Arginine 595 is duplicated in patients with acute leukemias carrying internal tandem duplications of FLT3 and modulates its transforming potential

Abstract

FLT3-internal tandem duplications (FLT3-ITDs IntroductionMutations in the FMS-like tyrosine-kinase 3 (FLT3) are one of the most frequently found genetic alterations in patients with acute myeloid leukemia (AML), 1-13 myelodysplastic syndromes (MDSs; 10%-15%), 2,14 and acute lymphoblastic leukemia (ALL; 1%-3%). 9,16,17 FLT3 belongs to the class III of receptor tyrosine kinases, which are characterized by the presence of an extracellular immunoglobulin-like domain, a transmembrane and the cytoplasmic juxtamembrane (JM) domain, and the tyrosine kinase domain (TKD). 18 The class III receptors also include KIT, CSF-1, PDGFRA, and PDGFRB. 19,20 Activation of FLT3 by FLT3 ligand (FL) leads to receptor oligomerization and transphosphorylation of specific tyrosine residues, 21 which activates the downstream signaling pathways including STAT5, Ras/mitogen-activated protein kinase (MAPK), and phosphatidylinositol 3-kinase (PI3K)/AKT. [22][23][24][25] FLT3 is highly expressed in CD34 ϩ hematopoietic progenitor cells and plays an important role in normal hematopoiesis. [26][27][28][29] Three distinct activating mutations of FLT3 in hematologic malignancies have been reported: point mutations (FLT3-JM-PM) 30,31 and internal tandem duplications (FLT3-ITD) in the JM domain and mutations in the tyrosine-kinase domain (FLT3-TKD). 1,8,9,12,16,17,32 The crystal structure of FLT3 has shown that the JM domain acts as an autoinhibitory domain in the inactive state. 33 The JM domain is highly conserved across all members of class III RTKs. Hence many tumors in humans show activating mutations of JM in class III RTKs. 34-37 FLT3-ITDs, found in a majority of acute leukemia patients, are in-frame duplications of a fragment of the JM domain. FLT3-ITDs are highly heterogeneous and vary in length from 2 to 68 AAs. These duplications are thought to disrupt the autoinhibitory mechanism and result in constitutive activation of the catalytic domain of FLT3. Activated FLT3 mutants promote cell proliferation and inhibit apoptosis, leading to factorindependent growth of murine hematopoietic cells in vitro and a myeloproliferative phenotype in vivo. 38 FLT3-ITDs are present in the leukemic blasts of 20% to 30% of all AML patients. Recent studies have also shown that FLT3-ITDs are found in the leukemic stem cells. 39 Furthermore, the presence of a FLT3-ITD has been recognized as an independent poor prognostic factor in AML and is associated with a decreased survival due to an increased relapse rate. 8,9,11,12,[40][41][42][43] Several factors influence the poor prognosis seen in AML patients harboring FLT3-ITDs (eg, a high FLT3-ITD/wildtype ratio). 9,44 A recent study has reported that the detection of FLT3-ITD mutation in less mature progenitor populations, for example, CD34 ϩ /CD33 Ϫ , might be associated with drug resistance. 43 In the present study, we asked whether any common duplicated motif exists in AML patients carrying FLT3-ITDs, which might be responsible for the transforming potential. To address this question, we sequenced and analyzed th...

Insertion of FLT3 internal tandem duplication in the tyrosine kinase domain-1 is associated with resistance to chemotherapy and inferior outcome

“…Kiyoi et al hypothesized that the DNA sequence between amino acids 593 and 602 forms a palindromic intermediate that leads to a DNA replication error, thus inducing the tandem duplication. 37 Other investigators showed that 95% of the patients with FLT3-ITD mutations contained at least one amino acid of the tyrosine-rich stretch Y591 to Y599 30 and that the amino acid residue R595 may play an essential role in activation of STAT5. 30 In addition, tyrosine to phenylalanine substitution of residues 589 and 591 abrogates STAT5 activation of FLT3-ITD, and FLT3-ITD-Y589/591F is incapable of inducing a myeloproliferative phenotype.…”

“…37 Other investigators showed that 95% of the patients with FLT3-ITD mutations contained at least one amino acid of the tyrosine-rich stretch Y591 to Y599 30 and that the amino acid residue R595 may play an essential role in activation of STAT5. 30 In addition, tyrosine to phenylalanine substitution of residues 589 and 591 abrogates STAT5 activation of FLT3-ITD, and FLT3-ITD-Y589/591F is incapable of inducing a myeloproliferative phenotype. 31 Interestingly, we showed that in 96.1% of all analyzed FLT3-ITDs at least one amino acid of this motif (Y591-Y599) was affected, and this was independent of ITD length and insertion site.…”

“…Since a recent study highlighted the particular role of the specific tyrosine-rich amino acid stretch Y591 to Y599 (YVDFREYEY) for intracellular signaling, 30 we specifically determined how often this specific amino acid motif was involved in an ITD. Duplication of at least one residue in this specific stretch was seen in 271 (96.1%) of the 282 ITDs.…”

FMS-Like Tyrosine Kinase 3 Internal Tandem Duplication and the Patterns of Its Gene Sequence in 207 Chinese Patients With De Novo Acute Myeloid Leukemia

“…The latter may involve a frameshift and affect transcriptional activity, especially for some critical codons. 23 In summary, our study indicates that FLT3-ITD is a common genetic alteration in Chinese adult patients with AML. Multivariate analysis shows that the ITD to wildtype ratio, the gene patterns, and CD7 expression status appear to be independent prognostic indices for patients with AML.…”

FLT3 inhibitors for acute myeloid leukemia: a review of their efficacy and mechanisms of resistance

“…The most common abnormalities, occurring in approximately 23 % of newly diagnosed de novo AML, are the FLT3/ITD mutations [6]. These mutations consist of in-frame tandem duplications of coding sequence, almost always encompassing arginine residue 595 within the juxtamembrane domain [16]. The juxtamembrane domain of RTKs often has an autoinhibitory function [17].…”