Skip to main content

Advertisement

SpringerLink
Log in

Validation of the BRCA1 antibody MS110 and the utility of BRCA1 as a patient selection biomarker in immunohistochemical analysis of breast and ovarian tumours

  • Original Article
  • Published:
Virchows Archiv Aims and scope Submit manuscript

An Erratum to this article was published on 19 April 2013

Abstract

BRCA1 protein measurement has previously been evaluated as a potential diagnostic marker without reaching a conclusive recommendation. In this study, we applied current best practice in antibody validation to further characterize MS110, a widely used antibody targeting BRCA1. Antibody specificity was investigated using different biochemical validation techniques. We found that BRCA1 could not be reliably detected using immunoprecipitation and Western blot in endogenously expressing cells. We used immunohistochemistry on formalin-fixed paraffin-embedded cell pellets to establish compatibility with formalin-fixed paraffin-embedded samples. We demonstrated that in transfected cells and cell lines with known genetic BRCA1 status, MS110 successfully detected BRCA1 giving the expected level of staining in immunohistochemistry. Following this, we investigated the use of BRCA1 protein measurement by immunohistochemistry in a cohort of triple negative breast and serous ovarian tumour samples to explore the use of BRCA1 protein measurement by immunohistochemistry for patient stratification. Using MS110 in repeated standardized experiments, on serial sections from a panel of patient samples, results demonstrated considerable run-to-run variability. We concluded that in formalin-fixed tissue samples, MS110 does detect BRCA1; however, using standard methodologies, BRCA1 expression levels in tissue samples is incompatible with the use of this protein as a statistically robust patient selection marker in immunohistochemistry. These results demonstrate the need for further development to deliver BRCA1 protein quantification by immunohistochemistry as a patient stratification marker.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Miki Y, Swensen J, Shattuck-Eidens D, Futreal PA, Harshman K, Tavtigian S, Liu Q, Cochran C, Bennett LM, Ding W (1994) A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1. Science 266:66–71

    Article  PubMed  CAS  Google Scholar 

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

    Article  PubMed  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

    Article  PubMed  CAS  Google Scholar 

  4. Hall JM, Lee MK, Newman B, Morrow JE, Anderson LA, Huey B, King MC (1990) Linkage of early-onset familial breast cancer to chromosome 17q21. Science 250:1684–1689

    Article  PubMed  CAS  Google Scholar 

  5. Frank TS, Manley SA, Olopade OI, Cummings S, Garber JE, Bernhardt B, Antman K, Russo D, Wood ME, Mullineau L, Isaacs C, Peshkin B, Buys S, Venne V, Rowley PT, Loader S, Offit K, Robson M, Hampel H, Brener D, Winer EP, Clark S, Weber B, Strong LC, Thomas A (1998) Sequence analysis of BRCA1 and BRCA2: correlation of mutations with family history and ovarian cancer risk. J Clin Oncol 16:2417–2425

    PubMed  CAS  Google Scholar 

  6. Bansal A, Critchfield GC, Frank TS, Reid JE, Thomas A, Deffenbaugh AM, Neuhausen SL (2000) The predictive value of BRCA1 and BRCA2 mutation testing. Genet Test 4:45–48

    Article  PubMed  CAS  Google Scholar 

  7. Anonymous (2011) StatBite lifetime risk of breast cancer subtypes by race. J Natl Cancer Inst 103:537

    Article  Google Scholar 

  8. Vorrius TR, Snyder K, Pica-Mendez A, Tan C, Laterza O, Toniatti C, Carpenter C, Lee H, Tanaka W, Zhang Z (2002) Immunohistochemical detection of BRCA-1 and BRCA-2 expression in human breast and ovarian tumors. J Histotechnol 32:202–203

    Google Scholar 

  9. Perez-Valles A, Martorell-Cebollada M, Nogueira-Vazquez E, Garcia-Garcia JA, Fuster-Diana E (2001) The usefulness of antibodies to the BRCA1 protein in detecting the mutated BRCA1 gene. An immunohistochemical study. J Clin Pathol 54:476–480

