scholarly journals Antitumor Activity of Phosphoinositide 3-Kinases Inhibitor, Copanlisib in Multiple Myeloma Cells

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5755-5755
Author(s):  
Yuko Tanaka ◽  
Seiichi Okabe ◽  
Tetsuzo Tauchi ◽  
Yoshikazu Ito ◽  
Kazuma Ohyashiki
Keyword(s):  
Cell Growth ◽  
Growth Inhibition ◽  
Signaling Pathway ◽  
Proteasome Inhibitor ◽  
Proteasome Inhibitors ◽  
Pi3k Signaling ◽  
Dependent Manner ◽  
Cell Growth Inhibition ◽  
Pi3k Signaling Pathway ◽  
Dose Dependent

Abstract Multiple myeloma (MM) is one of the common hematological malignancies and is a uniformly fatal disorder of B cells characterized by accumulation of abnormal plasma cells in the bone marrow. Proteasome inhibitor, bortezomib, and immunomodulatory drugs such as thalidomide and lenalidomide play important roles in the treatment of MM patients. Although novel agents including, e.g. bortezomib, have significantly improved the response and survival of patients with MM, a large number of patients eventually have relapsed. For the patients who relapse after treatment with novel agents, the prognosis is still poor. Thus circumstanced, alternative strategies are required for continued disease control. Phosphoinositide 3-kinases (PI3Ks) are a family of proteins involved in the regulator of cell growth, metabolism and proliferation. PI3K signaling pathway also plays a critical regulatory role in MM pathology, including survival, cellular proliferation, migration and angiogenesis. Therefore, PI3K signaling pathway may present attractive targets for MM treatment. Copanlisib also known as BAY80-6946 is a potent and highly selective reversible PI3K inhibitor. Copnalisib is currently investigated in a pivotal phase 2 clinical trial against hematological malignancy such as malignant lymphoma. We hypothesized that treatment with PI3K inhibitor and proteasome inhibitors together would result in enhanced therapeutic activity in MM cells. In this study, we investigated the efficacy of copanlisib by using the MM cell lines, RPMI8226, MM1.S and MM1.R and primary sample. 72 h treatment of copanlisib exhibits cell growth inhibition of MM cell lines in a dose dependent manner. The treatment of proteasome inhibitors, bortezomib and carfilzomib exhibits cell growth inhibition partially against RPMI8226 cells in the presence of feeder cell line, HS-5. We examined the intracellular signaling in the presence of HS-5. Phosphorylation of Akt and activation of caspase 3 and poly (ADP-ribose) polymerase (PARP) was partially reduced by carfilzomib or bortezomib in the presence of HS-5. We found that the treatment of copanlisib abrogated the protective effects of HS-5 in RPMI8226 cells. We examined the intracellular signaling after treatment of copanlisib. Activity of caspase 3 and poly (ADP-ribose) polymerase (PARP) was increased after copnlisib treatment in a dose dependent manner. Because PI3K signaling pathway regulates MM cell migration, we next evaluated the chemotactic response of MM cells to stromal cell-derived factor 1α (SDF-1α). We found that 4 h treatment of SDF-1α significantly induced the migration of MM cells compared to control medium. Treatment of copanlisib inhibited SDF-1α-stimulated chemotaxis in a dose dependent manner. We found that phosphorylation of Akt was reduced after copanlisib treatment suggesting that intracellular PI3K signaling pathway may play the important role in SDF-1α induced chemotaxis of MM cells. We investigated the copanlisib activity against MM cells. Combined treatment of MM cells with proteasome inhibitor, carfilzomib or bortezomib, and copanlisib caused significantly more cytotoxicity than each drugs alone. Phosphorylation of Akt was reduced and cleaved PARP was increased after copanlisib with or without proteasome inhibitor. We also found that copanlisib which was combinaed with carfilzomib or borteomib exhibited cell growth inhibition against MM primary sample. Data from this study suggested that administration of the PI3K inhibitor, copanlisib may be a powerful strategy against stroma-associated drug resistance of MM cells and enhance cytotoxic effects of proteasome inhibitors in those residual MM cells. Disclosures No relevant conflicts of interest to declare.

