The Heme Oxygenase-1-Targeting Compound PEG-ZnPP Inhibits Growth of Imatinib-Resistant BCR/ABL-Transformed Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1986-1986 ◽  
Author(s):  
Matthias Mayerhofer ◽  
Karl J. Aichberger ◽  
Stefan Florian ◽  
Maria-Theresa Krauth ◽  
Sophia Derdak ◽  
...  
Keyword(s):  
Heme Oxygenase ◽  
Kinase Inhibitor ◽  
K562 Cells ◽  
Pi3 Kinase ◽  
Leukemic Cells ◽  
Heme Oxygenase 1 ◽  
Zinc Protoporphyrin ◽  
Inhibitory Effects ◽  
Growth Inhibitory ◽  
Imatinib Resistant

Abstract Chronic myeloid leukemia (CML) is a myeloproliferative disease characterized by the BCR/ABL oncogene and an increased survival of leukemic cells. The BCR-ABL tyrosine kinase inhibitor imatinib has successfully been introduced as a treatment of CML. However, resistance after an intitial response is common in patients with advanced disease, and it is not yet clear if responses in early disease phases will be durable. Therefore, current studies focus on novel potential drug-targets in CML cells. We have recently identified heme oxygenase-1 (HO-1) as a novel BCR/ABL-dependent survival-molecule in primary CML cells. In this study, we analyzed signal transduction pathways underlying BCR/ABL-induced expression of HO-1 and evaluated the role of HO-1 as a potential new target of drug therapy. We found that the PI3-kinase inhibitor LY294002 and MEK inhibitor PD98059 downregulate expression of HO-1 in CML cells. In addition, constitutively active Ras- and Akt -mutants were found to promote expression of HO-1 in Ba/F3 cells, further supporting the involvement of the PI3-kinase/Akt as well as the MAPK pathway in regulating HO-1 expression. To establish a role for HO-1 in survival of CML cells, expression of HO-1 was silenced by siRNAs which resulted in apoptosis of K562 cells. Next, HO-1 was targeted in CML cells by pegylated zinc protoporphyrin (PEG-ZnPP), a competitive inhibitor of HO-1. Exposure to PEG-ZnPP resulted in growth inhibition and induction of apoptosis in primary CML cells as well as in the CML-derived cell lines K562 and KU812 with IC50 values ranging between 1–10 μM. The growth-inhibitory effects of PEG-ZnPP were not only observed in CML cells responsive to imatinib, but also in imatinib-resistant K562 cells and Ba/F3 cells expressing various imatinib-resistant mutants of BCR/ABL (T315I, E255K, M351T, Y253F, Q252H, H396P). Moreover, imatinib and PEG-ZnPP were found to exert synergistic growth inhibitory effects on imatinib-resistant leukemic cells. Together, these data suggest that HO-1 represents a novel drug target in cells expressing BCR/ABL, including those with resistance to imatinib.

Effects of Bosutinib (SKI-606) in CML: Kinase Target Profile, Effects on BCR/ABL Mutants, and Synergism with Dasatinib in T315I+ Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3195-3195
Author(s):  
Karoline Veronika Gleixner ◽  
Lily L Remsing Rix ◽  
Christian Baumgartner ◽  
Uwe Rix ◽  
Alexander Gruze ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Synergistic Effects ◽  
K562 Cells ◽  
Leukemic Cells ◽  
Kinase Assay ◽  
Thymidine Uptake ◽  
Dependent Manner ◽  
Growth Inhibitory ◽  
Imatinib Resistant

