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Updated: Sep 26, 2026
Formation of Covalent DNA Adducts by Enzymatically Activated Carcinogens and Drugs In Vitro and Their Determination by 32P-postlabeling
Published on: March 20, 2018
Interactive effects of nrf2 genotype and oltipraz on benzo[a]pyrene-DNA adducts and tumor yield in mice
Minerva Ramos-Gomez1, Patrick M Dolan, Ken Itoh
1Department of Environmental Health Sciences, Bloomberg School of Public Health, The Johns Hopkins University, Baltimore, MD 21202, USA.
Abstract:
The cancer chemopreventive actions of oltipraz (4-methyl-5-[2-pyrazinyl]-1,2-dithiole-3-thione) have been primarily associated with the induction of phase 2 detoxifying enzymes through transcriptional activation of the antioxidant response element (ARE) in the promoter regions of these genes. The transcription factor Nrf2 has been shown to bind to and activate AREs. Previously, we demonstrated that nrf2-deficient mice had low basal expression of phase 2 enzymes and were substantially more susceptible to benzo[a]pyrene (B[a]P)-induced neoplasia of the forestomach than wild-type. Moreover, loss of Nrf2 abrogated the chemopreventive action of oltipraz, when administered 48 h before B[a]P, an interval allowing maximal induction of many phase 2 enzymes. Oltipraz also inhibits some cytochrome P450s involved in the bioactivation of B[a]P. In the present study we observed that oltipraz had no protective effect on tumor burden in the forestomach of nrf2-deficient mice when administered 1 h before B[a]P, a timeline that selectively optimizes for possible inhibitory effects on cytochrome P450s. To evaluate the role of nrf2 genotype on B[a]P disposition, levels of B[a]P-DNA adducts were measured as tetrols released from DNA isolated from target (forestomach) and non-target tissues (liver) of wild-type and nrf2-deficient mice treated with either vehicle or oltipraz 1 or 48 h before B[a]P. Levels of B[a]P-DNA adducts in forestomach were significantly higher in nrf2-deficient compared with wild-type mice. Oltipraz treatment at 1 or 48 h before B[a]P had no protective effect on forestomach tetrol levels in nrf2-deficient mice, whereas a significant reduction was observed in wild-type mice treated with oltipraz 48 h, but not 1 h, before carcinogen. Combining all treatments and genotypes, there was a strong correlation (R(2) = 0.91) between levels of B[a]P-DNA adducts in forestomach and subsequent yield of tumors. In contrast to the results in forestomach, nrf2 genotype did not modify hepatic B[a]P-DNA adduct levels while both oltipraz treatments were protective, suggesting that Nrf2-independent mechanisms (e.g. P450 inhibition) for oltipraz can also occur in vivo in some tissues.
Insights
The chemopreventive effects of oltipraz depend on the Nrf2 pathway for forestomach cancer prevention, as Nrf2-deficient mice showed no protection. Oltipraz also inhibits P450 enzymes, offering protection independent of Nrf2 in the liver.
Area of Science:
- Chemoprevention
- Cancer Research
- Molecular Toxicology
Background:
- Oltipraz (4-methyl-5-[2-pyrazinyl]-1,2-dithiole-3-thione) is known for cancer chemoprevention, primarily via inducing phase 2 detoxifying enzymes through the antioxidant response element (ARE) and Nrf2 transcription factor.
- Nrf2-deficient mice exhibit lower basal phase 2 enzyme expression and increased susceptibility to benzo[a]pyrene (B[a]P)-induced forestomach tumors compared to wild-type.
- Previous studies showed that Nrf2 deficiency abrogates oltipraz's chemopreventive action against B[a]P-induced neoplasia when administered 48 hours prior, a timeframe optimal for enzyme induction.
Purpose of the Study:
- To investigate the role of Nrf2 genotype in oltipraz's chemopreventive efficacy against B[a]P-induced forestomach tumors.
- To determine if oltipraz's inhibitory effects on cytochrome P450s, involved in B[a]P bioactivation, are Nrf2-dependent or independent.
- To assess the impact of oltipraz administration timing (1h vs. 48h pre-B[a]P) on chemoprevention in both wild-type and Nrf2-deficient mice.
Main Methods:
- Measurement of B[a]P-DNA adducts (as tetrols) in forestomach and liver DNA of wild-type and Nrf2-deficient mice treated with oltipraz (1h or 48h prior) or vehicle, followed by B[a]P administration.
- Evaluation of forestomach tumor burden in relation to B[a]P-DNA adduct levels and Nrf2 genotype.
- Comparison of oltipraz's protective effects on B[a]P disposition and tumorigenesis under different Nrf2 genotypes and treatment timings.
Main Results:
- Oltipraz failed to protect Nrf2-deficient mice from forestomach tumors when given 1h before B[a]P, indicating Nrf2 dependence for this effect.
- Forestomach B[a]P-DNA adduct levels were significantly higher in Nrf2-deficient mice than wild-type mice.
- Oltipraz administration 48h before B[a]P significantly reduced forestomach B[a]P-DNA adducts in wild-type mice but not in Nrf2-deficient mice.
- A strong correlation (R(2) = 0.91) was observed between forestomach B[a]P-DNA adduct levels and tumor yield across all treatment groups and genotypes.
- In contrast to forestomach results, Nrf2 genotype did not affect hepatic B[a]P-DNA adduct levels, and oltipraz provided protection in both genotypes, suggesting Nrf2-independent mechanisms.
Conclusions:
- The chemopreventive action of oltipraz against B[a]P-induced forestomach neoplasia is critically dependent on the Nrf2 pathway.
- Nrf2-independent mechanisms, likely involving cytochrome P450 inhibition, contribute to oltipraz's chemopreventive effects in tissues like the liver.
- The study highlights the tissue-specific roles of Nrf2 in mediating oltipraz's chemopreventive effects and carcinogen disposition.
