The Carcinogen-DNA Interactions Section investigates both the extent of DNA adduct formation and persistence, and the biological consequences of DNA damage. We utilize experimental models and human subjects, with the intention of applying the knowledge gained to elucidate mechanisms of cancer causation and approaches to cancer prevention in humans. One goal is to ascertain an association between DNA adduct formation and human cancer risk. Ongoing studies in 3 different populations are focused on the validation of human polycyclic aromatic hydrocarbon (PAH)-DNA adduct formation as a biomarker for risk of human cancer. In US Navy officers a 2-fold increased risk of colon adenoma was found in individuals with the highest leukocyte PAH-DNA adduct level. Using esophageal samples from Linxian, China, where the esophageal cancer mortality rate is 20%, we have validated PAH-DNA adduct semi-quantitative measurements by immunohistochemistry in biopsies taken in 1985 and 1994, and currently we are investigating a possible association between PAH-DNA adduct formation and cancer risk within a prospective nested case-control study design. Finally, we measured PAH-DNA adducts in cervix biopsies from women infected with human papilloma virus (HPV), and we are examining the association between HPV status, PAH-DNA adducts in cervix and a documented smoking-related increase in cervical cancer risk. The potential formation of tamoxifen (TAM)-DNA adducts in human endometrium is a controversial topic of interest, as TAM-exposed women are at risk for endometrial cancer. Modeling of human TAM exposures in female cynomolgus monkeys revealed TAM-DNA adduct formation in ovary and uterus, but our current efforts to measure TAM-DNA adducts in human endometrium have been inconclusive. To validate methods used for TAM-DNA determination in human samples we organized an interlaboratory study using rat liver DNA, and demonstrated similarity for all the methods. Hypothesizing that TAM-induced gene expression changes may provide insight into breast tissue response, TAM-exposed cultured normal human mammary epithelial cell (NHMEC) strains were subjected to microarray. We found up-regulation of immune response genes in 3 strains, suggesting a novel mechanism of TAM activity in the breast.DNA damage response was explored in mouse keratinocytes ablated for the p21WAF1/Cip1 gene, and the data suggest that this gene may have a DNA repair-like function in mouse skin cells. Drug combinations that include one or two antiretroviral nucleoside reverse transcriptase inhibitors (NRTIs) are standard of care therapy for individuals infected with the human immunodeficiency virus (HIV). However, mitochondrial toxicities can limit NRTI use, and genotoxicity studies suggest that exposure may confer a long-term cancer risk. Currently we are investigating genotoxicity and mitochondrial toxicity associated with NRTIuse, with special focus on fetal consequences of maternal NRTI exposures during pregnancy. Since the last Site Visit unique methods have been developed to measure incorporation of the NRTI Lamivudine (3TC) into DNA, to quantify mtDNA levels in human and monkey tissues, and to determine mitochondrial gene expression by mitochondrial microarray. In cell culture studies that employed transformed human cell lines (HeLa and MOLT-3) and NHMEC strains, exposure to the NRTI Zidovudine (AZT) induced accumulation of cells in S-phase, accompanied by cell cycle gene expression changes consistent with the S-phase arrest. In addition, exposure of MOLT-3 cells to AZT for ?r5 passages resulted in loss of thymidine kinase 1 (TK1), the enzyme responsible for AZT mono-phosphorylation, suggesting a mechanism for drug resistance. Long-term mitochondrial toxicity was examined in HeLa cells exposed to AZT for 71 passages.
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