Abstract Chronic myeloid leukemia (CML) is a stem cell disease characterized by the BCR/ABL oncoprotein. The ABL kinase inhibitor imatinib is effective in most patients and considered standard first line therapy. However, not all patients show a long-lasting response. Treatment failure is usually associated with the occurrence of imatinib-resistant mutants of BCR/ABL. For these patients, novel multi-kinase inhibitors such as dasatinib represent alternative treatment options. Still, however, not all patients respond to these drugs, especially when leukemic cells bear the BCR/ABL mutant T315I that confers resistance against most kinase-blockers. Bosutinib is a novel multi-kinase inhibitor that has been described to act growth-inhibitory in ABL-transformed leukemias. In the current study, we examined the effects of bosutinib alone and in combination with dasatinib on growth and survival of CML cells. Bosutinib was found to inhibit 3H-thymidine uptake and thus proliferation in imatinib-sensitive and imatinib-resistant K562 cells in a dose-dependent manner, with identical IC50 values (10–100 nM). Moreover, bosutinib was found to inhibit the growth of primary CML cells and Ba/F3 cells bearing various imatinibresistant mutants of BCR/ABL, except the T315I mutant (IC50>1 μM). The growth-inhibitory effects of bosutinib were found to be associated with signs of apoptosis. Dasatinib showed similar effects on CML cells, and again did not block the growth of subclones bearing BCR/ABL T315I. Unexpectedly, however, we found that bosutinib and dasatinib synergize with each other in producing growth inhibition in primary CML cells exhibiting BCR/ABL T315I at pharmacologic concentrations (0.01–1 μM). Clear synergistic effects were also observed in imatinib-sensitive and imatinib-resistant K562 cells as well as in Ba/F3 cells bearing BCR/ABL T315I. In parallel, we performed multiplexed kinase assays as well as chemical proteomics analysis and mass spectrometry using K562 cells and primary CML cells and coupleable dasatinib and bosutinib analogues. In these experiments, dasatinib and bosutinib were found to express an overlapping, but non-identical profile of target kinases. As expected, both drugs were found to bind to wt ABL, SRC kinases, and TEC-family kinases including BTK. Specific targets preferentially bound and inhibited by bosutinib were STE20s, the FES/FER family, CAMKIIG, PYK2 and TBK1. We were also able to confirm that the dasatinib-targets KIT and PDGFRA are not recognized by bosutinib. Interestingly, whereas wt ABL (IC50<0.5 nM) and most of the ABL mutants tested (H396P, M351T, Q252H, and Y253F) were all completely inhibited by both drugs at 1 μM in the kinase assay, the ABL T315I mutant was inhibited by bosutinib (IC50=26 nM) almost 70 times more potently than by dasatinib. Together, these data show that bosutinib and dasatinib synergize with each other in producing antileukemic effects on CML cells including BCR/ABL T315I+ subclones. These synergistic effects may be explained by differential target kinase profiles and by the fact that bosutinib retains some activity against the BCR/ABL T315I mutant kinase.

The Heat Shock Protein 32 (Hsp32)/HO-1-Targeting Drug SMA-ZnPP and the Triterpenoid CDDO-Me Exert Synergistic Growth-Inhibitory Effects on TKI-Resistant Leukemic Cells in Ph+ CML

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4414-4414
Author(s):  
Karoline V. Gleixner ◽  
Harald Herrmann ◽  
Katharina Blatt ◽  
Winfried F Pickl ◽  
Marina Konopleva ◽  
...  
Keyword(s):  
Heat Shock ◽  
Growth Inhibition ◽  
Synergistic Effects ◽  
K562 Cells ◽  
Leukemic Cells ◽  
Drug Resistant ◽  
Inhibitory Effects ◽  
Growth Inhibitory ◽  
Imatinib Resistant ◽  
Ic50 Values

