DNA Repair Gene Expression Levels as Indicators of Breast Cancer in the Breast Cancer Family Registry.

AIM: The expression level of DNA repair-related genes and their association with breast cancer status among participants of the New York site of the Breast Cancer Family Registry was investigated. MATERIALS AND METHODS: RNA from mononuclear cells in 194 sister sets (n=475 women) were assayed for ATM, BRCA1, MSH2, MUTYH and XPC gene expression levels and analyzed using generalized estimating equations (GEE). RESULTS: Individuals with decreased ATM and MSH2 expression had significantly higher odds for breast cancer compared to individuals with higher levels of expression (odds ratio (OR)=1.1, 95% confidence interval (CI)=1.02, 1.18) and (OR=1.90, 95% CI=1.21, 2.97), respectively. Upon stratifying the GEE model, reductions in ATM and MSH2 expression levels was heightened among women with an extended family history (FH) of breast cancer. CONCLUSION: Reduced expression of ATM and MSH2 compromises DNA repair capacity and, thereby, increases breast cancer prevalence.

Genome-wide aberrant DNA methylation of microRNA host genes in hepatocellular carcinoma.

Mature microRNAs (miRNAs) are a class of small non-coding RNAs involved in posttranslational gene silencing. Previous studies found that downregulation of miRNAs is a common feature observed in solid tumors, including hepatocellular carcinoma (HCC). We employed a genome-wide approach to test the hypothesis that DNA methylation alterations in miRNA host genes may cause deregulated miRNA expression in HCC. We analyzed tumor and adjacent non-tumor tissues from 62 Taiwanese HCC cases using Infinium HumanMethylation27 DNA Analysis BeadChips that include 254 CpG sites covering 110 miRNAs from 64 host genes. Expression levels of three identified miRNAs (miR-10a, miR-10b and miR-196b) were measured in a subset of 37 HCC tumor and non-tumor tissues. After Bonferroni adjustment, a total of 54 CpG sites from 27 host genes significantly differed in DNA methylation levels between tumor and adjacent non-tumor tissues with 53 sites significantly hypermethylated in tumor tissues. Among the 54 significant CpG sites, 15 sites had more than 2-fold tumor/non-tumor changes, 17 sites had differences > 10%, and 10 sites had both features [including 8 significantly hypermethylated CpG sites in the host genes of miR-10a, miR-10b and miR-196b (HOXB4, HOXD4 and HOXA9, respectively)]. Significant downregulation of miR-10a was observed in tumor compared with non-tumor tissues (0.50 vs. 1.73, p = 0.031). The concordance for HOXB4 methylation alteration and dysregulation of miR-10a was 73.5%. No significant change was observed for miR-10b expression. Unexpectedly, miR-196b was significantly upregulated in tumor compared with non-tumor tissues (p = 0.0001). These data suggest that aberrant DNA methylation may lead to dysregulation of miR-10a in HCC tumor tissues.

Global DNA methylation levels in white blood cell DNA from sisters discordant for breast cancer from the New York site of the Breast Cancer Family Registry.

Lower global DNA methylation is associated with genomic instability and it is one of the epigenetic mechanisms relevant to carcinogenesis. Emerging evidence for several cancers suggests that lower overall levels of global DNA methylation in blood are associated with different cancer types, although less is known about breast cancer. We examined global DNA methylation levels using a sibling design in 273 sisters affected with breast cancer and 335 unaffected sisters from the New York site of the Breast Cancer Family Registry. We measured global DNA methylation in total white blood cell (WBC) and granulocyte DNA by two different methods, the [ ( 3) H]-methyl acceptance assay and the luminometric methylation assay (LUMA). Global methylation levels were only modestly correlated between sisters discordant for breast cancer (Spearman correlation coefficients ranged from -0.08 to 0.24 depending on assay and DNA source). Using conditional logistic regression models, women in the quartile with the lowest DNA methylation levels (as measured by the [ ( 3) H]-methyl acceptance assay) had a 1.8-fold (95% CI = 1.0-3.3) higher relative association with breast cancer than women in the quartile with the highest DNA methylation levels. When we examined the association on a continuous scale, we also observed a positive association (odds ratio, OR = 1.3, 95% CI = 1.0-1.7, for a one unit change in the natural logarithm of the DPM/µg of DNA). We observed no association between measures by the LUMA assay and breast cancer risk. If replicated in prospective studies, this study suggests that global DNA methylation levels measured in WBC may be a potential biomarker of breast cancer risk even within families at higher risk of cancer.

White blood cell global methylation and IL-6 promoter methylation in association with diet and lifestyle risk factors in a cancer-free population.

