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Ionizing radiation or mitomycin-induced micronuclei in lymphocytes of BRCA1 or BRCA2 mutation carriers

Breast Cancer Research and Treatment volume 127, pages 611–622 (2011)Cite this article

Abstract

BRCA1 and BRCA2 genes are essential in preserving the integrity of genome, and it is not unambiguously clear whether the heterozygosity status may affect BRCA1 or BRCA2 functions. This may have implications for the clinical management of BRCA1 and BRCA2 mutation carriers both in breast cancer (BC) screening modality and in cancer treatment based on DNA-damaging or DNA-repair-inhibiting drugs. We investigated whether lymphocytes carrying BRCA1 or BRCA2 mutations displayed an increased sensitivity to radiation or mitomycin C (MMC) in vitro treatments. Peripheral blood from 21 BRCA1 mutation carriers (12 with BC and 9 healthy), 24 BRCA2 carriers (13 with BC and 11 healthy), 15 familial BC patients without detected mutation in BRCA1 or BRCA2 and 16 controls without familial history of cancer (5 with BC and 11 healthy) were irradiated or treated with MMC. Chromosomal damage was measured using the cytokinesis-block micronucleus assay. We evaluated micronuclei (MN) and nucleoplasmic bridges (NPBs). The BRCA2 mutation carriers and familial BC patients without detected mutation in BRCA1 or BRCA2 showed less basal NPB than BRCA1 carriers and controls. The BRCA1+/− or BRCA2+/− lymphocytes did not have increased frequencies of MN or NPB after irradiation. In contrast, BRCA2+/− lymphocytes presented higher levels of MN after MMC exposure than BRCA1 carriers and controls. The monoallelic BRCA1 or BRCA2 pathogenic mutations seem not to be associated with an enhanced radiosensitivity. The mutation of one BRCA2 allele conferred an increased sensitivity to MMC, presumably because of the role of this gene in the repair of MMC-induced DNA damage. This finding indicates that the MMC-induced MN analysis could be useful in identifying functional deficiencies of BRCA2 or genes related to BRCA2. Since MMC can be used as an anti-cancer drug, these data may be relevant for the management and follow-up of BRCA2 mutation carriers.

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References

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

    PubMed  Article  CAS  Google Scholar 

  2. Boulton SJ (2006) Cellular functions of the BRCA tumour-suppressor proteins. Biochem Soc Trans 34:633–645

    PubMed  Article  CAS  Google Scholar 

  3. Gudmundsdottir K, Ashworth A (2006) The roles of BRCA1 and BRCA2 and associated proteins in the maintenance of genomic stability. Oncogene 25:5864–5874

    PubMed  Article  CAS  Google Scholar 

  4. Cousineau I, Belmaaza A (2007) BRCA1 haploinsufficiency, but not heterozygosity for a BRCA1-truncating mutation, deregulates homologous recombination. Cell Cycle 6:962–971

    PubMed  Article  CAS  Google Scholar 

  5. Venkitaraman AR (2009) Linking the cellular functions of BRCA genes to cancer pathogenesis and treatment. Annu Rev Pathol 4:461–487

    PubMed  Article  CAS  Google Scholar 

  6. Arnold K, Kim MK, Frerk K, Edler L, Savelyeva L, Schmezer P, Wiedemeyer R (2006) Lower level of BRCA2 protein in heterozygous mutation carriers is correlated with an increase in DNA double strand breaks and an impaired DSB repair. Cancer Lett 243:90–100

    PubMed  Article  CAS  Google Scholar 

  7. Speit G, Trenz K (2004) Chromosomal mutagen sensitivity associated with mutations in BRCA genes. Cytogenet Genome Res 104:325–332

    PubMed  Article  CAS  Google Scholar 

  8. Fenech M (2007) Cytokinesis-block micronucleus cytome assay. Nat Protoc 2:1084–1104

    PubMed  Article  CAS  Google Scholar 

  9. Thomas P, Umegaki K, Fenech M (2003) Nucleoplasmic bridges are a sensitive measure of chromosome rearrangement in the cytokinesis-block micronucleus assay. Mutagenesis 18:187–194

