ABSTRACT
Next-generation sequencing of samples from patients with acute myeloid leukemia (AML) has revealed several driver gene mutations in adult AML. However, unlike other cancers, AML is defined by relatively few mutations per patient, with a median of 4-5 depending on subtype. In this review, we will discuss the most common driver genes found in patients with AML and focus on the most clinically relevant ones that impact treatment strategies. The most common driver gene mutations in AML occur in NPM1 and FLT3, accounting for ~30% each. There are now targeted therapies being tested or already approved for these driver genes. Menin inhibitors, a novel targeted therapy that blocks the function of the menin protein, are in clinical trials for NPM1 driver gene mutant AML after relapse. A number of FLT3 inhibitors are now approved for FLT3 driver gene mutant AML in combination with chemotherapy in the frontline and also as single agent in relapse. Although mutations in IDH1/2 and TP53 only occur in around 10-20% of patients with AML each, they can affect the treatment strategy due to their association with prognosis and availability of targeted agents. While the impact of other driver gene mutations in AML is recognized, there is a lack of data on the actionable impact of those mutations.
Subject(s)
Leukemia, Myeloid, Acute , Mutation , Nucleophosmin , Humans , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/drug therapy , Mutation/genetics , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/metabolism , Molecular Targeted TherapyABSTRACT
FLT3 mutation is one of the most frequent genetic mutations in AML, identified in approximately 30% of patients, and FLT3-ITD mutation is considered a poor prognostic factor. Based on these molecular and clinical backgrounds, FLT3 mutations are considered promising therapeutic targets in AML, and intensive development of targeted therapeutics has been ongoing for more than two decades. Recently, combination of FLT3 inhibitors with intensive chemotherapy for untreated AML patients with FLT3 mutations and FLT3 inhibitor monotherapy for relapsed/refractory patients have been approved. In Japan, the combination of quizartinib and intensive chemotherapy for untreated FLT3-ITD-positive AML was approved in 2023. Clinical use of FLT3 inhibitors shows strong promise for improving the clinical outcomes of these AML patients with an extremely poor prognosis. Meanwhile, various resistance mechanisms to FLT3 inhibitors have been identified, including the emergence of resistance-associated mutations, and attenuated inhibitory effects of FLT3 inhibitors involving the bone marrow microenvironment surrounding AML cells. Thus, future efforts should aim to optimize combination therapy based on the characteristics of each FLT3 inhibitor, develop biomarkers that could inform treatment selection, and to better understand these resistance mechanisms and develop methods for overcoming them.
Subject(s)
Leukemia, Myeloid, Acute , Mutation , Protein Kinase Inhibitors , fms-Like Tyrosine Kinase 3 , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/genetics , Humans , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Protein Kinase Inhibitors/therapeutic use , Protein Kinase Inhibitors/pharmacology , Drug Resistance, NeoplasmABSTRACT
BACKGROUND: Approximately 25-30% of patients with acute myeloid leukemia (AML) have FMS-like receptor tyrosine kinase-3 (FLT3) mutations that contribute to disease progression and poor prognosis. Prolonged exposure to FLT3 tyrosine kinase inhibitors (TKIs) often results in limited clinical responses due to diverse compensatory survival signals. Therefore, there is an urgent need to elucidate the mechanisms underlying FLT3 TKI resistance. Dysregulated sphingolipid metabolism frequently contributes to cancer progression and a poor therapeutic response. However, its relationship with TKI sensitivity in FLT3-mutated AML remains unknown. Thus, we aimed to assess mechanisms of FLT3 TKI resistance in AML. METHODS: We performed lipidomics profiling, RNA-seq, qRT-PCR, and enzyme-linked immunosorbent assays to determine potential drivers of sorafenib resistance. FLT3 signaling was inhibited by sorafenib or quizartinib, and SPHK1 was inhibited by using an antagonist or via knockdown. Cell growth and apoptosis were assessed in FLT3-mutated and wild-type AML cell lines via Cell counting kit-8, PI staining, and Annexin-V/7AAD assays. Western blotting and immunofluorescence assays were employed to explore the underlying molecular mechanisms through rescue experiments using SPHK1 overexpression and exogenous S1P, as well as inhibitors of S1P2, ß-catenin, PP2A, and GSK3ß. Xenograft murine model, patient samples, and publicly available data were analyzed to corroborate our in vitro results. RESULTS: We demonstrate that long-term sorafenib treatment upregulates SPHK1/sphingosine-1-phosphate (S1P) signaling, which in turn positively modulates ß-catenin signaling to counteract TKI-mediated suppression of FLT3-mutated AML cells via the S1P2 receptor. Genetic or pharmacological inhibition of SPHK1 potently enhanced the TKI-mediated inhibition of proliferation and apoptosis induction in FLT3-mutated AML cells in vitro. SPHK1 knockdown enhanced sorafenib efficacy and improved survival of AML-xenografted mice. Mechanistically, targeting the SPHK1/S1P/S1P2 signaling synergizes with FLT3 TKIs to inhibit ß-catenin activity by activating the protein phosphatase 2 A (PP2A)-glycogen synthase kinase 3ß (GSK3ß) pathway. CONCLUSIONS: These findings establish the sphingolipid metabolic enzyme SPHK1 as a regulator of TKI sensitivity and suggest that combining SPHK1 inhibition with TKIs could be an effective approach for treating FLT3-mutated AML.
