Estrogen-DNA Adducts and Breast Cancer Risk in Premenopausal Asian Women.

The incidence of breast cancer among premenopausal women has been increasing rapidly in recent decades in East Asia. This case-control study investigated whether estrogen-DNA adducts were associated with breast cancer risk in Taiwan. The control group (n = 146) comprised healthy female volunteers and women with non-proliferative breast disease. The case group (n = 221) comprised women either with proliferative benign breast disease or breast cancer. The ratios of estrogen-DNA adducts to their respective metabolites and conjugates in plasma were analyzed using ultraperformance LC/MS-MS. The SNPs of CYP1A1, CYP1B1, and COMT were genotyped. Logistic regression model was used to compare the estrogen-DNA adduct ratios between the two groups. The estrogen-DNA adduct ratio in the case group was significantly higher than that in the control group (median ratio: 58.52 vs. 29.36, P = 0.004). A multiple logistic regression model demonstrated that a unit increase in the natural log of the estrogen-DNA adduct ratio in premenopausal women was a significant predictor of breast cancer risk, with an estimated hazard ratio of 1.718 (1.444-2.046, P < 0.001). However, the CYP1A1, CYP1B1, and COMT SNPs were not associated with the estrogen-DNA adduct ratios. In conclusion, plasma estrogen-DNA adduct ratio was associated with the presence of breast cancer or proliferating benign breast disease in premenopausal women in Taiwan. PREVENTION RELEVANCE: This study provides evidence that endogenous estrogen-induced genotoxicity may contribute to the carcinogenesis of breast cancer in premenopausal Asian women. This work could have important preventive implication for the emerging disease in East Asia.

Arsenic Exposure and Melanoma Among US Adults Aged 20 or Older, 2003-2016.

OBJECTIVES: Chronic exposure to arsenic has been reported as a risk factor for nonmelanoma skin cancer, notably squamous cell carcinoma. However, current knowledge is limited about the association between arsenic exposure and melanoma. Our objectives were to (1) measure the association between total urinary arsenic levels and melanoma compared with nonmelanoma skin cancer and no cancer and (2) analyze the association between water source and melanoma and nonmelanoma skin cancer. METHODS: We collected cross-sectional data from the 2003-2016 cycles of the National Health and Nutrition Examination Survey. We conducted univariate and multivariate logistic regressions. To evaluate the possible association of skin cancer with source of tap water, we calculated odds ratios for participants with melanoma and nonmelanoma skin cancer, compared with participants with no cancer. RESULTS: White race, higher education, higher socioeconomic status, and smoking history were associated with melanoma and nonmelanoma skin cancer in the full study population. After adjusting for age and race/ethnicity, the adjusted odds ratio of participants with >50 µg/L of total urinary arsenic for melanoma or nonmelanoma skin cancer was 1.87 (95% CI, 0.58-6.05) and 2.23 (95% CI, 1.12-4.45) times higher compared with no cancer, respectively. Participants with nonmelanoma skin cancer had 2.06 increased odds of reporting a nonmunicipal water source compared with participants without cancer. CONCLUSIONS: We did not find a relationship between the incidence of melanoma and exposure to arsenic among US adults. Nonmunicipal water sources were associated with nonmelanoma skin cancer and should be further investigated.

Low-Level Groundwater Atrazine in High Atrazine Usage Nebraska Counties: Likely Effects of Excessive Groundwater Abstraction.

Recent studies observed a correlation between estrogen-related cancers and groundwater atrazine in eastern Nebraska counties. However, the mechanisms of human exposure to atrazine are unclear because low groundwater atrazine concentration was observed in counties with high cancer incidence despite having the highest atrazine usage. We studied groundwater atrazine fate in high atrazine usage Nebraska counties. Data were collected from Quality Assessed Agrichemical Contaminant Nebraska Groundwater, Parameter-Elevation Regressions on Independent Slopes Model (PRISM), and water use databases. Descriptive statistics and cluster analysis were performed. Domestic wells (59%) were the predominant well type. Groundwater atrazine was affected by well depth. Clusters consisting of wells with low atrazine were characterized by excessive groundwater abstraction, reduced precipitation, high population, discharge areas, and metropolitan counties. Hence, low groundwater atrazine may be due to excessive groundwater abstraction accompanied by atrazine. Human exposure to atrazine in abstracted groundwater may be higher than the estimated amount in groundwater.

