Investigation of the in vitro metabolism profile of a phosphodiesterase-IVinhibitor, CDP-840: Leading to structural optimization

CDP-840 is a selective and potent phosphodiesterase type IV inhibitor, whos e in vitro metabolism profile was first investigated using liver microsomes from different species. At least 10 phase I oxidative metabolites (M1-M10) were detected in the microsomal incubations and characterized by capillary high-performance liquid chromatography continuous-flow liquid secondary io n mass spectrometry (CF-LSIMS). Significant differences in the microsomal m etabolism of CDP-840 were found between rat and other species. The major ro ute of metabolism in rat involved para-hydroxylation on the R4 phenyl. This pathway was not observed in human and several other species. The in vitro metabolism profile of CDP-840 was further examined using freshly isolated h epatocytes from rat, rabbit, and human. The hepatocyte incubations indicate d more extensive metabolism relative to that in microsomes. In addition to the phase I oxidative metabolites observed in microsomal incubations, sever al phase II conjugates were identified and characterized by CF-LSIMS. Inter species differences in phase II metabolism were also found in these hepatoc yte incubations. The major metabolite in human hepatocytes was identified a s the pyridinium glucuronide, which was not detected in rat hepatocytes. Si mple structural modification on R4, such as p-Cl substitution, greatly redu ced the species differences in microsomal metabolism. Furthermore, modifica tions on R3, such as the N-oxide, eliminated the N-glucuronide formation in human. These results not only helped in determining the suitability of ani mal species used in the preclinical safety studies but also provided valuab le directions for the synthetic efforts in finding backup compounds that ar e more metabolically stable.