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.

Biotechnical Methods Section (BTS)

Real-time quantitative PCR detection of WT1 gene expression in children with AML: prognostic significance, correlation with disease status and residual disease detection by flow cytometry

Leukemia volume 16, pages 1381–1389 (2002)Cite this article

Abstract

The clinical significance of WT1 gene expression at diagnosis and during therapy of AML has not yet been resolved. We analysed WT1 expression at presentation in an unselected group of 47 childhood AML patients using real-time quantitative reverse-transcription PCR. We also showed that within the first 30 h following aspiration RQ-RT-PCR results were not influenced by transportation time. We observed lower levels of WT1 transcript in AML M5 (P = 0.0015); no association was found between expression levels and sex, initial leukocyte count and karyotype-based prognostic groups. There was significant correlation between very low WT1 expression at presentation and excellent outcome (EFS P = 0.0014). Combined analysis of WT1 levels, three-colour flow cytometry residual disease detection and the course of the disease in 222 samples from 28 children with AML showed remarkable correlation. Fourteen patients expressed high WT1 levels at presentation. In eight of them, who suffered relapse or did not reach complete remission, dynamics of WT1 levels clearly correlated with the disease status and residual disease by flow cytometry. We conclude that very low WT1 levels at presentation represent a good prognostic factor and that RQ-RT-PCR-based analysis of WT1 expression is a promising and rapid approach for monitoring of MRD in approximately half of paediatric AML patients.

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

Access options

Rent or buy this article

Get just this article for as long as you need it

$39.95

Learn more

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

Figure 1
Figure 2
Figure 3

References

  1. Lee SB, Haber DA . Wilms tumor and the WT1 gene Exp Cell Res 2001 264: 74–99

    Article  CAS  PubMed  Google Scholar 

  2. Sekiya M, Adachi M, Hinoda Y, Imai K, Yachi A . Downregulation of Wilms’ tumor gene (wt1) during myelomonocytic differentiation in HL60 cells Blood 1994 83: 1876–1882

    CAS  PubMed  Google Scholar 

  3. Baird PN, Simmons PJ . Expression of the Wilms’ tumor gene (WT1) in normal hemopoiesis Exp Hematol 1997 25: 312–320

    CAS  PubMed  Google Scholar 

  4. Maurer U, Brieger J, Weidmann E, Mitrou PS, Hoelzer D, Bergmann L . The Wilms’ tumor gene is expressed in a subset of CD34+ progenitors and downregulated early in the course of differentiation in vitro Exp Hematol 1997 25: 945–950

    CAS  PubMed  Google Scholar 

  5. Menssen HD, Renkl HJ, Entezami M, Thiel E . Wilms’ tumor gene expression in human CD34+ hematopoietic progenitors during fetal development and early clonogenic growth Blood 1997 89: 3486–3487

    CAS  PubMed  Google Scholar 

  6. Inoue K, Tamaki H, Ogawa H, Oka Y, Soma T, Tatekawa T, Oji Y, Tsuboi A, Kim EH, Kawakami M, Akiyama T, Kishimoto T, Sugiyama H . Wilms’ tumor gene (WT1) competes with differentiation-inducing signal in hematopoietic progenitor cells Blood 1998 91: 2969–2976

    CAS  PubMed  Google Scholar 

  7. Svedberg H, Chylicki K, Baldetorp B, Rauscher FJ, Gullberg U . Constitutive expression of the Wilms’ tumor gene (WT1) in the leukemic cell line U937 blocks parts of the differentiation program Oncogene 1998 16: 925–932

    Article  CAS  PubMed  Google Scholar 

  8. Deuel TF, Guan LS, Wang ZY . Wilms’ tumor gene product WT1 arrests macrophage differentiation of HL-60 cells through its zinc-finger domain Biochem Biophys Res Commun 1999 254: 192–196

    Article  CAS  PubMed  Google Scholar 

  9. Smith SI, Weil D, Johnson GR, Boyd AW, Li CL . Expression of the Wilms’ tumor suppressor gene, WT1, is upregulated by leukemia inhibitory factor and induces monocytic differentiation in M1 leukemic cells Blood 1998 91: 764–773

    CAS  PubMed  Google Scholar 

  10. Ellisen LW, Carlesso N, Cheng T, Scadden DT, Haber DA . The Wilms tumor suppressor WT1 directs stage-specific quiescence and differentiation of human hematopoietic progenitor cells EMBO J 2001 20: 1897–1909

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Miwa H, Beran M, Saunders GF . Expression of the Wilms’ tumor gene (WT1) in human leukemias Leukemia 1992 6: 405–409

    CAS  PubMed  Google Scholar 

  12. Miyagi T, Ahuja H, Kubota T, Kubonishi I, Koeffler HP, Miyoshi I . Expression of the candidate Wilm's tumor gene, WT1, in human leukemia cells Leukemia 1993 7: 970–977