Risk of Breast Cancer Associated with Estrogen DNA Adduct Biomarker.

BACKGROUND: It is biologically plausible that genotoxic estrogens, namely estrogen DNA adducts (EDA), have a role in breast cancer development. Support comes from three prior studies that reported elevated concentrations of EDA relative to estrogen metabolites and conjugates (EDA:EMC) in women with breast cancer relative to control women. METHODS: In postmenopausal women in the Women's Health Initiative (WHI), EDA:EMC in 191 controls was compared with findings in 194 prediagnosis urine samples from breast cancer cases. EDA:EMC determinations were by mass spectrometry as previously described, and logistic regression was employed to estimate ORs. RESULTS: EDA:EMC did not differ in breast cancer cases compared with controls overall [0.93 (95% confidence interval, 0.71-1.23)], with a mean (SD) of 2.3 (0.8) and 2.4 (1.1) in cases and controls, respectively. Similarly, the ratio did not differ when examined by estrogen receptor or recency of biospecimen collection prior to breast cancer. CONCLUSIONS: Despite the demonstrated genotoxic properties of certain catechol estrogens resulting in EDAs, this analysis did not provide evidence for an increased breast cancer risk in relation to an elevated EDA:EMC. IMPACT: This analysis, conducted prospectively within postmenopausal women in the WHI study, suggests that a strong association between EDA:EMC and breast cancer could be ruled out, as this study was powered to detect an OR of 2.2 or greater.

Ultraviolet A light induces DNA damage and estrogen-DNA adducts in Fuchs endothelial corneal dystrophy causing females to be more affected.

Fuchs endothelial corneal dystrophy (FECD) is a leading cause of corneal endothelial (CE) degeneration resulting in impaired visual acuity. It is a genetically complex and age-related disorder, with higher incidence in females. In this study, we established a nongenetic FECD animal model based on the physiologic outcome of CE susceptibility to oxidative stress by demonstrating that corneal exposure to ultraviolet A (UVA) recapitulates the morphological and molecular changes of FECD. Targeted irradiation of mouse corneas with UVA induced reactive oxygen species (ROS) production in the aqueous humor, and caused greater CE cell loss, including loss of ZO-1 junctional contacts and corneal edema, in female than male mice, characteristic of late-onset FECD. UVA irradiation caused greater mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) damage in female mice, indicative of the sex-driven differential response of the CE to UVA, thus accounting for more severe phenotype in females. The sex-dependent effect of UVA was driven by the activation of estrogen-metabolizing enzyme CYP1B1 and formation of reactive estrogen metabolites and estrogen-DNA adducts in female but not male mice. Supplementation of N-acetylcysteine (NAC), a scavenger of reactive oxygen species (ROS), diminished the morphological and molecular changes induced by UVA in vivo. This study investigates the molecular mechanisms of environmental factors in FECD pathogenesis and demonstrates a strong link between UVA-induced estrogen metabolism and increased susceptibility of females for FECD development.

Loss of NQO1 generates genotoxic estrogen-DNA adducts in Fuchs Endothelial Corneal Dystrophy.

Fuchs Endothelial Corneal Dystrophy (FECD) is an age-related genetically complex disease characterized by increased oxidative DNA damage and progressive degeneration of corneal endothelial cells (HCEnCs). FECD has a greater incidence and advanced phenotype in women, suggesting a possible role of hormones in the sex-driven differences seen in the disease pathogenesis. In this study, catechol estrogen (4-OHE2), the byproduct of estrogen metabolism, induced genotoxic estrogen-DNA adducts formation, macromolecular DNA damage, and apoptotic cell death in HCEnCs; these findings were potentiated by menadione (MN)-mediated reactive oxygen species (ROS). Expression of NQO1, a key enzyme that neutralizes reactive estrogen metabolites, was downregulated in FECD, indicating HCEnC susceptibility to reactive estrogen metabolism in FECD. NQO1 deficiency in vitro exacerbated the estrogen-DNA adduct formation and loss of cell viability, which was rescued by the supplementation of N-acetylcysteine, a ROS scavenger. Notably, overexpression of NQO1 in HCEnCs treated with MN and 4-OHE2 quenched the ROS formation, thereby reducing the DNA damage and endothelial cell loss. This study signifies a pivotal role for NQO1 in mitigating the macromolecular oxidative DNA damage arising from the interplay between intracellular ROS and impaired endogenous estrogen metabolism in post-mitotic ocular tissue cells. A dysfunctional Nrf2-NQO1 axis in FECD renders HCEnCs susceptible to catechol estrogens and estrogen-DNA adducts formation. This novel study highlights the potential role of NQO1-mediated estrogen metabolite genotoxicity in explaining the higher incidence of FECD in females.

Role of Estrogen in Androgen-Induced Prostate Carcinogenesis in NBL Rats.

Androgens are thought to cause prostate cancer, but the underlying mechanisms are unclear. Data from animal studies suggest that for androgens to cause prostate cancer, they must be aromatized to estrogen and act in concert with estrogen metabolites. We tested the hypothesis that androgen-receptor and estrogen receptor-mediated effects of androgen and estrogen are necessary, as well as genotoxicity of estrogen metabolites. NBL rats were treated with androgenic and estrogenic compounds for 16-75 weeks through slow-release silastic implants or pellets. Testosterone alone induced cancer in the prostate of 37% of rats. 5α-Dihydrotestosterone, which cannot be converted to estradiol or testosterone, did not cause a significant prostate cancer incidence (4%). Addition of estradiol to 5α-dihydrotestosterone treatment did not markedly enhance prostate cancer incidence (14%), unlike adding estradiol to testosterone treatment which induced a 100% tumor incidence. Testosterone plus estradiol treatment induced a DNA adduct detectable by 32P-postlabeling, oxidative DNA damage (8-hydroxyguanosine), and lipid peroxidation at the site within the prostate where this treatment causes cancers, preceding later cancer formation. The non-estrogenic 4-hydroxy metabolite of estradiol, when combined with testosterone, induced prostatic dysplasia within 16 weeks and, after long-term treatment, a very low incidence of prostate cancer (21%). When an estrogen that cannot be hydroxylated (2-fluoroestradiol) was added to this combined treatment with testosterone and 4-hydroxyestradiol, dysplasia frequency after 16 weeks was doubled. These results strongly support the hypothesis, but additional definitive studies are needed which may identify new targets to interfere with these mechanisms that are clinically feasible in humans.

Estrogen Metabolism in African-American Women with and without Breast Cancer: A Pilot Study.

Studies in Caucasian women have shown that the formation of estrogen-DNA adducts is greater in women at high risk for breast cancer or already diagnosed with the disease. To begin investigating whether the role of estrogens in the etiology of breast cancer is similar in African-American (AA) women, a saliva sample and a spot urine sample were collected from 19 AA women with breast cancer and 23 AA women not diagnosed with breast cancer. In the urine samples, 20 estrogen metabolites, conjugates, and DNA adducts were analyzed by using ultraperformance liquid chromatography/tandem mass spectrometry, and then the ratio of adducts to metabolites and conjugates was calculated for each sample. The ratio of depurinating estrogen-DNA adducts to estrogen metabolites and estrogen conjugates was significantly greater in cases compared to controls (92.4 ± 46.4 vs 38.5 ± 18.9, p < 0.0001). From the saliva samples, genomic DNA was purified and analyzed for genetic polymorphisms in the genes for two estrogen-metabolizing enzymes, catechol- O-methyltransferase (rs4680) and cytochrome P450 1B1 (rs1056836). There was no association between rs4680 and rs1056836 genotypes and adduct ratios or breast cancer status. This pilot study found higher DNA adduct ratios in women with breast cancer, which suggests that estrogen metabolism is out of balance, and the formation of estrogen-DNA adducts may exert a critical role in breast cancer initiation in AA women.

Modulation of Cellular Response to Arsenic Trioxide Toxicity by Resveratrol.

Arsenic trioxide (As2O3) is an environmental carcinogen and a putative endocrine disruptor. Resveratrol has been shown to reverse As2O3-induced oxidative damage. In immortalized but nontransformed estrogen receptor α-negative human breast cells (MCF10A), we observed that 25 µM resveratrol ameliorated As2O3-induced cytotoxicity. As2O3, in the presence or absence of 25 µM resveratrol, induced quinone reductase (NAD(P)H quinone dehydrogenase 1), via the induction of NFE2-related factor 2. As2O3 caused a repression of cytochrome P450 (CYP)1B1, but the addition of 25 µM resveratrol rescued the expression of cytochrome P450 1B1 and kept it at a constant level. Therefore, 25 µM resveratrol can modulate the effects of As2O3 on enzymes involved in estrogen metabolism.

