In vitro glucuronidation of the cyclin-dependent kinase inhibitor flavopiridol by rat and human liver microsomes: Involvement of udpglucuronosyltransferases 1A1 and 1A9

The metabolism of flavopiridol (FLAP), a novel anticancer drug currently un dergoing clinical development, was investigated in rat and human liver micr osomes. In the presence of uridine 5'-diphosphoglucuronic acid, two biotran sformation products (M1 and M2) could be detected. Formation of metabolite M1 and M2 in terms of enzymatic efficacy (V-max/K-M) was about 50- and 5-fo ld higher in rat (1.58 +/- 2.23 and 7.22 +/- 1.17 mul/min/mg) as compared w ith human liver microsomes (0.032 +/- 0.016 and 1.52 +/- 0.93 mul/min/mg), indicating species-related differences in FLAP glucuronidation. Incubation in the presence of human recombinant UDP-glucuronosyl-transferases (UGTs) d emonstrated that M1 is almost exclusively catalyzed by UGT1A1, whereas M2 i s formed by UGT1A9 and only to a minor extent by UGT1A1 and UGT1A10. Chemic al inhibition experiments further prove the involvement of UGT1A1 and UGT1A 9 in the formation of M1 and M2, as the UGT1A1 substrate bilirubin preferab ly inhibited M1 over M2 (K-i : 36 and 258 muM, respectively), whereas the U GT1A9 substrate propofol showed a more pronounced decrease in M2 but not in M1 formation (K-i : 47 and 142 mM, respectively). Both conjugates were pur ified from rat liver microsomes and analyzed by mass spectrometry, NMR, and UV experiments. On the basis of these results, M1 was identified as 5-O-be ta -glucopyranuronosyl-flavopiridol and M2 as 7-O-beta -glucopyranuronosyl- flavopiridol. In conclusion, our results elucidate the enzymatic pathways o f FLAP in rat and human liver, which must be considered during cancer thera py of patients.