Abstract Abstract 4414 Resistance against one or more tyrosine kinase inhibitors (TKI) prevents eradication of Ph+ chronic myeloid leukemia (CML). In many patients BCR/ABL1 mutations are detectable. We have recently identified two targeted drugs that exert major growth-inhibitory effects on drug-resistant CML cells, the triterpenoid CDDO-Me (Bardoxolone-methyl, REATA Pharma) that blocks several signalling molecules including mTOR, Akt, and STAT3, and upregulates expression of heat shock protein 32 (Hsp32 = heme oxygenase 1, HO-1), and styrene-maleic acid-copolymer micelle-encapsulated ZnPP (SMA-ZnPP), a water-soluble inhibitor of Hsp32/HO-1. In the current project, we asked whether CDDO-Me exerts inhibitory effects on growth of TKI-resistant CML cells and whether the combination of CDDO-Me and SMA-ZnPP would produce synergistic effects in drug-resistant CML cells. As determined by 3H-thymidine incorporation, CDDO-Me was found to inhibit the proliferation of imatinib-responsive and imatinib-resistant K562, imatinib-resistant KCL-22, KU812, and Ba/F3 cells transfected with various TKI-resistant mutants of BCR/ABL1 (T315I, E255K, Y253F, H396P). In each case, IC50 values <1 μM were obtained without major differences between imatinib-responsive and imatinib-resistant cells. Growth-inhibition was accompanied by apoptosis as assessed by combined AnnexinV/PI staining as well as by an increase in expression of HO-1 in KU812 and KCL-22 cells. CDDO-Me was also found to inhibit proliferation of leukemic cells in all patients with TKI-resistant CML (n=4), with IC50 values ranging between <0.1 and 0.5 μM. No differences in IC50 values were observed between treatment-naïve and TKI-resistant cells. Next, we applied the combination CDDO-Me+SMA-ZnPP and found that this combination acts highly synergistically on imatinib-responsive and imatinib-resistant K562 cells as well as primary CML cells isolated from imatinib-naïve CML patients (n=2) or from patients with imatinib-resistant CML (n=2), including one patient in whom BCR/ABL1 T315I was detected. We also examined whether CDDO-Me would exert synergistic effects on CML cells when combined with BCR/ABL1 TKI. In these experiments, we applied the combinations CDDO-Me+dasatinib and CDDO-Me+nilotinib on K562 cells. Both combinations were found to synergistically induce growth inhibition. In conclusion, CDDO-Me inhibits the proliferation of imatinib-resistant BCR/ABL1+ cells, including primary CML cells isolated from untreated patients and cells derived from patients with TKI-resistant CML carrying the BCR/ABL1 mutant T315I. Our data also show that CDDO-Me + SMA-ZnPP and CDDO-Me + BCR/ABL1 TKI synergize in producing growth inhibition in CML cells. Whether these drug combinations also produce synergistic effects in vivo in patients with TKI-resistant CML remains to be evaluated. Disclosures: Valent: Novartis: Consultancy, Honoraria, Research Funding.

Protein Kinase C-βii-Dependent Up-Regulation Of Heme Oxygenase-1 In Chronic Lymphocytic Leukemia Cells Is Crucial For Resistance Against Imatinib

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5560-5560
Author(s):  
Dan Ma ◽  
Jishi Wang ◽  
Qin Fang ◽  
Yan Li
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Mrna Expression ◽  
Heme Oxygenase ◽  
K562 Cells ◽  
Leukemic Cells ◽  
Akt Pathway ◽  
Heme Oxygenase 1 ◽  
Kinase C ◽  
Imatinib Resistant