The Analysis of HDAC Inhibitor, Vorinostat Efficacy against Wild Type and BCR-ABL Mutant Positive Leukemia Cells in Monotherapy and in Combination with a Pan-Aurora Kinase Inhibitor, MK-0457

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 5025-5025 ◽  
Author(s):  
Seiichi Okabe ◽  
Tetsuzo Tauchi ◽  
Shinya Kimura ◽  
Taira Maekawa ◽  
Kazuma Ohyashiki
Keyword(s):  
Cell Growth ◽  
Growth Inhibition ◽  
Kinase Inhibitor ◽  
Aurora Kinase ◽  
Dependent Manner ◽  
Wild Type ◽  
Cell Growth Inhibition ◽  
Imatinib Resistant ◽  
T315i Mutation ◽  
Dose Dependent

Abstract The use of imatinib, an ABL tyrosine kinase inhibitor, has led to a dramatic change in the management of BCR-ABL positive leukemia patients. However, the resistance to imatinib mediated by mutations in the BCR-ABL domain has become a major problem in the treatment. Histone deacetylase (HDAC) inhibitors have been shown to mediate the regulation of gene expression, induce cell growth, cell differentiation and apoptosis of tumor cells. Vorinostat (suberoylamide hydroxamic acid:SAHA) is a hydroxamic acid based polar HDAC inhibitor. Vorinostat have shown efficacy in a wide range of cancers such as cutaneous T-cell lymphoma (CTCL). However, efficacy of vorinostat against the BCR-ABL mutants has fully not known. Here we report on the studies performed against murine Ba/F3 cell line which was transfected wild type (Wt) p210 and p185 BCR-ABL or imatinib resistant BCR-ABL mutants such as G250E, Q252H, Y253F, E255K, M294V, T315I, T315A, F317L, F317V, M351T, H396P and T315I(p185). 48 hours treatment of vorinostat exhibits cell growth inhibition and proapoptotic activity murine Ba/F3 cells ectopically expressing Wt and imatinib resistant BCR-ABL mutants including T315I mutation in a dose dependent manner. IC50 of these cell lines are Wt(720nM), G250E(625nM), Q252H(220nM), Y253F(525nM), E255K(685nM), M294V(785nM), T315I(500nM), T315A(715nM), F317L(560nM), F317V(565nM), M351T(375nM) and H396P(485nM). Aurora kinases play a pivotal role in the regulator of mitotic processes during cell division. MK-0457 is a small molecule inhibitor of the Aurora kinase family and was found to be active against the cells from BCR-ABL positive patients with T315I mutation in clinical trial. Because vorinostat also depleted BCR-ABL, as well as induced apoptosis and sensitized BCR-ABL-expressing leukemia cells, we examined whether vorinostat and MK-0457 enhances the apoptosis in imatinib resistant BCR-ABL-expressing cells. 48 hours treatment of MK-0457 exhibits cell growth inhibition of Ba/F3 cells ectopically expressing Wt and imatinib resistant BCR-ABL mutants including T315I mutation. IC50 of MK-0457 is Wt(215nM), G250E(205nM), Q252H(185nM), Y253F(245nM), E255K(185nM), M294V(238nM), T315I(205nM), T315A(165nM), F317L(200nM), F317V(200nM), M351T(225nM) and H396P(195nM). We examined the intracellular signaling by using these cell lines. We found that caspase 3, and poly (ADPribose) polymerase (PARP) were activated after MK-0457 treatment in a dose dependent manner. Phosphorylation of BCR-ABL and Crk-L which is downstream target of BCR-ABL was reduced after MK-0457 treatment. We found that combination of vorinostat and MK-0457 synergistically cell growth inhibition of Wt and BCR-ABL mutants Ba/F3 cells in 48 hours treatment. Phosphorylation of Crk-L was reduced after vorinostat and MK-0457 treatment. Caspase 3 and PARP activation were also synergistically increased after vorinostat and MK-0457 treatment. We evaluated the activity of MK-0457 and vorinostat in primary BCR-ABL positive acute lymphoblastic leukemia (ALL) cells with the T315I mutation. We found that MK-0457 potently induced cell growth inhibition of primary T315I cells in 48 hours treatment. Moreover, combination of vorinostat and MK-0457 synergistically increased the cell growth inhibition in primary T315I cells. This study demonstrate monotherapy of vorinostat and the combination of vorinostat and MK-0457 are more potent efficacy not only wild type BCR-ABL but also imatinib resistant BCR-ABL mutants cells and represents a promising new strategy for treatment of imatinib resistant BCR-ABL positive leukemias, including those harboring the T315I mutation.