Subject(s)
Glycogen Synthase Kinase 3 beta , Leukemia, Myeloid, Acute , Phosphotransferases (Alcohol Group Acceptor) , Protein Phosphatase 2 , beta Catenin , fms-Like Tyrosine Kinase 3 , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/metabolism , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Humans , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/pathology , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/metabolism , Glycogen Synthase Kinase 3 beta/metabolism , Glycogen Synthase Kinase 3 beta/genetics , beta Catenin/metabolism , beta Catenin/genetics , Phosphotransferases (Alcohol Group Acceptor)/metabolism , Phosphotransferases (Alcohol Group Acceptor)/genetics , Phosphotransferases (Alcohol Group Acceptor)/antagonists & inhibitors , Animals , Mice , Protein Phosphatase 2/metabolism , Protein Phosphatase 2/genetics , Protein Phosphatase 2/antagonists & inhibitors , Cell Line, Tumor , Sorafenib/pharmacology , Apoptosis/drug effects , Protein Kinase Inhibitors/pharmacology , Signal Transduction/drug effects , Cell Proliferation/drug effects , Drug Synergism , Xenograft Model Antitumor Assays , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/geneticsABSTRACT
Conventional dendritic cells subtype 1 (cDC1) play a vital role in the priming and expansion of tumor-specific CD8+ T cells and their recruitment to tumor microenvironment. However, cDC1s are often underrepresented in the microenvironment. Systemic administration of Fms-like tyrosine kinase 3 ligand, a hematopoietic growth factor that binds to FLT3 on myeloid and lymphoid progenitor cells, leads to cDC1 expansion in the periphery and recruitment into the microenvironment. FLT3 pathway stimulation using GS-3583, a novel FLT3 agonistic Fc fusion protein, has the potential to promote T-cell mediated antitumor activity. This was a first-in-human, placebo-controlled study of GS-3583 in healthy participants to evaluate the safety, pharmacokinetics (PK), and pharmacodynamic (PD) of escalating single doses (75-2000 µg) of GS-3583. Each dose cohort enrolled 8-12 healthy participants who received GS-3583 or placebo as single IV infusion at 3:1 ratio. As part of the PD evaluation, the changes in the number of cDC1 cells were investigated. GS-3583 was well-tolerated in healthy participants up to the highest evaluated dose (2000 µg). There have been no serious or grade III or higher adverse events. PK analysis suggested a dose-dependent increase in GS-3583 exposure with target-mediated disposition characteristics at low doses. PD analysis shows that administration of GS-3583 resulted in transient, dose-dependent increases in cDC1 cells that returned to baseline within 3 weeks of drug administration. The pharmacokinetics and pharmacodynamics of GS-3583 following single dosing were characterized in this study which enabled subsequent phase Ib assessments in patients with advanced solid tumors.
Subject(s)
Healthy Volunteers , Immunoglobulin Fc Fragments , Recombinant Fusion Proteins , fms-Like Tyrosine Kinase 3 , Humans , Adult , Male , Female , Middle Aged , Recombinant Fusion Proteins/pharmacokinetics , Recombinant Fusion Proteins/administration & dosage , Recombinant Fusion Proteins/adverse effects , Recombinant Fusion Proteins/pharmacology , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Young Adult , Immunoglobulin Fc Fragments/administration & dosage , Immunoglobulin Fc Fragments/pharmacology , Immunoglobulin Fc Fragments/adverse effects , Dose-Response Relationship, Drug , Dendritic Cells/drug effects , Dendritic Cells/immunology , Double-Blind Method , Infusions, IntravenousABSTRACT
INTRODUCTION: Despite the prognosis of patients affected by acute myeloid leukemia (AML) improved in the last decade, most patients relapse. Maintenance therapy after a chemotherapy approach with or without allogeneic stem cell transplantation could be a way to control the undetectable residual burden of leukemic cells. Several studies are being carried out as maintenance therapy in AML. Some critical points need to be defined, how the physician can choose among the various drugs available. AREAS COVERED: This review discusses the advances and controversies surrounding maintenance therapy for AML patients. EXPERT OPINION: Patients withFLT3-positive AML should receive midostaurin or quizartinib in the first-linesetting. For a patient initially receiving midostaurin, consider switching to sorafenib in the post-transplant setting. Because of the improved safety profile and potency, many experts will lean toward using a second-generation FLT3 inhibitor such as quizartinib or gilteritinib. Finally, no data indicate whether maintenance therapy should be prolonged until progression or for a defined period.
