FLT3 and dual Aurora B/FLT3 inhibitors have shown relevance in the search for promising new anticancer compounds, mainly for acute myeloid leukemia (AML). This study was designed to investigate the interactions between human FLT3 in the kinase domain with several indolin-2-one derivatives, structurally similar to Sunitinib. Molegro Virtual Docker (MVD) software was utilized in docking analyses. The predicted model of the training group, considering nineteen amino acid residues, performed in Chemoface, achieved an R2 of 0.82, suggesting that the binding conformations of the ligands with FLT3 are reasonable, and the data can be used to predict the interaction energy of other FLT3 inhibitors with similar molecular patterns. The MolDock Score for energy for compound 1 showed more stable interaction energy (-233.25 kcal mol-1) than the other inhibitors studied, while Sunitinib presented as one of the least stable (-160.94 kcal mol-1). Compounds IAF70, IAF72, IAF75, IAF80, IAF84, and IAF88 can be highlighted as promising derivatives for synthesis and biological evaluation against FLT3. Furthermore, IAF79 can be considered to be a promising dual Aurora B/FLT3 inhibitor, and its molecular pattern can be exploited synthetically to search for new indolin-2-one derivatives that may become drugs used in the treatment of cancers, including AML.

• Klíčová slova
• computational chemistry, dual Aurora B/FLT3 inhibitors, indolin−2-one derivatives,
• MeSH
• aktivace enzymů účinky léků   MeSH
• algoritmy MeSH
• aurora kinasa B antagonisté a inhibitory   chemie   MeSH
• inhibiční koncentrace 50 MeSH
• inhibitory proteinkinas chemie   farmakologie   MeSH
• lidé MeSH
• molekulární konformace MeSH
• molekulární modely MeSH
• molekulární struktura MeSH
• teoretické modely * MeSH
• tyrosinkinasa 3 podobná fms antagonisté a inhibitory   chemie   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• AURKB protein, human MeSH Prohlížeč
• aurora kinasa B MeSH
• FLT3 protein, human MeSH Prohlížeč
• inhibitory proteinkinas MeSH
• tyrosinkinasa 3 podobná fms MeSH

OBJECTIVES: Interaction of leukemia cells with the bone marrow extracellular matrix promotes cell survival and resistance to chemotherapy. In this work, we analyzed integrin expression and adhesivity to fibronectin in primary cells from patients with acute myeloid leukemia. METHODS: Surface expression of integrins β1 and αVβ3 on primary leukemia cells (N = 46) was correlated with the stem cell marker CD34, as well as with cell adhesivity to fibronectin. The results were analyzed with regard to the mutational status of NPM1 and FLT3 genes. RESULTS: The integrin β1 was omnipresent, whereas αVβ3 was often more expressed on CD34-positive cells. In particular, higher αVβ3 expression on CD34+ cells was associated with NPM1 mutation (P = .0018). Monocytic leukemias had significantly higher αVβ3 expression compared to less maturated cases (P = .0008). Cells from patients with internal tandem duplications in FLT3 (FLT3-ITD) had lower adhesivity to fibronectin compared to cells with wild-type FLT3 (P = .031), specifically in less differentiated myeloblasts. Inhibition of a putative FLT3-ITD target, EZH2, increased cell adhesivity in MV4-11 cell line (P = .024). CONCLUSIONS: The integrin αVβ3 is expressed in particular on CD34+ cells with NPM1 mutation and might have a prognostic value in patients with mutated NPM1. FLT3-ITD is associated with lower cell adhesivity, especially in patients with less differentiated leukemias.

• Klíčová slova
• AML, CD34, FLT3-ITD, Fms-like tyrosine kinase, integrin,
• MeSH
• akutní myeloidní leukemie genetika   metabolismus   MeSH
• buněčná adheze MeSH
• buněčná membrána metabolismus   MeSH
• duplikace genu MeSH
• exprese genu MeSH
• fibronektiny metabolismus   MeSH
• integriny genetika   metabolismus   MeSH
• jaderné proteiny genetika   metabolismus   MeSH
• lidé MeSH
• mutace MeSH
• nádorové buněčné linie MeSH
• nukleofosmin MeSH
• tyrosinkinasa 3 podobná fms genetika   metabolismus   MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fibronektiny MeSH
• FLT3 protein, human MeSH Prohlížeč
• integriny MeSH
• jaderné proteiny MeSH
• NPM1 protein, human MeSH Prohlížeč
• nukleofosmin MeSH
• tyrosinkinasa 3 podobná fms MeSH