Abstract Objective Resistance toward imatinib and other BCR/ABL tyrosine kinase inhibitors(TKI) remains an increasing clinical problem in the treatment of advanced stages of chronic myeloid leukemia (CML). Heme oxygenase-1 (HO-1) inhibition plus TKI were found to produce growth inhibition in imatinib-resistant K562/IM cells and in Ba/F3 cells harboring the T315I mutant of BCR/ABL. In this study, we aimed to explore the molecular mechanism of HO-1 explosive expression and relationship between HO-1 and Protein Kinase C-βII(PKC-βII) in imatinib-resistant CML cells. Methods We explored that mRNA expression of various drug-resistant-relative gene between K562 cells and K562/IM cells via high throughout screening assay detected by real-time PCR. According to the results mentioned above, relative protein expression and phosphorylation examined by western blot. And protein translocation was observed and recorded by fluorescence microscope. Cells apoptosis, reactive oxygen species(ROS) and intracellular calcium were analyzed by flow cytometry. Cells proliferation inhibition was detected by CCK-8 assay. The peripheral blood mononuclear cells(PBMNCs) were purified from 45 patients with freshly diagnosed CML or imatinib-resistant CML(including all kinds of drug-resistance involved in various of reasons). Compared the differences of mRNA expression between HO-1 and PKC-βII in those patients. Results Our results indicated that PKC-βII contributed to activation of HO-1 expression through PI3K/AKT pathway. PKC-βII phosphorylation localized to membrane from cytoplasim to activate PI3K/AKT pathway, meanwhile, the 7.8 folder higher (p<0.01) expression of HO-1 following the nucleus translocation of NF-E2–related factor 2(Nrf-2) was observed after pAKT up-regulation. The accumulation of ROS and intracellular calcium overload were reduced by HO-1 to protect cells against cytotoxicity, less apoptotic imatinib-resistant K562 cells were detected at the same concentration of imatinib(the total apoptotic rate of K562/IM:42.4%±3.3% vs. K562: 72.1%±4.1%, at the concentration of 1 μM, p<0.01). In addition, the results of HO-1 and PKC-βII mRNA expression in PBMNCs from 45 patients indicated that HO-1 activation kept positively correspondent with expression of PKC-βII in primary leukemic cells obtained from those patients. In addition, blockage of PI3K/AKT, silence of PKC-βII or HO-1 by small interfere RNA( or inhibited by targeted inhibitor) could significantly increase apoptotic rate of imatinib-resistant CML cells(the apoptotic rate of regulated expression/ natural expression: when blockage of PI3K/AKT: 1.54±0.17; silence of PKC-βII: 2.09±0.23; inhibition of HO-1: 1.87±0.12, p<0.05). Conclusion Selectively targeted-PKC-βII plus imatinib produced growth inhibition in primary leukemic cells. In summary, these data showed that HO-1 expression involved in PKC-βII activation in imatinib-treated CML cells, and targeted- PKC-βII may be the same promising novel strategy as HO-1 inhibition in imatinib resistant CML. Disclosures: No relevant conflicts of interest to declare.

Nilotinib and Dasatinib Produce Synergistic Growth-Inhibitory Effects In Imatinib-Resistant CML Cells, Including Subclones Bearing the Multi-Resistant BCR/ABL Mutant T315I

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2280-2280
Author(s):  
Karoline V. Gleixner ◽  
Harald Herrmann ◽  
Irina Sadovnik ◽  
Karina Schuch ◽  
Winfried F Pickl ◽  
...  
Keyword(s):  
Stem Cells ◽  
Research Funding ◽  
Synergistic Effects ◽  
Chronic Phase ◽  
Leukemic Cells ◽  
Inhibitory Effects ◽  
Blast Phase ◽  
Drug Concentrations ◽  
Growth Inhibitory ◽  
Imatinib Resistant

