scholarly journals Coronavirus S protein-induced fusion is blocked prior to hemifusion by Abl kinase inhibitors

2018 ◽  
Author(s):  
Jeanne M. Sisk ◽  
Matthew B. Frieman ◽  
Carolyn E. Machamer
Keyword(s):  
Host Cell ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Host Cells ◽  
Tissue Type ◽  
Host Cell Membrane ◽  
S Protein ◽  
Abl Kinase ◽  
Coronavirus Infection ◽  
Abl Kinase Inhibitors

ABSTRACTEnveloped viruses gain entry into host cells by fusing with cellular membranes, a step required for virus replication. Coronaviruses, including the severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and infectious bronchitis virus (IBV), fuse at the plasma membrane or use receptor-mediated endocytosis and fuse with endosomes depending on the cell or tissue type. The virus Spike (S) protein mediates fusion with the host cell membrane. We have shown previously that an Abl kinase inhibitor, imatinib, significantly reduces SARS-CoV and MERS-CoV viral titers and prevents endosomal entry by HIV SARS S and MERS S pseudotyped virions. SARS-CoV and MERS-CoV are classified as BSL-3 viruses, which can make experimentation into the cellular mechanisms involved in infection more challenging. Here, we use IBV, a BSL-2 virus, as a model for studying the role of Abl kinase activity during coronavirus infection. We found that imatinib and two specific Abl kinase inhibitors, GNF2 and GNF5, reduce IBV titers by blocking the first round of virus infection. Additionally, all three drugs prevented IBV S-induced syncytia formation prior to the hemifusion step. Our results indicate that membrane fusion (both virus-cell and cell-cell) is blocked in the presence of Abl kinase inhibitors. Studying the effects of Abl kinase inhibitors on IBV will be useful in identifying host cell pathways required for coronavirus infection. This will provide insight into possible therapeutic targets to treat infections by current as well as newly emerging coronaviruses.

Identification and Functional Significance of Genes Regulated by Structurally Different ABL Kinase Inhibitors.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2860-2860
Author(s):  
Kousuke Nunoda ◽  
Tetsuzo Tauchi ◽  
Tomoiku Takaku ◽  
Masahiko Sumi ◽  
Seiichi Okabe ◽  
...  
Keyword(s):  
Gene Expression ◽  
Growth Inhibition ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Specific Inhibitor ◽  
K562 Cells ◽  
Myelogenous Leukemia ◽  
Altered Expression ◽  
Abl Kinase ◽  
Abl Kinase Inhibitors

Abstract Imatinib is an ABL-specific inhibitor that binds with high affinity to the inactive conformation of the BCR-ABL tyrosine kinase and has been shown to be effective in the treatment of chronic myelogenous leukemia. Dasatinib is an ATP-competitive, dual-spesific SRC and ABL kinase inhibitor that can bind BCR-ABL in both the active and inactive conformations. From a clinical stand point, dasatinib is particular attractive because it has been shown to induce hematologic and cytogenetic responses in imatinib-resistant CML patients. In the view of the fact that the combination of imatinib and dasatinib shows the additive/synergistic growth inhibition on a wild type p210 BCR-ABL expressing cells, we reasoned that these ABL kinase inhibitors might induce the different molecular pathways. To address this question, we used DNA microarrays to identify genes whose transcription was altered by imatinib and dasatinib. K562 cells were cultured with imatinib or dasatinib for 16 hrs, and gene expression data was obtained from three independent microarray hybridizations. Almost all of the imatinib- and dasatinib- responsive genes appeared to be similarly increased or decreased in K562 cells; however, small subsets of genes were identified as selectively altered expression by either imatinib or dasatinib. The genes whose expression was affected by imatinib and dasatinib were categorized into different functional groups based on their biological function, and genes in the cell proliferation and apoptosis categories were examined in detail. Imatinib and dasatinib affected the expression of several cyclin-dependent kinases (CDK2, CDK4, CDK6, CDK8, and CDK9), cell division cycle genes (CDC6, CDC7, CDC25C, and CDC34), and cyclones (cyclin A2, C, D2, D3, E1, E2, F, G1, G2, and H). Imatinib and dasatinib also modulated the expression of apoptosis-related genes including APAF1, BAK1, BCL2, BCL10, MCL1, CASP3, and CASP6). One of the distinct genes which are selectively modulated by dasatinib are CDK2 and CDK8, which had a maximal fold reduction of <8-fold in microarray screen. Immunoblotting confirmed that gene expression changes induced only by dasatinib correlated with changes in protein expression. To assess the functional importance of dasatinib regulated genes, we used RNA interference to determine whether reduction of CDK2 and CDK8 affected the growth inhibition. The siRNA to CDK2 or CDK8 specifically reduced cdk2 or cdk8 in K562 cells. K562 cells pretreated with CDK2 or CDK8 siRNA showed the additive growth inhibition with imatinib but not with dasatinib. These finding demonstrate that the additive/synergistic growth inhibition by imatinib and dasatinib may be mediated by CDK2 and CDK8.

