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.

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.

The Plk-1 Inhibitor BI 2536 Counteracts the Growth of Neoplastic Mast Cells and Synergizes with the KIT D816V-Targeting Drug Midostaurin (PKC412) in Producing Growth-Inhibition.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3554-3554
Author(s):  
Veronika Ferenc ◽  
Karoline V. Gleixner ◽  
Alexander Gruze ◽  
Michael Kneidinger ◽  
Christian Baumgartner ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Mast Cells ◽  
Growth Inhibition ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Thymidine Uptake ◽  
Dependent Manner ◽  
Ic50 Values ◽  
Bi 2536 ◽  
Kit D816v

Abstract Systemic mastocytosis (SM) is a myeloid neoplasm characterized by abnormal growth and accumulation of mast cells (MC) in various internal organs. In most patients, the D816V-mutated variant of c-KIT, which mediates resistance against several tyrosine kinase (TK) inhibitors like imatinib, is found. In advanced SM, the response of neoplastic MC to conventional drugs is poor and the prognosis is grave. Therefore current research is attempting to identify novel targets in neoplastic MC. Polo-like kinase 1 (Plk-1) is a serine/threonine kinase that plays an essential role in mitosis and has recently been introduced as a new target in myeloid leukemias. In the present study, we analyzed expression and function of Plk-1 in neoplastic human MC, and asked whether Plk-1 can serve as a target of therapy in SM. As determined by immunohistochemistry, primary neoplastic MC were found to display activated/phosphorylated Plk-1 in all patients examined (n=5). The human MC leukemia cell line HMC-1 was also found to exhibit activated Plk-1. In addition, we found that primary neoplastic MC as well as HMC-1 cells express Plk-1 mRNA in RT-PCR experiments. As assessed by 3H-thymidine-uptake experiments, the Plk-1-targeting drug BI 2536 (Boehringer Ingelheim GmbH, Germany) was found to inhibit the proliferation of HMC-1 cells in a dose-dependent manner (IC50 5–15 nM). The effect of BI 2536 was seen in both subclones of HMC-1, i.e. in HMC-1.1 cells displaying KIT G560V (but not KIT D816V), and HMC-1.2 cells exhibiting both KIT G560V and KIT D816V, with comparable IC50 values. Moreover, BI 2536 was found to inhibit the proliferation of primary neoplastic cells, with IC50 values ranging between 5 and 50 nM. The growth-inhibitory effects of BI 2536 on HMC-1 cells were found to be associated with mitotic arrest and G2-M cell cycle arrest as well as consecutive apoptosis. In normal bone marrow or peripheral blood mononuclear cells, neither mitotic cell arrest nor apoptosis were observed after treatment with BI 2536. In a consecutive phase of the study, we asked whether combined targeting of KIT D816V and Plk-1 would lead to synergistic drug-interactions. For this purpose, HMC-1 cells and primary neoplastic MC were coincubated with BI 2536 and midostaurin (PKC412), a multitargeted kinase inhibitor that blocks KIT D816V TK activity. In these experiments, BI 2536 was found to synergize with midostaurin in counteracting the proliferation of HMC-1 cells and primary neoplastic MC. In conclusion, our data show that activated Plk-1 is detectable in MC neoplasms and plays a role in cell cycle progression and viability of neoplastic MC. Targeting of Plk-1 with BI 2536 leads to growth inhibition and apoptosis in neoplastic MC. Furthermore, BI 2536 synergizes with midostaurin in counteracting growth of neoplastic MC. Targeting of Plk-1 may be an attractive new pharmacologic concept 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.

Dasatinib Inhibits the Growth of Neoplastic Human Eosinophils (EOL-1) through Targeting of FIP1L1-PDGFRα.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3559-3559
Author(s):  
Christian Baumgartner ◽  
Karoline V. Gleixner ◽  
Alexander Gruze ◽  
Puchit Samorapoompichit ◽  
Harald Esterbauer ◽  
...  
Keyword(s):  
Kinase Inhibitor ◽  
Systemic Mastocytosis ◽  
Leukemia Cell ◽  
Malignant Cell ◽  
Organ Damage ◽  
Leukemia Cell Line ◽  
Dependent Manner ◽  
Growth And Survival ◽  
Chronic Eosinophilic Leukemia ◽  
Dose Dependent

