A Novel Tandem Duplication Assay to Detect Minimal Residual Disease in FLT3 /ITD AML
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ORIGINAL RESEARCH ARTICLE
A Novel Tandem Duplication Assay to Detect Minimal Residual Disease in FLT3/ITD AML Ming-Tseh Lin1 • Li-Hui Tseng1,2 • Jonathan C. Dudley1,3 • Stacey Riel1 Harrison Tsai1 • Gang Zheng1 • Keith W. Pratz4 • Mark J. Levis4 • Christopher D. Gocke1,4
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Ó Springer International Publishing Switzerland 2015
Abstract Background Internal tandem duplication (ITD) of the fms-related tyrosine kinase 3 (FLT3) gene is associated with a poor prognosis in acute myeloid leukemia (AML) patients with a normal karyotype. The current standard polymerase chain reaction (PCR) assay for FLT3/ITD detection is not sufficiently sensitive to monitor minimal residual disease (MRD). Clone-specific assays may have sufficient sensitivity but are not practical to implement, since each clone-specific primer/probe requires clinical validation. Objective To develop an assay for clinical molecular diagnostics laboratories to monitor MRD in FLT3/ITD AMLs. Methods We designed a simple novel assay, tandem duplication PCR (TD-PCR), and tested its sensitivity, specificity, and clinical utility in FLT3/ITD AML patients.
M.-T. Lin and L.-H. Tseng contributed equally to this article.
Electronic supplementary material The online version of this article (doi:10.1007/s40291-015-0170-3) contains supplementary material, which is available to authorized users. & Christopher D. Gocke [email protected] 1
Division of Molecular Pathology, Department of Pathology, Johns Hopkins University School of Medicine, Park SB202, 600 North Wolfe Street, Baltimore, MD 21287, USA
2
Department of Medical Genetics, National Taiwan University Hospital, Taipei, Taiwan
3
Department of Pathology, Massachusetts General Hospital, Boston, MA, USA
4
Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD, USA
Results TD-PCR was capable of detecting a single ITD molecule and was applicable to 75 % of ITD mutants tested. TD-PCR detected MRD in bone marrow prior to patient relapse. TD-PCR also identified low-level ITD mutants not only in FLT3/ITD AMLs but also in initial diagnostic specimens that were reportedly negative by the standard assay in patients who progressed with the same ITDs detected by the TD-PCR assay. Conclusion Detection of MRD by TD-PCR may guide patient selection for early clinical intervention. In contrast to clone-specific approaches, the TD-PCR assay can be more easily validated for MRD detection in clinical laboratories because it uses standardized primers and a universal positive control. In addition, our findings on multiclonality and low-level ITDs suggest that further studies are warranted to elucidate their clinical/biological significance.
Key Points Tandem duplication polymerase chain reaction (TDPCR) is a simple, ultra-sensitive assay to detect minimal residual disease in acute myeloid leukemia patients with fms-related tyrosine kinase 3 internal tandem duplication (FLT3/ITD) mutations. In contrast to assays using clone-specific primers/ probes, TD-PCR is amenable to clinical validation in molecular
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