Carfilzomib, An Irreversible Proteasome Inhibitor, Induces Long-Term Growth Inhibition of Mantle Cell Lymphoma In Vivo,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3740-3740
Author(s):  
Liang Zhang ◽  
Jianfei Qian ◽  
Zhishuo Ou ◽  
Luhong Sun ◽  
Kejie Zhang ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Proteasome Inhibitor ◽  
Cell Lymphoma ◽  
Proteasome Inhibitors ◽  
Dependent Manner ◽  
Mantle Cell ◽  
Induced Apoptosis ◽  
Dose Dependent

Abstract Abstract 3740 Mantle cell lymphoma (MCL) is an aggressive B-cell lymphoma with poor clinical outcome, thus, novel therapeutic agents are urgently needed. The proteasome inhibitors are small molecular agents which show significant anti-tumor effect in patients with relapsed/refractory MCL. Carfilzomib, an irreversible proteasome inhibitor with selectivity for the chymotrypsin-like active site, inhibits the proliferation of MCL cells in vitro, as well as the reversible proteasome inhibitor bortezomib. Unlike bortezomib, carfilzomib is good-tolerated and does not induce severe neuropathy in patients. Therefore, carfilzomib can be used in higher dose than bortezomib in vivo. Our study was undertaken to evaluate the therapeutic efficacy of carfilzomib on MCL cells both in vitro and in vivo compared with bortezomib. Four human MCL cell lines, MINO, Jeko-1, MAVER, and NCEB-1, freshly isolated primary MCL cells from the patients with relapsed/refractory MCL, were treated with carfilzomib or bortezomib. A 3H-thymidine incorporation assay showed that both carfilzomib and bortezomib displayed the same dose-dependent manner in inducing growth inhibition of the MCL cells. Similarly, flow cytometry analysis with fluorescence-labeled Annexin V and propidium iodide showed that carfilzomib induced apoptosis of MCL cells in the same dose-dependent manner with bortezomib. However, under the tolerable dose of each of the two proteasome inhibitors, they had different therapeutic effect in a MCL-bearing mouse model established in severe combined immunodeficient (SCID) mice. MINO cells (5 × 106) were inoculated subcutaneously into the right flank of SCID mice. Three weeks later, after palpable tumors developed, mice were treated intravenously with carfilzomib (5 mg/kg) on day 1 and day2, for 5 cycles, or treated intraperitoneally with bortezomib (1 mg/kg) on days 1, 4, 7 and 10, per 21 days. Tumor growth was almost abrogated after treatment with carfilzomib compared with bortezomib, and the survival time of tumor-bearing mice was significantly prolonged in the carfilzomib-treated mice versus bortezomib-treated mice. Notably, Increasing the frequency or dose of bortezomib treatment was unable because the mice were too suffered in toxicity to tolerate the treatment. Western blot analysis showed that carfilzomib induced apoptosis in caspase-dependent manner as well as bortezomib. Carfilzomib inhibited the phosphorylation of IκB, STAT3, and AKT and irreversibly blocked the release of NFκB to nuclei. In conclusion, carfilzomib displays the same anti-tumor effect and mechanism with bortezomib on MCL cells in vitro. However, carfilzomib but not bortezomib is well tolerated without severe side effect in vivo. Carfilzomib significantly inhibits tumor growth and prolongs survival indicating that carfilzomib is a potential agent in MCL chemotherapy. Disclosures: No relevant conflicts of interest to declare.

Therapeutic Potential Of Targeting Sphingosine-1-Phosuphate and Sphingosine Kinases In Multiple Myeloma

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1894-1894 ◽  
Author(s):  
Yuko Tanaka ◽  
Seiichi Okabe ◽  
Tetsuzo Tauchi ◽  
Yoshikazu Ito ◽  
Tomohiro Umezu ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Growth ◽  
Growth Inhibition ◽  
Cell Lines ◽  
Hematological Malignancies ◽  
Therapeutic Strategies ◽  
Dependent Manner ◽  
Cell Growth Inhibition ◽  
Sphingosine Kinases ◽  
Dose Dependent

