Motwani Hitesh V, Westberg Emelie, Lindh Christian, Abramsson-Zetterberg Lilianne, Törnqvist Margareta
Department of Environmental Science and Analytical Chemistry, Stockholm University, SE-106 91 Stockholm, Sweden.
Division of Occupational and Environmental Medicine, Institution of Laboratory Medicine, Lund University, SE-221 85 Lund, Sweden.
Mutat Res Genet Toxicol Environ Mutagen. 2020 Jan;849:503127. doi: 10.1016/j.mrgentox.2019.503127. Epub 2019 Nov 15.
The environmental and food contaminant, benzo[a]pyrene {B[a]P, a polycyclic aromatic hydrocarbon (PAH)}, is classified as a human carcinogen by the International Agency for Research on Cancer. The carcinogenicity of B[a]P is linked to the formation of electrophilic metabolites, namely B[a]P-diol epoxides (BPDEs) occurring as stereoisomers. In this work, we quantified the metabolic formation of BPDE isomers and the genotoxic effect in B[a]P-exposed mice, with an aim to estimate the genotoxic potency of B[a]P per in vivo dose of its most potent metabolite [i.e. (+)-anti-BPDE]. The increase in frequency of micronuclei (fMN) in erythrocytes was measured as a biomarker for genotoxic effect. Covalent adducts to serum albumin (SA) and those to DNA from the BPDEs were analysed using liquid chromatography tandem mass spectrometry (LC-MS/MS), as adducts to histidine (BPDE-His-Pro) and deoxyguanosine (BPDE-dG), respectively. For the first time in animal experiments it was possible to resolve adducts to SA from (+)-anti-, (-)-anti- and (±)-syn-BPDE isomers by LC-MS/MS. The adduct levels in the protein were about 16 fmol/mg SA, which was orders of magnitude lower than that in the nucleic acid, 28 pmol/mg DNA, in mice exposed to 100 mg B[a]P per kg body weight (bw). Using SA adduct levels, the in vivo dose of (+)-anti-BPDE was calculated to be approximately 50 nM·h per mg B[a]P per kg bw. This allowed to make a preliminary estimate of the genotoxic potency as 2‰ fMN per μM·h of (+)-anti-BPDE. This estimate was compared to that from another food toxicant, glycidol, studied with similar methods, which indicated that the BPDE has several orders of magnitude higher genotoxic potency. The demonstrated approach on integrating biomarkers of internal dose of a causative agent and that of genotoxic effect for assessing genotoxic potency, using B[a]P as a model, has a potential for improving cancer risk assessment procedures for PAHs.
环境和食品污染物苯并[a]芘{B[a]P，一种多环芳烃（PAH）}被国际癌症研究机构列为人类致癌物。B[a]P的致癌性与亲电代谢产物的形成有关，即作为立体异构体存在的苯并[a]芘二醇环氧化物（BPDEs）。在这项工作中，我们对B[a]P暴露小鼠中BPDE异构体的代谢形成和遗传毒性效应进行了定量，目的是估计B[a]P每体内剂量其最具活性代谢产物[即(+)-反式-BPDE]的遗传毒性效力。红细胞中微核频率（fMN）的增加作为遗传毒性效应的生物标志物进行测量。使用液相色谱串联质谱法（LC-MS/MS）分析BPDEs与血清白蛋白（SA）以及与DNA的共价加合物，分别作为与组氨酸（BPDE-His-Pro）和脱氧鸟苷（BPDE-dG）的加合物。在动物实验中，首次能够通过LC-MS/MS分辨(+)-反式、(-)-反式和(±)-顺式-BPDE异构体与SA的加合物。在每千克体重（bw）暴露于100 mg B[a]P的小鼠中，蛋白质中的加合物水平约为16 fmol/mg SA，这比核酸中的水平低几个数量级，核酸中的水平为28 pmol/mg DNA。利用SA加合物水平，计算出(+)-反式-BPDE的体内剂量约为每毫克B[a]P每千克bw 50 nM·h。这使得能够初步估计遗传毒性效力为每μM·h(+)-反式-BPDE产生2‰ fMN。将该估计值与用类似方法研究的另一种食品毒物缩水甘油的估计值进行比较，结果表明BPDE的遗传毒性效力高几个数量级。以B[a]P为模型，所展示的整合致病因子内部剂量生物标志物和遗传毒性效应生物标志物以评估遗传毒性效力的方法，具有改进PAHs癌症风险评估程序的潜力。
