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Review
. 2013 Sep;84(3):304-13.
doi: 10.1124/mol.113.086637. Epub 2013 Jun 12.

Oral benzo[a]pyrene: understanding pharmacokinetics, detoxication, and consequences--Cyp1 knockout mouse lines as a paradigm

Affiliations
Review

Oral benzo[a]pyrene: understanding pharmacokinetics, detoxication, and consequences--Cyp1 knockout mouse lines as a paradigm

Daniel W Nebert et al. Mol Pharmacol. 2013 Sep.

Abstract

Benzo[a]pyrene (BaP) is a prototypical polycyclic aromatic hydrocarbon (PAH); this ubiquitous environmental carcinogenic agent is found in tobacco smoke, charcoal-grilled foods, and PAH-contaminated surfaces of roofs, playgrounds, and highways. Cytochrome P450 1 wild-type, Cyp1a2(-/-), Cyp1b1(-/-), or Cyp1a2/1b1(-/-) knockouts, and mice with Cyp1a1 expression deleted in hepatocytes can ingest large oral BaP doses (125 mg/kg/d) without apparent toxicity. Cyp1a1(-/-) and Cyp1a1/1a2(-/-) knockouts and mice with Cyp1a1 expression deleted in gastrointestinal (GI) tract epithelial cells develop immunotoxicity and die within 32 days, indicating that GI tract inducible CYP1A1 is absolutely required for detoxication of oral BaP. Cyp1a1/1b1(-/-) and Cyp1a1/1a2/1b1(-/-) mice are rescued from immunosuppression and early death due to absent metabolic activation of BaP by CYP1B1 in immune cells. Ten-fold lower oral BaP doses result in adenocarcinoma of the proximal small intestine (PSI) in Cyp1a1(-/-) mice; Cyp1a1/1b1(-/-) double-knockout mice show no PSI cancer but develop squamous cell carcinoma of the preputial gland duct (PGD). BaP-metabolizing CYP1B1 in the PSI and CYP3A59 in the PGD are the most likely candidates to participate in tumor initiation in the epithelial cells of these two tissues; oncogenes and tumor-suppressor genes upregulated and downregulated during tumorigenesis are completely different between these tissues. This "oral BaP Cyp1" mouse paradigm represents a powerful teaching tool, showing that gene-environment interactions depend on route-of-administration: the same oral, but not intraperitoneal, BaP exposure leads to dramatic differences in target-organ toxicity and tumor type as a function of dose and Cyp1 genotype.

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Figures

Fig. 1.
Fig. 1.
Molecular structure of the prototypical polycyclic aromatic hydrocarbon, benzo[a]pyrene (BaP), with its standardized numbering system. Among 709 possible oxygenated BaP metabolites, which includes all syn- and anti-isomers, four products (or classes of products) of metabolic activation that are highly reactive intermediates are listed in the left column; four products (or classes of products) of detoxication that are negligibly reactive are listed in the right column. Electrophilic oxides and epoxides can react with nucleophilic groups of cellular macromolecules, can rearrange to become hydroxyl products, or can become conjugated with moieties such as glucuronide, glutathione, or sulfate. Phenols and the tetrol are generally nontoxic, and conjugation renders them even more hydrophilic and easy to excrete from the cell. Whereas the epoxide oxygen is derived from diatomic oxygen, the addition of a second oxygen atom across the same C—C bond to form a diol comes from cellular water. aBaP phenols are major nonreactive metabolites and therefore represent important detoxication pathways; all BaP phenols have been tested for carcinogenicity and are inactive except for 2-hydroxyBaP, which is active but not a known biologic metabolite. bOnly one of the four possible optically active diol-epoxide metabolites is carcinogenic. cAlthough the 7,8-diol is not “reactive,” it is highly carcinogenic because of its further metabolism. dWhereas BaP 4,5-oxide is chemically reactive and also mutagenic, it is not carcinogenic. In summary, some chemically reactive metabolites are not carcinogenic but can be toxic and/or mutagenic, whereas some nonreactive metabolites can be carcinogenic, toxic, and/or mutagenic due to further metabolism (reviewed in Conney et al., 1994).
Fig. 2.
Fig. 2.
Comparisons of the response of oral BaP (125 mg/kg/d) for 18 days in mice of different genotypes. BPO denotes the first step in oxygenated BaP; BPO can be the reactive intermediate as well as undergo all the detoxication possibilities, as detailed in Fig. 1. The ratio of reactive-intermediates-to-detoxified-products is most likely tissue or cell-type specific, depends on how “tightly coupled” phase II conjugation systems might be to the membrane-bound P450 enzymes (Nebert and Dalton, 2006), and is also likely to depend on the degree of CYP1 induction. The relative amounts of CYP1A1, CYP1A2, and CYP1B1 protein in each tissue or cell type and the route and rate of administration as well as the function of time during induction by BaP will also affect this ratio. (A) Cyp1(+/+) wild-type (right) versus Cyp1a1(−/−) single-knockout mice (left). Oral BaP in Cyp1(+/+) mice rapidly induces intestinal CYP1A1, leading to detoxication and rapid excretion of BaP metabolites. Absence of CYP1A1 in the GI tract of Cyp1a1(−/−) mice (left) results in negligible detoxication and therefore a 25-fold greater blood BaP level than in the wild-type mice; larger amounts of BaP reaching nonhepatic tissues then result in BaP-induced CYP1B1-mediated immunosuppression, wasting, and death. (B) Cyp1(+/+) (right) versus Cyp1a1/1b1(−/−) double-knockout mice. Lack of CYP1B1 in marrow and immune tissues (left) results in negligible metabolic activation of BaP and thus markedly diminished immunosuppression, toxicity, and prevention of early death—despite a 3-fold more BaP body burden than in the Cyp1a1(−/−) mice seen in panel A. (C) Alb>Cre>Cyp1a1(f/f) versus Vil>Cre>Cyp1a1(f/f) conditional knockout mice. Alb>Cre>Cyp1a1(f/f) mice (right) respond to oral BaP much like wild-type mice, whereas Vil>Cre>Cyp1a1(f/f) mice (left) respond much like the Cyp1a1(−/−) mice; these experiments demonstrate unequivocally that it is the CYP1A1 in the GI tract, not the liver, that is most important in oral BaP detoxication and hence protection from immunosuppression, immunotoxicity, and early death.

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