Abstract Multiple myeloma (MM) is one of the common hematological malignancies and is a uniformly fatal disorder of B cells characterized by accumulation of abnormal plasma cells in the bone marrow.Clinical progression of patients with MM is improved with the proteasome inhibitor (PI) (e.g. bortezomib) and the immunomodulatory drugs (IMiDs) such as thalidomide and lenalidomide. Although PI and IMiDs have considerably changed the treatment paradigm of MM, many patients show disease relapse due to developing into drug resistance of MM cells. Since the prognosis remains poor for patients with refractory disease, the new therapeutic strategies are required to treat against these patients. Sphingosine-1-phosphate (S1P) is a potent bioactive sphingolipid. Two isoforms of sphingosine kinases (SphKs), SK1 and SK2, catalyze the formation of the S1P in mammalian cells. SphKs have also been shown to be up-regulated in the variety of cancer types. SphKs/S1P/S1P receptor (S1PR) axis is involved in multiple biological processes. It has been reported that S1P is involved in cell proliferation, angiogenesis and inflammation. S1P is also involved in cancer progression including cell transformation, oncogenesis and cell survival in hematological malignancies such as multiple myeloma. Therefore, S1P and SphKs may present attractive targets for MM treatment. One of the S1P analog, fingolimod (FTY720), which is an orally active immunomodulatory drug, is developed for the treatment of multiple sclerosis. SKI-I, which is a non-lipid pan-SphK inhibitor and ABC294640, selective inhibitor of SK2, are currently investigated in a pivotal phase 1 clinical trial against solid tumors. In this study, we investigated the efficacy of fingolimod, SKI-I, and ABC294640 by using the MM cell lines, RPMI8226, MM1.S and MM1.R. 72 hours treatment of fingolimod exhibited cell growth inhibition of MM cell lines in a dose dependent manner. Treatment of SKI-I and ABC294640 also exhibited cell growth inhibition in a dose dependent manner. Since S1P is the ligand for a family of five G-protein-coupled receptors with distinct signaling pathways that regulate angiogenesis and chemotaxis, we next evaluated the chemotactic response of human umbilical vein endothelial cells (HUVEC). We found that 4 hours treatment of S1P significantly induced the migration of HUVECs compared to control medium. Treatment of HUVECs with fingolimod inhibited S1P-stimulated chemotaxis in a dose dependent manner. We also found that S1P-induced chemotaxis was abolished by the SKI-I and ABC294640. These results suggest that intracellular SK1 and SK2 may play the important role in S1P induced chemotaxis of HUVEC. We next investigated the S1P concentrations in MM patient by enzyme-linked immune sorbent assay (ELISA), because S1P is a potent tumorigenic growth factor that is likely released from tumor cells. We found that serum concentrations of S1P were significantly higher in patient with MM compared with normal samples. The average S1P levels of MM and normal control are 1503.431 and 1103.38 (p <0.05). We also found that conditioned medium from MM cell line had chemotactic activity for HUVECs. These results implicate that S1P may be a novel biomarker for early stage of MM and that S1P is an important bioactive sphingolipid involved in angiogenesis. In this study, we also demonstrate that fingolimod, SKI-I and ABC294640 have potent preclinical anti-tumor activity in MM. These agents possibly inhibit angiogenesis with relation to MM cell growth and offer unique opportunities for novel therapeutic strategies for the treatment of multiple myeloma. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Evidence for the critical role of the PI3K signaling pathway in particulate matter-induced dysregulation of the inflammatory mediators COX-2/PGE2 and the associated epithelial barrier protein Filaggrin in the bronchial epithelium

2019 ◽  
Vol 36 (4) ◽  
pp. 301-313
Author(s):  
Chenjian Song ◽  
Lingjing Liu ◽  
Junjie Chen ◽  
Yiran Hu ◽  
Jingli Li ◽  
...  
Keyword(s):  
Particulate Matter ◽  
Airway Inflammation ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Transmembrane Protein ◽  
Critical Role ◽  
Pi3k Signaling ◽  
Dependent Manner ◽  
Pi3k Signaling Pathway ◽  
Cox 2