    CAS  PubMed  Google Scholar 

  13. Brieger J, Weidmann E, Fenchel K, Mitrou PS, Hoelzer D, Bergmann L . The expression of the Wilms’ tumor gene in acute myelocytic leukemias as a possible marker for leukemic blast cells Leukemia 1994 8: 2138–2143

    CAS  PubMed  Google Scholar 

  14. Menssen HD, Renkl HJ, Rodeck U, Maurer J, Notter M, Schwartz S, Reinhardt R, Thiel E . Presence of Wilms’ tumor gene (wt1) transcripts and the WT1 nuclear protein in the majority of human acute leukemias Leukemia 1995 9: 1060–1067

    CAS  PubMed  Google Scholar 

  15. Inoue K, Sugiyama H, Ogawa H, Nakagawa M, Yamagami T, Miwa H, Kita K, Hiraoka A, Masaoka T, Nasu K et al. WT1 as a new prognostic factor and a new marker for the detection of minimal residual disease in acute leukemia Blood 1994 84: 3071–3079

    CAS  PubMed  Google Scholar 

  16. Inoue K, Ogawa H, Yamagami T, Soma T, Tani Y, Tatekawa T, Oji Y, Tamaki H, Kyo T, Dohy H, Hiraoka A, Masaoka T, Kishimoto T, Sugiyama H . Long-term follow-up of minimal residual disease in leukemia patients by monitoring WT1 (Wilms tumor gene) expression levels Blood 1996 88: 2267–2278

    CAS  PubMed  Google Scholar 

  17. Gaiger A, Linnerth B, Mann G, Schmid D, Heinze G, Tisljar K, Haas OA, Gadner H, Lion T . Wilms’ tumour gene (wt1) expression at diagnosis has no prognostic relevance in childhood acute lymphoblastic leukaemia treated by an intensive chemotherapy protocol Eur J Haematol 1999 63: 86–93

    Article  CAS  PubMed  Google Scholar 

  18. Gaiger A, Schmid D, Heinze G, Linnerth B, Greinix H, Kalhs P, Tisljar K, Priglinger S, Laczika K, Mitterbauer M, Novak M, Mitterbauer G, Mannhalter C, Haas OA, Lechner K, Jager U . Detection of the WT1 transcript by RT-PCR in complete remission has no prognostic relevance in de novo acute myeloid leukemia Leukemia 1998 12: 1886–1894

    Article  CAS  PubMed  Google Scholar 

  19. Elmaagacli AH, Beelen DW, Trenschel R, Schaefer UW . The detection of wt-1 transcripts is not associated with an increased leukemic relapse rate in patients with acute leukemia after allogeneic bone marrow or peripheral blood stem cell transplantation Bone Marrow Transplant 2000 25: 91–96

    Article  CAS  PubMed  Google Scholar 

  20. Brieger J, Weidmann E, Maurer U, Hoelzer D, Mitrou PS, Bergmann L . The Wilms’ tumor gene is frequently expressed in acute myeloblastic leukemias and may provide a marker for residual blast cells detectable by PCR Ann Oncol 1995 6: 811–816

    Article  CAS  PubMed  Google Scholar 

  21. Bergmann L, Miething C, Maurer U, Brieger J, Karakas T, Weidmann E, Hoelzer D . High levels of Wilms’ tumor gene (wt1) mRNA in acute myeloid leukemias are associated with a worse long-term outcome Blood 1997 90: 1217–1225

    CAS  PubMed  Google Scholar 

  22. Kreuzer KA, Saborowski A, Lupberger J, Appelt C, Na IK, le Coutre P, Schmidt CA . Fluorescent 5′-exonuclease assay for the absolute quantification of Wilms’ tumour gene (WT1) mRNA: implications for monitoring human leukaemias Br J Haematol 2001 114: 313–318

    Article  CAS  PubMed  Google Scholar 

  23. Schmid D, Heinze G, Linnerth B, Tisljar K, Kusec R, Geissler K, Sillaber C, Laczika K, Mitterbauer M, Zochbauer S, Mannhalter C, Haas OA, Lechner K, Jager U, Gaiger A . Prognostic significance of WT1 gene expression at diagnosis in adult de novo acute myeloid leukemia Leukemia 1997 11: 639–643

    Article  CAS  PubMed  Google Scholar 

  24. Neale GA, Coustan-Smith E, Pan Q, Chen X, Gruhn B, Stow P, Behm FG, Pui CH, Campana D . Tandem application of flow cytometry and polymerase chain reaction for comprehensive detection of minimal residual disease in childhood acute lymphoblastic leukemia Leukemia 1999 13: 1221–1226

    Article  CAS  PubMed  Google Scholar 

  25. van Dongen JJ, Seriu T, Panzer-Grumayer 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 leukaemia in childhood Lancet 1998 352: 1731–1738

