FLT3-TKD mutations are not inhibited at therapeutic concentrations. resistance in AML. Personalized treatment methods that address additional molecular focuses on beyond FLT3 could conquer resistance and facilitate molecular responses in AML. strong class=”kwd-title” Keywords: acute myeloid leukemia, AML, FMS-like tyrosine kinase 3, FLT3, FLT3-ITD, FLT3-TKD, resistance, midostaurin, quizartinib, gilteritinib, crenolanib 1. Introduction Activating FMS-like tyrosine kinase 3 (FLT3) mutations are detected in about one-third of patients with acute myeloid leukemia (AML) at diagnosis. The majority of these mutations are FLT3-internal tandem duplications (ITD) [1,2]. Approximately 25% of patients with AML are diagnosed as FLT3-ITD-mutated. The prognostic impact of FLT3-ITD depends on the allelic ratio (ITD/wildtype allele) and co-occurrence of a nucleophosmin-1 (NPM1) mutation. However, the European Leukemia Net (ELN) and National Comprehensive Malignancy Network (NCCN) guidelines use different methods for prognostic stratification of newly diagnosed patients with AML and FLT3-ITD mutations [3,4]. In general, the presence of co-occurring mutations has substantial effects on prognosis in AML [5]. Besides activating FLT3-ITD mutations, FLT3 tyrosine kinase domain name (TKD) mutations are detected in 7C11% of patients with AML [6,7]. TKD mutations predominantly occur at codons D835 or I836 and can be associated with main resistance to FLT3 inhibitors [8,9]. In contrast to the impact of FLT3-ITD, data concerning the prognostic significance of FLT3-TKD mutations controversially discussed and depend around the co-occurrence of other mutations, e.g., FLT3-TKD and NPM1 mutations [10,11]. FLT3-ITD mutations can be associated with adverse prognosis, particularly, in patients with a high allelic ratio of 0.5. Furthermore, FLT3-ITD mutations increase the relapse risk following rigorous induction chemotherapy, although most clinical trials failed to demonstrate impact of FLT3-ITD mutations on total remission rates following induction therapy. Patients with FLT3-ITD-positive AML have shown a higher probability of relapse following standard chemotherapy and allogeneic stem cell transplantation (ASCT) [12]. Even though prognostic benefit of ASCT for FLT3-ITD-positive AML is usually widely accepted, deeper insight regarding the underlying molecular mechanisms are clearly warranted. Aspects concerning depth of remission before ASCT are currently being evaluated in Tmem32 ongoing clinical trials with 2nd generation FLT3 inhibitors such as quizartinib or gilteritinib [13,14]. 2. FLT3-ITD Signaling Pathways and Diversity of ITD Mutations FLT3-ITD and FLT3-TKD mutations lead to constitutive activation of multiple downstream signaling pathways and this results in increased proliferation, reduced susceptibility to apoptosis, and inhibition of myeloid differentiation [15,16]. Importantly, activation patterns of downstream signaling are dependent on the presence of either FLT3-ITD or FLT3-TKD mutations and result in unique AML phenotypes [17,18]. ITD-mediated FLT3-activation is usually caused by destabilization of its autoinhibitory juxtamembrane (JM) domain name and results in constitutive activation of STAT, MAPK-ERK, and BPTES PI3K signaling [19,20]. Phosphorylation of STAT family members is an essential step that depends on activation of SRC kinase. Of notice, this signaling pathway is not observed downstream of FLT3-TKD mutations or ligand-activated FLT3-wildtype receptors. STAT5 activation has pleiotropic effects on cellular transformation. This includes the activation of the serine/threonine kinase PIM-1, which is responsible for stabilization of the 130 kDa FLT3-ITD variant and accelerates STAT5 activation [21]. Furthermore, PIM-1 can increase survival or mediate resistance to FLT3 inhibitors by activating the anti-apoptotic protein MCL-1 [22,23]. Interestingly, activation of MCL-1 is also a downstream effect of STAT5 signaling that contributes to the maintenance of FLT3-ITD-positive leukemic stem cells [24]. Impaired myeloid differentiation in FLT3-ITD-positive AML blasts is usually a consequence of deregulated gene expression and is induced by multiple signaling pathways. FLT3-ITD mutations lead to downregulation of myeloid transcription factors, such as PU.1 or CEBPA [25]; the latter has been shown to be phosphorylated and inhibited by FLT3-ITD-signaling [26]. Additionally, RGS2, an important regulator of myeloid differentiation, is usually repressed by FLT3-ITD [27]. In contrast, high expression of RUNX1 has been recognized in FLT3-ITD-positive AML cells, which may contribute to the development of AML and blockade of differentiation in FLT3-ITD blasts.It is tempting to speculate whether the combination of venetoclax with FLT3-TKI (and particularly next-generation