Abstract Chronic eosinophilic leukemia (CEL) is a myeloproliferative disorder characterized by molecular and/or cytogenetic evidence of monoclonality of eosinophils, sustained marked eosinophilia, and consecutive organ damage. In a majority of patients with CEL with or without associated mastocytosis, the transforming mutation FIP1L1-PDGFRα and the related CHIC2 deletion is found. The respective oncoprotein, FIP1L1-PDGFRα, is considered to play a major role in malignant cell growth in CEL. The tyrosine kinase (TK) inhibitor imatinib (STI571) has been described to counteract the TK activity of FIP1L1-PDGFRα in most patients, and has been introduced as a novel effective therapy in CEL. However, not all patients with CEL show a response to imatinib. Therefore, several attempts have been made to identify other TK inhibitors that counteract growth of neoplastic eosinophils in CEL. We provide evidence that dasatinib, a multi-targeted kinase inhibitor, blocks the growth and survival of EOL-1, an eosinophil leukemia cell line carrying FIP1L1-PDGFRα. The effects of dasatinib on proliferation of EOL-1 cells were dose-dependent, with an IC50 of 0.5–1 nM, that was found to be in the same range compared to IC50 values produced by imatinib. Dasatinib was also found to induce apoptosis in EOL-1 cells in a dose-dependent manner (IC-50: 1–10 nM). The apoptosis-inducing effects of dasatinib on EOL-1 cells were demonstrable by light microscopy, flow cytometry, and by a Tunel assay. To further examine the mechanism of growth inhibition induced by dasatinib in neoplastic eosinophils, Western blot experiments were performed using antibodies directed against phosphorylated or total PDGFRα. In these experiments, we were able to show that dasatinib at 1 μM completely blocks the phosphorylation of FIP1L1-PDGFRα in EOL-1 cells. In summary, our data show that dasatinib inhibits the growth of leukemic eosinophils through targeting of the TK activity of the disease-related oncoprotein FIP1L1-PDGFRα. Based on this observation, dasatinib may be considered as a new interesting treatment option for patients with CEL. As dasatinib is also known to block various KIT mutants as well as wild type KIT, such therapy may also be of interest for patients who have systemic mastocytosis (SM) with an associated CEL (SM-CEL).

The Multi-Kinase Inhibitor DCC-2618 Inhibits Proliferation and Survival of Neoplastic Mast Cells and Other Cell Types Involved in Systemic Mastocytosis

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1965-1965 ◽  
Author(s):  
Mathias A Schneeweiss ◽  
Barbara Peter ◽  
Katharina Blatt ◽  
Daniela Berger ◽  
Gabriele Stefanzl ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Mast Cells ◽  
Cell Line ◽  
Cell Lines ◽  
Research Funding ◽  
Kinase Inhibitor ◽  
Vascular Endothelial Cells ◽  
Systemic Mastocytosis ◽  
Leukemia Cell ◽  
Kit D816v