Abstract Abstract 2280 In most patients with chronic myeloid leukemia (CML), complete cytogenetic remission can be achieved with the BCR/ABL tyrosine kinase inhibitor (TKI) imatinib. However, not all patients are long-term responders. A major cause of acquired resistance against imatinib is the development of BCR/ABL mutations in subclones. In most of these patients, a second generation TKI is prescribed. However, the T315I mutant of BCR/ABL introduces resistance against most TKI, including nilotinib and dasatinib. One approach to overcome drug resistance in BCR/ABL T315I+ CML cells may be to apply drug combinations. Recent data suggest that the mechanisms through which dasatinib and nilotinib act on BCR/ABL differ from each other and that both drugs act on multiple additional targets in CML cells. Here, we show that dasatinib and nilotinib cooperate with each other in producing growth inhibition in imatinib-sensitive and imatinib-resistant CML cells, including subclones bearing BCR/ABL T315I. The drug combination was tested on leukemic cells obtained from 9 patients with chronic phase (CP) CML and 3 with blast phase (PB) of CML. Samples were assessed from 4 patients at the time of diagnosis, and against cells from 8 patients (CP, n=5; BP, n=3) who had developed resistance against one or more BCR/ABL TKI. In all 3 patients in PB, the T315I mutant was detectable. As expected, nilotinib and dasatinib failed to inhibit proliferation of cells harbouring BCR/ABL T315I when applied as single agents. However, the combination xnilotinib+dasatinibx produced synergistic effects in most samples, including primary CML cells and Ba/F3 cells harbouring BCR/ABL T315I. Interestingly, in all 3 patients with BP (BCR/ABL T315I+), strong cooperative or even synergistic growth-inhibitory effects were observed in primary CML cells, resulting in substantial anti-leukemic effects seen at reasonable (pharmacologic) drug concentrations (< 1 μ M) (figure). Based on these results, we treated one patient with TKI-resistant CML in hematologic relapse in whom 2 BCR/ABL mutant-bearing subclones, one clinically resistant against nilotinib (F359V) and one clinically resistant against dasatinib (F317L) had been detected, with a combination of nilotinb (800 mg p.o. daily) and dasatinib (50 mg/day p.o., days 1–5 every third week). A transient hematologic response was obtained in this patient, and except for mild bone pain, no side effects were recorded. Moreover, we were able to show that during treatment with xnilotinib+dasatinibx, the number of CD34+/CD38-/CD33+ CML stem cells decreased from clearly measurable levels (0.005%) to nearly undetectable levels (0.0002%). Finally, ex vivo analyses of leukemic blood cells confirmed, that the combination xnilotinib+dasatinibx produced strong cooperative growth-inhibitory effects in both disease-components, i.e. the F359V-bearing subclone and the F317L-bearing subclone. In summary, our data show that the combination of dasatinib and nilotinib can override acquired TKI resistance in CML, and can suppress growth of various imatinib-resistant subclones including cells that bear BCR/ABL T315I or other BCR/ABL mutants. Whether this combination can suppress imatinib-resistant subclones in CML for prolonged time periods or even can eradicate neoplastic stem cells remains in CML patients to be determined. Synergistic effects of nilotinib and dasatinib on primary leukemic cells obtained from a patient with a BCR/ABL T315I+ blast phase of CML Disclosures: Valent: Novartis: Research Funding; Bristol-Myers Squibb: Research Funding.

Targeting of heat shock protein 32 (Hsp32)/heme oxygenase-1 (HO-1) in leukemic cells in chronic myeloid leukemia: a novel approach to overcome resistance against imatinib

Blood ◽  
2008 ◽  
Vol 111 (4) ◽  
pp. 2200-2210 ◽  
Author(s):  
Matthias Mayerhofer ◽  
Karoline V. Gleixner ◽  
Julia Mayerhofer ◽  
Gregor Hoermann ◽  
Eva Jaeger ◽  
...  
Keyword(s):  
Heat Shock ◽  
Chronic Myeloid Leukemia ◽  
Heat Shock Protein ◽  
Heme Oxygenase ◽  
Myeloid Leukemia ◽  
Kinase Inhibitors ◽  
Leukemic Cells ◽  
Heme Oxygenase 1 ◽  
Abl Tyrosine Kinase ◽  
Imatinib Resistant