    Article  CAS  PubMed  Google Scholar 

  26. San Miguel JF, Martinez A, Macedo A, Vidriales MB, Lopez-Berges C, Gonzalez M, Caballero D, Garcia-Marcos MA, Ramos F, Fernandez-Calvo J, Calmuntia MJ, Diaz-Mediavilla J, Orfao A . Immunophenotyping investigation of minimal residual disease is a useful approach for predicting relapse in acute myeloid leukemia patients Blood 1997 90: 2465–2470

    CAS  PubMed  Google Scholar 

  27. San Miguel JF, Vidriales MB, Lopez-Berges C, Diaz-Mediavilla J, Gutierrez N, Canizo C, Ramos F, Calmuntia MJ, Perez JJ, Gonzalez M, Orfao A . Early immunophenotypical evaluation of minimal residual disease in acute myeloid leukemia identifies different patient risk groups and may contribute to postinduction treatment stratification Blood 2001 98: 1746–1751

    Article  CAS  PubMed  Google Scholar 

  28. Maurer U, Weidmann E, Karakas T, Hoelzer D, Bergmann L . Wilms tumor gene (wt1) mRNA is equally expressed in blast cells from acute myeloid leukemia and normal CD34+ progenitors Blood 1997 90: 4230–4232

    CAS  PubMed  Google Scholar 

  29. Fraizer GC, Patmasiriwat P, Zhang X, Saunders GF . Expression of the tumor suppressor gene WT1 in both human and mouse bone marrow Blood 1995 86: 4704–4706

    CAS  PubMed  Google Scholar 

  30. Zuna J, Hrusak O, Kalinova M, Muzikova K, Stary J, Trka J . TEL/AML1 positivity in childhood ALL: average or better prognosis? Czech Paediatric Haematology Working Group Leukemia 1999 13: 22–24

    Article  CAS  PubMed  Google Scholar 

  31. Chomczynski P, Sacchi N . Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction Anal Biochem 1987 162: 156–159

    Article  CAS  PubMed  Google Scholar 

  32. Pallisgaard N, Clausen N, Schroder H, Hokland P . Rapid and sensitive minimal residual disease detection in acute leukemia by quantitative real-time RT-PCR exemplified by t(12;21) TEL- AML1 fusion transcript Genes Chromosomes Cancer 1999 26: 355–365

    Article  CAS  PubMed  Google Scholar 

  33. Grimwade D, Walker H, Oliver F, Wheatley K, Harrison C, Harrison G, Rees J, Hann I, Stevens R, Burnett A, Goldstone A . The importance of diagnostic cytogenetics on outcome in AML: analysis of 1612 patients entered into the MRC AML 10 trial. The Medical Research Council Adult and Children's Leukaemia Working Parties Blood 1998 92: 2322–2333

    CAS  PubMed  Google Scholar 

  34. Lion T . Current recommendations for positive controls in RT-PCR assays Leukemia 2001 15: 1033–1037

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

Supported by grants from Grant Agency of the Czech Republic No. 312/P304, from the Czech Ministry of Health No. 6406–3 and 6512 and the Czech Ministry of Education No. 111300001, 111300003 and 111300005. We also acknowledge Dr E Coustan-Smith for critcial reading of the manuscript.

Author information

Authors and Affiliations

  1. CLIP – Childhood Leukaemia Investigation Prague, 2nd Medical School, Charles University, Prague, Czech Republic

    J Trka, M Kalinová, O Hrušák, J Zuna, O Krejčí & J Madžo

  2. 2 nd Department of Paediatrics, 2nd Medical School, Charles University, Prague, Czech Republic

    J Trka, M Kalinová, J Zuna, O Krejčí, J Madžo, P Sedláček, V Vávra & J Starý

  3. Institute of Immunology, 2nd Medical School, Charles University, Prague, Czech Republic

    O Hrušák

  4. 3 rd Department of Internal Medicine, 1st Medical School, Charles University, Prague, Czech Republic

    K Michalová

  5. Department of Haemato-oncology, Medical School, Palacky University, Olomouc, Czech Republic

    M Jarošová

Authors
  1. J Trka
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. M Kalinová
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. O Hrušák
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. J Zuna
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. O Krejčí
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. J Madžo
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. P Sedláček
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. V Vávra
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. K Michalová
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. M Jarošová
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. J Starý
    View author publications

    You can also search for this author in PubMed Google Scholar

Consortia

for Czech Paediatric Haematology Working Group

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Trka, J., Kalinová, M., Hrušák, O. et al. Real-time quantitative PCR detection of WT1 gene expression in children with AML: prognostic significance, correlation with disease status and residual disease detection by flow cytometry. Leukemia 16, 1381–1389 (2002). https://doi.org/10.1038/sj.leu.2402512

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.leu.2402512

Keywords

  • WT1
  • childhood AML
  • minimal residual disease
  • RQ-PCR
  • flow cytometry

This article is cited by

Search

Advanced search

Quick links