and adults with cutaneous leishmaniasis that affect drug exposure and could influence the outcome of treatment, and they provide bases for optimizing therapeutic regimens for CL in pediatric populations. (This study has been registered at ClinicalTrials.gov under identifier NCT01462500.).