    PubMed  Article  CAS  Google Scholar 

  10. Fell LJ, Paul ND, McMillan TJ (2002) Role for non-homologous end-joining in the repair of UVA-induced DNA damage. Int J Radiat Biol 78:1023–1027

    PubMed  Article  CAS  Google Scholar 

  11. Ouyang Y, Kwon YT, An JY, Eller D, Tsai SC, Diaz-Perez S, Troke JJ, Teitell MA, Marahrens Y (2006) Loss of Ubr2, an E3 ubiquitin ligase, leads to chromosome fragility and impaired homologous recombinational repair. Mutat Res 596:64–75

    PubMed  CAS  Google Scholar 

  12. Yue J, Wang Q, Lu H, Brenneman M, Fan F, Shen Z (2009) The cytoskeleton protein filamin-A is required for an efficient recombinational DNA double strand break repair. Cancer Res 69:7978–7985

    PubMed  Article  CAS  Google Scholar 

  13. Fenech M (2010) The lymphocyte cytokinesis-block micronucleus cytome assay and its application in radiation biodosimetry. Health Phys 98:234–243

    PubMed  Article  CAS  Google Scholar 

  14. Lee TK, Allison RR, O’Brien KF, Naves JL, Karlsson UL, Wiley AL (2002) Persistence of micronuclei in lymphocytes of cancer patients after radiotherapy. Radiat Res 157:678–684

    PubMed  Article  CAS  Google Scholar 

  15. Baria K, Warren C, Roberts SA, West CM, Scott D (2001) Chromosomal radiosensitivity as a marker of predisposition to common cancers? Br J Cancer 84:892–896

    PubMed  Article  CAS  Google Scholar 

  16. El-Zein RA, Schabath MB, Etzel CJ, Lopez MS, Franklin JD, Spitz MR (2006) Cytokinesis-blocked micronucleus assay as a novel biomarker for lung cancer risk. Cancer Res 66:6449–6456

    PubMed  Article  CAS  Google Scholar 

  17. El-Zein RA, Fenech M, Lopez MS, Spitz MR, Etzel CJ (2008) Cytokinesis-blocked micronucleus cytome assay biomarkers identify lung cancer cases amongst smokers. Cancer Epidemiol Biomarkers Prev 17:1111–1119

    PubMed  Article  CAS  Google Scholar 

  18. Baeyens A, Thierens H, Claes K, Poppe B, Messiaen L, de RL, Vral A (2002) Chromosomal radiosensitivity in breast cancer patients with a known or putative genetic predisposition. Br J Cancer 87:1379–1385

    PubMed  Article  CAS  Google Scholar 

  19. Burrill W, Barber JB, Roberts SA, Bulman B, Scott D (2000) Heritability of chromosomal radiosensitivity in breast cancer patients: a pilot study with the lymphocyte micronucleus assay. Int J Radiat Biol 76:1617–1619

    PubMed  Article  CAS  Google Scholar 

  20. Parshad R, Price FM, Bohr VA, Cowans KH, Zujewski JA, Sanford KK (1996) Deficient DNA repair capacity, a predisposing factor in breast cancer. Br J Cancer 74:1–5

    PubMed  Article  CAS  Google Scholar 

  21. Roberts SA, Spreadborough AR, Bulman B, Barber JB, Evans DG, Scott D (1999) Heritability of cellular radiosensitivity: a marker of low-penetrance predisposition genes in breast cancer? Am J Hum Genet 65:784–794

    PubMed  Article  CAS  Google Scholar 

  22. Scott D, Barber JB, Spreadborough AR, Burrill W, Roberts SA (1999) Increased chromosomal radiosensitivity in breast cancer patients: a comparison of two assays. Int J Radiat Biol 75:1–10

    PubMed  Article  CAS  Google Scholar 

  23. Wu X, Spitz MR, Amos CI et al (2006) Mutagen sensitivity has high heritability: evidence from a twin study. Cancer Res 66:5993–5996

