The Aurora-Kinase Inhibitor R763/AS703569 Exerts Major Growth-Inhibitory and Apoptosis-Inducing Effects on Neoplastic Mast Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3972-3972 ◽  
Author(s):  
Barbara Peter ◽  
Harald Herrmann ◽  
Karoline V. Gleixner ◽  
Emir Hadzijusufovic ◽  
Sylvia Laffer ◽  
...  
Keyword(s):  
Mast Cell ◽  
Research Funding ◽  
Aurora Kinase ◽  
Cell Leukemia ◽  
Aurora Kinase A ◽  
Aurora Kinase B ◽  
Growth Inhibitory ◽  
Mast Cell Leukemia ◽  
Kit D816v ◽  
Kinase A

Abstract Abstract 3972 Systemic mastocytosis (SM) is a myeloid neoplasm defined by abnormal growth and accumulation of neoplastic mast cells (MC) in one or more internal organs. In most patients, the D816V-mutated variant of KIT is detectable. This mutant supposedly confers resistance against several tyrosine kinase inhibitors including imatinib and masitinib. In aggressive SM (ASM) or mast cell leukemia (MCL) the response to conventional drugs is poor and the prognosis is grave. In these patients, additional KIT-independent signalling pathways and molecules, such as BTK and LYN may play an important role in disease evolution and MC proliferation. R763/AS703569 is a multikinase inhibitor that blocks the kinase activity of KIT, BTK, LYN, Aurora-Kinase-A, Aurora-Kinase-B, ABL, AKT, and FLT3. We analyzed the effects of R763/AS703569 on growth and survival of the human mast cell leukemia cell line HMC-1 and the canine mastocytoma cell line C2. Two subclones of HMC-1 were used, one expressing KIT D816V (HMC-1.2) and one lacking KIT D816V (HMC-1.1). Both HMC-1 subclones were found to express Aurora-Kinase-A mRNA and Aurora-Kinase-B mRNA in RT-PCR experiments. As assessed by 3H-thymidine uptake, R763/AS703569 was found to inhibit proliferation of HMC-1 cells in a dose-dependent manner, with lower IC50 values obtained in HMC-1.2 cells (1-5 nM) compared to HMC-1.1 cells (10-10-50 nM). Moreover, R763/AS703569 produced growth inhibition in C2 cells (IC50: 1–5 nM). As assessed by light microscopy and Tunel assay, the growth-inhibitory effects of R763/AS703569 were found to be accompanied by apoptosis in all three cell lines. Correspondingly, R763/AS703569 was found to induce cleavage of caspase-3, caspase-8, and caspase-9 in HMC-1 cells. Moreover, R763/AS703569 was found to induce a G2/M cell cycle arrest in HMC-1 cells and C2 cells after 24 hours. In order to define the target spectrum for R763/AS703569 in HMC-1 cells, Western blot experiments were performed. In these experiments, R763/AS703569 was found to inhibit the phosphorylation of KIT, Aurora-Kinase-A, and BTK in HMC-1.1 cells, whereas no effects of R763/AS703569 on phosphorylation of LYN were seen. We then combined R763/AS703569 with dasatinib, a drug known to block LYN activation in HMC-1 cells. In these experiments, we were able to show that both drugs cooperate with each other in inducing apoptosis in HMC-1.1 cells and HMC-1.2 cells. In summary, our data suggest that R763/AS703569 is a novel promising drug that should be tested for its anti-neoplastic effects in patients with ASM and MCL in clinical trials. Complete inhibition of growth of neoplastic MC may require drug combinations employing R763/AS703569 and other targeted or cytotoxic drugs. Disclosures: Sarno: Merck-Serono: Employment. Valent:Novartis: Research Funding; Bristol-Myers Squibb: Research Funding; Merck-Serono: Research Funding.

scholarly journals Mast Cell Leukemia: Clinical Heterogeneity, Molecular Aberrations, Treatment Responses, Survival, and Prognostic Factors