Acute myeloid leukaemia (AML) is a complex haematological malignancy characterised by diverse genetic alterations leading to abnormal proliferation of myeloid precursor cells. One of the most significant genetic alterations in AML involves mutations in the FLT3 gene, which plays a critical role in haematopoiesis and haematopoietic homeostasis. This review explores the current understanding of FLT3 gene mutations and isoforms and the importance of the FLT3 protein in AML. FLT3 mutations, including internal tandem duplications (FLT3-ITD) and point mutations in the tyrosine kinase domain (FLT3-TKD), occur in 25-30% in AML and are associated with poor prognosis. FLT3-ITD mutations lead to constitutive activation of the FLT3 signalling pathway, promoting cell survival and proliferation. FLT3-TKD mutations affect the tyrosine kinase domain and affect AML prognosis in various ways. Furthermore, FLT3 isoforms, including shorter variants, contribute to the complexity of FLT3 biology. Additionally, nonpathological polymorphisms in FLT3 are being explored for their potential impact on AML prognosis and treatment response. This review also discusses the development of molecular treatments targeting FLT3, including first-generation and next-generation tyrosine kinase inhibitors, highlighting the challenges of resistance that often arise during therapy. The final chapter describes FLT3 protein domain rearrangements and their relevance to AML pathogenesis.

• Klíčová slova
• Acute myeloid leukaemia, FLT3, Gene isoforms, Mutations,
• MeSH
• akutní myeloidní leukemie * genetika   MeSH
• lidé MeSH
• mutace genetika   MeSH
• protein - isoformy genetika   MeSH
• tyrosinkinasa 3 podobná fms genetika   MeSH
• tyrosinkinasy MeSH
• viabilita buněk MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• FLT3 protein, human MeSH Prohlížeč
• protein - isoformy MeSH
• tyrosinkinasa 3 podobná fms MeSH
• tyrosinkinasy MeSH

BACKGROUND: The study investigated FLT3 gene mutations in patients from eastern Slovakia using a simple molecular method. PATIENTS AND METHODS: We analyzed 141 patients with primary acute myeloid leukemia (AML) and 8 patients with AML that developed from myelodysplastic syndrome (MDS) who were aged 19-81 years. DNA isolated from peripheral blood and/or bone marrow was analyzed by PCR. FLT3 internal tandem duplication (FLT3-ITD) was detected by amplification of exons 14 and 15. Point mutations in the FLT3 tyrosine kinase domain (FLT3-TKD) were detected by digesting the PCR product of exon 20 with the restriction endonuclease EcoRV. Fragments were separated electrophoretically. PCR products of the positive samples were also analyzed using a microchip device (Bioanalyzer 2100). RESULTS: LT3-ITD and point mutations in the FLT-TKD were detected in 19 and 8% of patients, resp. Two patients (1%) harbored both types of mutations. Patients with and without FLT3 mutations were called FLT+ and FLT-, resp. Most FLT3+ patients had no chromosomal aberrations (59%) or harbored the t (15; 17) translocation in PML-RARA (15%). The mortality rate was 33% among FLT3+ patients and 10% among FLT3-patients. Among FLT3+ patients, the mortality rates of patients with FLT3-ITD and point mutations of the FLT-TKD were almost the same. A 77-year-old female patient with both FLT3-ITD and a point mutation in the FLT3-TKD was in remission. The eight patients who developed AML from MDS were assessed separately. Of these, three patients were FLT3+; two patients displayed FLT3-ITD, and one patient harbored a point mutation in the FLT3-TKD. No other genetic aberrations were detected. FLT3+ patients lived for longer than FLT3-patients. These analyses of FLT3 gene mutations in patients from eastern Slovakia are consistent with published data from other databases. CONCLUSION: The applied PCR method is reliable, relatively fast, and affordable, and can be used for routine monitoring of FLT3 gene mutations. FLT3 mutations can be verified using a microchip as an alternative to capillary electrophoresis. Key words: acute myelogenous leukemia - DNA - PCR - mutation - FLT3-ITD - FLT3-TKD The study was supported by the European Regional Development grant OPVaV-2009/2.2/05- -SORO (ITMS code: 26220220143). The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study. The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical paper Submitted: 19. 10. 2017 Accepted: 15. 2. 2018.

