Identification of Mcl-1 as a Novel Target in Neoplastic Mast Cells and Demonstration of Cooperative Growth-Inhibitory Effects of mcl-1 Antisense Oligonucleotides, PKC412, and AMN107.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3516-3516 ◽  
Author(s):  
Karl J. Aichberger ◽  
Matthias Mayerhofer ◽  
Karoline V. Gleixner ◽  
Maria-Theresa Krauth ◽  
Sophia Derdak ◽  
...  
Keyword(s):  
Mast Cells ◽  
Antisense Oligonucleotides ◽  
Kinase Inhibitors ◽  
Systemic Mastocytosis ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Apoptotic Cells ◽  
Kit Mutation ◽  
Myeloid Neoplasm ◽  
Novel Target

Abstract Mcl-1 is a Bcl-2 family-member that has been described to act anti-apoptotic in various myeloid neoplasms and therefore has been proposed as a potential therapeutic target. Systemic mastocytosis (SM) is a myeloid neoplasm involving myelomastocytic progenitors. We examined the expression and functional role of Mcl-1 in neoplastic mast cells (MC), to determine whether Mcl-1 could serve as a target in MC neoplasms. As assessed by RT-PCR and immunohistochemistry, primary neoplastic MC were found to express Mcl-1 mRNA and the Mcl-1 protein in a constitutive manner in all patients analyzed. Moreover, the human MC-leukemia cell line HMC-1 was found to express Mcl-1. Transfection of these cells with Mcl-1-specific antisense oligonucleotides (ASO) or an mcl-1-specific siRNA using lipofectin resulted in a reduced survival and increased percentage of apoptotic cells compared to control. The effects of mcl-1 ASO were seen with the HMC-1.1 subclone carrying the G560V c-kit mutation (mcl-1 ASO, 250 nM: 49±4% apoptotic cells compared to control: 3±2%, p<0.05; mcl-1 siRNA: 41±5% vs control: 5±3%, p<0.05) as well as with HMC-1.2 cells carrying both the G560V c-kit mutation and the D816V c-kit mutation (mcl-1 ASO, 250 nM: 36±2% apoptotic cells compared to control: 6±1%, p<0.05; mcl-1 siRNA: 30±6% vs control: 5±2%, p<0.05). Moreover, mcl-1 ASO were found to cooperate with the tyrosine kinase inhibitors (Novartis Pharma AG) imatinib, AMN107, and PKC412 in producing growth inhibition in HMC-1.2 cells. Together, these data show that Mcl-1 is a novel survival factor and attractive target in neoplastic human MC. Whether the Mcl-1-targeting concept can be developed far enough to reach clinical application remains to be elucidated.

Identification of MCL1 as a novel target in neoplastic mast cells in systemic mastocytosis: inhibition of mast cell survival by MCL1 antisense oligonucleotides and synergism with PKC412

Blood ◽  
2006 ◽  
Vol 109 (7) ◽  
pp. 3031-3041 ◽  
Author(s):  
Karl J. Aichberger ◽  
Matthias Mayerhofer ◽  
Karoline V. Gleixner ◽  
Maria-Theresa Krauth ◽  
Alexander Gruze ◽  
...  
Keyword(s):  
Mast Cells ◽  
Antisense Oligonucleotides ◽  
Kinase Inhibitors ◽  
Systemic Mastocytosis ◽  
Synergistic Effects ◽  
Kit Mutation ◽  
Myeloid Neoplasm ◽  
Novel Target ◽  
Kit D816v

Abstract MCL-1 is a Bcl-2 family member that has been described as antiapoptotic in various myeloid neoplasms. Therefore, MCL-1 has been suggested as a potential new therapeutic target. Systemic mastocytosis (SM) is a myeloid neoplasm involving mast cells (MCs) and their progenitors. In the present study, we examined the expression and functional role of MCL-1 in neoplastic MCs and sought to determine whether MCL-1 could serve as a target in SM. As assessed by RT-PCR and immunohistochemical examination, primary neoplastic MCs expressed MCL-1 mRNA and the MCL-1 protein in all SM patients examined. Moreover, MCL-1 was detectable in both subclones of the MC line HMC-1—HMC-1.1 cells, which lack the SM-related KIT mutation D816V, and HMC-1.2 cells, which carry KIT D816V. Exposure of HMC-1.1 cells or HMC-1.2 cells to MCL-1–specific antisense oligonucleotides (ASOs) or MCL-1–specific siRNA resulted in reduced survival and increased apoptosis compared with untreated cells. Moreover, MCL-1 ASOs were found to cooperate with various tyrosine kinase inhibitors in producing growth inhibition in neoplastic MCs, with synergistic effects observed with PKC412, AMN107, and imatinib in HMC-1.1 cells and with PKC412 in HMC-1.2 cells. Together, these data show that MCL-1 is a novel survival factor and an attractive target in neoplastic MCs.