Subject(s)
Leukemia, Myeloid, Acute , Maintenance Chemotherapy , Humans , Leukemia, Myeloid, Acute/therapy , Leukemia, Myeloid, Acute/drug therapy , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/metabolism , Hematopoietic Stem Cell Transplantation , Staurosporine/analogs & derivatives , Staurosporine/therapeutic use , Protein Kinase Inhibitors/therapeutic use , Antineoplastic Agents/therapeutic use , Sorafenib/therapeutic use , Phenylurea Compounds/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Aniline Compounds , Pyrazines , BenzothiazolesABSTRACT
BACKGROUND: FMS-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) is a common mutation type in acute myeloid leukemia (AML) and is usually associated with poor patient prognosis. With advancements in molecular diagnostics and the development of tyrosine kinase inhibitors (TKI), the overall survival (OS) of AML patients with FLT3-ITD mutations has been prolonged to some extent, but relapse and drug resistance are still substantial challenges. Ningetinib is a novel TKI against various kinases in relation to tumour pathogenesis and is undergoing clinical trials of lung cancer. In this study, we explored the antitumor activity of ningetinib against AML with FLT3 mutations both in vivo and in vitro. METHODS: Cell proliferation assays were performed in AML cell lines and Ba/F3 cells expressing various FLT3 mutations to validate the antileukemic activity of ningetinib in vitro. Immunoblot assays were used to verify the effect of ningetinib on the FLT3 protein and downstream pathways. Molecular docking and CETSA were used to validate the interaction of ningetinib with target proteins. The survival benefit of ningetinib in vivo was assessed in Ba/F3-FLT3-ITD-, MOLM13, Ba/F3-FLT3-ITD-F691L-, MOLM13-FLT3-ITD-F691L-induced leukemia mouse models. We also used patient-derived primary cells to determine the efficacy of ningetinib. RESULTS: Ningetinib inhibited cell proliferation, blocked the cell cycle, induced apoptosis and bound FLT3 to inhibit its downstream signaling pathways, including the STAT5, AKT and ERK pathways, in FLT3-ITD AML cell lines. In the mouse models with FLT3-ITD and FLT3-ITD-F691L mutation, ningetinib showed superior anti-leukemia activity to existing clinical drugs gilteritinib and quizartinib, significantly prolongating the survival of mice. In addition, ningetinib exhibited activity against patient-derived primary cells harboring FLT3-ITD mutations. CONCLUSION: Overall, our study confirmed the therapeutic role of ningetinib in AML with FLT3-ITD mutations, providing a potential new option for clinically resistant patients.
Subject(s)
Cell Proliferation , Drug Resistance, Neoplasm , Leukemia, Myeloid, Acute , Protein Kinase Inhibitors , fms-Like Tyrosine Kinase 3 , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/metabolism , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/pathology , Humans , Animals , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/genetics , Cell Line, Tumor , Protein Kinase Inhibitors/pharmacology , Cell Proliferation/drug effects , Mice , Xenograft Model Antitumor Assays , Apoptosis/drug effects , Mutation , Signal Transduction/drug effectsABSTRACT
Fms-like tyrosine kinase 3 (FLT3) mutations, present in over 30% of acute myeloid leukemia (AML) cases and dominated by FLT3-internal tandem duplication (FLT3-ITD), are associated with poor outcomes in patients with AML. While tyrosine kinase inhibitors (TKIs; e.g., gilteritinib) are effective, they face challenges such as drug resistance, relapse, and high costs. Here, we report that metformin, a cheap, safe, and widely used anti-diabetic agent, exhibits a striking synergistic effect with gilteritinib in treating FLT3-ITD AML. Metformin significantly sensitizes FLT3-ITD AML cells (including TKI-resistant ones) to gilteritinib. Metformin plus gilteritinib (low dose) dramatically suppresses leukemia progression and prolongs survival in FLT3-ITD AML mouse models. Mechanistically, the combinational treatment cooperatively suppresses polo-like kinase 1 (PLK1) expression and phosphorylation of FLT3/STAT5/ERK/mTOR. Clinical analysis also shows improved survival rates in patients with FLT3-ITD AML taking metformin. Thus, the metformin/gilteritinib combination represents a promising and cost-effective treatment for patients with FLT3-mutated AML, particularly for those with low income/affordability.
