Pharmacokinetics and metabolism in rats of 2,4-diamino-6-benzyloxy-5- nitrosopyrimidine, an inactivator of O6-alkylguanine-DNA alkyltransferase.
Inactivation of the human DNA repair protein, O6-alkylguanine-DNA-alkyltransferase (AGT), by exposure to O6-benzylguanine leads to a dramatic enhancement in the cytotoxic response of cells to chemotherapeutic alkylnitrosoureas. Benzylated pyrimidines identified as more potent inactivators than O6-benzylguanine in vitro include 2,4-diamino-6-benzyloxy-5-nitrosopyrimidine (5-nitroso-BP) and 2,4-diamino-6-benzyloxy-5-nitropyrimidine (5-nitro-BP). In efforts to determine the clinical usefulness of these benzylated pyrimidines, we examined the metabolism and pharmacokinetics of 5-nitroso-BP in Sprague-Dawley rats, together with its potency as an AGT inactivator in mice. The mean plasma half-life, clearance, and volume of distribution of 5-nitroso-BP in rats were, respectively, 3.8 min, 22 liters/hr/kg, and 2.1 liters/kg. Two metabolites were identified in rat plasma (i.e. 5-nitro-BP and 2,4,5-triamino-6-benzyloxypyrimidine) after intravenous administration of 5-nitroso-BP in rat. Reduction of 5-nitroso-BP (100 microM) occurred primarily in cytosol and was inhibited (> 95%) by 1 mM menadione. Dicumarol (100 microM), a DT-diaphorase inhibitor, did not significantly inhibit this reaction. This indicated a possible role of a dicumarol-resistant quinone reductase. At higher substrate and protein concentration, NADPH-dependent oxidation of 5-nitroso-BP to 5-nitro-BP primarily occurred in microsomes and was completely inhibited by 1-aminobenzotriazole (1 mM), a P450 inhibitor. Unfortunately, neither 5-nitroso-BP nor 5-nitro-BP was as effective as O6-benzylguanine at depleting AGT activity in mouse liver or spleen. At 1 hr after injection of 15 mg/kg O6-benzylguanine, 5-nitroso-BP, or 5-nitro-BP, AGT levels in liver fell to 1%, 66%, and 71% basal activity, respectively. Rapid cytosolic reduction of 5-nitroso-BP may explain the lack of potency of the pyrimidines in vivo.
Roy
S K
SK
Department of Medicine, University of Chicago, IL 60637, USA.
Moschel
R C
RC
Dolan
M E
ME
D000818:Animals; D004791:Enzyme Inhibitors / Q000097:blood / Q000378:metabolism / Q000493:pharmacokinetics*; D005260:Female; D006801:Humans; D008297:Male; D008780:Methyltransferases / Q000037:antagonists & inhibitors*; D051379:Mice; D019853:O(6)-Methylguanine-DNA Methyltransferase; D011743:Pyrimidines / Q000097:blood / Q000378:metabolism / Q000493:pharmacokinetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013347:Subcellular Fractions / Q000378:metabolism
C104367:2,4-diamino-6-benzyloxy-5-nitrosopyrimidine; D004791:Enzyme Inhibitors; D011743:Pyrimidines; D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase