Meenakshi Agrawal

Relationship between urinary 15-F2t-isoprostane and 8-oxodeoxyguanosine levels and breast cancer risk

To evaluate the role of oxidative stress in breast cancer, we measured urinary levels of 15-F2t-isoprostane (15-F2t-IsoP) and 8-oxodeoxyguanosine (8-oxodG) in 400 cases and 401 controls, participants of the Long Island Breast Cancer Study Project. We also analyzed the effect of different factors that are associated with oxidative stress and might influence 15-F2t-IsoP and 8-oxodG levels. We observed a statistically significant trend in breast cancer risk with increasing quartiles of 15-F2t-IsoP levels [odds ratio (OR), 1.25; 95% confidence interval (95% CI), 0.81-1.94; OR, 1.53; 95% CI, 0.99-2.35; OR, 1.88; 95% CI, 1.23-2.88, for the 2nd, 3rd, and 4th quartile relative to the lowest quartile, respectively; Ptrend = 0.002]. Although it is possible that increased levels may reflect the stress associated with recent treatment, the positive association was also observed when the analyses were restricted to case women for whom chemotherapy and radiation therapy had not yet been initiated at the time of the urine collection. The association with the highest quartile compared with lowest quartile of 15-F2t-IsoP was similar across strata of age, physical activity, fruit and vegetable intake, alcohol intake, cigarette smoking, body mass index, and menopausal status. We did not observe any association of breast cancer risk with 8-oxodG levels, but when cases with radiation treatment were removed from the analysis, a significant inverse trend (P = 0.04) was observed. Among controls, levels of 15-F2t-IsoP were higher among current cigarette smokers but did not differ by the amount of physical activity, fruit and vegetable intake, alcohol intake, body mass index, and menopausal status. Among controls, levels of 8-oxodG were higher among postmenopausal women and current and former cigarette smokers but did not differ by the other factors. In summary, our results suggest that urinary markers of lipid peroxidation and oxidative DNA damage may be associated with breast cancer risk. (Cancer Epidemiol Biomarkers Prev 2006;15(4):639-44)

Polymorphisms in Nucleotide Excision Repair Genes and DNA Repair Capacity Phenotype in Sisters Discordant for Breast Cancer

Interindividual differences in DNA repair capacity (DRC) may play a critical role in breast cancer risk. Previously, we determined that DRC measured via removal of in vitro–induced benzo[a]pyrene diolepoxide-DNA adducts in lymphoblastoid cell lines was lower in cases compared with controls among sisters discordant for breast cancer from the Metropolitan New York Registry of Breast Cancer Families. We have now determined genotypes for seven single nucleotide polymorphisms in five nucleotide excision repair genes, including Xeroderma pigmentosum complementation group A (XPA +62T>C), group C (XPC Lys939Gln and Ala499Val), group D (XPD Asp312Asn and Lys751Gln), and group G (XPG His1104Asp) and ERCC1 (8092 C>A) in a total of 160 sister pairs for whom DRC phenotype data were available. Overall, there were no statistically significant differences in average DRC for most of the genotypes. A final multivariate conditional logistic model, including three single nucleotide polymorphisms (XPA +62T>C, XPC Ala499Val, and XPG His1104Asp) and smoking status, only modestly predicted DRC after adjusting for case-control status and age of blood donation. The overall predictive accuracy was 61% in the model with a sensitivity of 78% and specificity of 39%. These findings suggest that those polymorphisms we have investigated to date in nucleotide excision repair pathway genes explain only a small amount of the variability in DRC. (Cancer Epidemiol Biomarkers Prev 2006;15(9):1614–20)

Aflatoxin B1 and polycyclic aromatic hydrocarbon adducts, p53 mutations and p16 methylation in liver tissue and plasma of hepatocellular carcinoma patients

Elevated aflatoxin B1‐albumin adducts (AFB1‐Alb) have been associated with an increased risk for HCC development. However, there are no studies in humans, correlating albumin adducts in blood with liver DNA adducts. Forty frozen tumor tissues and 39 paired plasma samples from HCC patients were collected in Taiwan, to determine the relationship between albumin adducts in blood and DNA adducts in liver tissue as well as mutations in p53 and methylation of p16. AFB1‐ and polycyclic aromatic hydrocarbon (PAH)‐DNA adducts in tissue and albumin adducts in plasma were determined by immunohistochemistry and competitive ELISA, respectively. Plasma AFB1‐Alb adducts in subjects with low, medium and high levels of AFB1‐DNA adducts in tumor tissues were 51.0 ± 36.5, 70.5 ± 48.1 and 84.9 ± 48.2 fmol/mg, respectively (ptrend = 0.05). No significant correlation was found for PAH. Fourteen of 40 (36%) tissues were positive for mutant p53 protein by immunohistochemistry; 11 of 40 tissue DNA samples (28%) were positive for p53 mutations, but not their corresponding plasma DNAs. p16 was methylated in 24 of 40 (62%) tissues and 12 of 39 (32%) plasma DNAs. Significant correlations were observed between AFB1‐Alb adducts and p53 mutations and p16 methylation. These data suggest that genetic, epigenetic and environmental exposure biomarkers in plasma may help in estimating the risk for the development of HCC.

Polymorphisms in Nucleotide Excision Repair Genes, Polycyclic Aromatic Hydrocarbon-DNA Adducts, and Breast Cancer Risk

Genes involved in the nucleotide excision repair (NER) pathway, which removes bulky DNA adducts, are potential low-penetrance cancer susceptibility genes. We recently reported an association between detectable polycyclic aromatic hydrocarbon (PAH)-DNA adducts and breast cancer risk. Using a population-based breast cancer case-control study on Long Island, New York, we examined whether polymorphisms in NER genes modified the association between PAH-DNA adducts and breast cancer risk. We examined polymorphisms in ERCC1 (3′-untranslated region 8092C/A), XPA (5′-untranslated region −4G/A), XPD (Asp312Asn in exon 10), XPF (Arg415Gln in exon 8), and XPG (Asp1104His in exon 15) in 1,053 breast cancer cases and 1,102 population-based controls. The presence of at least one variant allele in XPD was associated with a 25% increase in the odds ratio [OR, 1.25; 95% confidence interval (95% CI), 1.04-1.50] for breast cancer. The increase associated with homozygosity of the variant alleles for XPD and ERCC1 was stronger among those with detectable PAH-DNA adduct levels (OR, 1.83; 95% CI, 1.22-2.76 and OR, 1.92; 95% CI, 1.14-3.25 for detectable versus nondetectable adducts and homozygous wild-type genotype for XPD and ERCC1, respectively). We found no association between XPA, XPF, and XPG genotypes, PAH-DNA adducts, and breast cancer risk. When we combined genotypes for these NER pathway genes, there was a significant trend for increasing breast cancer risk with increasing number of putative high-risk alleles. Overall, this study suggests that the risk of breast cancer may be elevated among women with polymorphisms in NER pathway genes and detectable PAH-DNA adducts. (Cancer Epidemiol Biomarkers Prev 2007;16(10):2033–41)

Plasma protein carbonyl levels and breast cancer risk

To study the role of oxidative stress in breast cancer risk, we analysed plasma levels of protein carbonyls in 1050 cases and 1107 controls. We found a statistically significant trend in breast cancer risk in relation to increasing quartiles of plasma protein carbonyl levels (OR = 1.2, 95% CI = 0.9–1.5; OR = 1.5, 95% CI = 1.2–2.0; OR = 1.6, 95% CI = 1.2–2.1, for the 2nd, 3rd and 4th quartile relative to the lowest quartile, respectively, P for trend = 0.0001). The increase in risk was similar for younger (<50 years) and older women, more pronounced among women with higher physical activity levels (0.7 hrs/week for 4th quartile versus lowest quartile OR = 2.0, 95% CI = 1.4–3.0), higher alcohol consumption (≥15 grams/day for 4th quartile versus lowest quartile OR = 2.3, 95% CI = 1.1–4.7), and hormone replacement therapy use (HRT, OR = 2.6, 95% CI = 1.6–4.4 for 4th quartile versus lowest quartile). The multiplicative interaction terms were statistically significant only for physical activity and HRT. The positive association between plasma protein carbonyl levels and breast cancer risk was also observed when the analysis was restricted to women who had not received chemotherapy or radiation therapy prior to blood collection. Among controls, oxidized protein levels significantly increased with cigarette smoking and higher fruit and vegetable consumption, and decreased with alcohol consumption >30 grams per day. Women with higher levels of plasma protein carbonyl and urinary 15F2t‐isoprostane had an 80% increase in breast cancer risk (OR = 1.8, 95% CI = 1.2–2.6) compared to women with levels below the median for both markers of oxidative stress. In summary, our results suggest that increased plasma protein carbonyl levels may be associated with breast cancer risk.

Associations between Polycyclic Aromatic Hydrocarbon–Related Exposures and p53 Mutations in Breast Tumors

Background Previous studies have suggested that polycyclic aromatic hydrocarbons (PAHs) may be associated with breast cancer. However, the carcinogenicity of PAHs on the human breast remains unclear. Certain carcinogens may be associated with specific mutation patterns in the p53 tumor suppressor gene, thereby contributing information about disease etiology. Objectives We hypothesized that associations of PAH-related exposures with breast cancer would differ according to tumor p53 mutation status, effect, type, and number. Methods We examined this possibility in a population-based case–control study using polytomous logistic regression. As previously reported, 151 p53 mutations among 859 tumors were identified using Surveyor nuclease and confirmed by sequencing. Results We found that participants with p53 mutations were less likely to be exposed to PAHs (assessed by smoking status in 859 cases and 1,556 controls, grilled/smoked meat intake in 822 cases and 1,475 controls, and PAH–DNA adducts in peripheral mononuclear cells in 487 cases and 941 controls) than participants without p53 mutations. For example, active and passive smoking was associated with p53 mutation–negative [odds ratio (OR) = 1.55; 95% confidence interval (CI), 1.11–2.15] but not p53 mutation–positive (OR = 0.77; 95% CI, 0.43–1.38) cancer (ratio of the ORs = 0.50, p < 0.05). However, frameshift mutations, mutation number, G:C→A:T transitions at CpG sites, and insertions/deletions were consistently elevated among exposed subjects. Conclusions These findings suggest that PAHs may be associated with specific breast tumor p53 mutation subgroups rather than with overall p53 mutations and may also be related to breast cancer through mechanisms other than p53 mutation.

Plasma Protein Carbonyls and Breast Cancer Risk in Sisters Discordant for Breast Cancer from the New York Site of the Breast Cancer Family Registry

Reactive oxygen species are important in the pathogenesis of many diseases, including breast cancer. Several population-based case-control studies have shown that various biomarkers of oxidative stress are associated with an increase in breast cancer risk. We selected sisters discordant for breast cancer (n=645) from the New York site of the Breast Cancer Family Registry to explore factors that contribute to variation in plasma protein carbonyls, and to determine whether this biomarker is associated with an increase in breast cancer risk among those with a family history. Late age at menarche, hormone replacement therapy use, and Hispanic race were significantly associated with lower plasma protein carbonyl levels in unaffected sisters. Plasma protein carbonyls were associated with an increase in breast cancer risk [Q2 odds ratio (OR), 1.4; 95% confidence interval (CI), 0.8-2.7; Q3 OR, 2.4; 95% CI, 1.1-4.9; Q4 OR, 1.9; 95% CI, 0.8-4.2], although not in a dose-dependent manner. These data suggest that oxidative damage is a risk factor for breast cancer in high-risk women.

XRCC1 polymorphisms and breast cancer risk from the New York Site of the Breast Cancer Family Registry: A family-based case-control study

Background: XRCC1 is a scaffold protein involved in the early and late stages of Base Excision Repair (BER). Three DNA polymorphisms occur in XRCC1, resulting in non-synonymous amino acid changes, which could alter the binding or regulatory activities of XRCC1. Materials and Methods: We used a family-based case-control study design to evaluate the association between XRCC1 polymorphisms and breast cancer risk. Participants were breast cancer cases and their unaffected sisters enrolled in the New York Site of the Breast Cancer Family Registry. Conditional logistic regression was used to assess associations between genotype and breast cancer. XRCC1 mRNA levels and DNA nicking activity were measured in lymphoblastoid cell lines from unaffected sisters to determine whether the XRCC1 R399Q polymorphism has a functional effect on expression or protein activity. Results: XRCC1 194W was associated with a non-significant increase in breast cancer, while XRCC1 280H and XRCC1 399Q were associated with a non-significant decrease in breast cancer. We found a significant increase in XRCC1 expression in 399Q/Q lymphoblastoid cell lines from unaffected sisters (n=28, P=0.03). An increase in median nicking activity was not statistically significant. Conclusions: Our results suggest that XRCC1 399Q may alter mRNA expression and DNA repair phenotype, although the main effects of the genotype were not significantly associated with familial cancer risk. Additional research on the regulation of XRCC1 expression will contribute to an understanding of how this polymorphism may impact disease risk.

Mutations in p53, p53 protein overexpression and breast cancer survival

p53 is an important tumour suppressor gene that encodes p53 protein, a molecule involved in cell cycle regulation and has been inconsistently linked to breast cancer survival. Using archived tumour tissue from a population‐based sample of 859 women diagnosed with breast cancer between 1996 and 1997, we determined p53 mutations in exons 5–8 and p53 protein overexpression. We examined the association of p53 mutations with overexpression and selected tumour clinical parameters. We assessed whether either p53 marker was associated with survival through 2002, adjusting for other tumour markers and prognostic factors. The prevalence of protein overexpression in the tumour was 36% (307/859) and of any p53 mutation was 15% (128/859). p53 overexpression was positively associated with the presence of any p53 mutation (odds ratio [OR]= 2.2, 95% confidence interval [CI]= 1.5–3.2), particularly missense mutations (ER = 7.0, 95% CI = 3.6–13.7). Negative oestrogen and progesterone receptor (ER/PR) status was positively associated with both p53 protein overexpression (= 2.6, 95% CI = 1.7–4.0) and p53 mutation (OR = 3.9, 95% CI = 2.4–6.5). Any p53 mutation and missense mutations, but not p53 protein overexpression, were associated with breast cancer‐specific mortality (hazard ratio [HR]= 1.7, 95% CI = 1.0–2.8; HR = 2.0, 95% CI = 1.1–3.6, respectively) and all‐cause mortality (HR = 1.5, 95% CI = 1.0–2.4; HR = 2.0, 95% CI = 1.2–3.4, respectively); nonsense mutations were associated only with breast cancer‐specific mortality (HR = 3.0, 95% CI = 1.1–8.1). These associations however did not remain after adjusting for ER/PR status. Thus, in this population‐based cohort of women with breast cancer, although p53 protein overexpression and p53 mutations were associated with each other, neither independently impacted breast cancer‐specific or all‐causing mortality, after considering ER/PR status.