AbstractParticulate matter (PM) is an environmental pollutant closely associated with human airway inflammation. However, the molecular mechanisms of PM-related airway inflammation remains to be fully elucidated. It is known that COX-2/PGE2 play key roles in the pathogenesis of airway inflammation. Filaggrin is a transmembrane protein contributing to tight junction barrier function. As such, Filaggrin prevents leakage of transported solutes and is therefore necessary for the maintenance of epithelial integrity. The objective of the present study was to investigate the regulatory mechanisms of COX-2/PGE2 and Filaggrin upon PM exposure both in vivo and in vitro. C57BL/6 mice received intratracheal instillation of PM for two consecutive days. In parallel, human bronchial epithelial cells (HBECs) were exposed to PM for 24 h. PM exposure resulted in airway inflammation together with upregulation of COX-2/PGE2 and downregulation of Filaggrin in mouse lungs. Corresponding dysregulation of COX-2/PGE2 and Filaggrin was also observed in HBECs subjected to PM. PM exposure led to the phosphorylation of ERK, JNK, and PI3K signaling pathways in a time-dependent manner, while blockade of PI3K with the specific molecular inhibitor LY294002 partially reversed the dysregulation of COX-2/PGE2 and Filaggrin. Moreover, pretreatment of HBECs with NS398, a specific molecular inhibitor of COX-2, and AH6809, a downstream PGE2 receptor inhibitor, reversed the downregulation of Filaggrin upon PM exposure. Taken together, these data demonstrated that the PI3K signaling pathway upregulated COX-2 as well as PGE2 and acted as a pivotal mediator in the downregulation of Filaggrin.

Activity of the Aurora Kinase Inhibitor, MLN8237 (alisertib) Alone or in Combination with Ponatinib Against Imatinib-Resistant BCR-ABL-Positive Cells

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1333-1333
Author(s):  
Seiichi Okabe ◽  
Tetsuzo Tauchi ◽  
Seiichiro Katagiri ◽  
Yuko Tanaka ◽  
Kazuma Ohyashiki
Keyword(s):  
Cell Cycle ◽  
Cell Growth ◽  
Growth Inhibition ◽  
Kinase Inhibitor ◽  
K562 Cells ◽  
Aurora Kinase ◽  
Dependent Manner ◽  
Cell Growth Inhibition ◽  
Cycle Arrest ◽  
T315i Mutation

Abstract Abstract 1333 Chronic myeloid leukemia (CML) is characterized by cytogenetic aberration (Philadelphia chromosome: Ph) and chimeric tyrosine kinase BCR-ABL. ABL tyrosine kinase inhibitor, imatinib has demonstrated the potency against CML patients. However, resistance to imatinib can develop in CML patients due to BCR-ABL point mutations. One of T315I mutation is resistant to currently available ABL tyrosine kinase inhibitors. Therefore, new approach against T315I mutant may improve the outcome of Ph-positive leukemia patients. Aurora kinases are serine/threonine kinases and upregulated in many malignancies including leukemia, and play an important role in cell cycle control and tumor proliferations. Because Aurora kinases are overexpressed in leukemia cells, Aurora kinases may present attractive targets for leukemia treatment. One of Aurora kinase inhibitor, MLN8237 (alisertib) is an oral and selective Aurora kinase A inhibitor and is currently being investigated in a pivotal phase 3 clinical trial against hematological malignancies. We suggested that alisertib mediated inhibition Aurora kinase activity and in combination with ponatinib, also known as AP24534 may abrogate the proliferation and survival of Ph-positive cells including T315I mutation. In this study, we investigated the combination therapy with a ponatinib and an alisertib by using the BCR-ABL positive cell line, K562, murine Ba/F3 cell line which was transfected with T315I mutant, ponatinib resistant Ba/F3 cells and T315I primary sample. Protein expression of Aurora A and B were increased in Ph-positive leukemia cells. 72 hours treatment of alisertib exhibits cell growth inhibition and induced apoptosis against K562 cells in a dose dependent manner. Alisertib also induced cell cycle arrest. The treatment of ponatinib exhibits cell growth inhibition partially against K562 cells in the presence of feeder cell (HS-5) conditioned media. We found that the treatment of alisertib abrogated the protective effects of HS-5 conditioned media in K562 cells. We investigated the alisertib activity against T315I positive cells. Alisertib potently induced cell growth inhibition of Ba/F3 cells ectopically expressing T315I mutation and induced cell cycle arrest. We investigated the efficacy between ponatinib and alisertib by using these cell lines. Combined treatment of Ba/F3 T315I cells with ponatinib and alisertib caused significantly more cytotoxicity than each drug alone. Ponatinib and alisertib were also effective against T315I primary samples. We examined the intracellular signaling of alisertib. Phosphorylation of Aurora A was inhibited in a time dependent manner. We also found the phosphorylation of histone H3 was also reduced in a dose dependent manner suggested that high concentration of alisertib also inhibits Aurora B activity. We next investigated by using ponatinib resistant Ba/F3 cells. In the ponatinib resistant cell lines, IC50 of ponatinib was up to 200 nM. BCR-ABL triple point mutations (T315I, E255K and Y253H) were detected by direct sequence analysis. The treatment of alisertib exhibits cell growth inhibition against Ba/F3 ponatinib resistant cells in the dose dependent manner. Alisertib induced cell cycle arrest in ponatinib resistant cells. Combined treatment of Ba/F3 ponatinib resistant cells with ponatinib and alisertib caused significantly more cytotoxicity. To assess the activity of alisertib and ponatinib, we performed to test on CML tumor formation in mice. We injected nude mice subcutaneously with 1×107 Ba/F3 T315I cells. A dose of 30 mg/kg/day p.o of ponatinib and 30 mg/kg/day p.o of alisertib inhibited tumor growth and reduced tumor volume compared with control mice. The treatments were well tolerated with no animal health concerns observed indicating the feasibility of alisertib combination strategies in the clinic. Data from this study suggested that administration of the ponatinib and Aurora inhibitor, alisertib may be a powerful strategy against BCR-ABL mutant cells including T315I. Disclosures: No relevant conflicts of interest to declare.