Delineation of a KIT-Independent Oncogenic Pathway in Neoplastic Mast Cells That Involves Lyn and Btk, and Can Be Disrupted by the KIT/Lyn/Btk-Targeting Drug Dasatinib

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1541-1541 ◽  
Author(s):  
Karoline V. Gleixner ◽  
Matthias Mayerhofer ◽  
Uwe Rix ◽  
Gregor Hoermann ◽  
Alexander Gruze ◽  
...  
Keyword(s):  
Mast Cells ◽  
Potent Inhibitor ◽  
Systemic Mastocytosis ◽  
Synergistic Effects ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Growth And Survival ◽  
Myeloid Neoplasm ◽  
Oncogenic Pathway ◽  
Kit D816v

Abstract Systemic mastocytosis (SM) is a myeloid neoplasm characterized by increased growth and survival of neoplastic mast cells (MC). Aggressive SM (ASM) and MC leukemia (MCL) are advanced disease variants that usually are drug-resistant and have an unfavorable prognosis. In most patients, the D816V-mutated ′oncogenic′ variant of KIT is detectable. However, the mutant is also detectable in patients with indolent SM exhibiting a normal life-expectancy, and therefore is not considered to represent a fully transforming oncoprotein. This assumption is also supported by studies in Ba/F3 cells, and whether KIT D816V-targeting drugs are able to induce long-term remission in ASM or MCL, remains to be seen. Therefore, it has been hypothesized that in addition to KIT, other pro-oncogenic molecules and signaling pathways play a role in malignant transformation/progression in SM. We here describe a novel KIT D816V-independent oncogenic pathway in neoplastic MC that involves Lyn and Bruton’s tyrosine kinase (Btk). Western blotting and immunostaining revealed that neoplastic MC display the Btk- and Lyn protein. Both molecules were found to be constitutively phosphorylated in primary neoplastic MC and in the MC leukemia cell line HMC-1. Lyn/Btk-activation was not only detectable in KIT D816V-positive HMC-1.2 cells, but also in the KIT D816V-negative HMC-1.1 subclone. In studies employing Ba/F3 cells with doxycycline-inducible expression of KIT, we were able to show that KIT D816V induces activation of STAT5 and Akt, but does not induce activation of Btk. Correspondingly, pharmacologic deactivation/dephosphorylation of KIT in HMC-1 cells by midostaurin (PKC412) (Novartis, Basel, Switzerland) was not accompanied by a decrease in phosphorylation of Lyn or Btk. The functional significance of Btk expression/activation in neoplastic MC could be demonstrated by a Btk-specific siRNA that reduced the proliferation and survival in HMC-1 cells, and was found to cooperate with midostaurin in producing growth inhibition. In consecutive experiments, we identified the Src/Abl kinase-targeting drug dasatinib (BMS, Princeton, NJ) as a potent inhibitor of Lyn/Btk activation in neoplastic MC. In particular, dasatinib (1 μM) was found to block Lyn and Btk activity in HMC-1.1 cells as well as in HMC-1.2 cells, and corresponding results were obtained with primary neoplastic MC. Finally, as assessed by a chemical proteomics approach, we were able to show that dasatinib directly binds to Btk and Lyn in neoplastic MC. In summary, our data show that a KIT-independent Lyn/Btk-driven signaling pathway contributes to growth and survival of neoplastic MC, and possibly to disease progression in SM. Our study also identifies dasatinib as a potent inhibitor of the Lyn/Btk pathway, which may have clinical implications and may explain some of the synergistic effects obtained with combinations of dasatinib and other KIT-targeting TK inhibitors in neoplastic MC.

