Role of 1,N2-propanodeoxyguanosine adducts as endogenous DNA lesions in rodents and humans.

Results obtained in a number of studies in vitro and in vivo support the hypothesis that short- and long-chain enals and their epoxides derived from oxidized polyunsaturated fatty acids are potential endogenous sources of cyclic propano and etheno DNA adducts. We previously reviewed the evidence from some of these studies. Here, we describe the results of our more recent studies on the role of 1,N2-propanodeoxyguanosine adducts as endogenous DNA lesions. These studies include: the detection of distinct patterns of such adducts in various tissues of different species; the detection of long-chain trans-4-hydroxynonenal-derived deoxyguanosine adducts in vivo; the specificity of the formation of enal-derived propano adducts from omega-3 and omega-6 polyunsaturated fatty acids; and the detection of acrolein- and crotonaldehyde-derived adducts in human oral tissue DNA and their increased levels in smokers. Taken together, these studies further strengthen the hypothesis that enals produced by lipid peroxidation are the primary source for cyclic propano adducts in vivo, but these results cannot rule out the possible contribution of environmental and other sources. The mutagenicity of enals and their epoxides and the results of site-specific mutagenesis studies indicate that the cyclic adducts are potential promutagenic lesions; however, only circumstantial evidence is currently available for their role in carcinogenesis.

1,N2-propanodeoxyguanosine adducts: potential new biomarkers of smoking-induced DNA damage in human oral tissue.

Highly DNA-reactive alpha,beta-unsaturated aldehydes such as acrolein and crotonaldehyde are common environmental pollutants present in cigarette smoke and automobile exhaust and are also released endogenously by lipid peroxidation. Acrolein- and crotonaldehyde-derived 1,N2-propanodeoxyguanosine (AdG and CdG, respectively) have been detected in the tissues of carcinogen-treated rodents and as background lesions in DNA from humans and untreated rodents. To determine whether cigarette smoking increases the levels of AdG and CdG, gingival tissue DNA from 11 smokers (4 males and 7 females; 30-58 years old) and 12 nonsmokers (8 males and 4 females; 21-66 years old) was analyzed using a previously described 32P-postlabeling high-performance liquid chromatography method. The results showed that the mean AdG levels in smokers were significantly higher than those in nonsmokers (1.36 +/- 0.90 micromol/mol guanine in smokers versus 0.46 +/- 0.26 micromol/mol guanine in nonsmokers; P = 0.003). The mean CdG 1 levels in smokers and nonsmokers were 0.53 +/- 0.44 and 0.06 +/- 0.07 micromol/mol guanine, respectively, corresponding to an 8.8-fold increase for smokers (P = 0.0015). Similar to CdG 1, levels of CdG 2 were increased 5.5-fold in smokers as compared to nonsmokers, from 0.31 +/- 0.40 to 1.72 +/- 1.26 micromol/mol guanine (P = 0.0014). Furthermore, the total levels of cyclic adduct (AdG and CdG) in smokers were 4.4-fold greater than those in nonsmokers (P = 0.0003). This study shows the detection of the potentially promutagenic 1,N2-propanoguanine adducts in human oral tissues and demonstrates for the first time an increase of structurally identified adducts in oral tissue DNA by cigarette smoking.

Effect of glutathione depletion on exocyclic adduct levels in the liver DNA of F344 rats.

The effects of glutathione (GSH) depletion on the in vivo formation of cyclic 1,N2- propanodexoxyguanosine adducts (AdG and CdG) as background lesions in the liver DNA of F344 rats were investigated. A group of 5 male F344 rats were given drinking water containing 30 mM L-buthionine (S,R)-sulfoximine (BSO) for 21 days, and another group of 8 rats were given only drinking water as controls. The BSO-treated rats had significantly lower weight gain than control rats. The hepatic GSH levels in the BSO-treated group were reduced by 84% as compared with the control group, from 4.43 to 0.72 mumol/g of tissue. The isomeric AdG3, CdG1, and CdG2 were detected by the 32P-postlabeling/HPLC method in the liver DNA of rats without carcinogen treatment, as we reported previously [Nath, R. G., and Chung, F.-L. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 7491-7495. Nath, R. G., et al. (1996) Cancer Res. 56, 452-456]. The mean levels (mumol/mol of guanine) for AdG3, CdG1, and CdG2 were 0.57 +/- 0.25, 0.15 +/- 0.18, and 0.16 +/- 0.22 for the control group and 1.18 +/- 1.03, 3.16 +/- 3.26, and 2.50 +/- 2.59 for the BSO group, respectively. These increases correspond to approximately 2-fold for AdG and 15-21-fold for CdG adducts. The dramatic increase in the cyclic adduct levels in rat liver DNA could have resulted mainly from GSH depletion as a result of the BSO treatment, even though other unknown effects due to the toxicity of BSO cannot be ruled out. These results suggest that GSH plays an important role in protecting the liver against cyclic propano DNA adduction and provide further support for the endogenous origin of these adducts.

Detection of 1, N2-propanodeoxyguanosine adducts as potential endogenous DNA lesions in rodent and human tissues.

Our previous study (R.G. Nath and F-L. Chung; Proc. Natl. Acad. Sci. USA, 91: 7491-7495, 1994), using a 32P postlabeling method combined with high-performance liquid chromatography specifically developed for exocyclic adducts, has shown that acrolein- and crotonaldehyde-derived 1,N2-propanodeoxyguanosine adducts (AdG and CdG, respectively) are present in the liver DNA from humans and rodents without carcinogen treatment. Those findings raised important questions regarding their role as potential endogenous DNA lesions in carcinogenesis. In this study, using a similar assay, we examined a variety of tissues from untreated rats and mice (lung, kidney, brain, breast, prostate, colon, skin, and leukocytes) and detected AdG and CdG in the DNA of these tissues. More significantly, we also obtained evidence for the presence of these adducts in the DNA of human leukocytes and mammary glands. The identities of these adducts were verified by comigration of 3', 5' -bisphosphates of the 32P-labeled adduct from DNA with the synthetic standards in a reversed-phased high-performance liquid chromatography. Additional proof of identities was provided by enzymatic conversion of AdG and CdG 3',5' -bisphosphates to the corresponding 5'-monophosphates, followed by comigration with their synthetic standards. The estimated ranges of total AdG and CdG modifications in DNA of various tissues were from 0.10 to 1.60 mumol/mol guanine for humans, based on the recoveries of external standards. This study demonstrated the ubiquity of these adducts in various tissues, suggesting their potential role as endogeneous DNA lesions in rodents and humans.