Arylcarboxyamino-substituted diaryl ureas as potent and selective FLT3 inhibitors

Therapy resistance remains a major challenge in the management of multiple myeloma (MM). We evaluated the expression of FLT3 tyrosine kinase receptor (FLT3, CD135) in myeloma cells as a possible clonal driver. FLT3 expression was analyzed in bone marrow biopsies of patients with monoclonal gammopathy of undetermined significance or smoldering myeloma (MGUS, SMM), newly diagnosed MM (NDMM), and relapsed/refractory MM (RRMM) by immunohistochemistry (IHC). FLT3 gene expression was analyzed by RNA sequencing (RNAseq) and real-time PCR (rt-PCR). Anti-myeloma activity of FLT3 inhibitors (midostaurin, gilteritinib) was tested in vitro on MM cell lines and primary MM cells by 3H-tymidine incorporation assays or flow cytometry. Semi-quantitative expression analysis applying a staining score (FLT3 expression IHC-score, FES, range 1–6) revealed that a high FES (>3) was associated with a significantly shorter progression-free survival (PFS) in NDMM and RRMM patients (p = 0.04). RNAseq and real-time PCR confirmed the expression of FLT3 in CD138-purified MM samples. The functional relevance of FLT3 expression was corroborated by demonstrating the in vitro anti-myeloma activity of FLT3 inhibitors on FLT3-positive MM cell lines and primary MM cells. FLT3 inhibitors might offer a new targeted therapy approach in a subgroup of MM patients displaying aberrant FLT3 signaling.

Download Full-text

ATM/G6PD-driven redox metabolism promotes FLT3 inhibitor resistance in acute myeloid leukemia

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1603876113 ◽

2016 ◽

Vol 113 (43) ◽

pp. E6669-E6678 ◽

Cited By ~ 35

Author(s):

Mark A. Gregory ◽

Angelo D’Alessandro ◽

Francesca Alvarez-Calderon ◽

Jihye Kim ◽

Travis Nemkov ◽

...

Keyword(s):

Oxidative Stress ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Mitochondrial Oxidative Stress ◽

Flt3 Inhibitor ◽

A Genome ◽

Flt3 Inhibitors ◽

Acute Myeloid

Activating mutations in FMS-like tyrosine kinase 3 (FLT3) are common in acute myeloid leukemia (AML) and drive leukemic cell growth and survival. Although FLT3 inhibitors have shown considerable promise for the treatment of AML, they ultimately fail to achieve long-term remissions as monotherapy. To identify genetic targets that can sensitize AML cells to killing by FLT3 inhibitors, we performed a genome-wide RNA interference (RNAi)-based screen that identified ATM (ataxia telangiectasia mutated) as being synthetic lethal with FLT3 inhibitor therapy. We found that inactivating ATM or its downstream effector glucose 6-phosphate dehydrogenase (G6PD) sensitizes AML cells to FLT3 inhibitor induced apoptosis. Examination of the cellular metabolome showed that FLT3 inhibition by itself causes profound alterations in central carbon metabolism, resulting in impaired production of the antioxidant factor glutathione, which was further impaired by ATM or G6PD inactivation. Moreover, FLT3 inhibition elicited severe mitochondrial oxidative stress that is causative in apoptosis and is exacerbated by ATM/G6PD inhibition. The use of an agent that intensifies mitochondrial oxidative stress in combination with a FLT3 inhibitor augmented elimination of AML cells in vitro and in vivo, revealing a therapeutic strategy for the improved treatment of FLT3 mutated AML.

Download Full-text

The effects of lestaurtinib (CEP701) and PKC412 on primary AML blasts: the induction of cytotoxicity varies with dependence on FLT3 signaling in both FLT3-mutated and wild-type cases

Blood ◽

10.1182/blood-2006-04-015487 ◽

2006 ◽

Vol 108 (10) ◽

pp. 3494-3503 ◽

Cited By ~ 88

Author(s):

Steven Knapper ◽

Kenneth I. Mills ◽

Amanda F. Gilkes ◽

Steve J. Austin ◽

Val Walsh ◽

...

Keyword(s):