Resistance toward imatinib and other BCR/ABL tyrosine kinase inhibitors remains an increasing clinical problem in the treatment of advanced stages of chronic myeloid leukemia (CML). We recently have identified the heat shock protein 32 (Hsp32)/heme oxygenase-1 (HO-1) as a BCR/ABL-dependent survival molecule in CML cells. We here show that silencing Hsp32/HO-1 in CML cells by an siRNA approach results in induction of apoptosis. Moreover, targeting Hsp32/HO-1 by either pegylated zinc protoporphyrine (PEG-ZnPP) or styrene maleic acid-micelle–encapsulated ZnPP (SMA-ZnPP) resulted in growth inhibition of BCR/ABL-transformed cells. The effects of PEG-ZnPP and SMA-ZnPP were demonstrable in Ba/F3 cells carrying various imatinib-resistant mutants of BCR/ABL, including the T315I mutant, which exhibits resistance against all clinically available BCR/ABL tyrosine kinase inhibitors. Growth-inhibitory effects of PEG-ZnPP and SMA-ZnPP also were observed in the CML-derived human cell lines K562 and KU812 as well as in primary leukemic cells obtained from patients with freshly diagnosed CML or imatinib-resistant CML. Finally, Hsp32/HO-1–targeting compounds were found to synergize with either imatinib or nilotinib in producing growth inhibition in imatinib-resistant K562 cells and in Ba/F3 cells harboring the T315I mutant of BCR/ABL. In summary, these data show that HO-1 is a promising novel target in imatinib-resistant CML.

The Multi-Kinase/ABL Inhibitor R763/AS703569 Induces DNA Endoreduplication and Apoptosis In Imatinib-Resistant CML Cells and Synergizes with Nilotinib, Dasatinib, and the Plk-1 Inhibitor BI 2536, In Producing Growth Inhibition.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3394-3394
Author(s):  
Karoline V. Gleixner ◽  
Harald Herrmann ◽  
Barbara Peter ◽  
Katharina Blatt ◽  
Karina Schuch ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Research Funding ◽  
Kinase Inhibitors ◽  
K562 Cells ◽  
Aurora Kinase ◽  
Inhibitory Effects ◽  
Growth Inhibitory ◽  
Imatinib Resistant ◽  
Resistant Mutants ◽  
Bi 2536

Abstract Abstract 3394 Resistance to imatinib is a major clinical problem and challenge in advanced chronic myeloid leukemia (CML). In most patients, drug-resistant mutants of BCR/ABL are detectable. Although most of these mutants still are responsive to second generation BCR/ABL kinase inhibitors (KI) such as nilotinib or dasatinib, drug responses are often short-lived. The BCR/ABL mutant T315I confers resistance against all available BCR/ABL KI, including nilotinib and dasatinib. More recent data suggest that several Aurora kinase (AuK) inhibitors block the kinase activity of BCR/ABL T315I. We have examined the growth-inhibitory effects of the AuK/ABL inhibitor R763/AS703569 (Merck-Serono, Darmstadt, Germany) on primary CML cells (chronic phase, n=12), the CML cell line K562, and Ba/F3 cells transfected with various imatinib-resistant mutants of BCR/ABL. As assessed by 3H-thymidine-uptake, R763/AS703569 was found to inhibit proliferation in imatinib-sensitive and imatinib-resistant primary CML cells in all donors tested, in imatinib-resistant and imatinib-responsive K562 cells, and in Ba/F3 cells harbouring various mutants of BCR/ABL (E255K, Y253F, H396P, T315I). The effects of R763/AS703569 on BCR/ABL-transformed cells were dose-dependent with IC50 values ranging between 0.001–0.1 μ M in K562 cells, <0.001-1 μ M in primary CML cells, and 0.001–0.1 μ M in BCR/ABL+ Ba/F3 cells. In all three patients in whom an imatinib-resistant BCR/ABL mutation was detected (one with V379I, one with F359V, and one with T315), R763/AS703569 was found to inhibit growth of primary CML cells at pharmacologic concentrations (IC50: 0.5, 0.005, and 0.05 μ M, respectively). As assessed by flow cytometry, the growth-inhibitory effects of R763/AS703569 were accompanied by DNA endoreduplication and consecutive apoptosis. Western blot experiments using anti-pCrkL antibody were performed and confirmed that R763/AS703569 blocks BCR/ABL activity at 1 μ M in K562 cells and BCR/ABL-transformed Ba/F3 cells. In addition, R763/AS703569 was found to block Aurora kinase A and Lyn phosphorylation in CML cells. By contrast, no effect of R763/AS703569 on phosphorylation or expression of Polo-like kinase-1 (Plk-1) was seen. In a next step, we explored the effects of drug combinations on growth of K562 cells and BCR/ABL-transformed Ba/F3 cells. In these experiments, R763/AS703569 was found to synergize with the ABL/multi-kinase inhibitors nilotinib (Novartis, Basel Switzerland) and dasatinib (Bristol-Myers Squibb, Princeton, NJ) in producing growth inhibition in CML cells (figure). Moreover, R763/AS703569 and the Plk-1 inhibitor BI 2536 (Boehringer Ingelheim, Vienna, Austria) were found to produce synergistic growth-inhibitory effects on CML cells in all samples tested. Synergistic effects of the KI applied were also seen in Ba/F3 cells exhibiting BCR/ABL T315I. In summary, our data show that the novel AuK/ABL inhibitor R763/AS703569 produces growth inhibition and apoptosis in BCR/ABL-transformed cells including those harbouring BCR/ABL T315I or other imatinib-resistant BCR/ABL mutants. Moreover, our data show that R763/AS703569 synergize with other multi-kinase/ABL inhibitors as well as with the Plk-1 inhibitor BI 2536 in producing growth inhibition in imatinib-resistant CML cells. Whether R763/AS703569, applied alone or in combination with other targeted drugs, can produces anti-leukemic effects in patients with advanced CML remains to be determined in clinical trials. Synergistic growth-inhibitory effects of R763/AS703569 and nilotinib in BCR/ABL T315I+ Ba/F3 cells (left), and R763/AS703569 and dasatinib in K562 cells (right). Disclosures: Valent: Novartis: Research Funding; Bristol-Myers Squibb: Research Funding; Merck-Serono: Research Funding.

