Mechanisms of resistance against PKC412 in resistant FLT3-ITD positive human acute myeloid leukemia cells.
Treatment of acute myeloid leukemia (AML) remains challenging with many patients harboring unfavorable prognostic parameters such as FLT3 internal tandem duplication (FLT3-ITD) mutations leading to a constitutively activated FLT3-receptor tyrosine kinase (RTK). Activation of proteins by phosphorylation of tyrosine residues is a common mechanism in leukemia development. Therefore, specific tyrosine kinase inhibitors (TKI) have been developed for AML therapy and are currently under investigation. The staurosporine derivate PKC412 (Midostaurin) was found to be an effective inhibitor of the FLT3-RTK and is currently undergoing clinical trials for FLT3-mutated AML patients. Since resistance towards TKIs has been observed in vitro and in clinical trials, we have generated a PKC412-resistant clone (MV4-11r) of the human myelomonoblastic cell line MV4-11, which carries a homozygous FLT3-ITD mutation. MV4-11r displayed higher vitality after addition of PKC412 compared with MV4-11 with a pronounced reduction of apoptotic cells. Cytogenetic characterization revealed the acquisition of additional aberrations in the resistant cell line such as clonal alterations at chromosome 13q with additional FLT3 signals. Microarray analysis revealed significant expression changes in several genes prior to and after incubation with PKC412. The expression status of candidate genes being regulated by FLT-ITD like JAG1, p53, MCL-1, C-KIT, and FLT3/-L was confirmed by real-time PCR. In summary, resistance against PKC412 appears to be mediated by up-regulation of anti-apoptotic genes and down-regulation of proapoptotic signals as well as genes that are involved in normal and malignant hematopoiesis.
Stölzel
Friedrich
F
Medizinische Klinik und Poliklinik I, Universitätsklinikum Carl Gustav Carus Dresden der TU Dresden, Fetscherstr. 74, 01307 Dresden, Germany. friedrich.stoelzel@uniklinikum-dresden.de
Steudel
Christine
C
Oelschlägel
Uta
U
Mohr
Brigitte
B
Koch
Sina
S
Ehninger
Gerhard
G
Thiede
Christian
C
D045744:Cell Line, Tumor; D002882:Chromosomes, Human, Pair 13 / Q000235:genetics / Q000378:metabolism; D002986:Clinical Trials as Topic; D019008:Drug Resistance, Neoplasm / Q000187:drug effects* / Q000235:genetics; D020869:Gene Expression Profiling; D015973:Gene Expression Regulation, Leukemic / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000175:diagnosis / Q000188:drug therapy / Q000201:enzymology* / Q000235:genetics; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D011379:Prognosis; D047428:Protein Kinase Inhibitors / Q000031:analogs & derivatives / Q000494:pharmacology*; D019311:Staurosporine / Q000031:analogs & derivatives* / Q000494:pharmacology; D051941:fms-Like Tyrosine Kinase 3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
D047428:Protein Kinase Inhibitors; C497970:FLT3 protein, human; D051941:fms-Like Tyrosine Kinase 3; D019311:Staurosporine; C059539:midostaurin