Roy S K, Moschel R C, Dolan M E
Department of Medicine, University of Chicago, IL 60637, USA.
Drug Metab Dispos. 1996 Nov;24(11):1205-11.
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.
通过暴露于O6-苄基鸟嘌呤使人DNA修复蛋白O6-烷基鸟嘌呤-DNA烷基转移酶（AGT）失活，会导致细胞对化疗性烷基硝基脲的细胞毒性反应显著增强。在体外被鉴定为比O6-苄基鸟嘌呤更有效的失活剂的苄基化嘧啶包括2,4-二氨基-6-苄氧基-5-亚硝基嘧啶（5-亚硝基-BP）和2,4-二氨基-6-苄氧基-5-硝基嘧啶（5-硝基-BP）。为了确定这些苄基化嘧啶的临床实用性，我们研究了5-亚硝基-BP在Sprague-Dawley大鼠中的代谢和药代动力学，以及其在小鼠中作为AGT失活剂的效力。5-亚硝基-BP在大鼠中的平均血浆半衰期、清除率和分布容积分别为3.8分钟、22升/小时/千克和2.1升/千克。在大鼠静脉注射5-亚硝基-BP后，在大鼠血浆中鉴定出两种代谢物（即5-硝基-BP和2,4,5-三氨基-6-苄氧基嘧啶）。5-亚硝基-BP（100微摩尔）的还原主要发生在胞质溶胶中，并被1毫摩尔甲萘醌抑制（>95%）。双香豆素（100微摩尔），一种DT-黄递酶抑制剂，并未显著抑制该反应。这表明一种双香豆素抗性醌还原酶可能发挥作用。在较高的底物和蛋白质浓度下，5-亚硝基-BP向5-硝基-BP的NADPH依赖性氧化主要发生在微粒体中，并被1-氨基苯并三唑（1毫摩尔），一种P450抑制剂完全抑制。不幸的是，在耗尽小鼠肝脏或脾脏中的AGT活性方面，5-亚硝基-BP和5-硝基-BP都不如O6-苄基鸟嘌呤有效。在注射15毫克/千克O6-苄基鸟嘌呤、5-亚硝基-BP或5-硝基-BP后1小时，肝脏中的AGT水平分别降至基础活性的1%、66%和71%。5-亚硝基-BP在胞质溶胶中的快速还原可能解释了这些嘧啶在体内效力不足的原因。
