Metabolism of ochratoxin A: absence of formation of genotoxic derivatives by human and rat enzymes.
Ochratoxin A (OTA) is a potent renal carcinogen in male rats, although its mode of carcinogenicity is not known. The metabolism and covalent binding of OTA to DNA were investigated in vitro with cytochromes P450, glutathione S-transferases, prostaglandin H-synthase, and horseradish peroxidase. Incubation of OTA with rat or human liver microsomes fortified with NADPH resulted in formation of 4-(R)-hydroxyochratoxin A at low rates [10-25 pmol min(-1) (mg of protein)(-1)]. There was no evidence of OTA metabolism and glutathione conjugate formation with rat, mouse, or human kidney microsomes or postmitochondrial supernatants (S-9) [<5 pmol min(-1) (mg of protein)(-1)]. Recombinant human cytochromes P450 (P450) 1A1 and 3A4 formed 4-(R)-hydroxyochratoxin A at low rates [0.08 and 0.06 pmol min(-1) (pmol of P450)(-1), respectively]; no oxidation products of OTA were detected with recombinant human P450 1A2 or 2E1 or rat P450 1A2 or 2C11 [<0.02 pmol min(-1) (pmol of P450)(-1)]. Prostaglandin H-synthase produced small amounts of an apolar product [33 pmol min(-1) (mg of protein)(-1)], and OTA products were not formed with horseradish peroxidase. There was no evidence of DNA adduct formation when [(3)H]OTA was incubated with these enzyme systems in the presence of calf thymus DNA (<20 adducts/10(9) DNA bases); however, these enzymes catalyzed DNA adduct formation with the genotoxins aflatoxin B(1), 2-amino-3-methylimidazo[4,5-f]quinoline, benzo[a]pyrene, and pentachlorophenol. There was also no detectable [(3)H]OTA bound in vivo to kidney DNA of male Fischer-344 rats treated orally with [(3)H]OTA (1 mg/kg, 100 mCi/mmol, 24 h exposure, <2.7 adducts/10(9) DNA bases), based upon liquid scintillation counting. However, (32)P-postlabeling experiments did show evidence of DNA lesions with total adduct levels ranging from 31 to 71 adducts/10(9) DNA bases, while adducts in untreated rat kidney ranged from 6 to 24 adducts/10(9) DNA bases. These results do not support the premise that OTA or metabolically activated species covalently bind to DNA and suggest that the (32)P-postlabeled lesions are due to products derived from OTA-mediated cytotoxicity.
Gautier
J
J
Nestlé Research Center, Nestec Ltd., Vers-chez-les-Blanc, 1000 Lausanne 26, Switzerland.
Richoz
J
J
Welti
D H
DH
Markovic
J
J
Gremaud
E
E
Guengerich
F P
FP
Turesky
R J
RJ
D000818:Animals; D001711:Biotransformation; D002273:Carcinogens / Q000378:metabolism* / Q000493:pharmacokinetics / Q000633:toxicity; D002417:Cattle; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D005260:Female; D005982:Glutathione Transferase / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D007668:Kidney / Q000201:enzymology / Q000378:metabolism; D015227:Lipid Peroxidation / Q000187:drug effects; D008297:Male; D008315:Malondialdehyde / Q000378:metabolism; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism; D009183:Mycotoxins / Q000378:metabolism* / Q000493:pharmacokinetics / Q000633:toxicity; D009793:Ochratoxins / Q000378:metabolism* / Q000493:pharmacokinetics / Q000633:toxicity; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism; D051381:Rats; D011916:Rats, Inbred F344; D013268:Stimulation, Chemical
D002273:Carcinogens; D007527:Isoenzymes; D009183:Mycotoxins; D009793:Ochratoxins; D011506:Proteins; C025589:ochratoxin A; D008315:Malondialdehyde; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D011451:Prostaglandin-Endoperoxide Synthases; D005982:Glutathione Transferase