BACKGROUND: BRCA1 mutation carriers have an 85% risk of developing breast cancer but the risk of developing non-hereditary breast cancer is difficult to assess. Our objective is to test whether a DNA methylation (DNAme) signature derived from BRCA1 mutation carriers is able to predict non-hereditary breast cancer. METHODS: In a case/control setting (72 BRCA1 mutation carriers and 72 BRCA1/2 wild type controls) blood cell DNA samples were profiled on the Illumina 27 k methylation array. Using the Elastic Net classification algorithm, a BRCA1-mutation DNAme signature was derived and tested in two cohorts: (1) The NSHD (19 breast cancers developed within 12 years after sample donation and 77 controls) and (2) the UKCTOCS trial (119 oestrogen receptor positive breast cancers developed within 5 years after sample donation and 122 controls). RESULTS: We found that our blood-based BRCA1-mutation DNAme signature applied to blood cell DNA from women in the NSHD resulted in a receiver operating characteristics (ROC) area under the curve (AUC) of 0.65 (95% CI 0.51 to 0.78, P = 0.02) which did not validate in buccal cells from the same individuals. Applying the signature in blood DNA from UKCTOCS volunteers resulted in AUC of 0.57 (95% CI 0.50 to 0.64; P = 0.03) and is independent of family history or any other known risk factors. Importantly the BRCA1-mutation DNAme signature was able to predict breast cancer mortality (AUC = 0.67; 95% CI 0.51 to 0.83; P = 0.02). We also found that the 1,074 CpGs which are hypermethylated in BRCA1 mutation carriers are significantly enriched for stem cell polycomb group target genes (P <10(-20)). CONCLUSIONS: A DNAme signature derived from BRCA1 carriers is able to predict breast cancer risk and death years in advance of diagnosis. Future studies may need to focus on DNAme profiles in epithelial cells in order to reach the AUC thresholds required of preventative measures or early detection strategies.

Department of Women's Cancer UCL Elizabeth Garrett Anderson Institute for Women's Health University College London 74 Huntley Street London WC1E 6 AU UK

Department of Women's Cancer UCL Elizabeth Garrett Anderson Institute for Women's Health University College London 74 Huntley Street London WC1E 6 AU UK ; Faculty of Medical and Human Sciences The University of Manchester 46 Grafton Street Manchester M13 9NT UK

Gynecological Oncology Center Department of Obstetrics and Gynecology Charles University Prague 1st Faculty of Medicine and General University Hospital Apolinarska 18 128 00 Prague Czech Republic

MRC Unit for Lifelong Health and Ageing at UCL 33 Bedford Place London WC1B 5JU UK

Statistical Genomics Group Paul O'Gorman Building UCL Cancer Institute University College London 72 Huntley Street London WC1E 6BT UK ; CAS MPG Partner Institute for Computational Biology Chinese Academy of Sciences Shanghai Institute for Biological Sciences Shanghai 200031 China

See more in PubMed

Domchek SM, Friebel TM, Singer CF, Evans DG, Lynch HT, Isaacs C, Garber JE, Neuhausen SL, Matloff E, Eeles R, Pichert G, Van t’veer L, Tung N, Weitzel JN, Couch FJ, Rubinstein WS, Ganz PA, Daly MB, Olopade OI, Tomlinson G, Schildkraut J, Blum JL, Rebbeck TR. Association of risk-reducing surgery in BRCA1 or BRCA2 mutation carriers with cancer risk and mortality. JAMA. 2010;304:967–975. PubMed PMC

Fackenthal JD, Olopade OI. Breast cancer risk associated with BRCA1 and BRCA2 in diverse populations. Nat Rev Cancer. 2007;7:937–948. PubMed

Easton DF, Pooley KA, Dunning AM, Pharoah PD, Thompson D, Ballinger DG, Struewing JP, Morrison J, Field H, Luben R, Wareham N, Ahmed S, Healey CS, Bowman R, Meyer KB, Haiman CA, Kolonel LK, Henderson BE, Le Marchand L, Brennan P, Sangrajrang S, Gaborieau V, Odefrey F, Shen CY, Wu PE, Wang HC, Eccles D, Evans DG, Peto J. SEARCH collaborators et al.Genome-wide association study identifies novel breast cancer susceptibility loci. Nature. 2007;447:1087–1093. PubMed PMC