    Article  PubMed  CAS  Google Scholar 

  10. Bordeaux J, Welsh A, Agarwal S, Killiam E, Baquero M, Hanna J, Anagnostou V, Rimm D (2010) Antibody validation. Biotechniques 48:197–209

    Article  PubMed  CAS  Google Scholar 

  11. Dantzer F, de La Rubia G, Menissier-De Murcia J, Hostomsky Z, de Murcia G, Schreiber V (2000) Base excision repair is impaired in mammalian cells lacking Poly(ADP-ribose) polymerase-1. Biochemistry 39:7559–7569

    Article  PubMed  CAS  Google Scholar 

  12. Ame JC, Spenlehauer C, de Murcia G (2004) The PARP superfamily. Bioessays 26:882–893

    Article  PubMed  CAS  Google Scholar 

  13. Manders EM, Stap J, Brakenhoff GJ, van Driel R, Aten JA (1992) Dynamics of three-dimensional replication patterns during the S-phase, analysed by double labelling of DNA and confocal microscopy. J Cell Sci 103(Pt 3):857–862

    PubMed  CAS  Google Scholar 

  14. McCarty KS Jr, Szabo E, Flowers JL, Cox EB, Leight GS, Miller L, Konrath J, Soper JT, Budwit DA, Creasman WT, Seigler HF, McCarty KS Sr (1986) Use of a monoclonal anti-estrogen receptor antibody in the immunohistochemical evaluation of human tumors. Cancer Res (Suppl) 46:4244–4248

    Google Scholar 

  15. Lambie H, Miremadi A, Pinder SE, Bell JA, Wencyk P, Paish EC, Macmillan RD, Ellis IO (2003) Prognostic significance of BRCA1 expression in sporadic breast carcinomas. J Pathol 200:207–213

    Article  PubMed  CAS  Google Scholar 

  16. Lee WY, Jin YT, Chang TW, Lin PW, Su IJ (1999) Immunolocalization of BRCA1 protein in normal breast tissue and sporadic invasive ductal carcinomas: a correlation with other biological parameters. Histopathology 34:106–112

    Article  PubMed  CAS  Google Scholar 

  17. Sun X, Gong Y, Rao MS, Badve S (2002) Loss of BRCA1 expression in sporadic male breast carcinoma. Breast Cancer Res Treat 71:1–7

    Article  PubMed  CAS  Google Scholar 

  18. Yoshikawa K, Honda K, Inamoto T, Shinohara H, Yamauchi A, Suga K, Okuyama T, Shimada T, Kodama H, Noguchi S, Gazdar AF, Yamaoka Y, Takahashi R (1999) Reduction of BRCA1 protein expression in Japanese sporadic breast carcinomas and its frequent loss in BRCA1-associated cases. Clin Cancer Res 5:1249–1261

    PubMed  CAS  Google Scholar 

  19. Fraser JA, Reeves JR, Stanton PD, Black DM, Going JJ, Cooke TG, Bartlett JM (2003) A role for BRCA1 in sporadic breast cancer. Br J Cancer 88:1263–1270

    Article  PubMed  CAS  Google Scholar 

  20. Alamshah A, Springall R, Gillett CE, Solomon E, Morris JR (2008) Use of a BRCA1 peptide validates MS110 as a BRCA1-specific antibody in immunohistochemistry. Histopathology 53:117–120

    Article  PubMed  CAS  Google Scholar 

  21. Watson P, Townley AK, Koka P, Palmer KJ, Stephens DJ (2006) Sec16 defines endoplasmic reticulum exit sites and is required for secretory cargo export in mammalian cells. Traffic 7:1678–1687

    Article  PubMed  CAS  Google Scholar 

  22. Balczon R, Bao L, Zimmer WE (1994) PCM-1, A 228-kD centrosome autoantigen with a distinct cell cycle distribution. J Cell Biol 124:783–793

