Characterization of the metabolism of fenretinide by human liver microsomes, cytochrome P450 enzymes and UDP-glucuronosyltransferases.
OBJECTIVE
Fenretinide (4-HPR) is a retinoic acid analogue, currently used in clinical trials in oncology. Metabolism of 4-HPR is of particular interest due to production of the active metabolite 4'-oxo 4-HPR and the clinical challenge of obtaining consistent 4-HPR plasma concentrations in patients. Here, we assessed the enzymes involved in various 4-HPR metabolic pathways.


METHODS
Enzymes involved in 4-HPR metabolism were characterized using human liver microsomes (HLM), supersomes over-expressing individual human cytochrome P450s (CYPs), uridine 5'-diphospho-glucoronosyl transferases (UGTs) and CYP2C8 variants expressed in Escherichia coli. Samples were analysed by high-performance liquid chromatography and liquid chromatography/mass spectrometry assays and kinetic parameters for metabolite formation determined. Incubations were also carried out with inhibitors of CYPs and methylation enzymes.


RESULTS
HLM were found to predominantly produce 4'-oxo 4-HPR, with an additional polar metabolite, 4'-hydroxy 4-HPR (4'-OH 4-HPR), produced by individual CYPs. CYPs 2C8, 3A4 and 3A5 were found to metabolize 4-HPR, with metabolite formation prevented by inhibitors of CYP3A4 and CYP2C8. Differences in metabolism to 4'-OH 4-HPR were observed with 2C8 variants, CYP2C8*4 exhibited a significantly lower V(max) value compared with *1. Conversely, a significantly higher V(max) value for CYP2C8*4 versus *1 was observed in terms of 4'-oxo formation. In terms of 4-HPR glucuronidation, UGTs 1A1, 1A3 and 1A6 produced the 4-HPR glucuronide metabolite.


CONCLUSIONS
The enzymes involved in 4-HPR metabolism have been characterized. The CYP2C8 isoform was found to have a significant effect on oxidative metabolism and may be of clinical relevance.
Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne, UK.
N A
NA
Illingworth
A V
AV
Boddy
A K
AK
Daly
G J
GJ
Veal
D001189:Aryl Hydrocarbon Hydroxylases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D065727:Cytochrome P-450 CYP2C8; D051544:Cytochrome P-450 CYP3A / Q000235:genetics / Q000378:metabolism; D065692:Cytochrome P-450 CYP3A Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017313:Fenretinide / Q000031:analogs & derivatives / Q000378:metabolism*; D020719:Glucuronides / Q000378:metabolism; D014453:Glucuronosyltransferase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000187:drug effects; D007700:Kinetics; D008861:Microsomes / Q000187:drug effects / Q000378:metabolism; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism*; D050505:Mutant Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D014212:Tretinoin / Q000031:analogs & derivatives / Q000378:metabolism; D000099148:UGT1A1 Enzyme
D001189:Aryl Hydrocarbon Hydroxylases; D065727:Cytochrome P-450 CYP2C8; D051544:Cytochrome P-450 CYP3A; D065692:Cytochrome P-450 CYP3A Inhibitors; D003577:Cytochrome P-450 Enzyme System; D004791:Enzyme Inhibitors; D017313:Fenretinide; D020719:Glucuronides; D014453:Glucuronosyltransferase; D050505:Mutant Proteins; D011994:Recombinant Proteins; D014212:Tretinoin; D000099148:UGT1A1 Enzyme; C494898:4-oxofenretinide; C058346:N-(4-methoxyphenyl)retinamide; C450259:CYP2C8 protein, human; C510164:CYP3A5 protein, human; C510163:CYP3A4 protein, human; C122075:UDP-glucuronosyltransferase, UGT1A3; C110655:UDP-glucuronosyltransferase, UGT1A6