Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Manuscript
  • Published:

Minimal Residual Disease in B Cell Malignancies

Improved assessment of minimal residual disease in B cell malignancies using fluorogenic consensus probes for real-time quantitative PCR

Leukemia volume 14, pages 1419–1425 (2000)Cite this article

Abstract

PCR of clonally rearranged immunoglobulin heavy chain (IgH) gene sequences is increasingly used for detection of minimal residual disease (MRD) in lymphoid malignancies. Inherent quantitating problems are the main drawbacks of traditional PCR technologies. These limitations have been overcome by the recently developed real-time quantitative PCR (RQ PCR) technology. However, clinical application of the few published RQ PCR assays targeting immune gene rearrangements is hampered by the expensive and time-consuming need for individual hybridization probes for each patient. We have developed a new RQ PCR strategy targeting clonally rearranged IgH sequences that solves this problem. The method uses only two different JHhybridization probes and four downstream JH primers homologous to consensus germline JH gene segments. In combination with an allele-specific upstream (ASO) primer the consensus JH probes and primers allow quantitation of about 90% of possible IgH rearrangements. In a series of 22 B-lineage ALL the new assay allowed the detection of one to 10 blasts in a background of 105 normal cells. To prove the clinical utility we quantified MRD in 23 follow-up samples of six ALL patients with the new assay in comparison with a published RQ PCR technique that used individually designed primer/probe sets. We showed that the sensitivity of the new RQ PCR assay was slightly higher for four of the six cases and about 100-fold higher for one case, enabling detection of an increasing MRD level as an indicator of subsequent relapse 44 weeks earlier compared to the ASO probe assay in this particular patient. The results suggest, that the novel RQ PCR assay is a rapid, technically simple, reliable, and sensitive alternative to traditional quantification assays and simplifies current approaches of monitoring MRD in clinical trials.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2

Similar content being viewed by others

References

  1. Foroni L . Investigation of minimal residual disease in childhood and adult acute lymphoblastic leukemia by molecular analysis Br J Haematol 1999 105: 7–24

    CAS  PubMed  Google Scholar 

  2. Cave H, Guidal C, Rohrlich P, Delfau MH, Broyart A, Lescoeur B, Rahimy C, Fenneteau O, Monplaisir N, d'Auriol L, Elio J, Vilmer E, Grandchamp B . Prospective monitoring and quantitation of residual blasts in childhood acute lymphoblastic leukemia by polymerase chain reaction study of δ and γ T-cell receptor genes Blood 1994 83: 1892–1902

    CAS  PubMed  Google Scholar 

  3. Brisco MJ, Hughes E, Neoh SH, Sykes PJ, Bradstock K, Enno A, Szer J, McCaul K, Morley AA . Relationship between minimal residual disease and outcome in adult acute lymphoblastic leukemia Blood 1996 87: 5251–5256

    CAS  PubMed  Google Scholar 

  4. Roberts WM, Estrov Z, Ouspenskaia MV, Johnston DA, McClain KL, Zipf TF . Measurement of residual disease during remission in childhood acute lymphoblastic leukemia New Engl J Med 1997 336: 317–323

    Article  CAS  PubMed  Google Scholar 

  5. Cave H, van der Werff ten Bosch J, Suciu S, Guidal C, Waterkeyn C, Otten J, Bakkus M, Thielemans K, Grandchamp B, Vilmer E, Nelken B, Fournier M, Boutard P, Lebrun E, Mechinaud F, Garand R, Robert A, Dastugue N, Plouvier E, Racadot E, Ferster A, Gyselinck J, Fenneteau O, Duval M, Solbu G, Manel AM . Clinical significance of minimal residual disease in childhood acute lymphoblastic Leukemia New Engl J Med 1998 339: 591–598

    Article  CAS  PubMed  Google Scholar 

  6. Van Dongen JJ, Seriu T, Panzer-Grunmayer ER, Biondi A, Pongers-Willemse MJ, Corral L, Stolz F, Schrappe M, Masera G, Kamps WA, Gadner H, van Wering ER, Ludwig WD, Basso G, de Bruijn MA, Cazzaniga G, Hettinger K, van der Does-van den Berg A, Hop WC, Riehm H, Bartram CR . Prognostic value of minimal residual disease in acute lymphoblastic leukemia in childhood Lancet 1998 352: 1731–1738