Dual HDAC and PI3K Inhibitor, CUDC-907 Alone or in Combination with ABL Tyrosine Kinase Inhibitor Against ABL Tyrosine Kinase Inhibitor Resistant Leukemia Cells

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3693-3693
Author(s):  
Seiichi Okabe ◽  
Tetsuzo Tauchi ◽  
Yuko Tanaka ◽  
Juri Sakuta ◽  
Kazuma Ohyashiki
Keyword(s):  
Cell Growth ◽  
Growth Inhibition ◽  
Tyrosine Kinase Inhibitor ◽  
Kinase Inhibitor ◽  
K562 Cells ◽  
Leukemia Cells ◽  
Cell Growth Inhibition ◽  
Abl Tyrosine Kinase ◽  
Abl Tyrosine Kinase Inhibitor ◽  
Dose Dependent

Abstract Introduction: Chronic myeloid leukemia (CML) is characterized by the t(9:22) translocation known as the Philadelphia chromosome (Ph). ABL tyrosine kinase inhibitor (TKI), imatinib and second-generation ABL TKIs, nilotinib and dasatinib have demonstrated the potency against CML patients. However, resistance to ABL TKI can develop in CML patients due to BCR-ABL point mutations. Moreover, ABL TKIs do not eliminate the leukemia stem cells (LSCs). Therefore, new approach against BCR-ABL mutant cells and LSCs may improve the outcome of Ph-positive leukemia patients. In eukaryotic cells, histone acetylation/deacetylation is important in transcriptional regulation. Chromatin acetylation is controlled by the opposing effects of two families of enzymes: histone acetyltransferases (HAT) and histone deacetylases (HDACs). Deregulation of HDAC activity may be a cause of malignant disease in humans. Phosphoinositide 3-kinase (PI3K) pathway also regulates cell metabolism, proliferation and survival. Furthermore, aberrant activation of PI3K signaling pathway has been shown to be important in initiation maintenance of human cancers. CUDC-907 is an oral inhibitor of class I PI3K as well as class I and II HDAC enzymes. CUDC-907 is currently being investigated in a pivotal phase 1 clinical trial against hematological malignancies such as malignant lymphoma. We suggested that CUDC-907 mediated inhibition PI3K and HDAC activity and in combination with ABL TKIs may abrogate the proliferation and survival of Ph-positive leukemia cells including T315I mutation and ABL TKI resistant. Materials and methods: In this study, we investigated the combination therapy with a CUDC-907 and an ABL TKIs (imatinib, nilotinib and ponatinib) by using the BCR-ABL positive cell line, K562, murine Ba/F3 cell line which was transfected with T315I mutant, nilotinib resistant K562 and ponatinib resistant Ba/F3 cells and primary samples. Results: The treatment of imatinib, nilotinib and ponatinib exhibits cell growth inhibition partially against K562 cells in the presence of feeder cell (HS-5). We found that mRNA of PI3K subunit is significantly increased after a co-culture with HS-5 in K562 and primary CD34 positive CML samples. 72 h treatment of CUDC-907 exhibits cell growth inhibition and induced apoptosis against K562 cells in a dose dependent manner. We examined the intracellular signaling after treatment of CUDC-907. Phosphorylation of JNK, histone acetylation and activity of caspase 3, poly (ADP-ribose) polymerase (PARP) was increased. Anti-apoptotic protein, Mcl-1 was decreased in a dose dependent. We next investigated the efficacy between imatinib and CUDC-907 by using these cell line. Combined treatment of K562 cells with imatinib and CUDC-907 caused significantly more cytotoxicity than each drug alone. Caspase activity was increased and Akt activity was reduced. Phosphorylation of BCR-ABL, Crk-L was reduced and cleaved PARP was increased after imatinib and CUDC-907 treatment. We investigated the CUDC-907 activity against T315I positive cells. CUDC-907 potently induced cell growth inhibition of Ba/F3 T315I cells in a dose dependent manner. Combined treatment of Ba/F3 T315I cells with ponatinib and CUDC-907 caused significantly more cytotoxicity than each drug alone. Caspase activity was increased and Akt activity was reduced after ponatinib and CUDC-907 treatment. To assess the activity of ponatinib and CUDC-907, we performed to test on tumor formation in mice. We injected nude mice subcutaneously with Ba/F3 T315I mutant cells. A dose of 20 mg/kg/day p.o of ponatinib and 30 mg/kg/day p.o of CUDC-907 inhibited tumor growth and reduced tumor volume compared with control mice. The treatments were well tolerated with no animal health concerns observed. We also found that the treatment of CUDC-907 exhibits cell growth inhibition against Ba/F3 ponatinib resistant cells, K562 nilotinib resistant cells, T315I mutant primary samples and CD34 positive CML samples. Conclusion: These results indicated that administration of the dual PI3K and HDAC inhibitor, CUDC-907 may be a powerful strategy against ABL TKI resistant cells including T315I mutation and enhance cytotoxic effects of ABL TKI against those Ph-positive leukemia cells. Disclosures No relevant conflicts of interest to declare.