PKC412 Inhibits In Vitro Growth of Neoplastic Mast Cells Expressing the D816V-Mutated Variant of KIT: Comparison with AMN107 and Imatinib, and Evaluation of Drug-Interactions.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3523-3523
Author(s):  
Karoline V. Gleixner ◽  
Matthias Mayerhofer ◽  
Karl J. Aichberger ◽  
Sophia Derdak ◽  
Karoline Sonneck ◽  
...  
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
Drug Interactions ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Human Mast Cell ◽  
Cell Leukemia ◽  
Kit Mutation ◽  
Mast Cell Leukemia ◽  
Kit D816v

Abstract In most patients with systemic mastocytosis (SM) including aggressive SM and mast cell leukemia (MCL), neoplastic cells express the oncogenic c-KIT mutation D816V. KIT-D816V is associated with constitutive tyrosine kinase (TK) activity and thus represents an attractive target of drug therapy. However, most available TK inhibitors including STI571=imatinib, fail to block TK-activity of KIT D816V at pharmacologic concentrations. We provide evidence that the novel TK-targeting drugs PKC412 and AMN107 decrease TK-activity of D816V-mutated KIT and counteract growth of Ba/F3 cells with doxycycline-induced expression of KIT D816V as well as growth of the human mast cell leukemia cell line HMC-1 expressing this c-KIT mutation. PKC412 was found to be the superior drug with IC50 values of 50–250 nM and without differences seen between HMC-1 cells exhibiting or lacking KIT D816V. By contrast, AMN107 exhibited potent effects only in the absence of KIT D816V in HMC-1 cells. Corresponding results were obtained with Ba/F3 cells exhibiting wild-type or the D816V-mutated variant of KIT. Moreover, we found that PKC412 and AMN107 inhibit growth of primary neoplastic MC in a patient with KIT D816V+ SM. The growth-inhibitory effects of PKC412 and AMN107 on HMC-1 cells were associated with TK-inhibition of KIT and with induction of apoptosis. In addition, PKC412 was found to downregulate expression of CD2 and CD63, two cell surface antigens upregulated in SM. In co-incubation experiments, PKC412 was found to synergize with AMN107, imatinib, and 2CdA in producing growth inhibition in HMC-1 cells lacking KIT D816V, whereas in KIT D816V+ HMC-1 cells, drug-interactions were additive rather than synergistic. Together, PKC412 and AMN107 alone and in combination counteract growth of neoplastic mast cells. Both drugs may therefore be considered as novel promising agents for targeted therapy in patients with aggressive SM or MCL.

Effects of the Mcl-1/Bcl-2 Inhibitor GX015-070 (Obatoclax®) on Growth and Viability of Canine and Human Neoplastic Mast Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 861-861
Author(s):  
Barbara Peter ◽  
Karl J. Aichberger ◽  
Karoline V. Gleixner ◽  
Veronika Ferenc ◽  
Alexander Gruze ◽  
...  
Keyword(s):  
Mast Cells ◽  
Cell Line ◽  
Systemic Mastocytosis ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Thymidine Uptake ◽  
Dependent Manner ◽  
Growth And Survival ◽  
Kit D816v

Abstract Mcl-1 is a Bcl-2 family-member that has been described to act anti-apoptotic in various myeloid neoplasms. We and others have recently shown that neoplastic mast cells (MC) in patients with systemic mastocytosis (SM) display Mcl-1, Bcl-2, and Bcl-xL. In the present study, we examined the effects of the Mcl-1/Bcl-2-targeting drug GX015-070 (obatoclax®; GeminX, Montréal, Quebéc, Canada) on growth and viability of primary neoplastic MC obtained from patients with SM (n=3), the human MC leukemia cell line HMC-1, and the canine mastocytoma cell line C2. Two HMC-1 subclones, one lacking KIT D816V (HMC- 1.1) and one expressing KIT D816V (HMC-1.2) were examined. As assessed by RT-PCR and immunostaining, primary neoplastic MC as well as HMC-1 cells (both subclones) were found to express Mcl-1 mRNA and the Mcl-1 protein in a constitutive manner, but did not express significant amounts of proapoptotic Bim. Transfection of HMC-1 cells with Mcl-1-specific siRNA resulted in reduced proliferation and increased apoptosis compared to cells transfected with a control siRNA. GX015-070 was found to inhibit 3H-thymidine uptake and thus proliferation in HMC-1 cells in a dose-dependent manner, with higher IC50 values obtained in HMC-1.2 cells (0.5 μM) compared to HMC-1.1 cells (0.05 μM). GX015-070 also inhibited the growth and survival in the canine mastocytoma cell line C2 (IC50: 0.5-1 μM). Moreover, GX015-070 was found to inhibit the proliferation of primary human neoplastic MC in all SM patients tested (IC50: 0.05-0.1 μM). We next attempted to combine obatoclax with a modulator of Mcl-1/Bim expression in MC, in order to enhance drug effects. Since Bim is degraded via the proteasome, we applied the proteasome inhibitor bortezomib. Whereas GX015-07 did not modulate the production/expression of Mcl-1 or Bim in HMC-1 cells, bortezomib was found to promote the expression of Bim in our Western blot experiments. In addition, bortezomib was found to suppress 3H-thymidine uptake in both HMC-1 subclones. Finally, bortezomib was found to cooperate with GX015-070 in producing apoptosis in HMC-1.1 cells, HMC-1.2 cells, and C2 cells. Together, our data show that the Mcl-1/Bcl-2-targeting drug GX015-070 is a potent inhibitor of in vitro growth and survival of canine and human neoplastic MC. Targeting of Mcl-1 in neoplastic MC alone or in combination with a Bim-regulator may be an interesting pharmacologic approach in advanced SM.