    Article  CAS  PubMed  Google Scholar 

  7. Gruhn B, Hongeng S, Yi H, Hancock ML, Rubnitz JE, Neale GAM, Kitchingman GR . Minimal residual disease after intensive induction therapy in childhood acute lymphoblastic leukemia predicts outcome Leukemia 1998 12: 675–681

    CAS  PubMed  Google Scholar 

  8. Corradini P, Ladetto M, Pileri A, Tarella C . Clinical relevance of minimal residual disease monitoring in non-Hodgkin's lymphomas: a critical reappraisal of molecular strategies Leukemia 1999 13: 1691–1695

    Article  CAS  PubMed  Google Scholar 

  9. Gerard CJ, Olsson K, Ramanathan R, Rading C, Hanania EG . Improved quantitation of minimal residual disease in multiple myeloma using real-time polymerase chain reaction and plasmid-DNA complemetarity determining region III standards Cancer Res 1998 58: 3957–3964

    CAS  PubMed  Google Scholar 

  10. Elsworth AM, Evans PAS, Morgan GJ, Kinsey SE, Shiach CR . Quantitative PCR of the immunoglobulin heavy chain gene using genomic DNA Br J Haematol 1996 95: 700–703

    Article  CAS  PubMed  Google Scholar 

  11. Dölken L, Schüler F, Dölken G . Quantitative detection of t(14;18)-positive cells by real-time quantitative PCR using fluorogenic probes BioTechniques 1998 25: 1058–1064

    Article  PubMed  Google Scholar 

  12. Luthra R, McBride JA, Cabanillas F, Sarris A . Novel 5′ exonuclease-based real-time PCR assay for the detection of t(14;18)(q32;q21) in patients with follicular lymphoma Am J Pathol 1998 153: 63–68

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Pongers-Willemse MJ, Verhagen OJHM, Tibbe GJM, Wijkhuijs AJM, de Haas V, Roovers E, Van der Schoot CE, Van Dongen JJM . Real-time quantitative PCR for the detection of minimal residual disease in acute lymphoblastic leukemia using junctional region specific TaqMan probes Leukemia 1998 12: 2006–2014

    Article  CAS  PubMed  Google Scholar 

  14. Olsson K, Gerard CJ, Zehnder J, Jones C, Ramanathan R, Reading C, Hanania EG . Real-time t(11;14) and t(14;18) PCR assays provide sensitive and quantitative assessment of minimal residual disease (MRD) Leukemia 1999 13: 1833–1842

    Article  CAS  PubMed  Google Scholar 

  15. Holland PM, Abramson RD, Watson R, Gelfand DH . Detection of specific polymerase chain reaction product by utilizing the 5′–3′ exonuclease activity of Thermus aquaticus DNA polymerase Proc Natl Acad Sci USA 1991 88: 7276–7280

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Gibson UE, Heid CA, Williams PM . A novel method for real time quantitative RT-PCR Genome Res 1996 6: 995–1001

    Article  CAS  PubMed  Google Scholar 

  17. Heid CA, Stevens J, Livak KJ, Williams PM . Real time quantitative PCR Genome Res 1996 6: 321–328

    Article  Google Scholar 

  18. Linke B, Bolz I, Pott C, Hiddemann W, Kneba M . Use of UITma DNA polymerase improves the PCR detection of rearranged immunoglobulin heavy chain CDR3 junctions Leukemia 1995 9: 2133–2137

    CAS  PubMed  Google Scholar 

  19. Linke B, Bolz I, Fayyazi A, von Hofen M, Pott C, Bertram J, Hiddemann W, Kneba M . Automated high resolution PCR fragment analysis for identification of clonally rearranged immunoglobulin heavy chain genes Leukemia 1997 11: 1055–1062

    Article  CAS  PubMed  Google Scholar 

  20. Linke B, Pyttlich J, Tiemann M, Suttorp M, Parwaresch R, Hiddemann W, Kneba M . Identification and structural analysis of rearranged immunoglobulin heavy chain genes in lymphomas and leukemias Leukemia 1995 9: 840–847