Subject(s)
Aniline Compounds , Cell Cycle Proteins , Drug Synergism , Leukemia, Myeloid, Acute , Metformin , Mutation , Polo-Like Kinase 1 , Protein Serine-Threonine Kinases , Proto-Oncogene Proteins , Pyrazines , Signal Transduction , fms-Like Tyrosine Kinase 3 , Metformin/pharmacology , Metformin/therapeutic use , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/metabolism , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Proto-Oncogene Proteins/metabolism , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/antagonists & inhibitors , Humans , Animals , Cell Cycle Proteins/metabolism , Cell Cycle Proteins/genetics , Protein Serine-Threonine Kinases/metabolism , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/antagonists & inhibitors , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/pathology , Signal Transduction/drug effects , Pyrazines/pharmacology , Pyrazines/therapeutic use , Aniline Compounds/pharmacology , Aniline Compounds/therapeutic use , Mice , Mutation/genetics , Cell Line, Tumor , Thiophenes/pharmacology , Thiophenes/therapeutic use , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , STAT5 Transcription Factor/metabolism , STAT5 Transcription Factor/genetics , Female , Xenograft Model Antitumor Assays , Male , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/genetics , TOR Serine-Threonine Kinases/metabolismABSTRACT
Hyperactive FMS-like receptor tyrosine kinase-3 mutants with internal tandem duplications (FLT3-ITD) are frequent driver mutations of aggressive acute myeloid leukemia (AML). Inhibitors of FLT3 produce promising results in rationally designed cotreatment schemes. Since FLT3-ITD modulates DNA replication and DNA repair, valid anti-leukemia strategies could rely on a combined inhibition of FLT3-ITD and regulators of cell cycle progression and DNA integrity. These include the WEE1 kinase which controls cell cycle progression, nucleotide synthesis, and DNA replication origin firing. We investigated how pharmacological inhibition of FLT3 and WEE1 affected the survival and genomic integrity of AML cell lines and primary AML cells. We reveal that promising clinical grade and preclinical inhibitors of FLT3 and WEE1 synergistically trigger apoptosis in leukemic cells that express FLT3-ITD. An accumulation of single and double strand DNA damage precedes this process. Mass spectrometry-based proteomic analyses show that FLT3-ITD and WEE1 sustain the expression of the ribonucleotide reductase subunit RRM2, which provides dNTPs for DNA replication. Unlike their strong pro-apoptotic effects on leukemia cells with FLT3-ITD, inhibitors of FLT3 and WEE1 do not damage healthy human blood cells and murine hematopoietic stem cells. Thus, pharmacological inhibition of FLT3-ITD and WEE1 might become an improved, rationally designed therapeutic option.
Subject(s)
Apoptosis , Cell Cycle Proteins , DNA Damage , Leukemia, Myeloid, Acute , Protein Kinase Inhibitors , Protein-Tyrosine Kinases , fms-Like Tyrosine Kinase 3 , Humans , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/metabolism , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/pathology , Apoptosis/drug effects , Cell Cycle Proteins/metabolism , Cell Cycle Proteins/genetics , Cell Cycle Proteins/antagonists & inhibitors , Protein-Tyrosine Kinases/metabolism , Protein-Tyrosine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/genetics , Protein Kinase Inhibitors/pharmacology , Cell Line, Tumor , Drug Synergism , Animals , Pyrazoles , PyrimidinonesABSTRACT
Activating mutations of FLT3 contribute to deregulated hematopoietic stem and progenitor cell (HSC/Ps) growth and survival in patients with acute myeloid leukemia (AML), leading to poor overall survival. AML patients treated with investigational drugs targeting mutant FLT3, including Quizartinib and Crenolanib, develop resistance to these drugs. Development of resistance is largely due to acquisition of cooccurring mutations and activation of additional survival pathways, as well as emergence of additional FLT3 mutations. Despite the high prevalence of FLT3 mutations and their clinical significance in AML, there are few targeted therapeutic options available. We have identified 2 novel nicotinamide-based FLT3 inhibitors (HSN608 and HSN748) that target FLT3 mutations at subnanomolar concentrations and are potently effective against drug-resistant secondary mutations of FLT3. These compounds show antileukemic activity against FLT3ITD in drug-resistant AML, relapsed/refractory AML, and in AML bearing a combination of epigenetic mutations of TET2 along with FLT3ITD. We demonstrate that HSN748 outperformed the FDA-approved FLT3 inhibitor Gilteritinib in terms of inhibitory activity against FLT3ITD in vivo.