Heme Oxygenase-1 (HO-1): A Novel KIT D816V-Dependent Target in Neoplastic Human Mast Cells (HMC-1).

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3521-3521 ◽  
Author(s):  
Rudin Kondo ◽  
Matthias Mayerhofer ◽  
Karoline Gleixner ◽  
Anja Vales ◽  
Maria-Theresa Krauth ◽  
...  
Keyword(s):  
Mast Cells ◽  
Heme Oxygenase ◽  
Kinase Inhibitor ◽  
Northern Blotting ◽  
Heme Oxygenase 1 ◽  
Zinc Protoporphyrin ◽  
Thymidine Uptake ◽  
Dependent Manner ◽  
Survival Factor ◽  
Kit D816v

Abstract Systemic mastoyctosis is a myeloid neoplasm characterized by abnormal growth and accumulation of mast cells (MC). Most patients express the D816V-mutated variant of c-KIT, which mediates resistance against most available tyrosine kinase inhibitors. Therefore, current research is focusing on novel targets in MC. We analyzed expression and function of the survival factor heme oxygenase-1 (HO-1) in neoplastic MC. As assessed by Northern blotting and RT-PCR, the human MC line HMC-1 that exhibits KIT D816V, was found to express HO-1 mRNA. Expression of the HO-1 protein was demonstrable by immunocytochemistry and Western blotting. To examine the role of mutated KIT in expression of HO-1, Ba/F3 cells with doxycycline-inducible expression of c-KIT D816V were employed. In these experiments, KIT D816V was found to induce HO-1 promoter activity and expression of HO-1 mRNA and HO-1 protein in these cells. Moreover, the KIT-D816V-targeting compound PKC412 was found to downregulate expression of HO-1 in HMC-1 cells. The KIT D816V-induced upregulation of HO-1 in Ba/F3 cells was completely blocked by a combination of LY294002 (PI3-kinase inhibitor) and PD89059 (MEK inhibitor), but not by single agents, suggesting involvement of multiple signaling pathways. We next examined whether targeting of HO-1 is associated with decreased survival. As assessed by 3H-thymidine uptake, the pegylated HO-1 inhibitor zinc-protoporphyrin (PEG-ZnPP) reduced proliferation of HMC-1 cells in a dose-dependent manner (IC50: 5 μM). The PEG-ZnPP-induced inhibition of growth was found to be associated with induction of apoptosis (control: 1±0.6 vs PEG-ZnPP, 5 μM: 55±5% apoptotic cells, p&lt;0.05). The HO-1 inductor hemin (10 μM) increased the expression of HO-1 in HMC-1 cells and partly rescued these cells from PKC412-induced growth-inhibition (PKC412, 1 μM: 39±8% vs PKC412 + hemin: 23±11%). Finally, PKC412 and PEG-ZnPP were found to produce cooperative (additive) growth-inhibitory effects on HMC-1 cells. In summary, our data show that HO-1 is a novel survival factor and interesting target in neoplastic human MC exhibiting the D816V-mutated variant of KIT.