Abstract Systemic mastocytosis (SM) is a myeloid neoplasm defined by abnormal growth and pathologic accumulation of neoplastic mast cells (MC) in various internal organs. The indolent variant of SM (ISM) is associated with an almost normal life expectancy. By contrast, the prognosis in advanced SM, including SM with an associated hematologic neoplasm (AHN), aggressive SM (ASM), and MC leukemia (MCL) is poor with short survival times. Most patients with SM express the D816V-mutated variant of KIT, which confers resistance against several tyrosine kinase inhibitors (TKI), including imatinib. Midostaurin is a TKI that is effective against KIT D816V. However, despite encouraging clinical efficacy, this drug cannot produce continuous complete remission in all patients. One problem in advanced SM is that the AHN component of the disease, especially when progressing into acute myeloid leukemia (AML) is often drug-resistant. The aims of this study were to evaluate the effects of the multi-kinase inhibitor DCC-2618 on proliferation and survival of primary neoplastic mast cells, various mast cell lines and other malignant and non-malignant cell types that may play a role in advanced SM. As assessed by 3H-thymidine-uptake, DCC-2618 was found to inhibit the proliferation of all human MC lines tested, with lower IC50 values measured in HMC-1.1 cells lacking KIT D816V (11.2±4.3 nM) and ROSAKIT WT cells (61±11 nM) than in KIT D816V+ HMC-1.2 cells (147±68 nM) and ROSAKIT D816V cells (133±43 nM). DCC-2618 also produced growth inhibition in the multi-resistant MCL lines MCPV-1.1 (164±72 nM), MCPV-1.2 (256±167 nM), MCPV-1.3 (124±46 nM), and MCPV-1.4 (235±114 nM). In addition, DCC-2618 was found to inhibit the proliferation of primary neoplastic bone marrow MC obtained from patients with SM including MCL (Figure). We also found that DCC-2618 induces apoptosis in HMC-1 cells and ROSA cells, and to a lesser degree in MCPV-1 cells as determined by light microscopy and AnnexinV/PI staining. Moreover, DCC-2618 was found to block phosphorylation of KIT in all MC lines tested. In a next step, we explored the effects of DCC-2618 on growth of other leukemia cell lines as well on vascular endothelial cells. In these experiments, we were able to show that DCC-2618 inhibits the proliferation of the FIP1L1-PDGFRA+ eosinophilic leukemia cell line EOL-1 (IC50 2±0.6 nM) and the FLT3 ITD-mutated AML cell lines MV4-11 (IC50 130±18 nM) and MOLM-13 (IC50 110±26 nM). DCC-2618 also induced apoptosis in EOL-1, MV-411, and MOLM-13 cells. Moreover, DCC-2618 was found to inhibit the growth of cultured human vascular endothelial cells, suggesting that the drug may also counteract SM-related neo-angiogenesis in SM. DCC-2618 did not inhibit the proliferation of the immature AML cell line KG1 and the monoblastic cell line U937, but was found to block proliferation in primary leukemic monocytes in patients with monoblastic AML or chronic myelomonocytic leukemia (CMML) which may have clinical implications as CMML and AML are the most prevalent types of AHN in advanced SM. Finally, we were able to show that the major DCC-2618-metabolite, DP-5439, is equally effective in producing growth inhibition in all cell lines tested as well as in primary neoplastic MC compared to DCC-2618 (Figure). In summary, our data show that DCC-2618 is a new potent multi-targeted TKI that counteracts growth of neoplastic MC as well as growth and survival of leukemic monocytes, AML blasts, eosinophils, and endothelial cells in vitro. Whether DCC-2618 is also able to inhibit the growth of neoplastic MC and other leukemic (AHN) cells in vivo in patients with advanced SM remains to be determined in clinical trials. Indeed, a first Phase I clinical trial examining the effects of DCC-2618 in SM has recently been initiated. Figure Figure. Disclosures Valent: Novartis: Honoraria, Research Funding; Amgen: Honoraria; Celgene: Honoraria, Research Funding; Ariad: Honoraria, Research Funding; Deciphera Pharmaceuticals: Research Funding.

SL-401, a Targeted Therapy Directed To The Interleukin-3 Receptor (IL-3R), Possesses Preclinical Cytotoxic Activity Against Chronic Eosinophilic Leukemia

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4104-4104 ◽  
Author(s):  
Christopher Brooks ◽  
Vincent Macri ◽  
Arnaldo Albini ◽  
Ivan Bergstein ◽  
Eric Rowinsky
Keyword(s):  
Cell Line ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Severe Toxicity ◽  
Dependent Manner ◽  
Employment Equity ◽  
Chronic Eosinophilic Leukemia ◽  
Familial Form ◽  
Equity Ownership