Bosutinib Blocks Lyn and Btk Activation and Synergizes with the KIT D816V-Targeting Drug Midostaurin in Inducing Apoptosis in Neoplastic Human Mast Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1717-1717
Author(s):  
Karoline Veronika Gleixner ◽  
Matthias Mayerhofer ◽  
Gregor Hörmann ◽  
Karina Schuch ◽  
Sabine Cerny-Reiterer ◽  
...  
Keyword(s):  
Mast Cells ◽  
Growth Inhibition ◽  
Research Funding ◽  
Systemic Mastocytosis ◽  
Leukemia Cell ◽  
Target Function ◽  
Leukemia Cell Line ◽  
Thymidine Uptake ◽  
Organ Systems ◽  
Kit D816v

Abstract Abstract 1717 Poster Board I-743 Advanced systemic mastocytosis (SM) is a malignant hematopoietic neoplasm characterized by destructive growth of neoplastic mast cells (MC) in various organ systems. In these patients, the response to conventional cytoreductive therapy is poor and the prognosis is grave. The D816V-mutated variant of c-KIT is found in most patients and is considered to be a major transforming oncoprotein in SM that leads to abnormal survival and growth of neoplastic MC. Therefore, agents interfering with the kinase activity of KIT D816V have been developed. One promising agent is midostaurin (PKC412). However, in most patients with advanced SM, therapy with midostaurin is not sufficient to induce long term remissions. In addition, midostaurin is unable to block all pro-oncogenic signaling molecules, such as Lyn and Btk, in neoplastic MC, suggesting that additional oncoproteins and survival factors may play a role in malignant transformation in SM, and that novel therapeutic strategies are required to block such KIT-independent oncogenic pathways. Especially Lyn and Btk have attracted attention as potential new targets in neoplastic MC. Bosutinib (SKI-606) is a novel multikinase inhibitor that targets a broad spectrum of kinases including Lyn and Btk. The aim of the current study was to evaluate the effect of bosutinib on neoplastic MC, and potential cooperative drug interactions between bosutinib and midostaurin. As assessed by 3H-thymidine uptake, bosutinib was found to inhibit the growth of the MC leukemia cell line HMC-1, including the HMC-1.1 subclone that lacks KIT D816V and HMC-1.2 cells expressing KIT D816V, with similar IC50 values (1-5 μM). Furthermore, bosutinib was found to induce apoptosis in both HMC-1 subclones. Growth-inhibitory and apoptosis-inducing effects of bosutinib were also seen in primary neoplastic MC obtained from the bone marrow of patients with SM (n=3). As assessed by phosphoblotting, bosutinib did not inhibit the autophosphorylation of mutant KIT in HMC-1 cells, but was found to completely inhibit the phosphorylation of Lyn and Btk. To confirm the target-function of Lyn and Btk in neoplastic MC, siRNA experiments were performed. Knockdown of Lyn or Btk resulted in induction of apoptosis and growth-inhibition in HMC-1 cells. We next attempted to exploit target-specific and complementing effects of midostaurin and bosutinib by combining both substances. As expected, combined application of bosutinib and midostaurin resulted in a complete inhibition of phosphorylation of KIT, Lyn, and Btk in HMC-1.1 and HMC-1.2 cells. We were also able to show that bosutinib synergizes with midostaurin in inducing apoptosis in both HMC-1 subclones. Synergistic effects were also observed when combining midostaurin with Lyn- or Btk-siRNA. Together, we have identified Lyn and Btk as novel KIT-independent survival molecules in neoplastic MC. Inhibition of these kinases by siRNA-knockdown or by bosutinib leads to growth-inhibition and apoptosis. Synergistic pro-apoptotic effects were observed with the combination “bosutinib + midostaurin”, suggesting that simultaneous targeting of KIT and Lyn/Btk may be a powerful strategy to counteract the survival of neoplastic MC. This drug combination may therefore be an interesting approach to overcome drug-resistance in advanced forms of SM. Disclosures Valent: Bristol Myers Squibb: Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding.