HOXA10 Expression Induces by Abl Kinase Inhibitors Enhanced Apoptosis through PI3K Pathway in CML Cells.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4529-4529
Author(s):  
Isao Hirano ◽  
Yuya Sugimoto ◽  
Keiji Okinaka ◽  
Takaaki Ono ◽  
Kaori Shinjo ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Pi3k Pathway ◽  
Hematopoietic Progenitor ◽  
Leukemic Transformation ◽  
Hematopoietic Stem ◽  
Abl Kinase ◽  
Abl Kinase Inhibitors

Abstract [Background] Homeobox (Hox) genes are grouped together in 4 clusters, A to D. Recent studies has shown that the Hox proteins are important in the control of differentiation and proliferation in hematopoietic cells. We found that the Abl kinase inhibitors increased the expression of HoxA10 gene in CML cells. In this study, we analyzed the role of HoxA10 in CML cell lines and the hematopoietic progenitor cells derived from CML patients by inhibiting the expression of HoxA10. Moreover, we investigated whether the regulation of HoxA10 eradicate Ph+ hematopoietic stem/progenitor cells, which are the targets for leukemic transformation in CML. [Methods] We used AMN107 and BMS354825 for the Abl kinase inhibitors, LY294002 for a PI3K inhibitor, PP2 for a Src kinase inhibitor, and SB203580 for a p38 MAP kinase inhibitor. For analysis of HoxA10 mRNA and protein, RT-PCR and western blot were performed in K562, Meg-01 and U937 cells, which untreated or treated with AMN107, BMS354825, LY294002, PP2, or SB203580 respectively. We then attempted to localize the intracellular locations of HoxA10 in K562 and Meg01, which untreated or treated with AMN107, BMS354825, or LY294002 by using conforcal fluorescence microscopy. For analysis of proliferation in K562, Meg-01 and U937 transfected with siRNA HoxA10, MTT assays were performed in untransfected or transfected K562, Meg-01 and U937 treated with or without AMN107, BMS354825, or LY294002. Finally, we counted the colony numbers of CFU-GEMM, CFU-GM, and BFU-E in K562 and Meg-01 treated with the Abl kinase inhibitors or LY294002. Results Both K562 and Meg01 cells expressed HoxA10 mRNA and protein at lower level compared to U937 cells. Interestingly, treatment with AMN107, BMS354825, or LY294002 increased the expression of HoxA10 mRNA and protein in both K562 and Meg01 cells. The fluorescence of HoxA10 was more strongly observed in the area corresponding to the cell’s cytoplasm than nucleus, and the treatment with AMN107, BMS354825, or LY294002 increased the fluorescence in nucleus of K562 and Meg01 cells in a time-dependent manner. In K562 and Meg01 cells transfected with the siRNA HoxA10, treatment with AMN107 or BMS354825 slightly inhibited the proliferation compared to K562 and Meg01 transfected with control siRNA. Finally, we showed that the inhibition of HoxA10 expression by siRNA increased the numbers of CFU-GEMM, BFU-E, and CFU-GM when the cells were treated with the combination of BMS354825 and LY294002 compared to control cells. [Conclusions] In this study, we showed that the Abl kinase inhibitors induced the expression of HoxA10, and HoxA10 was regulated by PI3K pathway in CML cells. This finding indicates a new insight in the regulation of cell proliferation via the PI3K signal pathway in CML cells. Moreover, we found the role of HoxA10 in CML cell lines and the hematopoietic progenitor cells derived from CML patients by inhibiting the expression of HoxA10. We showed that the regulation of HoxA10 eradicated Bcr-Abl+ hematopoietic stem/progenitor cells, which are the targets for leukemic transformation in CML.

Depletion of PHLPP1 and 2 by Bcr-Abl Promotes CML Cell Proliferation through the Continuous Phosphorylation of Akt.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2182-2182
Author(s):  
Isao Hirano ◽  
Satoki Nakamura ◽  
Tomonari Takemura ◽  
Daisuke Yokota ◽  
Takaaki Ono ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Progenitor Cells ◽  
Cell Lines ◽  
Colony Formation ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Knock Down ◽  
Abl Kinase ◽  
Pcr Method ◽  
Abl Kinase Inhibitors