Wacholder S, Hartge P, Prentice R, Garcia-Closas M, Feigelson HS, Diver WR, Thun MJ, Cox DG, Hankinson SE, Kraft P, Rosner B, Berg CD, Brinton LA, Lissowska J, Sherman ME, Chlebowski R, Kooperberg C, Jackson RD, Buckman DW, Hui P, Pfeiffer R, Jacobs KB, Thomas GD, Hoover RN, Gail MH, Chanock SJ, Hunter DJ. Performance of common genetic variants in breast-cancer risk models. N Engl J Med. 2010;362:986–993. PubMed PMC

Independent UK Panel on Breast Cancer Screening. The benefits and harms of breast cancer screening: an independent review. Lancet. 2012;380:1778–1786. PubMed

Jorgensen KJ, Gotzsche PC. Overdiagnosis in publicly organised mammography screening programmes: systematic review of incidence trends. BMJ. 2009;339:b2587. PubMed PMC

Zahl PH, Gotzsche PC, Maehlen J. Natural history of breast cancers detected in the Swedish mammography screening programme: a cohort study. Lancet Oncol. 2011;12:1118–1124. PubMed

Cui H, Cruz-Correa M, Giardiello FM, Hutcheon DF, Kafonek DR, Brandenburg S, Wu Y, He X, Powe NR, Feinberg AP. Loss of IGF2 imprinting: a potential marker of colorectal cancer risk. Science. 2003;299:1753–1755. PubMed

Jones PA, Baylin SB. The epigenomics of cancer. Cell. 2007;128:683–692. PubMed PMC

Teschendorff AE, Jones A, Fiegl H, Sargent A, Zhuang JJ, Kitchener HC, Widschwendter M. Epigenetic variability in cells of normal cytology is associated with the risk of future morphological transformation. Genome Med. 2012;4:24. PubMed PMC

Widschwendter M, Jones PA. DNA methylation and breast carcinogenesis. Oncogene. 2002;21:5462–5482. PubMed

Baylin SB, Jones PA. A decade of exploring the cancer epigenome - biological and translational implications. Nat Rev Cancer. 2011;11:726–734. PubMed PMC

Maegawa S, Hinkal G, Kim HS, Shen L, Zhang L, Zhang J, Zhang N, Liang S, Donehower LA, Issa JP. Widespread and tissue specific age-related DNA methylation changes in mice. Genome Res. 2010;20:332–340. PubMed PMC

O’Hagan HM, Wang W, Sen S, Destefano Shields C, Lee SS, Zhang YW, Clements EG, Cai Y, Van Neste L, Easwaran H, Casero RA, Sears CL, Baylin SB. Oxidative damage targets complexes containing DNA methyltransferases, SIRT1, and polycomb members to promoter CpG Islands. Cancer Cell. 2011;20:606–619. PubMed PMC

Schlesinger Y, Straussman R, Keshet I, Farkash S, Hecht M, Zimmerman J, Eden E, Yakhini Z, Ben-Shushan E, Reubinoff BE, Bergman Y, Simon I, Cedar H. Polycomb-mediated methylation on Lys27 of histone H3 pre-marks genes for de novo methylation in cancer. Nat Genet. 2007;39:232–236. PubMed

Teschendorff AE, Menon U, Gentry-Maharaj A, Ramus SJ, Weisenberger DJ, Shen H, Campan M, Noushmehr H, Bell CG, Maxwell AP, Savage DA, Mueller-Holzner E, Marth C, Kocjan G, Gayther SA, Jones A, Beck S, Wagner W, Laird PW, Jacobs IJ, Widschwendter M. Age-dependent DNA methylation of genes that are suppressed in stem cells is a hallmark of cancer. Genome Res. 2010;20:440–446. PubMed PMC

Widschwendter M, Fiegl H, Egle D, Mueller-Holzner E, Spizzo G, Marth C, Weisenberger DJ, Campan M, Young J, Jacobs I, Laird PW. Epigenetic stem cell signature in cancer. Nat Genet. 2007;39:157–158. PubMed

Zhuang J, Jones A, Lee SH, Ng E, Fiegl H, Zikan M, Cibula D, Sargent A, Salvesen HB, Jacobs IJ, Kitchener HC, Teschendorff AE, Widschwendter M. The dynamics and prognostic potential of DNA methylation changes at stem cell gene loci in women's cancer. PLoS Genet. 2012;8:e1002517. PubMed PMC

Lee TI, Jenner RG, Boyer LA, Guenther MG, Levine SS, Kumar RM, Chevalier B, Johnstone SE, Cole MF, Isono K, Koseki H, Fuchikami T, Abe K, Murray HL, Zucker JP, Yuan B, Bell GW, Herbolsheimer E, Hannett NM, Sun K, Odom DT, Otte AP, Volkert TL, Bartel DP, Melton DA, Gifford DK, Jaenisch R, Young RA. Control of developmental regulators by Polycomb in human embryonic stem cells. Cell. 2006;125:301–313. PubMed PMC