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3109-3109 ◽  
Author(s):  
Mohamad Jawhar ◽  
Juliana Schwaab ◽  
Manja Meggendorfer ◽  
Nicole Naumann ◽  
Hans-Peter Horny ◽  
...  
Keyword(s):  
Mast Cell ◽  
Prognostic Factors ◽  
Mast Cells ◽  
Confidence Interval ◽  
Research Funding ◽  
Molecular Characteristics ◽  
Cell Leukemia ◽  
Mast Cell Leukemia ◽  
Treatment Responses ◽  
Kit D816v

Abstract Mast cell leukemia (MCL) is a rare variant of advanced systemic mastocytosis (advSM) characterized by ≥20% mast cells (MCs) in a bone marrow (BM) smear. Our current knowledge of MCL, including clinical and molecular characteristics, treatment options, survival, and prognostic factors is limited to case reports, small case-series and/or literature reviews. While the KIT D816V mutation is present in >80-90% of patients in other SM subtypes, it has only been reported in approximately 50% of patients with MCL. Of interest, recent data have highlighted that the molecular pathogenesis of advSM/MCL is complex. In particular, additional mutations in SRSF2, ASXL1 or RUNX1 (S/A/Rpos), seen in 60-70% of advSM patients, have a significant adverse impact on disease phenotype and prognosis (Jawhar et al., Leukemia, 2016). Here, we sought to evaluate a) relevant clinical and molecular characteristics, b) treatment responses, and c) survival and prognostic factors in 28 MCL patients (median age 67 years; range, 45-82; male 57%), enrolled in the 'German Registry of Disorders on Eosinophils and Mast Cells'. The median percentages of MC in BM smears and trephine biopsies were 25% (range, 20-95) and 65% (range, 20-100; 82% ≥50%), respectively. MC in peripheral blood (PB) ≥10% (leukemic MCL) were seen in only 2/28 patients. Median serum tryptase level was 550 µg/L (range, 160-1850; 93% ≥200, normal value <11.4). An associated hematologic neoplasm (AHN), e.g. CMML (n=7), MDS/MPNu (n=6), MDS (n=5) or CEL (n=2), was diagnosed in 20/28 (71%) patients. Primary MCL was diagnosed in 16/28 (57%) patients and secondary MCL evolving from other advSM subtypes (SM-AHN, n=10; aggressive SM, n=2) in 12/28 (43%) patients with a median of 18 months (range, 4-71) to transformation. Hematologic C-findings such as hemoglobin <10 g/dL and/or platelets <100x109/L were identified in 26/28 (93%) patients. Non-hematologic signs of organ dysfunction included elevated alkaline phosphatase (AP), seen in 20/28 patients (71%, median 181; range 59-548) and splenomegaly in 28/28 (100%) patients. Spleen volumetry results obtained by magnetic resonance imaging were available in 16 patients and showed marked splenomegaly (≥1200 mL) in 8/16 cases (50%). Mutations in KIT were identified in 25/28 (89%) patients (D816V, n=19; D816H, n=3; D816Y, n=2; F522C, n=1) with a median KIT D816V expressed allele burden of 43% (range 20-98) in peripheral blood as measured by quantitative RT-PCR (RT-qPCR). S/A/Rpos were identified in 13/25 (52%) patients (by NGS analyses of 18 myeloid genes). Median observation from the time of MCL diagnosis was 13 months (range, 2-86) and 18/28 patients (64%) died with a median OS of 17 months (95% confidence interval [CI], 10-24). Cytoreductive treatment included midostaurin (n=13), cladribine followed by midostaurin or vice versa (n=9), cladribine (n=3), midostaurin and/or cladribine followed by intensive chemotherapy (n=3) with (n=1) or without (n=2) allogeneic stem cell transplantation. The median overall survival (OS) was 17 months (95% confidence interval, CI [10-24]) with a 2-year OS probability of 24% for all patients. In univariate analyses of multiple clinical, laboratory and molecular variables only bicytopenia (hemoglobin <10 g/dL and platelets <100x109/L, n=13 vs. hemoglobin ≥10 g/dL or platelets ≥100x109/L, n=13, P=0.02, hazard ratio, HR 3.2 [1.2-8.9]), elevated AP (P=0.009, HR 3.3 [1.3-8.3]) and S/A/Rpos (P=0.007, HR 5.0 [1.8-18.1]) were significantly inferior regarding OS. In multivariate analyses, S/A/Rpos remained the only independent poor risk marker for OS (Figure). There was no significant difference regarding OS between primary vs. secondary MCL (Figure) or MCL with vs. without AHN. Of interest, no difference regarding OS was detected in comparison between patients treated with midostaurin (n=13) vs. patients treated with cladribine following midostaurin or vice versa (n=9). In summary, we have found that a) leukemic MCL and MCL without C-findings are rare, b) secondary MCL is frequent and evolves from other advSM subtypes but not ISM, c) KIT D816V mutations are more frequent than previously reported and KIT D816V negative patients should be tested for other KIT mutations d) the prognostically highly relevant mutations in the S/A/R gene panel are present in approximately 50% of patients with MCL, and e) median OS is approximately 1.5 years with significantly inferior survival in S/A/Rpos patients. Disclosures Meggendorfer: MLL Munich Leukemia Laboratory: Employment. Valent:Amgen: Honoraria; Novartis: Honoraria, Research Funding; Celegene: Honoraria, Research Funding.