Critical depurinating DNA adducts: Estrogen adducts in the etiology and prevention of cancer and dopamine adducts in the etiology and prevention of Parkinson's disease.

Endogenous estrogens become carcinogens when dangerous metabolites, the catechol estrogen quinones, are formed. In particular, the catechol estrogen-3,4-quinones can react with DNA to produce an excess of specific depurinating estrogen-DNA adducts. Loss of these adducts leaves apurinic sites in the DNA, generating subsequent cancer-initiating mutations. Unbalanced estrogen metabolism yields excessive catechol estrogen-3,4-quinones, increasing formation of depurinating estrogen-DNA adducts and the risk of initiating cancer. Evidence for this mechanism of cancer initiation comes from various types of studies. High levels of depurinating estrogen-DNA adducts have been observed in women with breast, ovarian or thyroid cancer, as well as in men with prostate cancer or non-Hodgkin lymphoma. Observation of high levels of depurinating estrogen-DNA adducts in high risk women before the presence of breast cancer indicates that adduct formation is a critical factor in breast cancer initiation. Formation of analogous depurinating dopamine-DNA adducts is hypothesized to initiate Parkinson's disease by affecting dopaminergic neurons. Two dietary supplements, N-acetylcysteine and resveratrol complement each other in reducing formation of catechol estrogen-3,4-quinones and inhibiting formation of estrogen-DNA adducts in cultured human and mouse breast epithelial cells. They also inhibit malignant transformation of these cells. In addition, formation of adducts was reduced in women who followed a Healthy Breast Protocol that includes N-acetylcysteine and resveratrol. When initiation of cancer is blocked, promotion, progression and development of the disease cannot occur. These results suggest that reducing formation of depurinating estrogen-DNA adducts can reduce the risk of developing a variety of types of human cancer.

Etiology and prevention of prevalent types of cancer.

Endogenous estrogens become carcinogens when excessive catechol estrogen quinone metabolites are formed. Specifically, the catechol estrogen-3,4-quinones can react with DNA to produce a large amount of specific depurinating estrogen-DNA adducts, formed at the N-3 of Ade and N-7 of Gua. Loss of these adducts leaves apurinic sites in the DNA, which can generate subsequent cancer-initiating mutations. Unbalanced estrogen metabolism yields excessive catechol estrogen-3,4-quinones, increasing formation of the depurinating estrogen-DNA adducts and the risk of initiating cancer. Evidence for this mechanism of cancer initiation comes from studies in vitro, in cell culture, in animal models and in human subjects. High levels of estrogen-DNA adducts have been observed in women with breast, ovarian or thyroid cancer, and in men with prostate cancer or non-Hodgkin lymphoma. Observation of high levels of depurinating estrogen-DNA adducts in high risk women before the presence of breast cancer indicates that adduct formation is a critical factor in breast cancer initiation. Two dietary supplements, N-acetylcysteine and resveratrol, complement each other in reducing formation of catechol estrogen-3,4-quinones and inhibiting formation of estrogen-DNA adducts in cultured human and mouse breast epithelial cells. They also inhibit malignant transformation of these epithelial cells. In addition, formation of adducts was reduced in women who followed a Healthy Breast Protocol that includes N-acetylcysteine and resveratrol. Blocking initiation of cancer prevents promotion, progression and development of the disease. These results suggest that reducing formation of depurinating estrogen-DNA adducts can reduce the risk of developing a variety of types of human cancer.

Depurinating estrogen-DNA adducts, generators of cancer initiation: their minimization leads to cancer prevention.

