Comparative metabolism of aschantin in human and animal hepatocytes

In Vitro Metabolism and Transport Characteristics of Zastaprazan

“…The experimental protocols were performed in accordance with FDA guidelines and the previous reports with slight modifications [ 13 , 16 ]. To investigate the inhibitory effect of CYP inhibitor, SKF-525A on JP-1366 metabolism, the reaction mixtures containing 80 μL of 50 mM potassium phosphate buffer (pH 7.4), 4 μL of 250 mM magnesium chloride, 1 μL of 500 μM JP-1366 (final concentration: 5 μM), 1 μL of 1 mM or 10 mM SKF-525A (final concentration: 10 μM and 100 μM), 10 μL of 2 mg/mL pooled human liver microsomes (final concentration: 0.2 mg/mL), and 5 μL of NADPH-generating system were incubated at 37 °C for 30 min.…”

“…To evaluate the enzyme kinetic parameters for the metabolism of JP-1366, the concentration-dependent metabolism of JP-1366 and M1 was conducted according to the previous reports [ 16 ]. Briefly, the reaction mixture consisted of 80 μL of 50 mM potassium phosphate buffer (pH 7.4); 4 μL of 250 mM magnesium chloride; 10 μL of 2 mg/mL pooled human liver microsomes (final concentration: 0.2 mg/mL) or human cDNA-expressed CYP1A2, 2C8, 2C9, 2C19, 2D6, 3A4, and 3A5 enzymes (final concentration: 4 pmol); and 1 μL of various concentrations of JP-1366 (final concentration: 1, 2, 5, 10, 20, 40, 60, or 80 μM).…”

“…The use of antibodies specific to individual CYP can be used to confirm the contribution of individual CYP in the metabolism of drugs or drug candidates according to the FDA guidelines [ 13 ]. To increase the selectivity of antibodies to a single CYP, we chose monoclonal antibodies, which bind to a single epitope of CYP and are treated with ultrapooled HLMs according to the previous reports in addition to the manufacturer’s protocols [ 16 , 17 , 18 , 19 ].…”