    PubMed  Article  CAS  Google Scholar 

  24. Baeyens A, Thierens H, Claes K, Poppe B, de RL, Vral A (2004) Chromosomal radiosensitivity in BRCA1 and BRCA2 mutation carriers. Int J Radiat Biol 80:745–756

    PubMed  Article  CAS  Google Scholar 

  25. Baria K, Warren C, Roberts SA, West CM, Evans DG, Varley JM, Scott D (2001) Correspondence re: A. Rothfuss et al., Induced micronucleus frequencies in peripheral blood lymphocytes as a screening test for carriers of a BRCA1 mutation in breast cancer families. Cancer Res., 60: 390–394, 2000. Cancer Res 61:5948–5949

    PubMed  CAS  Google Scholar 

  26. Barwell J, Pangon L, Georgiou A et al (2007) Lymphocyte radiosensitivity in BRCA1 and BRCA2 mutation carriers and implications for breast cancer susceptibility. Int J Cancer 121:1631–1636

    PubMed  Article  CAS  Google Scholar 

  27. Buchholz TA, Wu X, Hussain A et al (2002) Evidence of haplotype insufficiency in human cells containing a germline mutation in BRCA1 or BRCA2. Int J Cancer 97:557–561

    PubMed  Article  CAS  Google Scholar 

  28. Febrer E, Mestres M, Caballin MR, Barrios L, Ribas M, Gutierrez-Enriquez S, Alonso C, Cajal T, Francesc BJ (2008) Mitotic delay in lymphocytes from BRCA1 heterozygotes unable to reduce the radiation-induced chromosomal damage. DNA Repair (Amst) 7:1907–1911

    Article  CAS  Google Scholar 

  29. Kote-Jarai Z, Salmon A, Mengitsu T et al (2006) Increased level of chromosomal damage after irradiation of lymphocytes from BRCA1 mutation carriers. Br J Cancer 94:308–310

    PubMed  Article  CAS  Google Scholar 

  30. Kotsopoulos J, Chen Z, Vallis KA, Poll A, Ainsworth P, Narod SA (2007) DNA repair capacity as a possible biomarker of breast cancer risk in female BRCA1 mutation carriers. Br J Cancer 96:118–125

    PubMed  Article  CAS  Google Scholar 

  31. Kowalska E, Narod SA, Huzarski T, Zajaczek S, Huzarska J, Gorski B, Lubinski J (2005) Increased rates of chromosome breakage in BRCA1 carriers are normalized by oral selenium supplementation. Cancer Epidemiol Biomarkers Prev 14:1302–1306

    PubMed  Article  CAS  Google Scholar 

  32. Rothfuss A, Schutz P, Bochum S, Volm T, Eberhardt E, Kreienberg R, Vogel W, Speit G (2000) Induced micronucleus frequencies in peripheral lymphocytes as a screening test for carriers of a BRCA1 mutation in breast cancer families. Cancer Res 60:390–394

    PubMed  CAS  Google Scholar 

  33. Trenz K, Rothfuss A, Schutz P, Speit G (2002) Mutagen sensitivity of peripheral blood from women carrying a BRCA1 or BRCA2 mutation. Mutat Res 500:89–96

    PubMed  CAS  Google Scholar 

  34. Trenz K, Lugowski S, Jahrsdorfer U, Jainta S, Vogel W, Speit G (2003) Enhanced sensitivity of peripheral blood lymphocytes from women carrying a BRCA1 mutation towards the mutagenic effects of various cytostatics. Mutat Res 544:279–288

    PubMed  Article  CAS  Google Scholar 

  35. Varga D, Hoegel J, Maier C et al (2006) On the difference of micronucleus frequencies in peripheral blood lymphocytes between breast cancer patients and controls. Mutagenesis 21:313–320

    PubMed  Article  CAS  Google Scholar 

  36. Vogel W, Surowy H (2007) Reduced DNA repair in BRCA1 mutation carriers undetectable before onset of breast cancer? Br J Cancer 97:1184–1186

    PubMed  Article  CAS  Google Scholar 

  37. Bonassi S, Fenech M, Lando C et al (2001) HUman MicroNucleus project: international database comparison for results with the cytokinesis-block micronucleus assay in human lymphocytes: I. Effect of laboratory protocol, scoring criteria, and host factors on the frequency of micronuclei. Environ Mol Mutagen 37:31–45

