A Novel Tandem Duplication Assay to Detect Minimal Residual Disease in FLT3/ITD AML

Ming-Tseh Lin, Li Hui Tseng, Jonathan C. Dudley, Stacey Riel, Harrison Tsai, Gang Zheng, Keith Pratz, Mark J Levis, Christopher Gocke

Research output: Contribution to journalArticle

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. 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 multi-clonality and low-level ITDs suggest that further studies are warranted to elucidate their clinical/biological significance.

Original languageEnglish (US)
Pages (from-to)409-417
Number of pages9
JournalMolecular Diagnosis and Therapy
Volume19
Issue number6
DOIs
StatePublished - Dec 1 2015

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Residual Neoplasm
Acute Myeloid Leukemia
Polymerase Chain Reaction
Clone Cells
Molecular Pathology
Karyotype
Patient Selection
Tyrosine
Bone Marrow
Recurrence
Sensitivity and Specificity
Genes

ASJC Scopus subject areas

  • Genetics
  • Molecular Medicine
  • Medicine(all)
  • Pharmacology

Cite this

A Novel Tandem Duplication Assay to Detect Minimal Residual Disease in FLT3/ITD AML. / Lin, Ming-Tseh; Tseng, Li Hui; Dudley, Jonathan C.; Riel, Stacey; Tsai, Harrison; Zheng, Gang; Pratz, Keith; Levis, Mark J; Gocke, Christopher.

In: Molecular Diagnosis and Therapy, Vol. 19, No. 6, 01.12.2015, p. 409-417.

Research output: Contribution to journalArticle

Lin, Ming-Tseh ; Tseng, Li Hui ; Dudley, Jonathan C. ; Riel, Stacey ; Tsai, Harrison ; Zheng, Gang ; Pratz, Keith ; Levis, Mark J ; Gocke, Christopher. / A Novel Tandem Duplication Assay to Detect Minimal Residual Disease in FLT3/ITD AML. In: Molecular Diagnosis and Therapy. 2015 ; Vol. 19, No. 6. pp. 409-417.
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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. 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 multi-clonality and low-level ITDs suggest that further studies are warranted to elucidate their clinical/biological significance.",
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AU - Tsai, Harrison

AU - Zheng, Gang

AU - Pratz, Keith

AU - Levis, Mark J

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N2 - 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. 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 multi-clonality and low-level ITDs suggest that further studies are warranted to elucidate their clinical/biological significance.

AB - 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. 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 multi-clonality and low-level ITDs suggest that further studies are warranted to elucidate their clinical/biological significance.

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