GDC-0449, the Small Molecule Inhibitor of Hedgehog-Gli Pathway for the Treatment of BCR-ABL Positive Leukemia Cells and In Combination with Dasatinib

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2136-2136
Author(s):  
Seiichi Okabe ◽  
Tetsuzo Tauchi ◽  
Kazuma Ohyashiki
Keyword(s):  
Cell Growth ◽  
Tyrosine Kinase ◽  
Growth Inhibition ◽  
Cell Line ◽  
Kinase Inhibitors ◽  
Leukemia Cells ◽  
Dependent Manner ◽  
Feeder Cell ◽  
Cell Growth Inhibition ◽  
Abl Tyrosine Kinase

Abstract Abstract 2136 Imatinib has shown clinical efficacy against Philadelphia chromosome (Ph) positive leukemia cells and it is now the standard care for initial therapy. However, recent studies reported imatinib are not effective in quiescent primitive chronic myeloid leukemia (CML) stem cells. Moreover, many Ph-positive leukemia patients develop resistance or fail to respond to imatinib by mutation in the ABL kinase domain in clinically. These results indicated that alternative combination therapy such as BCR-ABL targeting tyrosine kinase inhibitors (TKIs) and nontoxic agents are required to cure the Ph positive leukemia patients. Hedgehog (Hh)- Glioma-associated oncogene homolog (Gli) signaling regulates self-renewal of stem cells and implicates in a large number of human cancers. One of the Hh inhibitor, GDC-0449 is a potent small molecule inhibitor of Hedgehog-Gli pathway. It has been reported GDC-0449 showed high target specificity and demonstrated antiproliferative activity against tumors and it is now in clinical trial. Therefore, combination therapy using a BCR-ABL tyrosine kinase inhibitors and a Hedgehog-Gli inhibitor, GDC-0449 may help prevent CML relapse and these approaches may be expected to improve the outcomes of Ph-positive leukemia patients. In this study, we investigated the GDC-0449 efficacy by using the BCR-ABL positive cell lines, OM9;22, K562 and primary samples when leukemic cells were protected by the feeder cell line, S9 cells. We examined a comprehensive drug combination experiment using GDC-0449 and dual Src/ABL tyrosine kinase inhibitor, dasatinib. Gli proteins (Gli1, Gli2 and Gli3) were existed in Ph-positive cell lines. We found the cell numbers of OM9;22 were significantly increased with the feeder cell line, S9 cells compared to without S9 cells. The treatment of dasatinib exhibits cell growth inhibition partially against OM9;22 cells in the presence of feeder cell line, S9 cells. Caspase-3 activity by 100 nM dasatinib treatment was also reduced in the presence of S9 cells. 72 h of combined treatment of Ph-positive leukemia cells with 10 μM of GDC-0449 and 100 nM of dasatinib in the presence of feeder cell line, caused significantly more cytotoxicity than each drug alone. We next investigated the efficacy and intracellular signaling of GDC-0449. The treatment of GDC-0449 exhibits cell growth inhibition and induced apoptosis against OM9;22 cells in a dose and time dependent manner. Expression of Gli1 and Gli2 proteins were reduced after GDC-0449 treatment. 10 μM of GDC-0449 also inhibited the growth of Ph-positive primary samples by colony assay. Another Hh inhibitor, SANT-2 also exhibits cell growth inhibition against OM9;22 cells in a dose dependent manner. Data from this study suggested that administration of the Hh inhibitor, GDC-0449 may be a powerful strategy against Ph-positive leukemia cells and enhance cytotoxic effects of dasatinib in the presence of feeder cell. Disclosures: Ohyashiki: Nippon Shinyaku Co., Ltd.: Research Funding.