Subject(s)
Drug Resistance, Neoplasm , Leukemia, Myeloid, Acute , Niacinamide , fms-Like Tyrosine Kinase 3 , Humans , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/pathology , Leukemia, Myeloid, Acute/metabolism , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/metabolism , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/genetics , Animals , Mice , Niacinamide/analogs & derivatives , Niacinamide/pharmacology , Cell Line, Tumor , Xenograft Model Antitumor Assays , Female , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Mutation , Mice, SCID , Mice, Inbred NODABSTRACT
Chromosomal translocations of the nucleoporin 98 (NUP98) gene are found in acute myeloid leukemia (AML) patients leading to very poor outcomes. The oncogenic activity of NUP98 fusion proteins is dependent on the interaction between Mixed Lineage Leukemia 1 and menin. NUP98-rearranged (NUP98-r) leukemia cells also rely on specific kinases, including CDK6 and/or FLT3, suggesting that simultaneous targeting of these kinases and menin could overcome limited sensitivity to single agents. Here, we found that combinations of menin inhibitor, MI-3454, with kinase inhibitors targeting either CDK6 (Palbociclib) or FLT3 (Gilteritinib) strongly enhance the anti-leukemic effect of menin inhibition in NUP98-r leukemia models. We found strong synergistic effects of both combinations on cell growth, colony formation and differentiation in patient samples with NUP98 translocations. These combinations also markedly augmented anti-leukemic efficacy of menin inhibitor in Patient Derived Xenograft models of NUP98-r leukemia. Despite inhibiting two unrelated kinases, when Palbociclib or Gilteritinib were combined with the menin inhibitor, they affected similar pathways relevant to leukemogenesis, including cell cycle regulation, cell proliferation and differentiation. This study provides strong rationale for clinical translation of the combination of menin and kinase inhibitors as novel treatments for NUP98-r leukemia, supporting the unexplored combinations of epigenetic drugs with kinase inhibitors.
Subject(s)
Nuclear Pore Complex Proteins , Protein Kinase Inhibitors , Proto-Oncogene Proteins , Translocation, Genetic , Humans , Animals , Nuclear Pore Complex Proteins/genetics , Mice , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , Proto-Oncogene Proteins/genetics , Xenograft Model Antitumor Assays , Piperazines/pharmacology , Piperazines/therapeutic use , Pyridines/pharmacology , Pyridines/therapeutic use , Cyclin-Dependent Kinase 6/antagonists & inhibitors , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/pathology , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Cell Proliferation/drug effects , Drug Synergism , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Cell Line, Tumor , Aniline Compounds , PyrazinesSubject(s)
Leukemia, Myeloid, Acute , Mutation , Protein Kinase Inhibitors , fms-Like Tyrosine Kinase 3 , Humans , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/drug therapy , Protein Kinase Inhibitors/therapeutic use , Veterans , United States , Male , United States Department of Veterans AffairsABSTRACT
AML is an aggressive malignancy of immature myeloid progenitor cells. Discovering effective treatments for AML through cell differentiation and anti-proliferation remains a significant challenge. Building on previous studies on CDK2 PROTACs with differentiation-inducing properties, this research aims to enhance CDKs degradation through structural optimization to facilitate the differentiation and inhibit the proliferation of AML cells. Compound C3, featuring a 4-methylpiperidine ring linker, effectively degraded CDK2 with a DC50 value of 18.73 ± 10.78 nM, and stimulated 72.77 ± 3.51 % cell differentiation at 6.25 nM in HL-60 cells. Moreover, C3 exhibited potent anti-proliferative activity against various AML cell types. Degradation selectivity analysis indicated that C3 could be endowed with efficient degradation of CDK2/4/6/9 and FLT3, especially FLT3-ITD in MV4-11 cells. These findings propose that C3 combined targeting CDK2/4/6/9 and FLT3 with enhanced differentiation and proliferation inhibition, which holds promise as a potential treatment for AML.
