|| Checking for direct PDF access through Ovid
The dried blood spot (DBS) assay has been developed and validated for E6005 and its metabolite (M11) in human blood.E6005 and M11 in DBS were quantifiable from 1ng/mL as free base using only 25μL blood.Typical validation parameters were within the acceptance criteria recommended by bioanalytical guidelines.DBS-associated specific validation parameters, including impacts by blood volume and hematocrit, were also evaluated.Stability assessments showed that E6005 and M11 were stable for 160days at ambient temperature, and below −15°C.E6005, a novel phosphodiesterase 4 inhibitor, is currently under clinical development for the treatment of atopic dermatitis. To support pediatric clinical trials, the dried blood spot assay for simultaneous determination of E6005 and its main metabolite, ER-392710 (M11), has been developed using ultra-performance liquid chromatography with tandem mass spectrometry. E6005 and M11, in 25μL blood spotted onto FTA™ DMPK-C cards, were extracted by water/acetonitrile (1:1, v/v), and then chromatographed on a reversed phase column under gradient elution. The mass transitions, m/z 473.1→163.0 for E6005 and m/z 459.1→149.0 for M11, with corresponding stable isotope internal standard, m/z 477.2→167.0, and m/z 463.2→153.0, were monitored. E6005 and M11 were quantifiable from 1 to 200ng/mL as free base. Accuracy and precision of the two analytes in the intra- and inter-batch reproducibility were within ±8.0% and 15.7%, respectively. Extraction recoveries of the analytes were 73% or more and hematocrit ranging from 26.9% to 51.8% did not impact the analytes’ accuracy. Various stability assessments, including possible conversion of E6005 to M11, were thoroughly performed, and bench-top stability was ensured up to 160days. The DBS method was applied to determine E6005 and M11 concentrations in blood samples supporting a pediatric clinical trial.