Lacroix D, Desrochers M, Castonguay A, Anderson A
Centre de recherche en cancérologie de l'Université Laval, L'Hôtel-Dieu de Québec, Canada.
Carcinogenesis. 1993 Aug;14(8):1639-42. doi: 10.1093/carcin/14.8.1639.
In all species where it has been tested, the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) has been shown to be a potent carcinogen, and NNK and other nitrosamines may play a role in human tobacco-related carcinogenesis. Purified rat CYP2B1 has been shown to metabolize NNK, and the CYP2B1 gene is expressed constitutively in rat lung. The objectives of this study were to test the capacity of CYP2B1, synthesized from a rat hepatic cDNA in Ad293 cells, to metabolize NNK, and to define the type and the proportions of the final metabolites produced. Ad293 cells were transfected with a CYP2B1 expression vector (pMT2-2B1), or with a control vector and incubated in culture medium containing [3H]NNK, after which alpha-carbon hydroxylation and pyridine N-oxidation metabolites were identified by HPLC analysis and quantitated by scintillation counting. pMT2-2B1-transfected cells were capable of catalyzing alpha-carbon hydroxylation and pyridine N-oxidation of NNK, although the reduction product 4-(methylnitrosamino)-1-(3-pyridyl)-1-butan-1-ol(NNAL) was the major metabolite formed in cells regardless of transfection treatment. The total amount of alpha-carbon hydroxylation metabolites produced by pMT2-2B1-transfected cells was greater than that of pyridine N-oxidation metabolites. However, pMT2-2B1 transfected cells produced approximately ten-fold more pyridine N-oxidation metabolites and only two-fold more alpha-carbon hydroxylation metabolites than control cells. Furthermore, the amount of NNAL-N-oxide was much lower than that of NNK-N-oxide in the medium of pMT2-2B1-transfected cells, even though the amount of available NNAL, resulting from carbonyl reduction of NNK, was very high; this suggests that NNAL is poorly N-oxidized by CYP2B1 compared to NNK. These results show that within living cells NNK was metabolized by CYP2B1 via both the pyridine N-oxidation and alpha-carbon hydroxylation pathways. However, CYP2B1 preferentially catalyzed pyridine N-oxidation, which is considered to be a deactivation reaction.
在所有已进行测试的物种中，烟草特有的亚硝胺4-（甲基亚硝胺基）-1-（3-吡啶基）-1-丁酮（NNK）已被证明是一种强效致癌物，并且NNK和其他亚硝胺可能在人类烟草相关的致癌过程中发挥作用。纯化的大鼠细胞色素P450 2B1（CYP2B1）已被证明可代谢NNK，并且CYP2B1基因在大鼠肺中组成性表达。本研究的目的是测试在Ad293细胞中由大鼠肝脏cDNA合成的CYP2B1代谢NNK的能力，并确定最终产生的代谢物的类型和比例。用CYP2B1表达载体（pMT2-2B1）或对照载体转染Ad293细胞，并在含有[3H]NNK的培养基中孵育，之后通过高效液相色谱分析鉴定α-碳羟基化和吡啶N-氧化代谢物，并通过闪烁计数进行定量。pMT2-2B1转染的细胞能够催化NNK的α-碳羟基化和吡啶N-氧化，尽管无论转染处理如何，还原产物4-（甲基亚硝胺基）-1-（3-吡啶基）-1-丁醇（NNAL）都是细胞中形成的主要代谢物。pMT2-2B1转染细胞产生的α-碳羟基化代谢物的总量大于吡啶N-氧化代谢物的总量。然而，pMT2-2B1转染的细胞产生的吡啶N-氧化代谢物比对照细胞多约十倍，而α-碳羟基化代谢物仅多两倍。此外，在pMT2-2B1转染细胞的培养基中，NNAL-N-氧化物的量远低于NNK-N-氧化物的量，尽管由NNK的羰基还原产生的可用NNAL的量非常高；这表明与NNK相比，CYP2B1对NNAL的N-氧化作用较弱。这些结果表明，在活细胞内，NNK通过吡啶N-氧化和α-碳羟基化途径被CYP2B1代谢。然而，CYP2B1优先催化吡啶N-氧化，这被认为是一种失活反应。