Subject(s)
Antineoplastic Agents , Cyclin-Dependent Kinases , Drug Discovery , Leukemia, Myeloid, Acute , Protein Kinase Inhibitors , Proteolysis Targeting Chimera , Proteolysis , fms-Like Tyrosine Kinase 3 , Humans , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/chemical synthesis , Cell Differentiation/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cyclin-Dependent Kinases/antagonists & inhibitors , Cyclin-Dependent Kinases/metabolism , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/metabolism , Leukemia, Myeloid, Acute/drug therapy , Molecular Structure , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/chemical synthesis , Structure-Activity Relationship , Proteolysis Targeting Chimera/chemistry , Proteolysis Targeting Chimera/pharmacology , Proteolysis Targeting Chimera/therapeutic useABSTRACT
Acute myeloid leukemia (AML) is often associated with poor prognosis and survival. Small molecule inhibitors, though widening the treatment landscape, have limited monotherapy efficacy. The combination therapy, however, shows suboptimal clinical outcomes due to low bioavailability, overlapping systemic toxicity and drug resistance. Here, we report that CXCR4-mediated codelivery of the BCL-2 inhibitor venetoclax (VEN) and the FLT3 inhibitor sorafenib (SOR) via T22 peptide-tagged disulfide cross-linked polymeric micelles (TM) achieves synergistic treatment of FLT3-ITD AML. TM-VS with a VEN/SOR weight ratio of 1/4 and T22 peptide density of 20% exhibited an extraordinary inhibitory effect on CXCR4-overexpressing MV4-11 AML cells. TM-VS at a VEN/SOR dosage of 2.5/10 mg/kg remarkably reduced leukemia burden, prolonged mouse survival, and impeded bone loss in orthotopic MV4-11-bearing mice, outperforming the nontargeted M-VS and oral administration of free VEN/SOR. CXCR4-mediated codelivery of BCL-2 and FLT3 inhibitors has emerged as a prospective clinical treatment for FLT3-ITD AML.
Subject(s)
Leukemia, Myeloid, Acute , Proto-Oncogene Proteins c-bcl-2 , Receptors, CXCR4 , Sorafenib , Sulfonamides , fms-Like Tyrosine Kinase 3 , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/metabolism , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/pathology , Leukemia, Myeloid, Acute/genetics , Animals , Receptors, CXCR4/antagonists & inhibitors , Receptors, CXCR4/genetics , Receptors, CXCR4/metabolism , Humans , Mice , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Proto-Oncogene Proteins c-bcl-2/genetics , Proto-Oncogene Proteins c-bcl-2/metabolism , Sulfonamides/pharmacology , Sulfonamides/administration & dosage , Sorafenib/pharmacology , Sorafenib/administration & dosage , Bridged Bicyclo Compounds, Heterocyclic/administration & dosage , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Cell Line, Tumor , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/administration & dosage , MicellesABSTRACT
The presence of FLT3 mutations, including the most common FLT3-ITD (internal tandem duplications) and FLT3-TKD (tyrosine kinase domain), is associated with an unfavorable prognosis in patients affected by acute myeloid leukemia (AML). In this setting, in recent years, new FLT3 inhibitors have demonstrated efficacy in improving survival and treatment response. Nevertheless, the development of primary and secondary mechanisms of resistance poses a significant obstacle to their efficacy. Understanding these mechanisms is crucial for developing novel therapeutic approaches to overcome resistance and improve the outcomes of patients. In this context, the use of novel FLT3 inhibitors and the combination of different targeted therapies have been studied. This review provides an update on the molecular alterations involved in the resistance to FLT3 inhibitors, and describes how the molecular monitoring may be used to guide treatment strategy in FLT3-mutated AML.
Subject(s)
Drug Resistance, Neoplasm , Leukemia, Myeloid, Acute , Mutation , Protein Kinase Inhibitors , fms-Like Tyrosine Kinase 3 , Humans , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/drug therapy , Drug Resistance, Neoplasm/genetics , Protein Kinase Inhibitors/therapeutic use , Protein Kinase Inhibitors/pharmacology , Molecular Targeted Therapy/methods , AdultABSTRACT
OPINION STATEMENT: The internal tandem duplication (ITD) mutation of the FMS-like receptor tyrosine kinase 3 (FLT3-ITD) is the most common mutation observed in approximately 30% of acute myeloid leukemia (AML) patients. It represents poor prognosis due to continuous activation of downstream growth-promoting signaling pathways such as STAT5 and PI3K/AKT. Hence, FLT3 is considered an attractive druggable target; selective small FLT3 inhibitors (FLT3Is), such as midostaurin and quizartinib, have been clinically approved. However, patients possess generally poor remission rates and acquired resistance when FLT3I used alone. Various factors in patients could cause these adverse effects including altered epigenetic regulation, causing mainly abnormal gene expression patterns. Epigenetic modifications are required for hematopoietic stem cell (HSC) self-renewal and differentiation; however, critical driver mutations have been identified in genes controlling DNA methylation (such as DNMT3A, TET2, IDH1/2). These regulators cause leukemia pathogenesis and affect disease diagnosis and prognosis when they co-occur with FLT3-ITD mutation. Therefore, understanding the role of different epigenetic alterations in FLT3-ITD AML pathogenesis and how they modulate FLT3I's activity is important to rationalize combinational treatment approaches including FLT3Is and modulators of methylation regulators or pathways. Data from ongoing pre-clinical and clinical studies will further precisely define the potential use of epigenetic therapy together with FLT3Is especially after characterized patients' mutational status in terms of FLT3 and DNA methlome regulators.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols , DNA Methylation , Leukemia, Myeloid, Acute , Molecular Targeted Therapy , Mutation , Protein Kinase Inhibitors , fms-Like Tyrosine Kinase 3 , Humans , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/therapy , Protein Kinase Inhibitors/therapeutic use , Protein Kinase Inhibitors/pharmacology , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Epigenesis, Genetic , Epigenome , Tandem Repeat Sequences , Biomarkers, Tumor , Prognosis , Disease ManagementABSTRACT