Abstract Abstract 2182 Poster Board II-159 Objective PHLPP1 and 2 (pleckstrin homology (PH) domain leucine-rich repeat protein phosphatase 1 and 2) is identified as the dephosphorylation enzyme of Akt, the same as that of PP2A of the dephosphorylation enzyme of Akt, and regulate the cell-growth signal of Akt through a dephosphorylation of the phosphorylated Akt (p-Akt). Previously, we reported the expression of PHLPP1 and 2 were suppressed in CML cell lines. In this study, we investigated the the CML and its progenitor cell proliferation through the phosphorylation of Akt by depletion of PHLPP1 and 2. Method CML cell lines (K562, Meg01, SHG3) and the CML clinical specimen (n= 10) were used for this research. The changes in the expression of PHLPP1 and 2 by treatment with Abl kinase inhibitors (STI571, AMN107 and BMS354825) or knock down with Bcr-Abl gene were evaluated by RT-PCR method. The influence on the expression of PHLPP1 and 2 in AML cell line transfected with Bcr-Abl also evaluated. CML progenitor cells derived from CML patients separated by ALDH activity, and the expression of PHLPP1 and 2 were investigated by quantitative RT-PCR method. p-Ser473 Akt1, 2 and 3 by Abl kinase inhibitor or knock down with PHLPP1 and 2 were measured, and the influence on the effect of cell growth inhibition by MTT assay. The expression of PHLPP1 and 2 and colony formation in colony forming unit-granulocyte, erythroid, macrophage, megakaryocyte (CFU-GEMM), colony forming unit-granulocyte, macrophage (CFU-GM) and burst forming unit-erythroid (BFU-E) derived from Bcr-Abl positive hematopoietic progenitor cells also evaluated. Result The expression of mRNA and protein of PHLPP1 and 2 were increased by treatment with Abl kinase inhibitor or Bcr-Abl knock down by siRNA in CML cell lines and AML cell line trasfected with Bcr-Abl gene. Moreover, p-Ser473 Akt 2 and 3 were decreased according to the expression of PHLPP1, and also p-Ser473 Akt1 and 3 according to the expression of PHLPP2. The Abl kinase inhibitors induced the expression of PHLPP1, 2 and the reduction of p-Ser473 Akt isoforms. The Abl kinase inhibitors inhibited the CML cell proliferation via the depletion of PHLPP1 and 2. In Bcr-Abl positive progenitor cells, the expression of PHLPP1 and 2 was increased, and colony formation was suppressed by Abl kinase inhibitor and by Bcr-Abl siRNA. The colony formation in progenitor cells knocked down PHLPP1 and 2 were decreased when treated with Abl kinase inhibitors. Conclusion These results suggest that Bcr-Abl might promote CML cell proliferarion through continuous phosphorylation of p-Ser473 Akt1, 2 and 3 by suppression of PHLPP1 and 2, and that the induction of PHLPP1 and 2 may be effective to regulate the cell proliferation in CML cells. It may be also that the induction of expression of PHLPPs has the potential possibility as the targets on the regulation of cell proliferation in Bcr-Abl positive progenitor cells. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Screening of Potential Inhibitors of Covid-19 with Repurposing Approach Via Molecular Docking

2020 ◽  
Author(s):  
Negin Alizadehmohajer ◽  
Bahman Sadeghi ◽  
Simin Najafgholian ◽  
Shabnam Moradi ◽  
Forogh Mohammadi ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Janus Kinase ◽  
New Drugs ◽  
Regulatory Function ◽  
Bacterial Biosensor ◽  
Abl Kinase ◽  
Dipeptidyl Peptidase 4 Inhibitors ◽  
Abl Kinase Inhibitors

Abstract Background: 2019-nCoV (COVID-19) is a pandemic disease with a high infectivity and mortality. The prevention and treatment of COVID-19 have become urgent matters for consideration. It often takes several years to develop new drugs, or vaccines, based on the usual clinical trial process. This dwell-time can be shortened by repurposing previously approved drugs.Methods: We have designed and evaluated a bacterial biosensor expressing a luciferase We aimed to assess several available small-molecule; Abl kinase inhibitors, Janus kinase inhibitor, Dipeptidyl peptidase 4 inhibitors, RNA-dependent RNA polymerase inhibitors, and Papin-like Protease inhibitors, using binding simulation with proteins that might prove to be effective in inhibiting COVID-19 infection. The efficiency of inhibitors was evaluated based on docking scores using auto dock vina software.Results: Strong ligand-protein interactions were predicted among some of these drugs, such as Imatinib, Remdesivir, and Telaprevir, and this may render these compounds promising candidates. Some candidate drugs might be efficient in disease control (directly and indirectly) or in viral proteins attenuation. It is worth to highlight the powerful immunomodulatory role of Abivertinib that inhibits pro-inflammatory cytokine production that are associated with cytokine release syndrome (CRS) or cytokine storm and progression of COVID-19 infection.Conclusions: COVID-19 is similar to SARS-CoV, the potential role of Abl kinase inhibitors such as Imatinib in reducing SARS-CoV and MERS-CoV viral titers, immune regulatory function and the development of acute respiratory distress syndrome (ARDS) may indicate that these drugs may be useful for COVID-19. Moreover, Remdesivir, and Telaprevir have the most efficiency with their docked proteins in-silico as well although clinical trials are needed to confirm the effect of these drugs.