Detection of Activated STAT5 in Neoplastic Mast Cells in Patients with Systemic Mastocytosis: Role of c-kit D816V.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3515-3515 ◽  
Author(s):  
Karoline Sonneck ◽  
Matthias Mayerhofer ◽  
Karoline V. Gleixner ◽  
Marc Kerenyi ◽  
Maria-Theresa Krauth ◽  
...  
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
Systemic Mastocytosis ◽  
Cell Leukemia ◽  
Oncogenic Signaling ◽  
Kit Mutation ◽  
Knock Out ◽  
Mast Cell Leukemia ◽  
Kit D816v

Abstract Recent data suggest that activated STAT5 contributes to growth and differentiation of mast cells (MC) and that STAT5-knock out mice are MC-deficient. We have recently shown that constitutively activated STAT5 acts as a potent oncogenic signaling molecule in hematopoietic progenitor cells (Cancer Cell2005;7:87–99). In the present study, we examined the expression of activated STAT5 in neoplastic MC in systemic mastocytosis (SM) and asked whether the SM-related oncogene c-kit D816V is involved in STAT5-activation. For the immunohistochemical detection of activated tyrosine phosphorylated STAT5 (P-Y-STAT5), we used the specific monoclonal antibody AX1 (Advantex) which does not react with inactive STAT5. In all patients with SM tested (indolent SM, n=11; smouldering SM, n=2; aggressive SM, n=1; mast cell leukemia, n=1; all exhibiting c-kit D816V), MC were found to display P-Y-STAT5. Expression of activated STAT5 was also demonstrable in the c-kit D816V-positive mast cell leukemia-derived cell line HMC-1. The reactivity of HMC-1 cells with AX1 antibody was abrogated by a STAT5-specific blocking-peptide. To define the role of c-kit D816V in STAT5-activation, Ba/F3 cells with doxycycline-inducible expression of c-kit D816V (Ton.kit) were employed. In these cells, induction of c-kit D816V was followed by a massive increase in phosphorylated STAT5 as determined by a specific DNA-binding assay, whereas the total amounts of STAT5-mRNA and of the STAT5-protein showed only a slight increase or remained unchanged. In summary, these data show that neoplastic MC in SM express activated STAT5 (P-Y-STAT5), and that the transforming c-kit mutation D816V leads to persistent activation of STAT5 in these cells.