Wang L, Zeng X, Chen S, Ding L, Zhong J, Zhao JC, Wang L, Sarver A, Koller A, Zhi J, Ma Y, Yu J, Chen J, Huang H. BRCA1 is a negative modulator of the PRC2 complex. EMBO J. 2013;32:1584–1597. PubMed PMC

Bosviel R, Garcia S, Lavediaux G, Michard E, Dravers M, Kwiatkowski F, Bignon YJ, Bernard-Gallon DJ. BRCA1 promoter methylation in peripheral blood DNA was identified in sporadic breast cancer and controls. Cancer Epidemiol. 2012;36:e177–e182. PubMed

Brennan K, Garcia-Closas M, Orr N, Fletcher O, Jones M, Ashworth A, Swerdlow A, Thorne H, Investigators KC, Riboli E, Vineis P, Dorronsoro M, Clavel-Chapelon F, Panico S, Onland-Moret NC, Trichopoulos D, Kaaks R, Khaw KT, Brown R, Flanagan JM. Intragenic ATM methylation in peripheral blood DNA as a biomarker of breast cancer risk. Cancer Res. 2012;72:2304–2313. PubMed

Choi JY, James SR, Link PA, McCann SE, Hong CC, Davis W, Nesline MK, Ambrosone CB, Karpf AR. Association between global DNA hypomethylation in leukocytes and risk of breast cancer. Carcinogenesis. 2009;30:1889–1897. PubMed PMC

Delgado-Cruzata L, Wu HC, Perrin M, Liao Y, Kappil MA, Ferris JS, Flom JD, Yazici H, Santella RM, Terry MB. 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. Epigenetics. 2012;7:868–874. PubMed PMC

Delgado-Cruzata L, Wu HC, Liao Y, Santella RM, Terry MB. Differences in DNA methylation by extent of breast cancer family history in unaffected women. Epigenetics. 2014;9:243–248. PubMed PMC

Flanagan JM, Munoz-Alegre M, Henderson S, Tang T, Sun P, Johnson N, Fletcher O, Dos Santos SI, Peto J, Boshoff C, Narod S, Petronis A. Gene-body hypermethylation of ATM in peripheral blood DNA of bilateral breast cancer patients. Hum Mol Genet. 2009;18:1332–1342. PubMed PMC

Ito Y, Koessler T, Ibrahim AE, Rai S, Vowler SL, Abu-Amero S, Silva AL, Maia AT, Huddleston JE, Uribe-Lewis S, Woodfine K, Jagodic M, Nativio R, Dunning A, Moore G, Klenova E, Bingham S, Pharoah PD, Brenton JD, Beck S, Sandhu MS, Murrell A. Somatically acquired hypomethylation of IGF2 in breast and colorectal cancer. Hum Mol Genet. 2008;17:2633–2643. PubMed PMC

Iwamoto T, Yamamoto N, Taguchi T, Tamaki Y, Noguchi S. BRCA1 promoter methylation in peripheral blood cells is associated with increased risk of breast cancer with BRCA1 promoter methylation. Breast Cancer Res Treat. 2011;129:69–77. PubMed

Widschwendter M, Apostolidou S, Raum E, Rothenbacher D, Fiegl H, Menon U, Stegmaier C, Jacobs IJ, Brenner H. Epigenotyping in peripheral blood cell DNA and breast cancer risk: a proof of principle study. PLoS One. 2008;3:e2656. PubMed PMC

Woo HD, Kim J. Global DNA hypomethylation in peripheral blood leukocytes as a biomarker for cancer risk: a meta-analysis. PLoS One. 2012;7:e34615. PubMed PMC

Wu HC, John EM, Ferris JS, Keegan TH, Chung WK, Andrulis I, Delgado-Cruzata L, Kappil M, Gonzalez K, Santella RM, Terry MB. Global DNA methylation levels in girls with and without a family history of breast cancer. Epigenetics. 2011;6:29–33. PubMed PMC

Xu X, Gammon MD, Hernandez-Vargas H, Herceg Z, Wetmur JG, Teitelbaum SL, Bradshaw PT, Neugut AI, Santella RM, Chen J. DNA methylation in peripheral blood measured by LUMA is associated with breast cancer in a population-based study. FASEB J. 2012;26:2657–2666. PubMed PMC

