Determination of Breast Cancer Dormancy: Analysis of Circulating Free DNA Using SNP 6.0 Arrays

Chapter
First Online:
Part of the Tumor Dormancy and Cellular Quiescence and Senescence book series (DOQU, volume 1)

Abstract

New biomarkers are needed in breast cancer to monitor minimal residual disease. Specific point mutations, promoter methylation and loss of heterozygosity have been demonstrated previously in paired tumor and circulating cell-free DNA (cfDNA) isolated from plasma. Moreover, acquired alterations unique to cfDNA have also been found, suggesting disease progression. This prompted us to characterize the “circulating” breast cancer genome, thus testing the hypothesis that cfDNA acts as a surrogate liquid biopsy of breast cancer. This was achieved using Affymetrix SNP 6.0 technology and bioinformatics to map single nucleotide polymorphism (SNP) and copy number variation (CNV), comparing cfDNA with matched normal leucocyte and primary tumor DNA in breast cancer patients and paired normal leucocytes and cfDNA in healthy female controls. Our results show that concordance of SNP genotype calls in paired leucocytes and cfDNA can distinguish between primary breast cancer patients and healthy controls (p < 0.0001), and between pre-surgical breast cancer patients and patients on follow-up after surgery and treatment (p = 0.0016). In 50 patients on follow-up, a significant difference (p = 0.0006) was seen between cfDNA samples taken an average of 3 years apart, suggesting disease progression. Considering CNVs, amplification was observed in matched tumor and cfDNA at multiple loci on different chromosome arms but was quiet or absent in normal DNA. Many of these tumor-specific CNVs contributed significantly to disease through binary logistic regression analysis. Furthermore these CNVs remained detectable in cfDNA up to 12 years after diagnosis and treatment despite no other evidence of disease. Taken together these cfDNA results suggest breast cancer dormancy in the majority of the patients on follow-up.

Keywords

Breast cancer • Cell-free DNA (cfDNA) • SNP 6.0 arrays • Copy number variation (CNV) 
This is a preview of subscription content, log in to check access.