Mast cell leukemia

Blood ◽  
2013 ◽  
Vol 121 (8) ◽  
pp. 1285-1295 ◽  
Author(s):  
Sophie Georgin-Lavialle ◽  
Ludovic Lhermitte ◽  
Patrice Dubreuil ◽  
Marie-Olivia Chandesris ◽  
Olivier Hermine ◽  
...  
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
Cell Activation ◽  
De Novo ◽  
Systemic Mastocytosis ◽  
Mast Cell Activation ◽  
Cell Leukemia ◽  
Mast Cell Leukemia ◽  
Kit D816v

Abstract Mast cell leukemia (MCL) is a very rare form of aggressive systemic mastocytosis accounting for < 1% of all mastocytosis. It may appear de novo or secondary to previous mastocytosis and shares more clinicopathologic aspects with systemic mastocytosis than with acute myeloid leukemia. Symptoms of mast cell activation—involvement of the liver, spleen, peritoneum, bones, and marrow—are frequent. Diagnosis is based on the presence of ≥ 20% atypical mast cells in the marrow or ≥ 10% in the blood; however, an aleukemic variant is frequently encountered in which the number of circulating mast cells is < 10%. The common phenotypic features of pathologic mast cells encountered in most forms of mastocytosis are unreliable in MCL. Unexpectedly, non-KIT D816V mutations are frequent and therefore, complete gene sequencing is necessary. Therapy usually fails and the median survival time is < 6 months. The role of combination therapies and bone marrow transplantation needs further investigation.

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.

Combination Therapy with the Histone Deacetylase Inhibitor Vorinostat Plus the Novel Aurora Kinase A Inhibitor MK-5108 Leads to Enhanced Lymphoma Cell Death Due to Acetylation of p53 and Repression of c-Myc, hTERT, and miRNA Levels.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1690-1690
Author(s):  
Leo Kretzner ◽  
Anna Scuto ◽  
Kowolik Claudia ◽  
Richard Jove ◽  
Stephen J Forman ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Cell Growth ◽  
Research Funding ◽  
Aurora Kinase ◽  
Aurora Kinase A ◽  
Non Hodgkin Lymphoma ◽  
Deacetylase Inhibitor ◽  
Kinase A

Abstract Abstract 1690 Poster Board I-716 Background Patients with relapsed or refractory Hodgkin (HL) and Non Hodgkin Lymphoma (NHL) have few options after salvage therapy and transplant, and new agents are thus needed. MK-5108 is a novel aurora kinase inhibitor (AKI) with specificity against aurora kinase A, that produces G2/M phase cell cycle arrest. We show that addition of vorinostat, a histone and protein deacetylase inhibitor, to AKI treatment results in reactivation of proapoptotic genes and enhanced lymphoma cell death. A panel of HL and NHL cell lines was studied with either drug or the combination, using cell growth, apoptosis, and flow cytometry assays, followed by molecular studies. Results MK-5108 alone at 0.1 – 3 mM results in significant growth inhibition and apoptosis in multiple cell lines representing Hodgkin, Burkitt, and Non-Hodgkin lymphoma types, interestingly,DHL-4 and DHL-6 cells were more sensitive to this agent than to the pan-AKI MK-0457. Vorinostat alone at a dose range of 0.5 – 3 mM reduces cell growth by 50% or more in all lines tested. The combination of 1.5 mM vorinostat and 100 nM MK-5108 results in over 85% apoptosis of multiple lymphoma lines tested at 72 hours. Cell cycle analyses by FACS of MK-5108 treated cells show an increased percentage of cells in G2/M with few cells in sub-G1, whereas in combination with vorinostat the G2/M peak decreases and there is a significant increase in the apoptotic sub-G1 population. Real-time PCR analysis and immunoblotting of L540 cells treated with either single agent or in combination revealed that vorinostat treatment leads to alteration in pro-apoptosis, growth arrest, and DNA damage response genes. Myc mRNA and protein levels are reduced by vorinostat, and repression of microRNAs (miRNAs) in the Myc-regulated polycistronic cluster of miRNAs of chromosome 13, such as miR-17.5p, -17.3p, and 18, occurs with vorinostat and TSA. Prosurvival genes such as bcl-XL and hTERT are downregulated five-fold by vorinostat treatment, while the proapoptotic BAK gene is upregulated 1.5 – 2-fold. Vorinostat treatment leads to enhanced acetylation of p53, with a corresponding increase in the p53 target genes p21 and Noxa. To analyze the role of Myc inhibition in the sensitization by vorinostat of lymphoma cells to MK-5108, siRNA-mediated knock-down of Myc expression in L540 cells was performed. The siRNA-Myc transfected L540 cells showed enhanced sensitivity to MK-5108 as compared to control siRNA-null cells, as well as decreased hTERT levels, confirming the role of Myc inhibition by vorinostat as an integral part of the sensitization of lymphoma cells to MK-5108. Conclusions The HDACi vorinostat leads to both transcriptional and post-transcriptional changes that create a pro-apoptotic milieu, sensitizing the cell to centrosome-acting agents such as the aurora kinase A inhibitor MK-5108. These preclinical data support clinical trials of MK-5108 plus vorinostat in patients with relapsed or refractory lymphomas. [We acknowledge Merck Inc for providing Vorinostat, MK-0457, MK-5108, and research support.] Disclosures Kretzner: Merck: Research Funding. Yen:Merck: Research Funding. Kirschbaum:Merck: Research Funding, Speakers Bureau.