Myeloid Leukemia ◽

Wild Type ◽

Mutation Status ◽

Activating Mutations ◽

Drug Concentrations ◽

Flt3 Inhibitors ◽

Show Evidence ◽

Flt3 Expression ◽

Molecular Therapeutic ◽

Cytotoxic Responses

Abstract The receptor tyrosine kinase FLT3 is a promising molecular therapeutic target in acute myeloid leukemia (AML). Activating mutations of FLT3 are present in approximately one-third of patients, while many nonmutants show evidence of FLT3 activation, which appears to play a significant role in leukemogenesis. We studied the effects of lestaurtinib (CEP701) and PKC412, 2 small molecule inhibitors of FLT3, on 65 diagnostic AML blast samples. Both agents induced concentration-dependent cytotoxicity in most cases, although responses to PKC412 required higher drug concentrations. Cytotoxic responses were highly heterogeneous and were only weakly associated with FLT3 mutation status and FLT3 expression. Importantly, lestaurtinib induced cytotoxicity in a synergistic fashion with cytarabine, particularly in FLT3 mutant samples. Both lestaurtinib and PKC412 caused inhibition of FLT3 phosphorylation in all samples. Translation of FLT3 inhibition into cytotoxicity was influenced by the degree of residual FLT3 phosphorylation remaining and correlated with deactivation of STAT5 and MAP kinase. FLT3 mutant and wild-type cases both varied considerably in their dependence on FLT3 signaling for survival. These findings support the continued clinical assessment of FLT3 inhibitors in combination with cytotoxic chemotherapy: Entry to future clinical trials should include FLT3 wild-type patients and should remain unrestricted by FLT3 expression level.

Download Full-text

FLT3 inhibitors in acute myeloid leukemia

Journal of Hematology & Oncology ◽

10.1186/s13045-018-0675-4 ◽

2018 ◽

Vol 11 (1) ◽

Cited By ~ 40

Author(s):

Mei Wu ◽

Chuntuan Li ◽

Xiongpeng Zhu

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Flt3 Inhibitors ◽

Acute Myeloid

Download Full-text

Blocking DNA Damage Repair May Be Involved in Stattic (STAT3 Inhibitor)-Induced FLT3-ITD AML Cell Apoptosis

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.637064 ◽

2021 ◽

Vol 9 ◽

Author(s):

Yuxuan Luo ◽

Ying Lu ◽

Bing Long ◽

Yansi Lin ◽

Yanling Yang ◽

...

Keyword(s):

Dna Damage ◽

Myeloid Leukemia ◽

Tandem Duplication ◽

Dna Damage Repair ◽

Underlying Mechanism ◽

Internal Tandem Duplication ◽

Mrna Sequencing ◽

Damage Repair ◽

Flt3 Inhibitors ◽

Flt3 Internal Tandem Duplication

The FMS-like tyrosine kinase 3 (FLT3)- internal tandem duplication (ITD) mutation can be found in approximately 25% of all acute myeloid leukemia (AML) cases and is associated with a poor prognosis. The main treatment for FLT3-ITD-positive AML patients includes genotoxic therapy and FLT3 inhibitors, which are rarely curative. Inhibiting STAT3 activity can improve the sensitivity of solid tumor cells to radiotherapy and chemotherapy. This study aimed to explore whether Stattic (a STAT3 inhibitor) affects FLT3-ITD AML cells and the underlying mechanism. Stattic can inhibit the proliferation, promote apoptosis, arrest cell cycle at G0/G1, and suppress DNA damage repair in MV4-11cells. During the process, through mRNA sequencing, we found that DNA damage repair-related mRNA are also altered during the process. In summary, the mechanism by which Stattic induces apoptosis in MV4-11cells may involve blocking DNA damage repair machineries.

Download Full-text

Genome-Wide CRISPR Screening in Acute Myeloid Leukemia Cells Identifies Genes in the PI3K/AKT/mTOR and MEK/ERK Pathways That Define Sensitivity to Trametinib

Blood ◽

10.1182/blood-2019-129117 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 1395-1395

Author(s):

Tamilla Nechiporuk ◽

Alisa Damnernsawad ◽

Daniel Bottomly ◽

Quinlan Morrow ◽

Suyoun Choi ◽

...

Keyword(s):