    PubMed  Article  CAS  Google Scholar 

  38. Patino-Garcia B, Hoegel J, Varga D, Hoehne M, Michel I, Jainta S, Kreienberg R, Maier C, Vogel W (2006) Scoring variability of micronuclei in binucleated human lymphocytes in a case-control study. Mutagenesis 21:191–197

    PubMed  Article  CAS  Google Scholar 

  39. Fenech M, Bonassi S, Turner J et al (2003) Intra- and inter-laboratory variation in the scoring of micronuclei and nucleoplasmic bridges in binucleated human lymphocytes. Results of an international slide-scoring exercise by the HUMN project. Mutat Res 534:45–64

    PubMed  CAS  Google Scholar 

  40. Burma S, Chen BP, Chen DJ (2006) Role of non-homologous end joining (NHEJ) in maintaining genomic integrity. DNA Repair (Amst) 5:1042–1048

    Article  CAS  Google Scholar 

  41. Frankenberg-Schwager M, Gebauer A, Koppe C, Wolf H, Pralle E, Frankenberg D (2009) Single-strand annealing, conservative homologous recombination, nonhomologous DNA end joining, and the cell cycle-dependent repair of DNA double-strand breaks induced by sparsely or densely ionizing radiation. Radiat Res 171:265–273

    PubMed  Article  CAS  Google Scholar 

  42. Koch K, Wrona A, Dikomey E, Borgmann K (2009) Impact of homologous recombination on individual cellular radiosensitivity. Radiother Oncol 90:265–272

    PubMed  Article  CAS  Google Scholar 

  43. Baldeyron C, Jacquemin E, Smith J, Jacquemont C, De OI, Gad S, Feunteun J, Stoppa-Lyonnet D, Papadopoulo D (2002) A single mutated BRCA1 allele leads to impaired fidelity of double strand break end-joining. Oncogene 21:1401–1410

    PubMed  Article  CAS  Google Scholar 

  44. Bau DT, Mau YC, Shen CY (2006) The role of BRCA1 in non-homologous end-joining. Cancer Lett 240:1–8

    PubMed  Article  CAS  Google Scholar 

  45. Ernestos B, Nikolaos P, Koulis G, Eleni R, Konstantinos B, Alexandra G, Michael K (2010) Increased chromosomal radiosensitivity in women carrying BRCA1/BRCA2 mutations assessed with the G2 assay. Int J Radiat Oncol Biol Phys 76:1199–1205

    PubMed  Article  CAS  Google Scholar 

  46. de Berrington GA, Berg CD, Visvanathan K, Robson M (2009) Estimated risk of radiation-induced breast cancer from mammographic screening for young BRCA mutation carriers. J Natl Cancer Inst 101:205–209

    Article  Google Scholar 

  47. Shanley S, McReynolds K, Ardern-Jones A et al (2006) Late toxicity is not increased in BRCA1/BRCA2 mutation carriers undergoing breast radiotherapy in the United Kingdom. Clin Cancer Res 12:7025–7032

    PubMed  Article  CAS  Google Scholar 

  48. Broeks A, Braaf LM, Huseinovic A et al (2007) Identification of women with an increased risk of developing radiation-induced breast cancer: a case only study. Breast Cancer Res 9:R26

    PubMed  Article  Google Scholar 

  49. Shorrocks J, Tobi SE, Latham H, Peacock JH, Eeles R, Eccles D, McMillan TJ (2004) Primary fibroblasts from BRCA1 heterozygotes display an abnormal G1/S cell cycle checkpoint following UVA irradiation but show normal levels of micronuclei following oxidative stress or mitomycin C treatment. Int J Radiat Oncol Biol Phys 58:470–478

    PubMed  Article  CAS  Google Scholar 

  50. Lee YJ, Park SJ, Ciccone SL, Kim CR, Lee SH (2006) An in vivo analysis of MMC-induced DNA damage and its repair. Carcinogenesis 27:446–453