Xu Z, Bolick SC, DeRoo LA, Weinberg CR, Sandler DP, Taylor JA. Epigenome-wide association study of breast cancer using prospectively collected sister study samples. J Natl Cancer Inst. 2013;105:694–700. PubMed PMC

Wadsworth M, Kuh D, Richards M, Hardy R. Cohort Profile: The 1946 National Birth Cohort (MRC National Survey of Health and Development) Int J Epidemiol. 2006;35:49–54. PubMed

Kuh D, Pierce M, Adams J, Deanfield J, Ekelund U, Friberg P, Ghosh AK, Harwood N, Hughes A, Macfarlane PW, Mishra G, Pellerin D, Wong A, Stephen AM, Richards M, Hardy R. on behalf of the NSHD scientific and data collection team. Cohort profile: updating the cohort profile for the MRC National Survey of Health and Development: a new clinic-based data collection for ageing research. Int J Epidemiol. 2011;40:e1–e9. PubMed PMC

Rousseau K, Vinall LE, Butterworth SL, Hardy RJ, Holloway J, Wadsworth ME, Swallow DM. MUC7 haplotype analysis: results from a longitudinal birth cohort support protective effect of the MUC7*5 allele on respiratory function. Ann Hum Genet. 2006;70:417–427. PubMed

Hologic Gen-Probe – DNA extraction. [ http://www.gen-probe.com]

The National Survey of Health and development (NSHD) data archive. [ http://www.nshd.mrc.ac.uk/data.aspx]

Zou H, Hastie T. Regularization and variable selection via the elastic net. J R Stat Soc. 2005;B.67:301–320.

Friedman JH, Hastie T, Tibshirani R. Regularization paths for generalized linear models via coordinate descent. J Stat Softw. 2010;33:1–22. PubMed PMC

Newson R. Confidence intervals for rank statistics: Percentile slopes, differences, and ratios. Stata J. 2006;6:497–520.

Teschendorff AE, Menon U, Gentry-Maharaj A, Ramus SJ, Gayther SA, Apostolidou S, Jones A, Lechner M, Beck S, Jacobs IJ, Widschwendter M. An epigenetic signature in peripheral blood predicts active ovarian cancer. PLoS One. 2009;4:e8274. PubMed PMC

Mootha VK, Lindgren CM, Eriksson KF, Subramanian A, Sihag S, Lehar J, Puigserver P, Carlsson E, Ridderstrale M, Laurila E, Houstis N, Daly MJ, Patterson N, Mesirov JP, Golub TR, Tamayo P, Spiegelman B, Lander ES, Hirschhorn JN, Altshuler D, Groop LC. PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat Genet. 2003;34:267–273. PubMed

Subramanian A, Tamayo P, Mootha VK, Mukherjee S, Ebert BL, Gillette MA, Paulovich A, Pomeroy SL, Golub TR, Lander ES, Mesirov JP. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc Natl Acad Sci U S A. 2005;102:15545–15550. PubMed PMC

Huzarski T, Byrski T, Gronwald J, Gorski B, Domagala P, Cybulski C, Oszurek O, Szwiec M, Gugala K, Stawicka M, Morawiec Z, Mierzwa T, Janiszewska H, Kilar E, Marczyk E, Kozak-Klonowska B, Siołek M, Surdyka D, Wiśniowski R, Posmyk M, Sun P, Lubiński J, Narod SA. Ten-year survival in patients with BRCA1-negative and BRCA1-positive breast cancer. J Clin Oncol. 2013;31:3191–3196. PubMed

Narod SA, Foulkes WD. BRCA1 and BRCA2: 1994 and beyond. Nat Rev Cancer. 2004;4:665–676. PubMed

Menon U, Gentry-Maharaj A, Ryan A, Sharma A, Burnell M, Hallett R, Lewis S, Lopez A, Godfrey K, Oram D, Herod J, Williamson K, Seif M, Scott I, Mould T, Woolas R, Murdoch J, Dobbs S, Amso N, Leeson S, Cruickshank D, McGuire A, Campbell S, Fallowfield L, Skates S, Parmar M, Jacobs I. Recruitment to multicentre trials–lessons from UKCTOCS: descriptive study. BMJ. 2008;337:a2079. PubMed PMC

DNA methylation at quantitative trait loci (mQTLs) varies with cell type and nonheritable factors and may improve breast cancer risk assessment

The WID-BC-index identifies women with primary poor prognostic breast cancer based on DNA methylation in cervical samples

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