• MeSH
• akutní myeloidní leukemie genetika   MeSH
• dospělí MeSH
• kostní dřeň MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mutace * MeSH
• polymerázová řetězová reakce metody   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• tyrosinkinasa 3 podobná fms genetika   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Slovenská republika MeSH
• Názvy látek
• FLT3 protein, human MeSH Prohlížeč
• tyrosinkinasa 3 podobná fms MeSH

Oncogenic mutations in gene encoding FLT3 kinase are often detected in acute myeloid leukaemia (AML) patients, and several potent kinase inhibitors have been developed. However, the FLT3 inhibitor treatment often leads to the resistance development and subsequent relapse. Targeted degradation of oncogenic protein kinases has emerged as a feasible pharmacological strategy, providing more robust effect over traditional competitive inhibitors. Based on previously developed competitive inhibitor of FLT3 and CDK9, we have designed and prepared a novel pomalidomide-based PROTAC. A series of biochemical and cellular experiments showed selectivity towards FLT3-ITD bearing AML cells and confirmed proteasome-dependent mechanism of action. Dual FLT3-ITD and CDK9 protein degradation resulted in the block of FLT3-ITD downstream signalling pathways, apoptosis activation and cell cycle arrest of FLT3-ITD AML cells. Moreover, transcriptional repression caused by CDK9 degradation significantly reduced expression of crucial genes involved in AML pathogenesis. The obtained results indicate the beneficial impact of simultaneous FLT3-ITD/CDK9 degradation for AML therapy.

• Klíčová slova
• Acute myeloid leukaemia, CDK9, FLT3, Proteolysis targeting chimera (PROTAC),
• MeSH
• akutní myeloidní leukemie * patologie   MeSH
• apoptóza MeSH
• cyklin-dependentní kinasa 9 metabolismus   MeSH
• inhibitory proteinkinas farmakologie   terapeutické užití   MeSH
• lidé MeSH
• mutace MeSH
• proteolýza MeSH
• tyrosinkinasa 3 podobná fms genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• CDK9 protein, human MeSH Prohlížeč
• cyklin-dependentní kinasa 9 MeSH
• FLT3 protein, human MeSH Prohlížeč
• inhibitory proteinkinas MeSH
• tyrosinkinasa 3 podobná fms MeSH

Allogeneic hematopoietic cell transplantation (alloHCT) as a postremission therapy in patients with FLT3-ITD-positive intermediate-risk acute myeloid leukemia (AML) remains controversial. FLT3-ITD mutations are heterogeneous with respect to allelic ratio, location, and length of the insertion, with a high mutant-to-wild-type ratio consistently associated with inferior prognosis. We retrospectively analyzed the role of alloHCT in first remission in relationship to the allelic ratio and presence or absence of nucleophosmin 1 mutations (NPM1) in the Study Alliance Leukemia AML2003 trial. FLT3-ITD mutations were detected in 209 patients and concomitant NPM1 mutations in 148 patients. Applying a predefined cutoff ratio of .8, AML was grouped into high- and low-ratio FLT3-ITD AML (HR(FLT3-ITD) and LR(FLT3-ITD)). Sixty-one patients (29%) were transplanted in first remission. Overall survival (OS) (HR, .3; 95% CI, .16 to .7; P = .004) and event-free survival (EFS) (HR, .4; 95% CI, .16 to .9; P = .02) were significantly increased in patients with HR(FLT3-ITD) AML who received alloHCT as consolidation treatment compared with patients who received consolidation chemotherapy. Patients with LR(FLT3-ITD) AML and wild-type NPM1 who received alloHCT in first remission had increased OS (HR, .3; 95% CI, .1 to .8; P = .02) and EFS (HR, .2; 95% CI, .1 to .8; P = .02), whereas alloHCT in first remission did not have a significant impact on OS and EFS in patients with LR(FLT3-ITD) AML and concomitant NPM1 mutation. In conclusion, our results provide additional evidence that alloHCT in first remission improves EFS and OS in patients with HR(FLT3-ITD) AML and in patients with LR(FLT3-ITD) AML and wild-type NPM1.