    PubMed  Article  CAS  Google Scholar 

  51. van der Heijden MS, Brody JR, Dezentje DA et al (2005) In vivo therapeutic responses contingent on Fanconi anemia/BRCA2 status of the tumor. Clin Cancer Res 11:7508–7515

    PubMed  Article  Google Scholar 

  52. Chalasani P, Kurtin S, Dragovich T (2008) Response to a third-line mitomycin C (MMC)-based chemotherapy in a patient with metastatic pancreatic adenocarcinoma carrying germline BRCA2 mutation. JOP 9:305–308

    PubMed  Google Scholar 

  53. Beetstra S, Salisbury C, Turner J, Altree M, McKinnon R, Suthers G, Fenech M (2006) Lymphocytes of BRCA1 and BRCA2 germ-line mutation carriers, with or without breast cancer, are not abnormally sensitive to the chromosome damaging effect of moderate folate deficiency. Carcinogenesis 27:517–524

    PubMed  Article  CAS  Google Scholar 

  54. Daniels MJ, Wang Y, Lee M, Venkitaraman AR (2004) Abnormal cytokinesis in cells deficient in the breast cancer susceptibility protein BRCA2. Science 306:876–879

    PubMed  Article  CAS  Google Scholar 

  55. Jonsdottir AB, Vreeswijk MP, Wolterbeek R, Devilee P, Tanke HJ, Eyfjord JE, Szuhai K (2009) BRCA2 heterozygosity delays cytokinesis in primary human fibroblasts. Cell Oncol 31:191–201

    PubMed  CAS  Google Scholar 

  56. Lekomtsev S, Guizetti J, Pozniakovsky A, Gerlich DW, Petronczki M (2010) Evidence that the tumor-suppressor protein BRCA2 does not regulate cytokinesis in human cells. J Cell Sci 123:1395–1400

    PubMed  Article  CAS  Google Scholar 

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Acknowledgments

This study was supported by two grants of the “Fondo de Investigación Sanitaria” from the Spanish Health Ministry: FIS 04/1832 and FIS 05/2181 and by a grant of the “Fundación Mutua Madrileña” (2008–2011).

Conflict of interest statement

The authors declare that they have no competing interests.

Author information

Affiliations

  1. Vall d’Hebron Institute of Oncology (VHIO), Barcelona, Spain

    Sara Gutiérrez-Enríquez & Orland Diez

  2. Medical Oncology Program, Vall d’Hebron University Hospital Research Institute, Barcelona, Spain

    Sara Gutiérrez-Enríquez

  3. Medical Oncology Department, Santa Creu and Sant Pau Hospital, Barcelona, Spain

    Teresa Ramón y Cajal, Carmen Alonso & Judith Sanz

  4. Group of Mutagenesis, Genetic and Microbiology Department, Autonomous University of Barcelona, Cerdanyola del Vallès, Spain

    Anna Corral

  5. Radiophysics and Radioprotection Department, Santa Creu and Sant Pau Hospital, Barcelona, Spain

    Pablo Carrasco & Montserrat Ribas

  6. Genetics Department, Santa Creu and Sant Pau Hospital, Barcelona, Spain

    Mónica Cornet & Montserrat Baiget

  7. Oncogenetics Laboratory, Clinical Laboratories, University Hospital Vall d’Hebron, Passeig Vall d’Hebron 119-129, Barcelona, 08035, Spain

    Sara Gutiérrez-Enríquez & Orland Diez

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  1. Sara Gutiérrez-Enríquez
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  2. Teresa Ramón y Cajal
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Correspondence to Sara Gutiérrez-Enríquez.

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Gutiérrez-Enríquez, S., Ramón y Cajal, T., Alonso, C. et al. Ionizing radiation or mitomycin-induced micronuclei in lymphocytes of BRCA1 or BRCA2 mutation carriers. Breast Cancer Res Treat 127, 611–622 (2011). https://doi.org/10.1007/s10549-010-1017-6

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  • DOI: https://doi.org/10.1007/s10549-010-1017-6

Keywords

  • In vitro radiation
  • Mitomycin C
  • Micronuclei
  • BRCA1 and BRCA2