The Midostaurin (PKC412) Metabolite CGP52421 Shows Little Growth-Inhibitory Activity Against Against Neoplastic Mast Cells but Retains Inhibitory Effects on IgE-Dependent Activation and Histamine Release

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1417-1417 ◽  
Author(s):  
Barbara Peter ◽  
Katharina Blatt ◽  
Gabriele Stefanzl ◽  
Emir Hadzijusufovic ◽  
Catherine Dutreix ◽  
...  
Keyword(s):  
Mast Cells ◽  
Histamine Release ◽  
Clinical Symptoms ◽  
Systemic Mastocytosis ◽  
Leukemia Cell ◽  
Latency Period ◽  
Leukemia Cell Line ◽  
Active Metabolites ◽  
Multikinase Inhibitor ◽  
Ic50 Values

Abstract Abstract 1417 The multikinase inhibitor midostaurin (PKC412) is currently tested in clinical trials in patients with advanced systemic mastocytosis (SM). Although clinical symptoms improve in many patients and sometimes the proliferation of neoplastic mast cells (MC) can be kept under control for some time, most patients progress after a variable latency period, even if their mediator-related symptoms are still completely suppressed. In vivo, midostaurin is metabolized to two major and active metabolites, namely CGP62221 and CGP52421. Whereas the in vitro effects of midostaurin on growth and activation of MC are well documented, only little data on effects of midostaurin-metabolites are available. We examined the effects of midostaurin and its pharmacologically relevant metabolites CGP52421 and CGP62221, on IgE-dependent mediator secretion in basophils as well as growth of primary neoplastic MC and the human MC leukemia cell line HMC-1. All three compounds were found to inhibit IgE-dependent secretion of histamine in blood basophils, with comparable IC50 values (<1 μM). Midostaurin and CGP62221 were also found to inhibit the proliferation in HMC-1.1 cells and HMC-1.2 cells with IC50 values ranging between 50 and 250 nM. However, the second metabolite of PKC412, CGP52421, did not produce comparable anti-proliferative effects, even when applied at concentrations up to 1 μM. Corresponding results were obtained when analyzing drug-effects on primary neoplastic MC obtained from two patients, one with indolent SM and one with aggressive SM. Furthermore, whereas midostaurin and CGP62221 induced apoptosis in HMC-1.1 and HMC-1.2 cells as evidenced by light microscopy, caspase 3 staining and TUNEL assay, CGP52421 did not induce apoptosis in neoplastic MC. Finally, midostaurin and CGP62221 were found to induce dephosphorylation of KIT V560G and KIT D816V in HMC-1 cells, whereas no effects of CGP52421 on KIT activation were seen. In drug competition experiments, the metabolite CGP52421 did not interfere with midostaurin- or CGP62221-induced growth inhibition in HMC-1 cells. In conclusion, the pharmacologically relevant midostaurin metabolite CGP52421 inhibits IgE-mediated histamine release but does not efficiently block KIT-dependent proliferation of neoplastic MC. This observation may help to explain clinical responses including improvement of mediator-related symptoms seen in midostaurin-treated patients with advanced SM, and would favor the development of new treatment concepts employing midostaurin as an inhibitor of both MC activation and MC growth in patients with SM. Disclosures: Dutreix: Novartis: Employment. Gross:Novartis: Employment. Roesel:Novartis Pharma AG: Employment. Valent:Novartis: Consultancy, Honoraria, Research Funding.