Aleukemic Mast Cell Leukemia (aMCL) - Clinical Characteristics and Outcomes

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4255-4255 ◽  
Author(s):  
Preetesh Jain ◽  
Sa Wang ◽  
Hagop M. Kantarjian ◽  
Nawid Sarwari ◽  
Keyur P. Patel ◽  
...  
Keyword(s):  
Overall Survival ◽  
Mast Cell ◽  
Mast Cells ◽  
Research Funding ◽  
Median Overall Survival ◽  
Systemic Mastocytosis ◽  
Cell Transplant ◽  
Cell Leukemia ◽  
Mast Cell Leukemia

Abstract ABSTRACT Introduction: Systemic mastocytosis (SM) is a complex and rare disease of clonal mast cells. Mast cell leukemia (MCL) is a very rare form of SM seen in <1% patients. We describe here our experience with MCL patients treated at our institution. Methods: We reviewed the medical records of 218 patients with mastocytosis who presented to our institution between 1994 and 2016. The survival of patients was calculated from the date of initial presentation to the date of last follow up. Kaplan-Meier product limit method was used to estimate the median survival. Results: From the 218 patients with mastocytosis, we identified 13 patients with MCL (6.5%). All 13 had aleukemic variant of MCL (aMCL; no presence of mast cells in blood measured by standard CBC technique but with ≥ 20% atypical mast cells in the marrow aspirate). In addition, in 4 of 13 patients, associated hematologic neoplasm (AHN) was present: 2 with myelodysplastic syndrome, 1 with chronic myelomonocytic leukemia and 1 with multiple myeloma. Median age of 13 patients was 62 years (range 24-75 years). Baseline features are summarized in Table-1. During their initial clinical presentation, 85% had constitutional symptoms, 54% had skin rash and other cutaneous symptoms, 31% gastrointestinal symptoms and 23% had joint pains. Imaging studies were performed in 6 patients: 5/6 had enlarged liver and/or spleen, 3 patients had nodal involvement, 2 had sclerotic bony lesions and 1 pt had adrenal involvement. Serum tryptase was >200 ng/ml in all the patients. Testing for c-KITD816V mutation was performed in 9 patients using mutation-specific quantitative (real-time) PCR, of which mutation was undetectable in 6 and detected in 3 patients. The sensitivity of detection was approximately 1 in 1000 mutation-bearing cells. None had FIP1L1-PDGFRA mutations. Treatments given were heterogeneous. Two patients each received imatinib with no response, 2 received dasatinib (one [positive for KITD816V mutation] had significant improvement in hepatosplenomegaly and pruritus but progressed after 1 year of dasatinib therapy), 2 cladribine based therapy with no response, and 2 with stem cell transplant and progression after it. Overall, median follow up was 111 months (1.4-131); 3 patients were alive and 9 died at the time of last follow up. Interestingly, among our 13 aMCL patients, those 4 with AHN appear to have had worse outcome: Figure-1A, shows the median overall survival (OS) of aMCL vs aMCL-AHN (32 vs 25 months; p=0.22). Overall, the median overall survival (OS) of aMCL without AHN was significantly inferior compared to aggressive systemic mastocytosis (ASM) and indolent systemic mastocytosis (ISM) without AHN: 31 months and not reached respectively (Figure-1B; p<0.001). Conclusions: In this analysis, we have shown that aMCL is very rare and these patients have very poor outcome. Based on the recent data from Gotlib et al NEJM 2016, role of midostaurin in the treatment of MCL should be explored. Further studies to characterize the genomic profile of patients with MCL are needed to identify potential therapeutic targets and disease resistance pathways. Table Survival of patients with aleukemic mast cell leukemia (aMCL) with/without AHN (A) and survival comparison of aMCL to other types of systemic mastocytosis, all without AHN (B) Table. Survival of patients with aleukemic mast cell leukemia (aMCL) with/without AHN (A) and survival comparison of aMCL to other types of systemic mastocytosis, all without AHN (B) Figure Figure. Disclosures Daver: Sunesis: Consultancy, Research Funding; Kiromic: Research Funding; Ariad: Research Funding; Pfizer: Consultancy, Research Funding; Otsuka: Consultancy, Honoraria; Karyopharm: Honoraria, Research Funding; BMS: Research Funding. Cortes:ARIAD: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Teva: Research Funding. Verstovsek:Celgene: Research Funding; Bristol-Myers Squibb: Research Funding; AstraZeneca: Research Funding; Incyte Corporation: Membership on an entity's Board of Directors or advisory committees, Research Funding; Galena BioPharma: Research Funding; Gilead: Research Funding; CTI BioPharma Corp: Research Funding; Lilly Oncology: Research Funding; NS Pharma: Research Funding; Geron: Research Funding; Promedior: Research Funding; Seattle Genetics: Research Funding; Roche: Research Funding; Pfizer: Research Funding; Genentech: Research Funding.

