GSK2251052 [(S)-3-(aminomethyl)-7-(3-hydroxypropoxy)1-hydroxy-1,3-dihydro-2,1-benzoxaborole hydrochloride], formerly known as AN3365, a novel boron-containing leucyl-tRNA synthetase inhibitor, is currently being developed for the treatment of serious Gram-negative bacterial infections. Distribution, metabolism, and excretion of GSK2251052 were investigated following a single intravenous administration of 14C-GSK2251052 in male and female Sprague Dawley rats, male Long Evans rats, at 50 mg/kg (150 μCi/kg), and male and female cynomolgus monkeys at 25 mg/kg (150 μCi/kg). QWBA in rats showed radioactivity was widely and rapidly distributed to most tissues with insignificant retention in pigmented ocular tissues by 35 days, and the majority of radioactivity was eliminated from most tissues by 168 h. Potentially preferential distribution of radioactivity into the cellular component of blood with blood to plasma ratio of 1.3 for 24 h was observed. The overall mean recoveries of radioactivity in male and female rats and monkeys were 95% and 82%, respectively. Most of the total 14C‑GSK2251052‑derived radioactivity was excreted within 48 hours, with urinary excretion as the primary route of elimination, representing 67.5% and 51.7% of the total dose in rats and monkeys by 168 h, respectively. Metabolism of GSK2251052 was moderate in rats and extensive in monkeys. Significant quantities of an oxidative metabolite (M3), whereby the hydroxypropyl moiety of GSK2251052 was oxidized to a propanoic acid moiety, were present in monkey plasma, while moderate amounts of N-acetylated M3 (M7) were evident in rat plasma. Differences in urinary excretion profiles were demonstrated, with GSK2251052 as the major component excreted in rat urine accounting for 46% of the radioactivity dose, in comparison to less than 10% of the dose present in urine from monkeys. Metabolite M3 was the major component excreted in urine from monkeys. No gender difference on metabolic profile and excretion of GSK2251052 was observed in rats and monkeys. The species differences in the metabolic profiles suggest renal clearance of GSK2251052 predominates in rats, while the metabolic clearance is higher in monkeys than rats.