Bcr-Abl ubiquitination and Usp9x inhibition block kinase signaling and promote CML cell apoptosis

Blood ◽  
2011 ◽  
Vol 117 (11) ◽  
pp. 3151-3162 ◽  
Author(s):  
Hanshi Sun ◽  
Vaibhav Kapuria ◽  
Luke F. Peterson ◽  
Dexing Fang ◽  
William G. Bornmann ◽  
...  
Keyword(s):  
Cell Apoptosis ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Therapeutic Option ◽  
Myelogenous Leukemia ◽  
Kinase Signaling ◽  
Abl Kinase ◽  
Novel Approach ◽  
Abl Kinase Inhibitors

Abstract Although chronic myelogenous leukemia (CML) is effectively controlled by Bcr-Abl kinase inhibitors, resistance to inhibitors, progressive disease, and incomplete eradication of Bcr-Abl–expressing cells are concerns for the long-term control and suppression of this disease. We describe a novel approach to targeting key proteins in CML cells with a ubiquitin-cycle inhibitor, WP1130. Bcr-Abl is rapidly modified with K63-linked ubiquitin polymers in WP1130-treated CML cells, resulting in its accumulation in aggresomes, where is it unable to conduct signal transduction. Induction of apoptosis because of aggresomal compartmentalization of Bcr-Abl was observed in both imatinib-sensitive and -resistant cells. WP1130, but not Bcr-Abl kinase inhibitors, directly inhibits Usp9x deubiquitinase activity, resulting in the down-regulation of the prosurvival protein Mcl-1 and facilitating apoptosis. These results demonstrate that ubiquitin-cycle inhibition represents a novel and effective approach to blocking Bcr-Abl kinase signaling and reducing Mcl-1 levels to engage CML cell apoptosis. This approach may be a therapeutic option for kinase inhibitor–resistant CML patients.

scholarly journals Coronavirus S protein-induced fusion is blocked prior to hemifusion by Abl kinase inhibitors

2018 ◽  
Vol 99 (5) ◽  
pp. 619-630 ◽  
Author(s):  
Jeanne M. Sisk ◽  
Matthew B. Frieman ◽  
Carolyn E. Machamer
Keyword(s):  
Kinase Inhibitors ◽  
S Protein ◽  
Abl Kinase ◽  
Abl Kinase Inhibitors

Invasion of Toxoplasma gondii occurs by active penetration of the host cell

1995 ◽  
Vol 108 (6) ◽  
pp. 2457-2464 ◽  
Author(s):  
J.H. Morisaki ◽  
J.E. Heuser ◽  
L.D. Sibley
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Tyrosine Phosphorylation ◽  
Host Cells ◽  
The Novel ◽  
Host Proteins ◽  
Host Cell Membrane ◽  
Membrane Ruffling ◽  
Obligate Intracellular ◽  
Obligate Intracellular Parasite

Toxoplasma gondii is an obligate intracellular parasite that infects a wide variety of vertebrate cells including macrophages. We have used a combination of video microscopy and fluorescence localization to examine the entry of Toxoplasma into macrophages and nonphagocytic host cells. Toxoplasma actively invaded host cells without inducing host cell membrane ruffling, actin microfilament reorganization, or tyrosine phosphorylation of host proteins. Invasion occurred rapidly and within 25–40 seconds the parasite penetrated into a tight-fitting vacuole formed by invagination of the plasma membrane. In contrast, during phagocytosis of Toxoplasma, extensive membrane ruffling captured the parasite in a loose-fitting phagosome that formed over a period of 2–4 minutes. Phagocytosis involved both reorganization of the host cytoskeleton and tyrosine phosphorylation of host proteins. In some cases, parasites that were first internalized by phagocytosis, were able to escape from the phagosome by a process analogous to invasion. These studies reveal that active penetration of the host cell by Toxoplasma is fundamentally different from phagocytosis or induced endocytic uptake. The novel ability to penetrate the host cell likely contributes to the capability of Toxoplasma-containing vacuoles to avoid endocytic processing.

scholarly journals Intracellular Retention of ABL Kinase Inhibitors Determines Commitment to Apoptosis in CML Cells

PLoS ONE ◽  
2012 ◽  
Vol 7 (7) ◽  
pp. e40853 ◽  
Author(s):  
Daniel B. Lipka ◽  
Marie-Christine Wagner ◽  
Marek Dziadosz ◽  
Tina Schnöder ◽  
Florian Heidel ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Abl Kinase ◽  
Abl Kinase Inhibitors
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