Estrogens can initiate cancer by reacting with DNA. Specific metabolites of endogenous estrogens, the catechol estrogen-3,4-quinones, react with DNA to form depurinating estrogen-DNA adducts. Loss of these adducts leaves apurinic sites in the DNA, generating mutations that can lead to the initiation of cancer. A variety of endogenous and exogenous factors can disrupt estrogen homeostasis, which is the normal balance between estrogen activating and protective enzymes. In fact, if estrogen metabolism becomes unbalanced and generates excessive catechol estrogen 3,4-quinones, formation of depurinating estrogen-DNA adducts increases and the risk of initiating cancer is greater. The levels of depurinating estrogen-DNA adducts are high in women diagnosed with breast cancer and those at high risk for the disease. High levels of depurinating estrogen-DNA adducts before the presence of breast cancer indicates that adduct formation is a critical factor in breast cancer initiation. Women with thyroid or ovarian cancer also have high levels of estrogen-DNA adducts, as do men with prostate cancer or non-Hodgkin lymphoma. Depurinating estrogen-DNA adducts are initiators of many prevalent types of human cancer. These findings and other discoveries led to the recognition that reducing the levels of estrogen-DNA adducts could prevent the initiation of human cancer. The dietary supplements N-acetylcysteine and resveratrol inhibit formation of estrogen-DNA adducts in cultured human breast cells and in women. These results suggest that the two supplements offer an approach to reducing the risk of developing various prevalent types of human cancer. Graphical abstract Major metabolic pathway in cancer initiation by estrogens.

IDH2R172 mutations define a unique subgroup of patients with angioimmunoblastic T-cell lymphoma.

Angioimmunoblastic T-cell lymphoma (AITL) is a common subtype of peripheral T-cell lymphoma (PTCL) with a poor prognosis. We performed targeted resequencing on 92 cases of PTCL and identified frequent mutations affecting RHOA, TET2, DNMT3A, and isocitrate dehydrogenase 2 (IDH2). Although IDH2 mutations are largely confined to AITL, mutations of the other 3 can be found in other types of PTCL, although at lower frequencies. These findings indicate a key role of epigenetic regulation in the pathogenesis of AITL. However, the epigenetic alterations induced by these mutations and their role in AITL pathogenesis are still largely unknown. We correlated mutational status with gene expression and global DNA methylation changes in AITL. Strikingly, AITL cases with IDH2(R172) mutations demonstrated a distinct gene expression signature characterized by downregulation of genes associated with TH1 differentiation (eg, STAT1 and IFNG) and a striking enrichment of an interleukin 12-induced gene signature. Ectopic expression of IDH2(R172K) in the Jurkat cell line and CD4(+) T cells led to markedly increased levels of 2-hydroxyglutarate, histone-3 lysine methylation, and 5-methylcytosine and a decrease of 5-hydroxymethylcytosine. Correspondingly, clinical samples with IDH2 mutations displayed a prominent increase in H3K27me3 and DNA hypermethylation of gene promoters. Integrative analysis of gene expression and promoter methylation revealed recurrently hypermethylated genes involved in T-cell receptor signaling and T-cell differentiation that likely contribute to lymphomagenesis in AITL.

Breast health and reducing breast cancer risk: a functional medicine approach.

BACKGROUND/OBJECTIVE: A functional medicine approach to reduce breast cancer risk is preferable to early detection and treatment in maintaining breast health. Estrogens are implicated in breast cancer initiation through conversion to metabolites that react with DNA to form specific adducts associated with the development of breast cancer. The purpose of this study was to determine the ability of a defined clinical intervention, the AVERTi-Healthy Breast Program (AHBP), to reduce breast cancer risk conditions likely to develop into breast disease. METHODS: To obtain evidence that risk conditions in breast tissue can be reduced with a defined, multifaceted approach, this small clinical trial of 21 women measured indicators of breast health. A detailed clinical evaluation was conducted with all participants, including identification of physical symptoms, such as areas of tenderness upon palpation. Two laboratory assessments were conducted to determine the efficacy of the AHBP. First, 31 estrogen metabolites, estrogen conjugates, and depurinating estrogen-DNA adducts in urine samples taken before intervention were analyzed. The ratio of DNA adducts to metabolites and conjugates was calculated for each sample. Second, oxidative stress was analyzed by measuring the redox potential of glutathione and cysteine in blood plasma. All assessments were conducted before and after participation. RESULTS: The estrogen adduct ratio and redox potential were improved after 90 days on the AHBP. A significant mean reduction of 3.31 (p=0.03) was observed in the adduct ratio, along with a significant improvement in the redox potential of 3.80 (p=0.05). The significant change in the adduct ratio occurred in women whose oxidative stress profile also improved. CONCLUSION: These significant within-individual decreases suggest that the AHBP can reduce the risk for breast cancer in a relatively short time.