    CAS  PubMed  Google Scholar 

  21. Flanagan JG, Rabbitts TH . The sequence of a human immunoglobulin epsilon heavy chain constant region gene, and evidence for three non-allelic genes EMBO J 1982 1: 655–660

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Mattila PS, Schugk HW, Makela O . Extensive allelic sequence variation in the J region of the human immunoglobulin heavy chain gene locus Eur J Immunol 1995 25: 2578–2582

    Article  CAS  PubMed  Google Scholar 

  23. Ichihara Y, Matsuoka H, Kurosawa Y . Organization of human immunoglobulin heavy chain diversity gene loci EMBO J 1988 7: 4141–4150

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Huang C, Stewart AK, Schwartz RS, Stollar BD . Immunoglobulin heavy chain gene expression in peripheral blood lymphocytes J Clin Invest 1992 89: 1331–1343

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Mortari F, Wang JY, Schroeder HW . Mixed population of VH gene segments and CDR3 distribution in the expressed Cα and Cγ repertoires J Immunol 1993 150: 1348–1357

    CAS  PubMed  Google Scholar 

  26. Corbett SJ, Tomlinson IM, Sonnhammer, Buck D, Winter G . Sequence of the human immunoglobulin diversity (D) segment locus: a systematic analysis provides no evidence for the use of DIR segments, inverted D segments, ‘minor’ D segments or D–D recombination J Mol Biol 1997 270: 587–597

    Article  CAS  PubMed  Google Scholar 

  27. Brezinschek HP, Brezinschek RI, Lipsky PE . Analysis of the heavy chain repertoire of human peripheral B cells using single-cell polymerase chain reaction J Immunol 1995 155: 190–202

    CAS  PubMed  Google Scholar 

  28. Nizet Y, Martiat P, Vaerman JL, Philippe M, Wildmann C, Staelens JP, Cornu G, Ferrant A, Michaux JL Sokal G . Follow-up of residual disease (MRD) in B lineage akute leukemia using a simplified PCR strategy: evolution of MRD rather than its detection is correlated with clinical outcome Br J Haematol 1991 79: 205–210

    Article  CAS  PubMed  Google Scholar 

  29. Foroni L, Coyle LA, Papaioannou M, Yaxley JC, Cole Sinclai MF, Chim JS, Cannel P, Secker-Walker LM, Mehta AB, Prentice HG, Hoffbrand AV . Molecular detection of minimal residual disease in adult and childhood acute lymphoblastic leukemia reveals differences in treatment response Leukemia 1997 11: 1732–1741

    Article  CAS  PubMed  Google Scholar 

  30. Sykes PJ, Neoh SH, Brisco MJ, Hughes E, Condon J, Morley AA . Quantitation of targets for PCR by use of limiting dilution BioTechniques 1992 13: 444–449

    CAS  PubMed  Google Scholar 

  31. Mayer SP, Giamelii J, Sandoval C, Roach AS, Ozkaynak MF, Tugal O, Rovera G, Jayabose S . Quantitation of leukemia clone-specific antigen gene rearrangements by a single-step PCR and fluorescence-based detection method Leukemia 1999 13: 1843–1852

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by Deutsche Forschungsgemeinschaft Grant KN 422/1-1.

Author information

Authors and Affiliations

  1. II. Medizinische Klinik der Christian-Albrechts-Universität zu Kiel, Germany

    M Brüggemann, J Droese, I Bolz, P Lüth, C Pott, N von Neuhoff & M Kneba

  2. III. Medizinische Klinik der Johann Wolfgang Goethe-Universität Frankfurt am Main, Germany

    U Scheuering

Authors
  1. M Brüggemann
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. J Droese
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. I Bolz
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. P Lüth
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. C Pott
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. N von Neuhoff
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. U Scheuering
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. M Kneba
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Brüggemann, M., Droese, J., Bolz, I. et al. Improved assessment of minimal residual disease in B cell malignancies using fluorogenic consensus probes for real-time quantitative PCR. Leukemia 14, 1419–1425 (2000). https://doi.org/10.1038/sj.leu.2401831

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.leu.2401831

Keywords

This article is cited by

Search

Advanced search

Quick links