Effects of Tetrahydrouridine Pre-dosing on Absorption, Metabolism, and Excretion of Decitabine in the Mouse

• THU pre-treatment increases DEC plasma exposure. • THU pre-treatment shifts the pathways for DEC metabolism. • DEC and its metabolites are primarily excreted via the urinary tract, independent of THU. Elevated fetal hemoglobin (HbF) levels improve outcomes in sickle cell disease (SCD). Decitabine (DEC), a DNA methyltransferase 1 inhibitor, reactivates silenced HbF but is rapidly degraded by cytidine deaminase (CDA). Co-administered tetrahydrouridine (THU) inhibits CDA, enhancing DEC bioavailability. NDec, a novel oral, fixed-dose combination of DEC and THU, has been developed as treatment for people with SCD. NDec is currently undergoing clinical evaluation, beginning with the Phase II ASCENT1 study (NCT05405114). This study aimed to evaluate the impact of THU pre-dosing on the pharmacokinetics (PK), metabolism, and excretion profile of DEC in mice. Mice were orally administered with either radiolabeled DEC ([ 14 C]-DEC; 1 mg/kg) or with two different doses of THU (10 and 170 mg/kg) one hour before a fixed-dose of [ 14 C]-DEC (0.4 mg/kg). Liquid scintillation counting (LSC), high-performance liquid chromatography (HPLC), LC-mass spectrometry (LC-MS), and LC-radioactivity monitor-MS (LC-RAM-MS) analyses were used to detect the presence of [ 14 C]-DEC and its metabolites in plasma and urine at different time points after dosing. DEC metabolism occurs by either deamination (CDA metabolism) or by competitive hydrolysis and oxidation pathways (non-CDA metabolism), generating six DEC metabolites. THU pre-treatment increased DEC plasma exposure and shifted the metabolite profile towards non-CDA mediated metabolism. Finally, DEC and its metabolites were primarily excreted via the urinary tract, independently of THU pre-dosing. These findings provide crucial insights into DEC metabolism, highlighting the impact of THU pre-treatment on DEC plasma exposure, metabolism, and excretion.