Jiang Hao, Shen Yu-Min, Quinn Amy M, Penning Trevor M
Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6084, USA.
Chem Res Toxicol. 2005 Feb;18(2):365-74. doi: 10.1021/tx0497245.
(+/-)-7,8-Dihydroxy-7,8-dihydrobenzo[a]pyrene (BP-7,8-diol), a proximate carcinogen derived from benzo[a]pyrene (BP) requires further metabolic activation to exert its carcinogenic effects. Two principal pathways have been implicated, and these involve either the formation of (+/-)-trans-7,8-dihydroxy-9alpha,10alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (anti-BPDE) catalyzed by P450 1A1/P450 1B1 (NADPH-dependent monoxygenases) or the formation of benzo[a]pyrene-7,8-dione (BP-7,8-dione) catalyzed by human aldo-keto reductases AKR1A1 and AKR1C1-AKR1C4 [NAD(P)(H)-dependent oxidoreductases]. The relative contributions of the two pathways to PAH activation are unknown. In this study, BP-7,8-diol metabolism was studied in human bronchoalveolar H358 cell extracts. Parental H358 cells do not constitutively express P450 1A1/P450 1B1 or AKRs but were manipulated by induction with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to express P450 1A1/P450 1B1 or by stable transfection to express AKR1A1 (aldehyde reductase). TCDD induction of AKR1A1 transfectants provided a cell line that expressed both pathways. Extracts derived from parental H358 cells plus TCDD (P450 induction) produced electrophilic anti-BPDE, which hydrolyzed to benzo[a]pyrene tetrahydrotetrols (BP-tetrols), extracts derived from AKR1A1-transfected cells (AKR1A1 expression) produced reactive and redox-active BP-7,8-dione, which was trapped in situ as its mono(thioether) conjugate, and extracts derived from AKR1A1 transfectants plus TCDD (coexpression of P450 1A1/P450 1B1 and AKR1A1) produced both anti-BPDE and BP-7,8-dione. The competing activation of BP-7,8-diol by P450 1A1/P450 1B1 and AKR1A1 was studied with varied NADPH:NAD+ ratios. The system with a relatively higher concentration of NADPH favored formation of anti-BPDE via P450 1A1/P450 1B1, while the system with the higher concentration of NAD+ favored formation of BP-7,8-dione via AKR1A1. Under conditions that mimic the cellular redox state, 10 microM NADPH and 1 mM NAD+, equal amounts of BP-tetrols and BP-7,8-dione were formed. This suggests that P450 1A1/P450 1B1 and AKR1A1 play competing roles in the metabolic activation of BP-7,8-diol and that the dominant pathway of BP-7,8-diol activation depends on the redox state of the cells. These model systems provide a cellular context in which the dominant DNA adducts/lesions formed by either pathway may be compared.
（±）-7,8-二羟基-7,8-二氢苯并[a]芘（BP-7,8-二醇）是一种由苯并[a]芘（BP）衍生而来的近致癌物，需要进一步的代谢活化才能发挥其致癌作用。已经涉及两条主要途径，这些途径涉及由P450 1A1/P450 1B1（NADPH依赖性单加氧酶）催化形成（±）-反式-7,8-二羟基-九α,10α-环氧-7,8,9,10-四氢苯并[a]芘（反式-BPDE），或者由人醛糖酮还原酶AKR1A1和AKR1C1-AKR1C4（NAD(P)(H)依赖性氧化还原酶）催化形成苯并[a]芘-7,8-二酮（BP-7,8-二酮）。这两条途径对多环芳烃活化的相对贡献尚不清楚。在本研究中，在人支气管肺泡H358细胞提取物中研究了BP-7,8-二醇的代谢。亲代H358细胞不组成性表达P450 1A1/P450 1B1或醛糖酮还原酶，但通过用2,3,7,8-四氯二苯并对二恶英（TCDD）诱导来操纵以表达P450 1A1/P450 1B1或通过稳定转染来表达AKR1A1（醛还原酶）。AKR1A1转染子的TCDD诱导提供了一个表达两条途径的细胞系。来自亲代H358细胞加TCDD（P450诱导）的提取物产生亲电的反式-BPDE，其水解为苯并[a]芘四氢四醇（BP-四醇）；来自AKR1A1转染细胞（AKR1A1表达）的提取物产生具有反应性和氧化还原活性的BP-7,8-二酮，其以其单（硫醚）共轭物的形式原位捕获；来自AKR1A1转染子加TCDD（P450 1A1/P450 1B1和AKR1A1共表达）的提取物产生反式-BPDE和BP-7,8-二酮。用不同的NADPH:NAD+比率研究了P450 1A1/P450 1B1和AKR1A1对BP-7,8-二醇的竞争性活化。具有相对较高浓度NADPH的系统有利于通过P450 1A1/P450 1B1形成反式-BPDE，而具有较高浓度NAD+的系统有利于通过AKR1A1形成BP-7,8-二酮。在模拟细胞氧化还原状态的条件下，10μM NADPH和1 mM NAD+，形成等量的BP-四醇和BP-7,8-二酮。这表明P450 1A1/P450 1B1和AKR1A1在BP-7,8-二醇的代谢活化中起竞争作用，并且BP-7,8-二醇活化的主要途径取决于细胞的氧化还原状态。这些模型系统提供了一个细胞环境，在其中可以比较由任一途径形成的主要DNA加合物/损伤。