Internal tandem duplication mutations of FLT3 (FLT3/ITD) confer poor prognosis in AML. FLT3 tyrosine kinase inhibitors (TKIs) alone have limited and transient clinical efficacy thus calling for new targets for more effective combination therapy. In a loss-of-function RNAi screen, we identified NOTCH4 as one such potential target whose inhibition proved cytotoxic to AML cells, and also sensitized them to FLT3 inhibition. Further investigation found increased NOTCH4 expression in FLT3/ITD AML cell lines and primary patient samples. Inhibition of NOTCH4 by shRNA knockdown, CRISPR-Cas9-based knockout or γ-secretase inhibitors synergized with FLT3 TKIs to kill FLT3/ITD AML cells in vitro. NOTCH4 inhibition sensitized TKI-resistant FLT3/ITD cells to FLT3 TKI inhibition. The combination reduced phospho-ERK and phospho-AKT, indicating inhibition of MAPK and PI3K/AKT signaling pathways. It also led to changes in expression of genes involved in regulating cell cycling, DNA repair and transcription. A patient-derived xenograft model showed that the combination reduced both the level of leukemic involvement of primary human FLT3/ITD AML cells and their ability to engraft secondary recipients. In summary, these results demonstrate that NOTCH4 inhibition synergizes with FLT3 TKIs to eliminate FLT3/ITD AML cells, providing a new therapeutic target for AML with FLT3/ITD mutations.
Subject(s)
Leukemia, Myeloid, Acute , Protein Kinase Inhibitors , Receptor, Notch4 , Xenograft Model Antitumor Assays , fms-Like Tyrosine Kinase 3 , Humans , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/pathology , Leukemia, Myeloid, Acute/metabolism , Animals , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , Mice , Receptor, Notch4/genetics , Mutation , Cell Line, Tumor , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/genetics , Signal Transduction/drug effectsABSTRACT
INTRODUCTION: Acute myeloid leukemia (AML) encompasses a heterogeneous group of aggressive myeloid malignancies, where FMS-like tyrosine kinase 3 (FLT3) mutations are prevalent, accounting for approximately 25-30% of adult patients. The presence of this mutation is related to a dismal prognosis and high relapse rates. In the lasts years many FLT3 inhibitors have been developed. AREAS COVERED: This review provides a comprehensive overview of FLT3mut AML, summarizing the state of art of current treatment and available data about combination strategies including an FLT3 inhibitor. EXPERT OPINION: In addition, the review discusses the emergence of drug resistance and the need for a nuanced approaches in treating patients who are ineligible for or resistant to intensive chemotherapy. Specifically, it explores the historical context of FLT3 inhibitors (FLT3Is) and their impact on treatment outcomes, emphasizing the pivotal role of midostaurin, as well as gilteritinib and quizartinib, and providing detailed insights into ongoing trials exploring the safety and efficacy of novel triplet combinations involving FLT3Is in different AML settings.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols , Leukemia, Myeloid, Acute , Mutation , Protein Kinase Inhibitors , fms-Like Tyrosine Kinase 3 , Humans , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/therapy , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Protein Kinase Inhibitors/therapeutic use , Drug Resistance, Neoplasm , Staurosporine/analogs & derivatives , Staurosporine/therapeutic use , Treatment Outcome , Aniline Compounds , PyrazinesABSTRACT
BACKGROUND: Acute myeloid leukemia (AML) is a complex disease with diverse mutations, including prevalent mutations in the FMS-like receptor tyrosine kinase 3 (FLT3) gene that lead to poor prognosis. Recent advancements have introduced FLT3 inhibitors that have improved outcomes for FLT3-mutated AML patients, however, questions remain on their application in complex conditions such as relapsed/refractory (R/R) disease. Therefore, we aimed to evaluate the clinical effectiveness of second-generation FLT3 inhibitors in treating patients with R/R AML. METHODS: A systematic literature search of PubMed, MEDLINE, SCOPUS and Google Scholar databases was made to identify relevant studies up to January 30, 2024. This study was conducted following the guidelines of the PRISMA. RESULTS: The ADMIRAL trial revealed significantly improved overall survival and complete remission rates with gilteritinib compared to salvage chemotherapy, with manageable adverse effects. Ongoing research explores its potential in combination therapies, showing synergistic effects with venetoclax and promising outcomes in various clinical trials. The QuANTUM-R trial suggested longer overall survival with quizartinib compared to standard chemotherapy, although concerns were raised regarding trial design and cardiotoxicity. Ongoing research explores combination therapies involving quizartinib, such as doublet or triplet regimens with venetoclax, showing promising outcomes in FLT3-mutated AML patients. CONCLUSION: These targeted therapies offer promise for managing this subgroup of AML patients, but further research is needed to optimize their use. This study underscores the importance of personalized treatment based on genetic mutations in AML, paving the way for more effective and tailored approaches to combat the disease.