Associations between Dietary Intake of Fruits and Vegetables in relation to Urinary Estrogen DNA Adduct Ratio.

BACKGROUND: Estrogen exposure plays a role in breast cancer (BC) development. A novel estrogen biomarker, the estrogen DNA adduct (EDA) ratio, was shown to be elevated in women at high-risk of BC and among BC cases. Modifiable factors may impact the EDA ratio, with studies demonstrating that resveratrol reduces EDA ratio in vitro. We sought to examine the hypothesis that dietary intake of fruits and vegetables is inversely associated with EDA ratio. METHODS: This analysis was conducted in 53 pre-menopausal, healthy women aged 40-45 years from a cross-sectional study in which participants provided first-void urine samples and 3-day food records. Urine samples were analyzed using ultraperformance liquid chromatography/tandem mass spectrometry. The EDA ratio was calculated as the estrogen-DNA adducts divided by estrogen metabolites and conjugates. A trend test was used to assess associations between tertiles of dietary intake using linear regression. RESULTS: After adjustment for age, total energy, percent adiposity, serum estradiol and estrone-sulfate, we observed inverse associations of EDA ratio with carbohydrate consumption (P=0.01) and vegetable intake (P =0.01). EDA ratio was inversely associated with 5 botanical groups (Chenopodiaceae: P=0.02; Umbelliferae: P=0.03; Compositae: P=0.01; Ericaceae: P=0.01; Musaceae: P=0.03) but not fruit intake overall. CONCLUSION: Although these data require replication before conclusions are drawn, this report suggests an inverse association between vegetable and carbohydrate consumption and EDA ratio. IMPACT: While more information is still needed, these findings suggest a link between dietary intake and a biomarker that is both associated with high-risk BC status and associated with modifiable factors.

Unbalanced estrogen metabolism in ovarian cancer.

Greater exposure to estrogens is a risk factor for ovarian cancer. To investigate the role of estrogens in ovarian cancer, a spot urine sample and a saliva sample were obtained from 33 women with ovarian cancer and 34 age-matched controls. Thirty-eight estrogen metabolites, conjugates and DNA adducts were analyzed in the urine samples using ultraperformance liquid chromatography/tandem mass spectrometry, and the ratio of adducts to metabolites and conjugates was calculated for each sample. The ratio of depurinating estrogen-DNA adducts to estrogen metabolites and conjugates was significantly higher in cases compared to controls (p < 0.0001), demonstrating high specificity and sensitivity. DNA was purified from the saliva samples and analyzed for genetic polymorphisms in the genes for two estrogen-metabolizing enzymes. Women with two low-activity alleles of catechol-O-methyltransferase plus one or two high-activity alleles of cytochrome P450 1B1 had higher levels of estrogen-DNA adducts and were more likely to have ovarian cancer. These findings indicate that estrogen metabolism is unbalanced in ovarian cancer and suggest that formation of estrogen-DNA adducts plays a critical role in the initiation of ovarian cancer.

Reduced formation of depurinating estrogen-DNA adducts by sulforaphane or KEAP1 disruption in human mammary epithelial MCF-10A cells.

