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Minimal Residual Disease

Standardization of WT1 mRNA quantitation for minimal residual disease monitoring in childhood AML and implications of WT1 gene mutations: a European multicenter study

Leukemia volume 23, pages 1472–1479 (2009)Cite this article

Abstract

A standardized, sensitive and universal method for minimal residual disease (MRD) detection in acute myeloid leukemia (AML) is still pending. Although hyperexpression of Wilms' tumor (WT1) gene transcript has been frequently proposed as an MRD marker in AML, wide comparability of the various methods used for evaluating WT1 expression has not been given. We established and standardized a multicenter approach for quantifying WT1 expression by quantitative reverse transcriptase PCR (qRT-PCR), on the basis of a primer/probe set combination at exons 6 and 7. In a series of quality-control rounds, we analyzed 69 childhood AML samples and 47 normal bone marrow (BM) samples from 4 participating centers. Differences in the individual WT1 expressions levels ranged within <0.5 log of the mean in 82% of the cases. In AML samples, the median WT1/1E+04 Abelson (ABL) expression was 3.5E+03 compared with that of 2.3E+01 in healthy BM samples. As 11.5% of childhood AML samples in this cohort harbored WT1 mutations in exon 7, the effect of mutations on WT1 expression has been investigated, showing that mutated cases expressed significantly higher WT1 levels than wild-type cases. Hence, our approach showed high reproducibility and applicability, even in patients with WT1 mutations; therefore, it can be widely used for the quantitation of WT1 expression in future clinical trials.

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Acknowledgements

This work was supported by the ‘Deutsche José Carreras Leukämie Stiftung e.V. (DJCLS R04/09)’, Munich, Germany (PB, HK, AMW), by the ‘Research Project of the Czech Ministry of Education (No. 0021620813)’, Prague, Czech Republic (JT, MK), by the ‘Fondazione Tettamanti, Fondazione Cariplo and AIRC’, Italy (AB, GC) and by the ‘KOCR Foundation’, Rotterdam, The Netherlands (IHIMH, CMZ). We thank Vida Meyer (Frankfurt), Nadine Pfaffendorf (Jena), Vincenzo Rossi (Monza) and Patricia Hoogeveen (Rotterdam) for their excellent technical assistance.

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Author notes

  1. G Cazzaniga, P Bader and J Trka: These authors contributed equally to the manuscript

Authors and Affiliations

  1. Department of Pediatric Hematology, Oncology and Hemostaseology, Goethe University Frankfurt, Hospital for Children and Adolescents III, Frankfurt am Main, Germany

    A M Willasch, H Kreyenberg, G Weber & P Bader

  2. Department of Pediatric Hematology and Oncology, Schiller University Jena, University Children's Hospital, Jena, Germany

    B Gruhn

  3. Department of Pediatric Hematology and Oncology, Universitá Milano, Bicocca Clinica Pediatrica, Ospedale San Gerardo, Monza, Italy

    T Coliva, A Biondi & G Cazzaniga

  4. Department of Pediatric Hematology and Oncology, Charles University and University Children's Hospital, Motol, Prague, Czech Republic

    M Kalinova & J Trka

  5. Department of Computer Science and Mathematics, Goethe University Frankfurt, Frankfurt am Main, Germany

    G Schneider

  6. Department of Pediatric Hematology and Oncology, University Ulm, University Children's Hospital, Ulm, Germany

    D Steinbach

  7. Department of Pediatric Hematology and Oncology, Erasmus MC, Sophia Children's Hospital, Rotterdam, The Netherlands

    I H I M Hollink & C M Zwaan

  8. Department of Immunology, Erasmus MC, Rotterdam, The Netherlands

    V H J van der Velden

  9. Department of Pediatric Oncology and Hematology, AML-BFM Study Group, Medical High School, Hannover, Germany

    D Reinhardt

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on behalf of the European Study Group on WT1 Expression in Childhood AML. Frankfurt, Jena (Germany), Monza (Italy), Prague (Czech Republic)

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Correspondence to A M Willasch.

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Willasch, A., Gruhn, B., Coliva, T. et al. Standardization of WT1 mRNA quantitation for minimal residual disease monitoring in childhood AML and implications of WT1 gene mutations: a European multicenter study. Leukemia 23, 1472–1479 (2009). https://doi.org/10.1038/leu.2009.51

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