scholarly journals De novo Leukemic Variant of Mast Cell Leukemia With KIT D816V

2015 ◽  
Vol 35 (2) ◽  
pp. 260-262
Author(s):  
Hae In Bang ◽  
Rojin Park ◽  
Eun Su Park ◽  
In Ho Choi ◽  
Kyoung Ha Kim ◽  
...  
Keyword(s):  
Mast Cell ◽  
De Novo ◽  
Cell Leukemia ◽  
Mast Cell Leukemia ◽  
Kit D816v

scholarly journals Aurora kinase B inhibits Aurora kinase A to control maternal mRNA translation in mouse oocytes

Development ◽  
2021 ◽  
Author(s):  
Mansour Aboelenain ◽  
Karen Schindler
Keyword(s):  
Mrna Translation ◽  
Aurora Kinase ◽  
Cytoplasmic Polyadenylation ◽  
Aurora Kinase A ◽  
Aurora Kinase B ◽  
Maternal Mrna ◽  
Kinase C ◽  
Maternal Mrnas ◽  
Mrna Polyadenylation ◽  
Kinase A

Mammalian oocytes are transcriptionally quiescent, and meiosis and early embryonic divisions rely on translation of stored maternal mRNAs. Activation of these mRNAs is mediated by polyadenylation. Cytoplasmic polyadenylation binding element 1 (CPEB1) regulates activates mRNA polyadenylation. One message is Aurora kinase C (Aurkc), encoding a protein that regulates chromosome segregation. We previously demonstrated that AURKC levels are upregulated in oocytes lacking Aurora kinase B (AURKB), and this upregulation caused increased aneuploidy rates, a role we investigate here. Using genetic and pharmacologic approaches, we found that AURKB negatively regulates CPEB1-dependent translation of many messages. To determine why translation is increased, we evaluated Aurora kinase A (AURKA), a kinase that activates CPEB1 in other organisms. We find that AURKA activity is increased in Aurkb knockout oocytes and demonstrate that this increase drives the excess translation. Importantly, removal of one copy of Aurka from the Aurkb knockout strain background, reduces aneuploidy rates. This study demonstrates that AURKA is required for CPEB1-dependent translation, and it describes a new AURKB requirement to maintain translation levels through AURKA, a function critical to generating euploid eggs.