• Klíčová slova
• Acute myeloid leukemia, Allogeneic transplantation, FLT3-ITD, NPM1 mutation,
• MeSH
• akutní myeloidní leukemie * enzymologie   genetika   mortalita   terapie   MeSH
• alely * MeSH
• alografty MeSH
• dospělí MeSH
• jaderné proteiny genetika   MeSH
• lidé středního věku MeSH
• lidé MeSH
• míra přežití MeSH
• mladiství MeSH
• mutace * MeSH
• nukleofosmin MeSH
• přežití po terapii bez příznaků nemoci MeSH
• transplantace kmenových buněk * MeSH
• tyrosinkinasa 3 podobná fms genetika   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH
• randomizované kontrolované studie MeSH
• Názvy látek
• FLT3 protein, human MeSH Prohlížeč
• jaderné proteiny MeSH
• NPM1 protein, human MeSH Prohlížeč
• nukleofosmin MeSH
• tyrosinkinasa 3 podobná fms MeSH

Therapy of FLT3-positive acute myeloid leukemia still remains complicated, despite the availability of newly approved kinase inhibitors. Various strategies to avoid the reduced efficacy of therapy have been explored, including the development of dual targeting compounds, which inhibit FLT3 and another kinase necessary for the survival and proliferation of AML cells. We have designed new 2,7,9-trisubstituted 8-oxopurines as FLT3 inhibitors and report here the structure-activity relationship studies. We demonstrated that substituents at positions 7 and 9 modulate activity between CDK4 and FLT3 kinase, and the isopropyl group at position 7 substantially increased the selectivity toward FLT3 kinase, which led to the discovery of compound 15a (9-cyclopentyl-7-isopropyl-2-((4-(piperazin-1-yl)phenyl)amino)-7,9-dihydro-8H-purin-8-one). Cellular analyses in MV4-11 cells revealed inhibition of autophosphorylation of FLT3 kinase in nanomolar doses, including the suppression of downstream STAT5 and ERK1/2 phosphorylation. We also describe mechanistic studies in cell lines and activity in a mouse xenograft model in vivo.

• Klíčová slova
• FLT3, acute myeloid leukemia, cyclin-dependent kinase, inhibitor, purine,
• MeSH
• akutní myeloidní leukemie * farmakoterapie   MeSH
• apoptóza MeSH
• fosforylace MeSH
• inhibitory proteinkinas terapeutické užití   MeSH
• lidé MeSH
• mutace MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• proliferace buněk MeSH
• tyrosinkinasa 3 podobná fms MeSH
• vztahy mezi strukturou a aktivitou MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• FLT3 protein, human MeSH Prohlížeč
• inhibitory proteinkinas MeSH
• tyrosinkinasa 3 podobná fms MeSH

BACKGROUND: Patients with internal tandem duplication in fms-related tyrosine kinase receptor gene 3 (FLT3-ITD)-mutated acute myeloid leukemia (AML) have a dismal prognosis and the only curative option seems to be allogeneic stem cell transplantation (alloSCT). However, its timing is still matter of debate. PATIENTS AND METHODS: We retrospectively analyzed 73 consecutive AML patients with FLT3-ITD (median age 53, range 20-68 years) allografted with consistent policy to try to refer them all for upfront alloSCT in first complete remission (CR1). RESULTS: With a median follow-up of 44 (range, 5-135) months the 5-year overall survival (OS)/disease-free survival (DFS) probabilities were 49%/47%. The cumulative incidence of relapse and nonrelapse mortality (NRM) were 37% and 14%, respectively. The estimated 5-year OS for patients who received transplantation in CR1 was 62% versus 0% for patients who received transplantation beyond CR1. Multivariable analysis identified stem cell transplantation beyond CR1 as the key factor for poor OS (hazard ratio [HR], 5.41; P < .0001), DFS (HR, 4.41; P = .0002), and high relapse incidence (HR, 8.08; P < .0001). Acute graft versus host disease Grade ≥3 predicted higher NRM (HR, 3.80; P = .059) as well as inferior OS (HR, 2.04; P = .0079). No association of patient age, nucleophosmin status, donor type, conditioning, and other variables on the survival was detected. CONCLUSION: AlloSCT should be regarded with urgency as soon as CR1 is achieved in this subset of AML patients.

