Modification of Nucleotide Excision Repair Activity by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and Sulforaphane

Abstract

The studies described in this thesis investigated the relationship between the DNA repair pathway nucleotide excision repair (NER, which is critical for protecting against carcinogenesis), and two exogenous chemicals, the tobacco carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and the nutraceutical sulforaphane. NER activity in female A/J mouse liver nuclear extracts decreased by 46% 12 h post NNK treatment and increased by 48% 24 h post NNK. These changes in NER activity were not attributed to alterations in levels of the NER proteins XPC, XPA, XPB and p53. However, the binding of hepatic XPA and XPB to damaged DNA at both timepoints was increased, while the binding of XPC to damaged DNA was unchanged at 12 h and decreased at 24 h. A 63% decrease in NER activity in female A/J mouse lung nuclear protein extracts 24 h post NNK treatment was not attributed to changes in levels of XPC, XPA, XPB or p53. However, the binding of lung XPA and XPB to damaged DNA was decreased and the binding of XPC to damaged DNA was increased at 24 h. Hence, NNK-induced NER changes are time and organ dependent and are associated with changes in the binding activities, but not the levels, of specific recognition and early excision NER proteins. A dose (100 mg/kg) and timepoint (6 h) of maximal effect of sulforaphane on female CD-1 mouse hepatic and pulmonary mRNA levels of two genes regulated by Nrf2, a major signalling pathway of sulforaphane mediated-effects, were found. A 50% increase in NER activity was observed in liver nuclear extracts 12 h post sulforaphane treatment, while lung NER at 12 h and liver NER at 6 h were not affected by sulforaphane. The increase in hepatic NER activity at 12 h was not associated with changes in levels of XPC, XPA, XPB or p53 or in the binding of hepatic XPC, XPA and XPB to damaged DNA. These results suggest that the impact of sulforaphane on repair-associated processes is time and organ dependent and is not associated with changes in the levels or activities of these specific recognition and early excision NER proteins.