Identification of proapoptotic Bim as a tumor suppressor in neoplastic mast cells: role of KIT D816V and effects of various targeted drugs

Blood ◽  
2009 ◽  
Vol 114 (26) ◽  
pp. 5342-5351 ◽  
Author(s):  
Karl J. Aichberger ◽  
Karoline V. Gleixner ◽  
Irina Mirkina ◽  
Sabine Cerny-Reiterer ◽  
Barbara Peter ◽  
...  
Keyword(s):  
Mast Cells ◽  
Tumor Suppressor ◽  
Systemic Mastocytosis ◽  
Leukemia Cell ◽  
Myeloid Neoplasm ◽  
Myeloid Neoplasms ◽  
Leukemia Cell Lines ◽  
Survival And Growth ◽  
Kit D816v

Abstract Systemic mastocytosis (SM) is a myeloid neoplasm involving mast cells (MCs) and their progenitors. In most cases, neoplastic cells display the D816V-mutated variant of KIT. KIT D816V exhibits constitutive tyrosine kinase (TK) activity and has been implicated in increased survival and growth of neoplastic MCs. Recent data suggest that the proapoptotic BH3-only death regulator Bim plays a role as a tumor suppressor in various myeloid neoplasms. We found that KIT D816V suppresses expression of Bim in Ba/F3 cells. The KIT D816–induced down-regulation of Bim was rescued by the KIT-targeting drug PKC412/midostaurin. Both PKC412 and the proteasome-inhibitor bortezomib were found to decrease growth and promote expression of Bim in MC leukemia cell lines HMC-1.1 (D816V negative) and HMC-1.2 (D816V positive). Both drugs were also found to counteract growth of primary neoplastic MCs. Furthermore, midostaurin was found to cooperate with bortezomib and with the BH3-mimetic obatoclax in producing growth inhibition in both HMC-1 subclones. Finally, a Bim-specific siRNA was found to rescue HMC-1 cells from PKC412-induced cell death. Our data show that KIT D816V suppresses expression of proapoptotic Bim in neoplastic MCs. Targeting of Bcl-2 family members by drugs promoting Bim (re)-expression, or by BH3-mimetics such as obatoclax, may be an attractive therapy concept in SM.

PKC412 inhibits in vitro growth of neoplastic human mast cells expressing the D816V-mutated variant of KIT: comparison with AMN107, imatinib, and cladribine (2CdA) and evaluation of cooperative drug effects

Blood ◽  
2006 ◽  
Vol 107 (2) ◽  
pp. 752-759 ◽  
Author(s):  
Karoline V. Gleixner ◽  
Matthias Mayerhofer ◽  
Karl J. Aichberger ◽  
Sophia Derdak ◽  
Karoline Sonneck ◽  
...  
Keyword(s):  
Mast Cells ◽  
Systemic Mastocytosis ◽  
Drug Effects ◽  
Kit Mutation ◽  
Growth Inhibitory ◽  
Ic50 Values ◽  
Tk Inhibitors ◽  
Mast Cell Leukemia ◽  
Kit D816v

AbstractIn most patients with systemic mastocytosis (SM), including aggressive SM and mast cell leukemia (MCL), neoplastic cells express the oncogenic KIT mutation D816V. KIT D816V is associated with constitutive tyrosine kinase (TK) activity and thus represents an attractive drug target. However, imatinib and most other TK inhibitors fail to block the TK activity of KIT D816V. We show that the novel TK-targeting drugs PKC412 and AMN107 counteract TK activity of D816V KIT and inhibit the growth of Ba/F3 cells with doxycycline-inducible expression of KIT D816V as well as the growth of primary neoplastic mast cells and HMC-1 cells harboring this KIT mutation. PKC412 was a superior agent with median inhibitory concentration (IC50) values of 50 to 250 nM without differences seen between HMC-1 cells exhibiting or lacking KIT D816V. By contrast, AMN107 exhibited more potent effects in KIT D816V- HMC-1 cells. Corresponding results were obtained with Ba/F3 cells exhibiting wild-type or D816V-mutated KIT. The growth-inhibitory effects of PKC412 and AMN107 on HMC-1 cells were associated with induction of apoptosis and down-regulation of CD2 and CD63. PKC412 was found to cooperate with AMN107, imatinib, and cladribine (2CdA) in producing growth inhibition in HMC-1, but synergistic drug interactions were observed only in cells lacking KIT D816V. Together, PKC412 and AMN107 represent promising novel agents for targeted therapy of SM. (Blood. 2006;107: 752-759)

scholarly journals (Phenylamino)pyrimidine-1,2,3-triazole derivatives as analogs of imatinib: searching for novel compounds against chronic myeloid leukemia