Subject(s)
Leukemia, Myeloid, Acute , Protein Kinase Inhibitors , fms-Like Tyrosine Kinase 3 , Humans , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/genetics , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/mortality , Protein Kinase Inhibitors/therapeutic use , Drug Resistance, Neoplasm , Mutation , Aniline Compounds/therapeutic use , Phenylurea Compounds/therapeutic use , Neoplasm Recurrence, Local/drug therapy , Neoplasm Recurrence, Local/pathology , Pyrazines/therapeutic use , BenzothiazolesABSTRACT
Fms-like tyrosine receptor kinase 3 (FLT3) proteolysis targeting chimeras (PROTACs) emerge as a promising approach to overcome the limitations of FLT3 inhibitors, while the development of orally bioavailable FLT3-PROTACs faces great challenges. Here, we report the rational design and evaluation of a series of Gilteritinib-based FLT3-PROTACs. Among them, B3-2 exhibited the strongest antiproliferative activity against FLT3-ITD mutant AML cells, and significantly induced FLT3-ITD protein degradation. Mechanistic investigations demonstrated that B3-2 induced FLT3-ITD degradation in a ubiquitin-proteasome-dependent manner. More importantly, B3-2 exhibited an oral bioavailability of 5.65%, and oral administration of B3-2 showed good antitumor activity in MV-4-11 xenograft models. Furthermore, B3-2 showed strong antiproliferative activity against FLT3 resistant mutations, highlighting its potential in overcoming drug resistance.
Subject(s)
Antineoplastic Agents , Cell Proliferation , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Leukemia, Myeloid, Acute , Protein Kinase Inhibitors , Pyrazines , fms-Like Tyrosine Kinase 3 , fms-Like Tyrosine Kinase 3/antagonists & inhibitors , fms-Like Tyrosine Kinase 3/metabolism , Humans , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/pathology , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/chemical synthesis , Pyrazines/chemistry , Pyrazines/pharmacology , Pyrazines/chemical synthesis , Cell Proliferation/drug effects , Animals , Structure-Activity Relationship , Molecular Structure , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/chemical synthesis , Mice , Drug Discovery , Thiophenes/chemistry , Thiophenes/pharmacology , Thiophenes/chemical synthesis , Proteolysis/drug effects , Aniline Compounds/chemistry , Aniline Compounds/pharmacology , Aniline Compounds/chemical synthesis , Cell Line, Tumor , Neoplasms, Experimental/drug therapy , Neoplasms, Experimental/pathology , Neoplasms, Experimental/metabolismABSTRACT
Acute myeloid leukemia (AML) patients carrying Fms-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD) mutations often face a poor prognosis. While some FLT3 inhibitors have been used clinically, challenges such as short efficacy and poor specificity persist. Proteolytic targeting chimera (PROTAC), with its lower ligand affinity requirement for target proteins, offers higher and rapid targeting capability. Gilteritinib, used as the ligand for the target protein, was connected with different E3 ligase ligands to synthesize several series of PROTAC targeting FLT3-ITD. Through screening and structural optimization, the optimal lead compound PROTAC Z29 showed better specificity than Gilteritinib. Z29 induced FLT3 degradation through the proteasome pathway and inhibited tumor growth in subcutaneous xenograft mice. We verified Z29's minimal impact on platelets in a patient-derived xenografts (PDX) model compared to Gilteritinib. The combination of Z29 and Venetoclax showed better anti-tumor effects, lower platelet toxicity, and lower hepatic toxicity in FLT3-ITD+ models. The FLT3-selective PROTAC can mitigate the platelet toxicity of small molecule inhibitors, ensuring safety and efficacy in monotherapy and combination therapy with Venetoclax. It is a promising strategy for FLT3-ITD+ patients, especially those with platelet deficiency or liver damage.