    Article  PubMed  CAS  Google Scholar 

  23. Chodosh LA (1998) Expression of BRCA1 and BRCA2 in normal and neoplastic cells. J Mammary Gland Biol Neoplasia 3:389–402

    Article  PubMed  CAS  Google Scholar 

  24. Favy DA, Lafarge S, Rio P, Vissac C, Bignon YJ, Bernard-Gallon D (2000) Real-time PCR quantification of full-length and exon 11 spliced BRCA1 transcripts in human breast cancer cell lines. Biochem Biophys Res Commun 274:73–78

    Article  PubMed  CAS  Google Scholar 

  25. Taron M, Rosell R, Felip E, Mendez P, Souglakos J, Ronco MS, Queralt C, Majo J, Sanchez JM, Sanchez JJ, Maestre J (2004) BRCA1 mRNA expression levels as an indicator of chemoresistance in lung cancer. Hum Mol Genet 13:2443–2449

    Article  PubMed  CAS  Google Scholar 

  26. Wenger CR, Clark GM (1998) S-phase fraction and breast cancer—a decade of experience. Breast Cancer Res Treat 51:255–265

    Article  PubMed  CAS  Google Scholar 

  27. McGarry TJ, Kirschner MW (1998) Geminin, an inhibitor of DNA replication, is degraded during mitosis. Cell 93:1043–1053

    Article  PubMed  CAS  Google Scholar 

  28. Gonzalez MA, Tachibana KE, Chin SF, Callagy G, Madine MA, Vowler SL, Pinder SE, Laskey RA, Coleman N (2004) Geminin predicts adverse clinical outcome in breast cancer by reflecting cell-cycle progression. J Pathol 204:121–130

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

All research in the manuscript was funded by AstraZeneca PLC. We would like to thank Dr. Madhuri Warren of Pathology Diagnostics, Ltd. for the meticulous work of second pathologist scoring of all immunohistochemical samples.

Conflicts of interest

At the time of data collection and writing of the manuscript all authors, with the exception of Erica Van Dorp, were employees of AstraZeneca R&D. Erica Van Dorp declares she has no conflict of interest.

Author information

Authors and Affiliations

  1. AstraZeneca, Alderley Park, Macclesfield, Cheshire, UK

    Roy Milner, Helen Wombwell, Sonia Eckersley, Donna Barnes, Juli Warwicker, Erica Van Dorp, Simon Dearden, Glen Hughes, Chris Harbron, Bob Wellings, Darren Hodgson, Chris Womack, Neil Gray, Alan Lau, Mark J. O’Connor, Catherine Marsden & Alexander J. Kvist

  2. 19 F7 Mereside, Alderley Park, Macclesfield, SK10 4TG, Cheshire, England, UK

    Alexander J. Kvist

Authors
  1. Roy Milner
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Helen Wombwell
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sonia Eckersley
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Donna Barnes
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Juli Warwicker
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Erica Van Dorp
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Simon Dearden
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Glen Hughes
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Chris Harbron
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Bob Wellings
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Darren Hodgson
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Chris Womack
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Neil Gray
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Alan Lau
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Mark J. O’Connor
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Catherine Marsden
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. Alexander J. Kvist
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Alexander J. Kvist.

Additional information

All work was performed at AstraZeneca.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary Fig. 1

Immunohistochemical staining of normal breast tissue for BRCA1 and PARP1. Panel a shows staining for BRCA1 on normal breast tissue with MS110. Panel b shows staining for PARP on normal breast tissue with Serotec antibody MCA1522. Scale bar, 100 μm (JPEG 2717 kb)

High resolution (EPS 5620 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Milner, R., Wombwell, H., Eckersley, S. et al. Validation of the BRCA1 antibody MS110 and the utility of BRCA1 as a patient selection biomarker in immunohistochemical analysis of breast and ovarian tumours. Virchows Arch 462, 269–279 (2013). https://doi.org/10.1007/s00428-012-1368-y

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00428-012-1368-y

Keywords

Search

Navigation