Effect of 5-fluoro-2′-deoxycytidine (FdCyd) on p16INK4a, p14ARF, p15INK4b, and DNA methyltransferase 1, 3a, and 3b Genes Expression, Apoptosis Induction, and Cell Growth Inhibition in Pancreatic Cancer AsPC-1 and Hepatocellular Carcinoma LCL-PI 11 Cell Li

2020 ◽  
Author(s):  
Masumeh Sanaei ◽  
Fraidoon Kavoosi
Keyword(s):  
Cell Growth ◽  
Growth Inhibition ◽  
Cell Apoptosis ◽  
Dna Methyltransferase ◽  
Dna Methyltransferase 1 ◽  
Dependent Manner ◽  
Apoptosis Induction ◽  
Cell Growth Inhibition ◽  
Genes Expression ◽  
Methyltransferase 1

Background: Aberrant DNA methylation of the promoter region is one of the most epigenetic changes in numerous cancers. DNA methyltransferase inhibitors (DNMTIs) can revert DNA hypermethylation in tumor suppressor genes (TSGs). The present study was designed to investigate the effect of 5-fluoro-2′-deoxycytidine (FdCyd) on p16INK4a, p14ARF, p15INK4b, and DNA methyltransferase 1, 3a, and 3b genes expression, apoptosis induction, cell growth inhibition in pancreatic cancer AsPC-1 and hepatocellular carcinoma LCL-PI 11 cell lines. Materials and Methods: The cells were treated with FdCyd at different periods. Then, the MTT assay, cell apoptosis assay, and qRT-PCR were done to determine cell viability, cell apoptosis, and the relative gene expression level respectively. Results: The FdCyd decreased DNA methyltransferase 1, 3a, and 3b and increased p16INK4a, p14ARF, and p15INK4b genes expression significantly (P<0.001). Besides, LCL-PI 11 cell was more sensitive to FdCyd in comparison to AsPC-1 cell. FdCyd induced significant cell growth inhibition with a dose- and time-dependent manner (P<0.001). The IC50 value of FdCyd was obtained with approximately 1μM. Further, FdCyd induced cell apoptosis significantly as a time-dependent manner. The number of apoptotic cells was significantly increased in all groups. The percentage of apoptotic cells after 24 and 48 h were 13.86 and 29.6 % in AsPC-1 and 21.04 and 41.52 % in LCL-PI 11 cell line respectively (P<0.001). Conclusion: The FdCyd can reactivate the p16INK4a, p14ARF, and p15INK4b through inhibition of DNA methyltransferase 1, 3a, and 3b gene expression.

Cell growth inhibition by 3-deoxysappanchalcone is mediated by directly targeting the TOPK signaling pathway in colon cancer

Phytomedicine ◽  
2019 ◽  
Vol 61 ◽  
pp. 152813 ◽  
Author(s):  
Ran Zhao ◽  
Hai Huang ◽  
Bu Young Choi ◽  
Xuejiao Liu ◽  
Man Zhang ◽  
...  
Keyword(s):  
Colon Cancer ◽  
Cell Growth ◽  
Growth Inhibition ◽  
Signaling Pathway ◽  
Cell Growth Inhibition
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