• Klíčová slova
• AML, Allogeneic SCT, Complete remission, Donor, FLT3-ITD,
• MeSH
• akutní myeloidní leukemie diagnóza   genetika   mortalita   terapie   MeSH
• dárci tkání MeSH
• dospělí MeSH
• duplikace genu * MeSH
• homologní transplantace MeSH
• indukce remise MeSH
• kombinovaná terapie MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mutace * MeSH
• prognóza MeSH
• recidiva MeSH
• retrospektivní studie MeSH
• senioři MeSH
• transplantace hematopoetických kmenových buněk * škodlivé účinky   metody   MeSH
• tyrosinkinasa 3 podobná fms genetika   MeSH
• výsledek terapie MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• FLT3 protein, human MeSH Prohlížeč
• tyrosinkinasa 3 podobná fms MeSH

FLT3 tyrosine kinase is a potential drug target in acute myeloid leukemia (AML) because patients with FLT3-ITD mutations respond poorly to standard cytotoxic agents and there is a clear link between the disease and the oncogenic properties of FLT3. We present novel 2,6,9-trisubstituted purine derivatives with potent FLT3 inhibitory activity. The lead compound 7d displays nanomolar activity in biochemical assays and selectively blocks proliferation of AML cell lines harboring FLT3-ITD mutations, whereas other transformed and normal human cells are several orders of magnitude less sensitive. The MV4-11 cells treated with 7d suppressed the phosphorylation of FLT3 and its downstream signaling pathways, with subsequent G1 cell cycle arrest and apoptosis. Additionally, a single dose of 7d in mice with subcutaneous MV4-11 xenografts caused sustained inhibition of FLT3 and STAT5 phosphorylation over 48 h, in contrast to the shorter effect observed after administration of the reference FLT3 inhibitor quizartinib.

• MeSH
• akutní myeloidní leukemie genetika   patologie   MeSH
• antitumorózní látky chemie   metabolismus   farmakologie   MeSH
• diaminy chemie   metabolismus   farmakologie   MeSH
• inhibitory proteinkinas chemie   metabolismus   farmakologie   MeSH
• konformace proteinů MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• objevování léků * MeSH
• simulace molekulového dockingu MeSH
• tyrosinkinasa 3 podobná fms antagonisté a inhibitory   chemie   genetika   metabolismus   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• xenogenní modely - testy antitumorózní aktivity MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antitumorózní látky MeSH
• diaminy MeSH
• FLT3 protein, human MeSH Prohlížeč
• inhibitory proteinkinas MeSH
• tyrosinkinasa 3 podobná fms MeSH

BACKGROUND: Vaccinia virus, one of the best known members of poxvirus family, has a wide host range both in vivo and in vitro. The expression of Flt3 ligand (FL) by recombinant vaccinia virus (rVACV) highly influenced properties of the virus in dependence on the level of expression. RESULTS: High production of FL driven by the strong synthetic promoter decreased the growth of rVACV in macrophage cell line J774.G8 in vitro as well as its multiplication in vivo when inoculated in mice. The inhibition of replication in vivo was mirrored in low levels of antibodies against vaccinia virus (anti-VACV) which nearly approached to the negative serum level in non-infected mice. Strong FL expression changed not only the host range of the recombinant but also the basic protein contents of virions. The major proteins - H3L and D8L - which are responsible for the virus binding to the cells, and 28 K protein that serves as a virulence factor, were changed in the membrane portion of P13-E/L-FL viral particles. The core virion fraction contained multiple larger, uncleaved proteins and a higher amount of cellular proteins compared to the control virus. The overexpression of FL also resulted in its incorporation into the viral core of P13-E/L-FL IMV particles. In contrary to the equimolar ratio of glycosylated and nonglycosylated FL forms found in cells transfected with the expression plasmid, the recombinant virus incorporated mainly the smaller, nonglycosylated FL. CONCLUSIONS: It has been shown that the overexpression of the Flt3L gene in VACV results in the attenuation of the virus in vivo.

• MeSH
• buněčné linie MeSH
• exprese genu * MeSH
• lidé MeSH
• membránové proteiny genetika   metabolismus   MeSH
• myši MeSH
• replikace viru MeSH
• vakcínie genetika   metabolismus   virologie   MeSH
• virus vakcinie genetika   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• flt3 ligand protein MeSH Prohlížeč
• membránové proteiny MeSH