Sulforaphane (SFN) is a potent inducer of detoxication enzymes such as NAD(P)H:quinone oxidoreductase 1 (NQO1) and glutathione-S-transferase (GST) via the Kelch-like erythroid-derived protein with CNC homology-associated protein 1 (Keap1)-NF-E2-related factor 2 (Nrf2) signaling pathway. NQO1 reduces the carcinogenic estrogen metabolite, catechol estrogen-3,4-quinone, whereas GSTs detoxify it through conjugation with glutathione. These 3,4-quinones can react with DNA to form depurinating DNA adducts. Thus, SFN may alter estrogen metabolism and thus protect against estrogen-mediated DNA damage and carcinogenesis. Human breast epithelial MCF-10A cells were treated with either vehicle or SFN and either estradiol (E2) or its metabolite 4-hydroxyestradiol (4-OHE2). 4-Hydroxy-derived estrogen metabolites and depurinating DNA adducts formed from E2 and its interconvertable metabolite estrone (E1) were analyzed by mass spectrometry. Levels of the depurinated adducts, 4-OHE1/2-1-N3Adenine and 4-OHE1/2-1-N7Guanine, were reduced by 60% in SFN-treated cells, whereas levels of 4-OCH3E1/2 and 4-OHE1/2-glutathione conjugates increased. To constitutively enhance the expression of Nrf2-regulated genes, cells were treated with either scrambled or siKEAP1 RNA. Following E2 or 4-OHE2 treatments, levels of the adenine and guanine adducts dropped 60-70% in siKEAP1-treated cells, whereas 4-OHE1/2-glutathione conjugates increased. However, 4-OCH3E1/2 decreased 50% after siKEAP1 treatment. Thus, treatment with SFN or siKEAP1 has similar effects on reduction of depurinating estrogen-DNA adduct levels following estrogen challenge. However, these pharmacologic and genetic approaches have different effects on estrogen metabolism to O-methyl and glutathione conjugates. Activation of the Nrf2 pathway, especially elevated NQO1, may account for some but not all of the protective effects of SFN against estrogen-mediated DNA damage.

Unbalanced estrogen metabolism in thyroid cancer.

Well-differentiated thyroid cancer most frequently occurs in premenopausal women. Greater exposure to estrogens may be a risk factor for thyroid cancer. To investigate the role of estrogens in thyroid cancer, a spot urine sample was obtained from 40 women with thyroid cancer and 40 age-matched controls. Thirty-eight estrogen metabolites, conjugates and DNA adducts were analyzed by using ultraperformance liquid chromatography/tandem mass spectrometry and the ratio of adducts to metabolites and conjugates was calculated for each sample. The ratio of depurinating estrogen-DNA adducts to estrogen metabolites and conjugates significantly differed between cases and controls (p < 0.0001), demonstrating high specificity and sensitivity. These findings indicate that estrogen metabolism is unbalanced in thyroid cancer and suggest that formation of estrogen-DNA adducts might play a role in the initiation of thyroid cancer.

Evaluation of serum estrogen-DNA adducts as potential biomarkers for breast cancer risk.

This study was conducted to determine whether the ratio of estrogen-DNA adducts to their respective metabolites and conjugates in serum differed between women with early-onset breast cancer and those with average or high risk of developing breast cancer. Serum samples from women at average risk (n=63) or high risk (n=80) for breast cancer (using Gail model) and women newly diagnosed with early breast cancer (n=79) were analyzed using UPLC-MS/MS. Adduct ratios were statistically compared among the three groups, and the Area Under the Receiver Operating Characteristic Curve (AUC) was used to identify a diagnostic cut-off point. The median adduct ratio in the average-risk group was significantly lower than that of both the high-risk group and the breast cancer group (p values<0.0001), and provided good discrimination between those at average versus high risk of breast cancer (AUC=0.84, 95% CI 0.77-0.90). Sensitivity and specificity were maximized at an adduct ratio of 77. For women in the same age and BMI group, the odds of being at high risk for breast cancer was 8.03 (95% CI 3.46-18.7) times higher for those with a ratio of at least 77 compared to those with a ratio less than 77. The likelihood of being at high risk for breast cancer was significantly increased for those with a high adduct ratio relative to those with a low adduct ratio. These findings suggest that estrogen-DNA adducts deserve further study as potential biomarkers for risk of developing breast cancer.

The etiology and prevention of breast cancer.

Metabolism of estrogens via the catechol estrogen pathway is characterized by a balanced set of activating and protective enzymes (homeostasis). Disruption of homeostasis, with excessive production of catechol estrogen quinones, can lead to reaction of these quinones with DNA to form depurinating estrogen-DNA adducts. Some of the mutations generated by these events can lead to initiation of breast cancer. A wealth of evidence, from studies of metabolism, mutagenicity, cell transformation and carcinogenicity, demonstrates that estrogens are genotoxic. Women at high risk for breast cancer, or diagnosed with the disease, have relatively high levels of depurinating estrogen-DNA adducts compared to normal-risk women. The dietary supplements N-acetylcysteine and resveratrol can inhibit formation of catechol estrogen quinones and their reaction with DNA to form estrogen-DNA adducts, thereby preventing initiation of breast cancer.