Myeloid Leukemia ◽

Research Funding ◽

Molecular Pathways ◽

Refractory Disease ◽

Loss Of Function ◽

Ras Pathway ◽

Genetics Research ◽

Genome Wide ◽

Flt3 Inhibitors ◽

Acute Myeloid

Acute myeloid leukemia (AML), the most common acute leukemia in adults, remains a challenging disease to treat due to its heterogeneity and high level of relapsed/refractory disease. Exploration of molecular pathways that drive AML have implicated broad activation of the RAS/RAF/MEK/ERK and PI3K/AKT/mTOR pro-survival pathways in the pathogenesis of the disease. Among the most frequent genetic perturbations, direct mutation of N- or K-RAS as well as mutation of nearby pathway regulators (PTPN11, NF1) are identified in ~20% of patients with AML. An additional ~35% of AML patients exhibit mutations in FLT3 or KIT, upstream receptor tyrosine kinases known to activate the RAS pathway. Moreover, relapsed/refractory disease in response to novel molecularly targeted FLT3 inhibitors often results in RAS pathway mutations underpinning the connection between these molecular pathways in AML. These findings resulted in several preclinical studies and ongoing clinical trials testing the efficacies of MEK1/2 inhibitors in AML either as a single agent or in combination with FLT3 inhibitors. To elucidate pathways leading to changes in trametinib sensitivity and resistance in a FLT3-ITD genomic landscape, we performed a genome-wide CRISPR screen for trametinib sensitivity in MOLM13 AML cells, which carry a FLT3-ITD mutation. Results from both the genome-wide screen and independently generated cell lines with decreased sensitivity to trametinib indicated involvement of a diversity of genes and pathways, including the tumor-suppressor, PTEN (a negative regulator of PI3K/AKT/mTOR), AMBRA1 (an autophagy regulator via the mTOR pathway), and DUSP7 (a phosphatase negatively regulating downstream ERK activity). Cells engineered to have loss-of-function for these genes as well as cells cultured for resistance to trametinib showed perturbed signaling in downstream PI3K/AKT/mTOR and MEK/ERK signaling cascades. Our work identified genes whose loss of function in the disease-implicated pathways confer trametinib resistance in AML and provide a rationale for selecting combinatorial trametinib/FLT3 inhibitors treatment based on unique patient mutational and gene expression landscapes. Disclosures Tyner: Incyte: Research Funding; Janssen: Research Funding; Incyte: Research Funding; Gilead: Research Funding; Janssen: Research Funding; Gilead: Research Funding; Takeda: Research Funding; Takeda: Research Funding; Genentech: Research Funding; Constellation: Research Funding; Aptose: Research Funding; Genentech: Research Funding; Syros: Research Funding; AstraZeneca: Research Funding; Seattle Genetics: Research Funding; Seattle Genetics: Research Funding; Array: Research Funding; Agios: Research Funding; Agios: Research Funding; Aptose: Research Funding; Array: Research Funding; AstraZeneca: Research Funding; Constellation: Research Funding; Petra: Research Funding; Syros: Research Funding; Petra: Research Funding.

Download Full-text

Genome-wide CRISPR screen identifies regulators of MAPK and MTOR pathways mediating sorafenib resistance in acute myeloid leukemia

Haematologica ◽

10.3324/haematol.2020.257964 ◽

2020 ◽

Author(s):

Alisa Damnernsawad ◽

Daniel Bottomly ◽

Stephen E. Kurtz ◽

Christopher A. Eide ◽

Shannon K. McWeeney ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Ex Vivo ◽

Mek Inhibitors ◽

Genome Wide ◽

A Genome ◽

Flt3 Mutations ◽

Flt3 Inhibitors ◽

Crispr Screen ◽

Acute Myeloid

Drug resistance impedes the long-term effect of targeted therapies in acute myeloid leukemia (AML), necessitating the identification of mechanisms underlying resistance. Approximately 25% of AML patients carry FLT3 mutations and develop post-treatment insensitivity to FLT3 inhibitors, including sorafenib. Using a genome-wide CRISPR screen, we identified LZTR1, NF1, TSC1 or TSC2, negative regulators of the MAPK and MTOR pathways, as mediators of sorafenib resistance. Analyses of ex vivo drug sensitivity assays in FLT3-ITD AML patient samples revealed lower expression of LZTR1, NF1, and TSC2 correlated with sorafenib sensitivity. Importantly, MAPK and/or MTOR complex1 (MTORC1) activity were upregulated in AML cells made resistant to several FLT3 inhibitors, including crenolanib, quizartinib, or sorafenib. These cells were sensitive to MEK inhibitors, and the combination of FLT3 and MEK inhibitors showed enhanced efficacy, suggesting its effectiveness in AML patients with FLT3 mutations and those with resistance to FLT3 inhibitors.

Download Full-text

MUC1-C oncoprotein promotes FLT3 receptor activation in acute myeloid leukemia cells

Blood ◽

10.1182/blood-2013-04-493858 ◽

2014 ◽

Vol 123 (5) ◽

pp. 734-742 ◽

Cited By ~ 13

Author(s):

Suiyang Liu ◽

Li Yin ◽

Dina Stroopinsky ◽

Hasan Rajabi ◽

Alexandre Puissant ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Receptor Activation ◽

Leukemia Cells ◽

Receptor Targeting ◽

Potential Approach ◽

Key Points ◽

Flt3 Inhibitors ◽

Acute Myeloid Leukemia Cells ◽

Acute Myeloid

Key Points The MUC1-C oncoprotein is aberrantly expressed in AML cells and contributes to activation of the mutant FLT3 receptor. Targeting MUC1-C thus inhibits FLT3 signaling and represents a potential approach for AML cells resistant to FLT3 inhibitors.

Download Full-text

FLT3 Inhibitors in AML

Clinical Lymphoma Myeloma & Leukemia ◽

10.1016/j.clml.2018.06.033 ◽

2018 ◽

Vol 18 ◽

pp. S1-S2

Author(s):

Alexander Perl

Keyword(s):

Flt3 Inhibitors

Download Full-text