inhibitors) may be a promising strategy for future clinical development. myeloid leukemia (AML) at diagnosis. The majority of these mutations are FLT3-internal tandem duplications (ITD) [1,2]. Approximately 25% of patients with AML are diagnosed as FLT3-ITD-mutated. The prognostic impact of FLT3-ITD depends on the allelic ratio (ITD/wildtype allele) and co-occurrence of a nucleophosmin-1 (NPM1) mutation. However, the European Leukemia Net (ELN) and National Comprehensive Malignancy Network (NCCN) guidelines use different methods for prognostic stratification of newly diagnosed patients with AML and FLT3-ITD mutations [3,4]. In general, the presence of co-occurring mutations has substantial effects on prognosis in AML [5]. Besides activating FLT3-ITD mutations, FLT3 tyrosine kinase domain name (TKD) mutations are detected in 7C11% of patients with AML [6,7]. BPTES TKD mutations predominantly occur at codons D835 or I836 and can be associated with main resistance to FLT3 inhibitors [8,9]. In contrast to the impact of FLT3-ITD, data concerning the prognostic significance of FLT3-TKD mutations controversially discussed and depend around the co-occurrence of other mutations, e.g., FLT3-TKD and NPM1 mutations [10,11]. FLT3-ITD mutations can be associated with adverse prognosis, particularly, in patients with a high allelic ratio of 0.5. Furthermore, FLT3-ITD mutations increase the relapse risk following rigorous induction chemotherapy, although most clinical trials failed to demonstrate impact of FLT3-ITD mutations on total remission rates following induction therapy. Patients with FLT3-ITD-positive AML have shown a higher probability of relapse following standard chemotherapy and allogeneic stem cell transplantation (ASCT) [12]. Even though prognostic benefit of ASCT for FLT3-ITD-positive AML is usually widely accepted, deeper insight regarding the underlying molecular mechanisms are clearly warranted. Aspects concerning depth of remission before ASCT are currently being evaluated in ongoing clinical trials with 2nd generation FLT3 inhibitors such as quizartinib or gilteritinib [13,14]. 2. FLT3-ITD Signaling Pathways and Diversity of ITD Mutations FLT3-ITD and FLT3-TKD mutations lead to constitutive activation of multiple downstream signaling pathways and this results in increased proliferation, reduced susceptibility to apoptosis, and inhibition of myeloid differentiation [15,16]. Importantly, activation patterns of downstream signaling are dependent on the presence of either FLT3-ITD or FLT3-TKD mutations and result in unique AML phenotypes [17,18]. ITD-mediated FLT3-activation is usually caused by destabilization of its autoinhibitory juxtamembrane (JM) domain name and results in constitutive activation of STAT, MAPK-ERK, and PI3K signaling [19,20]. Phosphorylation of STAT family members is an essential step that depends on activation of SRC kinase. Of notice, this signaling pathway is not observed downstream of FLT3-TKD mutations or ligand-activated FLT3-wildtype receptors. STAT5 activation has pleiotropic effects on cellular transformation. This includes the activation of the serine/threonine kinase PIM-1, which is responsible for stabilization of the 130 kDa FLT3-ITD variant and accelerates STAT5 activation [21]. Furthermore, PIM-1 can increase survival or mediate resistance to FLT3 inhibitors by activating the anti-apoptotic protein MCL-1 [22,23]. Interestingly, activation of MCL-1 is also a downstream effect of STAT5 signaling that contributes to the maintenance of FLT3-ITD-positive leukemic stem cells [24]. Impaired myeloid differentiation in FLT3-ITD-positive AML blasts is usually a consequence of deregulated gene expression and is induced by BPTES multiple signaling pathways. FLT3-ITD mutations lead to downregulation of myeloid transcription factors, such as PU.1 or CEBPA [25]; the latter has been shown to be phosphorylated and inhibited by FLT3-ITD-signaling [26]. Additionally, RGS2, an important regulator of myeloid differentiation, is usually repressed by FLT3-ITD [27]. In contrast, high expression of RUNX1 has been recognized in FLT3-ITD-positive AML cells, which may contribute to the development of AML and blockade of differentiation in FLT3-ITD blasts [28]. Thus, profound changes in transcriptional programs may block differentiation in the presence of FLT3-ITD mutations. Maturation of FLT3-ITD receptor represents a complex process of post-translational modifications that includes multiple actions of glycosylation occurring in the Golgi apparatus (GA) and endoplasmic reticulum (ER). Importantly, the majority of FLT3-ITD molecules can be detected in the ER of AML cells, while a relatively small amount of FLT3-ITD protein is located at the cell membrane [29]. The hypoglycosylated 130 kDa species of FLT3-ITD that is retained in the ER can.