Mutations in the tyrosine kinase domain of FLT3 define a new molecular mechanism of acquired drug resistance to PTK inhibitors in FLT3-ITD-transformed hematopoietic cells

Ksenia Bagrintseva; Ruth Schwab; Tobias M Kohl; Susanne Schnittger; Sabine Eichenlaub; Joachim W Ellwart; Wolfgang Hiddemann; Karsten Spiekermann

doi:10.1182/blood-2003-05-1653

Mutations in the tyrosine kinase domain of FLT3 define a new molecular mechanism of acquired drug resistance to PTK inhibitors in FLT3-ITD-transformed hematopoietic cells

Blood. 2004 Mar 15;103(6):2266-75. doi: 10.1182/blood-2003-05-1653. Epub 2003 Nov 6.

Authors

Ksenia Bagrintseva¹, Ruth Schwab, Tobias M Kohl, Susanne Schnittger, Sabine Eichenlaub, Joachim W Ellwart, Wolfgang Hiddemann, Karsten Spiekermann

Affiliation

¹ Department of Medicine III, University Hospital Grosshadern, Luwig-Maximilians University, Munich, Germany.

PMID: 14604974
DOI: 10.1182/blood-2003-05-1653

Abstract

Activating mutations in the juxtamembrane domain (FLT3-length mutations, FLT3-LM) and in the protein tyrosine kinase domain (TKD) of FLT3 (FLT3-TKD) represent the most frequent genetic alterations in acute myeloid leukemia (AML) and define a molecular target for therapeutic interventions by protein tyrosine kinase (PTK) inhibitors. We could show that distinct activating FLT3-TKD mutations at position D835 mediate primary resistance to FLT3 PTK inhibitors in FLT3-transformed cell lines. In the presence of increasing concentrations of the FLT3 PTK inhibitor SU5614, we generated inhibitor resistant Ba/F3 FLT3-internal tandem duplication (ITD) cell lines (Ba/F3 FLT3-ITD-R1-R4) that were characterized by a 7- to 26-fold higher IC50 (concentration that inhibits 50%) to SU5614 compared with the parental ITD cells. The molecular characterization of ITD-R1-4 cells demonstrated that specific TKD mutations (D835N and Y842H) on the ITD background were acquired during selection with SU5614. Introduction of these dual ITD-TKD, but not single D835N or Y842H FLT3 mutants, in Ba/F3 cells restored the FLT3 inhibitor resistant phenotype. Our data show that preexisting or acquired mutations in the PTK domain of FLT3 can induce drug resistance to FLT3 PTK inhibitors in vitro. These findings provide a molecular basis for the evaluation of clinical resistance to FLT3 PTK inhibitors in patients with AML.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Acute Disease
Animals
Antimetabolites, Antineoplastic / pharmacology
Antineoplastic Agents / pharmacology
Apoptosis / drug effects
Apoptosis / physiology
Cell Division / drug effects
Cell Division / physiology
Cell Line, Transformed / drug effects
Cell Line, Transformed / physiology
Cytarabine / pharmacology
DNA-Binding Proteins / metabolism
Drug Resistance, Neoplasm
Enzyme Inhibitors / pharmacology*
Genistein / pharmacology
Humans
Indoles / pharmacology*
Leukemia, Myeloid*
MAP Kinase Signaling System / physiology
Milk Proteins*
Mutagenesis
Phenotype
Phosphorylation / drug effects
Protein Structure, Tertiary
Proto-Oncogene Proteins / chemistry
Proto-Oncogene Proteins / genetics*
Proto-Oncogene Proteins / metabolism*
Receptor Protein-Tyrosine Kinases / chemistry
Receptor Protein-Tyrosine Kinases / genetics*
Receptor Protein-Tyrosine Kinases / metabolism*
STAT5 Transcription Factor
Staurosporine / analogs & derivatives*
Staurosporine / pharmacology
Trans-Activators / metabolism
Tyrphostins / pharmacology
fms-Like Tyrosine Kinase 3

Substances

6,7-dimethoxy-2-phenylquinoxaline
Antimetabolites, Antineoplastic
Antineoplastic Agents
DNA-Binding Proteins
Enzyme Inhibitors
Indoles
Milk Proteins
Proto-Oncogene Proteins
STAT5 Transcription Factor
SU 5614
Trans-Activators
Tyrphostins
Cytarabine
Genistein
FLT3 protein, human
Receptor Protein-Tyrosine Kinases
fms-Like Tyrosine Kinase 3
Staurosporine
midostaurin