References

  1. Alix-Panabières C, Schwarzenbach H, Pantel K (2012) Circulating tumor cells and circulating tumor DNA. Annu Rev Med 63:199–215PubMedCrossRefGoogle Scholar
  2. Anker P, Stroun M, Maurice PA (1975) Spontaneous release of DNA by human blood lymphocytes as shown in an in vitro system. Cancer Res 35:2375–2382PubMedGoogle Scholar
  3. Chen X, Bonnefoi H, Diebold-Berger S, Lyautey J, Lederrey C, Faltin-Traub E, Stroun M, Anker P (1999) Detecting tumor-related alterations in plasma or serum DNA of patients diagnosed with breast cancer. Clin Cancer Res 5:2297–2303PubMedGoogle Scholar
  4. Conrad DF, Pinto D, Redon R, Feuk L, Gokcumen O, Zhang Y, Aerts J, Andrews TD, Barnes C, Campbell P, Fitzgerald T, Hu M, Ihm CH, Kristiansson K, Macarthur DG, Macdonald JR, Onyiah I, Pang AW, Robson S, Stirrups K, Valsesia A, Walter K, Wei J, Wellcome Trust Case Control Consortium, Tyler-Smith C, Carter NP, Lee C, Scherer SW, Hurles ME (2009) Origins and functional impact of copy number variation in the human genome. Nature 464:704–712PubMedCrossRefGoogle Scholar
  5. Gangnus R, Langer S, Breit E, Pantel K, Speicher MR (2004) Genomic profiling of viable and proliferative micrometastatic cells from early-stage breast cancer patients. Clin Cancer Res 10:3457–3464PubMedCrossRefGoogle Scholar
  6. Haimovich AD (2011) Methods, challenges, and promise of next-generation sequencing in cancer biology. Yale J Biol Med 84:439–446PubMedGoogle Scholar
  7. Jahr S, Hentze H, Englisch S, Hardt D, Fackelmayer FO, Hesch RD, Knippers R (2001) DNA fragments in the blood plasma of cancer patients: quantitations and evidence for their origin from apoptotic and necrotic cells. Cancer Res 61:1659–1665PubMedGoogle Scholar
  8. Jasmine F, Ahsan H, Andrulis IL, John EM, Chang-Claude J, Kibriya MG (2008) Whole-genome amplification enables accurate genotyping for microarray-based high-density single nucleotide polymorphism array. Cancer Epidemiol Biomarkers Prev 17:3499–3508PubMedCrossRefGoogle Scholar
  9. Jin G, Sun J, Liu W, Zhang Z, Chu LW, Kim ST, Sun J, Feng J, Duggan D, Carpten JD, Wiklund F, Grönberg H, Isaacs WB, Zheng SL, Xu J (2011) Genome-wide copy-number variation analysis identifies common genetic variants at 20p13 associated with aggressiveness of prostate cancer. Carcinogenesis 32:1057–1062PubMedCrossRefGoogle Scholar
  10. Jung K, Fleischhacker M, Rabien A (2010) Cell-free DNA in the blood as a solid tumour biomarker – a critical appraisal of the literature. Clin Chim Acta 411:1611–1624PubMedCrossRefGoogle Scholar
  11. Karrison TG, Ferguson DJ, Meier P (1999) Dormancy of mammary carcinoma after mastectomy. J Natl Cancer Inst 91:80–85PubMedCrossRefGoogle Scholar
  12. Korn JM, Kuruvilla FG, McCarroll SA, Wysoker A, Nemesh J, Cawley S, Hubbell E, Veitch J, Collins PJ, Darvishi K, Lee C, Nizzari MM, Gabriel SB, Purcell S, Daly MJ, Altshuler D (2008) Integrated genotype calling and association analysis of SNPs, common copy number polymorphisms and rare CNVs. Nat Genet 40:1253–1260PubMedCrossRefGoogle Scholar
  13. LaFramboise T (2009) Single nucleotide polymorphism arrays: a decade of biological, computational and technological advances. Nucleic Acids Res 37:4181–4193PubMedCrossRefGoogle Scholar
  14. Leon SA, Shapiro B, Sklaroff DM, Yaros MJ (1977) Free DNA in the serum of cancer patients and the effect of therapy. Cancer Res 37:646–650PubMedGoogle Scholar
  15. Lo YM, Zhang J, Leung TN, Lau TK, Chang AM, Hjelm NM (1999) Rapid clearance of fetal DNA from maternal plasma. Am J Hum Genet 64:218–224PubMedCrossRefGoogle Scholar
  16. Mandel P, Métais P (1947) Les acides nucleiques du plasma sanguin l’homme. C R Acad Sci Paris 142:241–243Google Scholar
  17. Meng S, Tripathy D, Frenkel EP, Shete S, Naftalis EZ, Huth JF, Beitsch PD, Leitch M, Hoover S, Euhus D, Haley B, Morrison L, Fleming TP, Herlyn D, Terstappen LW, Fehm T, Tucker TF, Lane N, Wang J, Uhr JW (2004) Circulating tumor cells in patients with breast cancer dormancy. Clin Cancer Res 10:8152–8162PubMedCrossRefGoogle Scholar
  18. Nawroz H, Koch W, Anker P, Stroun M, Sidransky D (1996) Microsatellite alterations in serum DNA of head and neck cancer patients. Nat Med 2:1035–1037PubMedCrossRefGoogle Scholar
  19. Page K, Powles T, Slade MJ, De Bella MT, Walker RA, Coombes RC, Shaw JA (2006) The importance of careful blood processing in isolation of cell-free DNA. Ann N Y Acad Sci 1075:313–317PubMedCrossRefGoogle Scholar