Altered levels of global DNA methylation and gene silencing through methylation of promoter regions can impact cancer risk, but little is known about their environmental determinants. We examined the association between lifestyle factors and levels of global genomic methylation and IL-6 promoter methylation in white blood cell DNA of 165 cancer-free subjects, 18-78 years old, enrolled in the COMIR (Commuting Mode and Inflammatory Response) study, New York, 2009-2010. Besides self-administrated questionnaires on diet and physical activity, we measured weight and height, white blood cell (WBC) counts, plasma levels of high sensitivity C-reactive protein (hs-CRP), and genomic (LINE-1) and gene-specific methylation (IL-6) by pyrosequencing in peripheral blood WBC. Mean levels of LINE-1 and IL-6 promoter methylation were 78.2% and 57.1%, respectively. In multivariate linear regression models adjusting for age, gender, race/ethnicity, body mass index, diet, physical activity, WBC counts and CRP, only dietary folate intake from fortified foods was positively associated with LINE-1 methylation. Levels of IL-6 promoter methylation were not significantly correlated with age, gender, race/ethnicity, body mass index, physical activity or diet, including overall dietary patterns and individual food groups and nutrients. There were no apparent associations between levels of methylation and inflammation markers such as WBC counts and hs-CRP. Overall, among several lifestyle factors examined in association with DNA methylation, only dietary folate intake from fortification was associated with LINE-1 methylation. The long-term consequence of folate fortification on DNA methylation needs to be further evaluated in longitudinal settings.

Biologic and epigenetic impact of commuting to work by car or using public transportation: a case-control study.

BACKGROUND AND AIMS: Commuting by public transportation (PT) entails more physical activity and energy expenditure than by cars, but its biologic consequences are unknown. METHODS: In 2009-2010, we randomly sampled New York adults, usually commuting either by car (n=79) or PT (n=101). Measures comprised diet and physical activity questionnaires, weight and height, white blood cell (WBC) count, C reactive protein, (CRP) gene-specific methylation (IL-6), and global genomic DNA methylation (LINE-1 methylation). RESULTS: Compared to the 101 PT commuters, the 79 car drivers were about 9 years older, 2 kg/m(2) heavier, more often non-Hispanic whites, and ate more fruits and more meats. The 2005 guidelines for physical activity were met by more car drivers than PT users (78.5% vs. 65.0%). There were no differences in median levels of CRP (car vs. PT: 0.6 vs. 0.5mg/dl), mean levels of WBC (car vs. PT: 6.7 vs. 6.5 cells/mm(3)), LINE-1 methylation (car vs. PT: 78.0% vs. 78.3%), and promoter methylation of IL-6 (car vs. PT: 56.1% vs. 58.0%). CONCLUSIONS: PT users were younger and lighter than car drivers, but their commute mode did not translate into a lower inflammatory response or a higher DNA methylation, maybe because, overall, car drivers were more physically active.

Tissue inhibitor of metalloproteinases-1 stimulates gene expression in MDA-MB-435 human breast cancer cells by means of its ability to inhibit metalloproteinases.

Tissue inhibitor of metalloproteinases-1 (TIMP-1) is a widely expressed, secreted protein that functions primarily to inhibit members of a large family of metalloproteinases (MPs). Because of the ability of TIMP-1 to inhibit MPs, it functions in many of the same pathophysiological processes as these enzymes, e.g. wound healing, ovulation, angiogenesis, and cancer cell metastasis. TIMP-1 can also stimulate proliferation ([3H]thymidine incorporation) and cellular anabolic processes (Alamar Blue reduction). This stimulation has been shown to be dependent on the MP-inhibitory ability of TIMP-1 in the human breast cancer cell line MDA-MB-435 (Porter et al., Br J Cancer 90: 463, 2004). To shed light on the mechanism by which TIMP-1 stimulates cellular anabolic processes, an oligonucleotide microarray analysis was performed over a time course of TIMP-1 treatment of MDA-MB-435 cells. Fifteen genes whose mRNAs were differentially regulated were identified. Six (Importin-7, MGC10471, FOXC1, subunit p20 of Arp2/3 complex, mitochondrial ribosomal protein L32, and the serine/threonine kinase-4 (MST1)) of these genes were confirmed by quantitative real time PCR. These same mRNAs were shown to be regulated by the synthetic hydroxamate MP-inhibitor GM6001 but not by its inactive derivative GM6001*, suggesting that the differential regulation occurs through the MP-inhibitory ability of TIMP-1. These results suggest a complex action of TIMP-1 on cancer cells mediated by constitutively active cell surface metalloproteinases that release factors regulating cell signaling pathways; they may account for the paradoxical observation that elevated levels of TIMP-1 in tumors can correlate with an adverse prognosis.