A Kinome shRNA Screen to Identify Pathways That Regulate Megakaryocyte Polyploidization and New Targets for Differentiation Therapy

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 89-89
Author(s):  
Qiang Jeremy Wen ◽  
Serena Silver ◽  
Tim Lewis ◽  
Sridevi Ponduru ◽  
Vlado Dancik ◽  
...  
Keyword(s):  
Cell Line ◽  
Ex Vivo ◽  
Terminal Differentiation ◽  
Aurora Kinase ◽  
Cyclin B1 ◽  
Leukemia Cell Line ◽  
Aurora Kinase A ◽  
Aurora Kinase B ◽  
Shrna Screen ◽  
Kinase A

Abstract Abstract 89 Megakaryocytes undergo a unique mode of cell cycle called endomitosis, which is characterized by repeated rounds of DNA synthesis without cell division. Normally, polyploidization is tightly coupled with expression of megakaryocyte specific genes and terminal differentiation. Although there have been several reports of differences between the proliferative and endomitotic cell cycles, the precise mechanisms that regulate the switch to polyploidization and terminal differentiation remain unknown. To identify genes that play a role in this process, we performed a high throughput shRNA screen assaying for genes whose decreased expression induce polyploidization in CMK megakaryocytic leukemia cell line. We used a kinase biased library of shRNAs, reasoning that targeting the kinome would identify pathways or complexes important for this switch. After transduction of with the lentiviral shRNA library, CMK cells were selected with puromycin, fixed and stained with Hoechst dye. A high content imaging assay using CellProfiler software was employed to analyze the DNA content of microscopic images of cells. By this approach, we screened 775 genes, 650 encoding kinases, with at least 5 hairpins per gene. In parallel, we performed an otherwise identical screen (i.e., modifier screen) that included sub-maximal doses of dimethylfasudil, a drug that we have previously shown to promote megakaryocyte polyploidization. To reduce the off-target effects of shRNA, we only considered genes that scored with two or more shRNAs. In comparison to non-targeted shRNA controls, we considered the top 5% of genes in either the direct or modifier screens to be hits. By this analysis, we identified 71 genes whose loss promoted polyploidization. Pathway analysis revealed that the majority of these genes clustered into five groups. Among these clusters, one was notable for containing AURKA (aurora kinase A), AURKB (aurora kinase B), CCNB1 (cyclin B1), CDK2 and PLK1 (polo-like kinase). The screen revealed that knockdown of Aurora B kinase significantly increased polyploidization in the absence of diMF (p<0.001), but that knockdown of Aurora A kinase only increased polyploidization when diMF was included (p<0.01). Of note, although several studies have examined the expression and function of AURKB in polyploidization, its role in human versus mouse megakaryopoiesis is controversial. Moreover, whether inhibition of Aurora kinases would be a useful therapy for megakaryocytic leukemias, characterized by defects in polyploidization and terminal differentiation, has not been addressed. We discovered that MLN8237, a selective Aurora kinase A inhibitor, and AZD1152, a selective Aurora kinase B inhibitor dose dependently inhibited proliferation while inducing polyploidization, upregulation of megakaryocyte specific lineage markers, and apoptosis of CMK cells and the 6133-MPL murine cell line, which expreses the AMKL fusion protein OTT-MAL and the W515L constitutively active allele of MPL. MLN8237 and AZD1152 induced polyploidization with an EC50 of 22 nM and 210 nM in CMK cells respectively. Both compounds also induced robust polyploidization of human and mouse primary megakaryocytes cultured ex vivo. At the doses utilized, neither drug induced polyploidization of the non-megakaryocyte fractions of the ex vivo cultures, indicating that these inhibitors are selective for megakaryocytes at the doses used in this study. Finally, since we found that MLN8237 was a more potent inducer of polyploidization, we next assayed the ability of the compound to block progression of AMKL in an animal model. MLN8237 significantly increased the survival of 6133-MPL transplanted mice compared to vehicle (p=0.02), demonstrating the anti-leukemic activity of this small molecule. Together, our data suggest that Aurora kinase inhibitors may be useful as differentiation therapeutic agents for AMKL. Disclosures: No relevant conflicts of interest to declare.

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