2021 ◽  
Vol 17 ◽  
pp. 2260-2269
Author(s):  
Luiz Claudio Ferreira Pimentel ◽  
Lucas Villas Boas Hoelz ◽  
Henayle Fernandes Canzian ◽  
Frederico Silva Castelo Branco ◽  
Andressa Paula de Oliveira ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Tyrosine Kinase ◽  
Myeloid Leukemia ◽  
Kinase Inhibitors ◽  
Competitive Inhibition ◽  
Leukemia Cell ◽  
Docking Studies ◽  
Leukemia Cell Line ◽  
Inhibition Mechanism ◽  
Triazole Derivatives

The enzyme tyrosine kinase BCR-Abl-1 is the main molecular target in the treatment of chronic myeloid leukemia and can be competitively inhibited by tyrosine kinase inhibitors such as imatinib. New potential competitive inhibitors were synthesized using the (phenylamino)pyrimidine-pyridine (PAPP) group as a pharmacophoric fragment, and these compounds were biologically evaluated. The synthesis of twelve new compounds was performed in three steps and assisted by microwave irradiation in a 1,3-dipolar cycloaddition to obtain 1,2,3-triazole derivatives substituted on carbon C-4 of the triazole nucleus. All compounds were evaluated for their inhibitory activities against a chronic myeloid leukemia cell line (K562) that expresses the enzyme tyrosine kinase BCR-Abl-1 and against healthy cells (WSS-1) to observe their selectivity. Three compounds showed promising results, with IC50 values between 1.0 and 7.3 μM, and were subjected to molecular docking studies. The results suggest that such compounds can interact at the same binding site as imatinib, probably sharing a competitive inhibition mechanism. One compound showed the greatest interaction affinity for BCR-Abl-1 in the docking studies.

scholarly journals Induction of apoptosis by Kola nut extract as a recent and promising treatment strategy for Leukemia

Bionatura ◽  
2021 ◽  
Vol 6 (2) ◽  
pp. 1725-1732
Author(s):  
Hamdah Alsaeedi ◽  
Rowaid Qahwaji ◽  
Talal Qadah
Keyword(s):  
Cell Cycle ◽  
Cell Line ◽  
Leukemia Cell ◽  
K562 Cells ◽  
Myelogenous Leukemia ◽  
Time Dependent ◽  
Leukemia Cell Line ◽  
Apoptotic Cells ◽  
Dependent Manner ◽  
Anti Cancer

Kola nut extracts have recently been reported to contain chemopreventive compounds providing several pharmacological benefits. This study investigated Kola nut extracts' anti-cancer activity on human immortalized myelogenous leukemia cell line K562 through apoptosis and cell cycle arrest. Fresh Kola nuts were prepared as powder and dissolved in DMSO. Different concentrations (50, 100, 150, 200, and 250 μg/ml) of working solutions were prepared. The K562 cells were treated with the different concentrations of Kola nut extract or vehicle control (10% DMSO) followed by incubation at 37°C for 24, 48, and 72 hours, respectively. Treatment activity was investigated in K562 cells; by Resazurin, and FITC/Propidium Iodide and 7-AAD stained cells to evaluate apoptotic cells and the cell cycle's progression. Inhibition of leukemia cell proliferation was observed. The extract effectively induced cell death, early and late apoptosis by approximately 30% after 24 and 48 hours incubation, and an increase in the rate of dead cells by 50% was observed after 72 hours of incubation. Also, cell growth reduction was seen at high dose concentrations (150 and 200 µg/ml), as evident by cell count once treated with Kola nut extract. The total number of apoptotic cells increased from 5.8% of the control group to 27.4% at 250 µg/ml concentration. Moreover, Kola nut extracts' effects on K562 cells increased gradually in a dose and time-dependent manner. It was observed that Kola nut extracts could arrest the cell cycle in the G2/M phase as an increase in the number of cells by 29.8% and 14.6 % were observed from 9.8% and 5.2% after 24 and 48 hours of incubation, respectively. This increase was detected in a dose and time-dependent manner. Kola nut extracts can be used as a novel anti-cancer agent in Leukemia treatment as it has shown significant therapeutic potential and therefore provides new insights in understanding the mechanisms of its action. Keywords: Kola nut extracts, Leukemia, K562 cell line, Apoptosis, Cancer.

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