  20. Page K, Hava N, Ward B, Brown J, Guttery DS, Ruangpratheep C, Blighe K, Sharma A, Walker RA, Coombes RC, Shaw JA (2011) Detection of HER2 amplification in circulating free DNA in patients with breast cancer. Br J Cancer 104:1342–1348PubMedCrossRefGoogle Scholar
  21. Payne R, Hava N, Page K, Blighe K, Ward B, Slade M, Brown J, Guttery D, Zaidi SAA, Stebbing J, Jacob J, Tat T, Yagüe E, Shaw JA, Coombes RC (2012) The presence of disseminated tumour cells in the bone marrow is inversely related to circulating-free DNA in plasma in breast cancer dormancy. Br J Cancer 106:375–382PubMedCrossRefGoogle Scholar
  22. Quackenbush J (2002) Microarray data normalization and transformation. Nat Genet 32:496–501PubMedCrossRefGoogle Scholar
  23. Redon R, Ishikawa S, Fitch KR, Feuk L, Perry GH, Andrews TD, Fiegler H, Shapero MH, Carson AR, Chen W, Cho EU, Dallaire S, Freeman JL, González JR, Gratacós M, Huang J, Kalaitzopoulos D, Komura D, MacDonald JR, Marshall CR, Mei R, Montgomery L, Nishimura K, Okamura K, Shen F, Somerville MJ, Tchinda J, Valsesia A, Woodwark C, Yang F, Zhang J, Zerjal T, Zhang J, Armengols L, Conrad DF, Estivill X, Tyler-Smith C, Carter NP, Aburatani H, Lee C, Jones KW, Scherer SW, Hurles ME (2006) Global variation in copy number in the human genome. Nature 444:444–454PubMedCrossRefGoogle Scholar
  24. Rook MS, Delach SM, Deyneko G, Worlock A, Wolfe JL (2004) Whole genome amplification of DNA from laser capture-microdissected tissue for high-throughput single nucleotide polymorphism and short tandem repeat genotyping. Am J Pathol 164:23–33PubMedCrossRefGoogle Scholar
  25. Schwarzenbach H, Hoon DS, Pantel K (2011) Cell-free nucleic acids as biomarkers in cancer patients. Nat Rev Cancer 11:426–437PubMedCrossRefGoogle Scholar
  26. Sebat J, Lakshmi B, Troge J, Alexander J, Young J, Lundin P, Månér S, Massa H, Walker M, Chi M, Navin N, Lucito R, Healy J, Hicks J, Ye K, Reiner A, Gilliam TC, Trask B, Patterson N, Zetterberg A, Wigler M (2004) Large-scale copy number polymorphism in the human genome. Science 305:525–528PubMedCrossRefGoogle Scholar
  27. Shapiro B, Chakrabarty M, Cohn EM, Leon SA (1983) Determination of circulating DNA levels in patients with benign or malignant gastrointestinal disease. Cancer 51:2116–2120PubMedCrossRefGoogle Scholar
  28. Shaw JA, Smith BM, Walsh T, Johnson S, Primrose L, Slade MJ, Walker RA, Coombes RC (2000) Microsatellite alterations in plasma DNA of primary breast cancer patients. Clin Cancer Res 6:1119–1124PubMedGoogle Scholar
  29. Shaw JA, Page K, Blighe K, Hava N, Guttery D, Ward B, Brown J, Ruangpratheep C, Stebbing J, Payne R, Palmieri C, Cleator S, Walker RA, Coombes RC (2012) Genomic analysis of circulating cell-free DNA infers breast cancer dormancy. Genome Res 22:220–231PubMedCrossRefGoogle Scholar
  30. Silva JM, Garcia JM, Dominguez G, Silva J, Miralles C, Cantos B, Coca S, Provencio M, España P, Bonilla F (2002) Persistence of tumor DNA in plasma of breast cancer patients after mastectomy. Ann Surg Oncol 9:71–76PubMedCrossRefGoogle Scholar
  31. Tuefferd M, De Bondt A, Van Den Wyngaert I, Talloen W, Verbeke T, Carvalho B, Clevert DA, Alifano M, Raghavan N, Amaratunga D, Göhlmann H, Broët P, Camilleri-Broët S (2008) Genome-wide copy number alterations detection in fresh frozen and matched FFPE samples using SNP 6.0 arrays. Genes Chromosomes Cancer 47:957–964PubMedCrossRefGoogle Scholar
  32. Uchida K (2006) Gene amplification and cancer, encyclopedia of life sciences. Wiley Online Library. http://onlinelibrary.wiley.com/doi/10.1038/npg.els.0006047/full
  33. Van de Wiel MA, Picard F, van Wieringen WN, Ylstra B (2011) Preprocessing and downstream analysis of microarray DNA copy number profiles. Brief Bioinform 12:10–21PubMedCrossRefGoogle Scholar
  34. Wang BG, Huang HY, Chen YC, Bristow RE, Kassauei K, Cheng CC, Roden R, Sokoll LJ, Chan DW, Shih IM (2003) Increased plasma DNA integrity in cancer patients. Cancer Res 63:3966–3968PubMedGoogle Scholar
  35. Yoshihara K, Tajima A, Adachi S, Quan J, Sekine M, Kase H, Yahata T, Inoue I, Tanaka K (2011) Germline copy number variations in BRCA1-associated ovarian cancer patients. Genes Chromosomes Cancer 50:167–177PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  1. 1.Cancer Studies and Molecular MedicineRobert Kilpatrick CSB (Room 305), Leicester Royal InfirmaryLeicesterUK
  2. 2.Department of Cancer Studies and Molecular Medicine, Infirmary Close, Leicester Royal InfirmaryUniversity of Leicester, Robert Kilpatrick Clinical Sciences BuildingLeicesterUK
  3. 3.Cancer Research UK Laboratories, Department of OncologyImperial College, Hammersmith Hospital CampusLondonUK

Personalised recommendations