Abstract Background Hypereosinophilic syndrome (HES), a rare myeloproliferative disorder characterized by a persistently elevated eosinophil count, results in damage to the heart, lungs, peripheral nervous system, and other organs. An acquired (non-familial) form of HES is particularly aggressive and debilitating. Acquired forms of HES are subclassified as secondary (reactive), idiopathic, and clonal HES, the latter often transitioning into chronic eosinophilic leukemia (CEL), or hypereosinophilic leukemia, which can result in myocardial fibrosis and congestive heart failure. Patients may respond to corticosteroids, the monoclonal antibodies mepolizumab and alemtuzumab, and the tyrosine kinase inhibitor imatinib, which is registered for HES patients who express the FIP1L1-PDGFRA fusion protein. However, several of the aforementioned agents also induce severe toxicity. Since eosinophils ubiquitously express the IL-3R, SL-401, a novel IL-3R-targeted therapeutic, which is comprised of IL-3 conjugated to a truncated diphtheria toxin, represents a rational and selective agent for development in hypereosinophilic disorders. Methods Expression of the IL-3R (CD123) on a human eosinophilic leukemia cell line (EOL-1) was determined by flow cytometry. Cells were washed with phosphate-buffered saline (PBS) and stained with anti-CD123 PE (BD Biosciences) and control IgG for 20 minutes at 4oC and then washed with PBS again. Stained cells were acquired using the LSR II (BD) cytometer and data were analyzed using Flow Jo (Tree Star). To analyze patient samples, PBMCs were prepared and stained using a similar method and then analyzed using Cytopaint software. The sensitivity of the cells to SL-401 was assessed using a CellTiter Glo in vitro cytotoxicity assay. A CD123 positive leukemia cell line (TF-1), which is known to be sensitive to SL-401, was used as a positive control. The cells were cultured in the presence of absence of SL-401 for 48 hours and assessed for viability at concentrations ranging from 5x10-6 to 21 µg/ml (8.7x10-5 to 368 nM). Results Primary eosinophils, elevated in number and CD16 negative, were harvested from a patient with HES and found to be positive for IL-3R. EOL-1 cells, similarly, were found to express high levels of IL-3R, with greater than 98% of cells expressing the receptor compared to isotype control. Based on the IL-3R expression pattern, the sensitivity of EOL-1 cells to SL-401 was then tested. Cells were shown to be highly sensitive to SL-401 after 48-hour incubation in a concentration-dependent manner, with IC50 values of 1, 0.6, and 0.47 pM from triplicate experiments (mean: 0.69 pM). Cell viability was also reduced by 96% at 48 hours post-treatment with a picomolar concentration of SL-401 (350 pM). Conclusion These results indicate SL-401 possesses potent in vitro anti-cancer activity against CEL, which expresses high levels of the IL-3R. Importantly, SL-401 exhibited potent activity against these cells at concentrations that were significantly lower (>10-fold) than peak plasma concentrations achieved in clinical studies. These findings, together with the need for novel clinical strategies to treat CEL and HES, warrant further exploration of SL-401 for the treatment of patients with hypereosinophilic diseases. Disclosures: Brooks: Stemline Therapeutics: Employment, Equity Ownership. Macri:Stemline Therapeutics, Inc., New York, NY USA : Employment. Bergstein:Stemline Therapeutics: Employment, Equity Ownership, Patents & Royalties. Rowinsky:Stemline Therapeutics: Employment, Equity Ownership, Membership on an entity’s Board of Directors or advisory committees.

The Human Leukemic HL-60 Cell Line: An in Vitro Model for Cell Death Endpoint Identification

1996 ◽  
Vol 24 (4) ◽  
pp. 581-587
Author(s):  
Cristiana Zanetti ◽  
Arrnalaura Stammati ◽  
Orazio Sapora ◽  
Flavia Zucco
Keyword(s):  
Cell Death ◽  
Cell Line ◽  
In Vitro Model ◽  
Leukemia Cell ◽  
Promyelocytic Leukemia ◽  
Leukemia Cell Line ◽  
Cell Type ◽  
Vitro Model ◽  
Promyelocytic Leukemia Cell Line

The aim of this study was to investigate the endpoints related to cell death, either necrosis or apoptosis, induced by four chemicals in the promyelocytic leukemia cell line, HL-60. Cell morphology, DNA fragmentation, cytofluorimetric analysis and oxygen consumption were used to classify the type of cell death observed. In our analysis, we found that not all the selected parameters reproduced the differences observed in the cell death caused by the four chemicals tested. As cell death is a very complex phenomenon, several factors should be taken into account (cell type, exposure time and chemical concentration), if chemicals are to be classified according to differences in the mechanisms more directly involved in cell death.

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 Human Platelets Effectively Kill K-562 Cells, a Chronic Myelogenic Leukemia Cell Line,in vitro

1990 ◽  
Vol 81 (5) ◽  
pp. 449-453 ◽  
Author(s):  
Terutaka Sagawa ◽  
Takeshi Kodama ◽  
Akio Tominaga ◽  
Mariko Okada
Keyword(s):  
Cell Line ◽  
Leukemia Cell ◽  
Human Platelets ◽  
Leukemia Cell Line ◽  
K 562 Cells

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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