BCR/ABL Induces Expression of Histidine Decarboxylase and Synthesis of Histamine in CML Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4835-4835
Author(s):  
Karl J. Aichberger ◽  
Matthias Mayerhofer ◽  
Maria-Theresa Krauth ◽  
Anna Vales ◽  
Sophia Derdak ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Histidine Decarboxylase ◽  
Chronic Phase ◽  
K562 Cells ◽  
Mek Inhibitor ◽  
Pi3 Kinase ◽  
Leukemic Cells ◽  
Biochemical Basis ◽  
X Cells

Abstract Basophil numbers are typically elevated in patients with chronic myeloid leukemia (CML) and characteristically increase during disease progression. As a consequence, blood histamine levels are highly upregulated in CML. We examined the biochemical basis of production of histamine in CML cells and analyzed the effects of the CML-related oncoprotein BCR/ABL on the generation of this mediator. Expression of histamine and of histidine decarboxylase (HDC), the major enzyme involved in histamine synthesis, were examined in primary CML cells obtained from patients with chronic phase (CP) CML or accelerated phase (AP) CML with basophilia, in the CML-derived cell lines K562 and KU812, and in Ba/F3 cells inducibly expressing BCR/ABL on exposure to doxycycline (TonB.210-X cells). In all patients with AP-CML and basophilia, high levels of HDC mRNA and of histamine were detected, whereas in CP-CML cells only expressed low amounts of HDC and of histamine. HDC mRNA and histamine were detectable in the basophil-committed CML cell line KU812, but not in K562 cells. Exposure of primary CML cells or KU812 cells to the BCR/ABL tyrosine kinase inhibitors imatinib (1 μM; Novartis Pharma AG) or AMN107 (100 nM; Novartis Pharma AG) decreased the levels of histamine and expression of HDC in BCR/ABL-transformed cells. Moreover, BCR/ABL was found to promote the expression of HDC mRNA and to increase the levels of histamine in TonB.210-X cells. The BCR/ABL-induced synthesis of histamine in these cells was blocked by the PI3-kinase inhibitor LY294002, whereas neither the MEK inhibitor PD98059, nor the mTOR inhibitor rapamycin affected histamine levels. In conclusion, our data show that the CML-specific oncoprotein BCR/ABL induces expression of HDC and synthesis of histamine in leukemic cells through a pathway involving the PI3-kinase.

scholarly journals 6-Shogaol protects against ischemic acute kidney injury by modulating NF-κB and heme oxygenase-1 pathways

2019 ◽  
Vol 317 (3) ◽  
pp. F743-F756 ◽  
Author(s):  
Sang Jun Han ◽  
Mihwa Kim ◽  
Vivette D. D’Agati ◽  
H. Thomas Lee
Keyword(s):  
Acute Kidney Injury ◽  
Proximal Tubule ◽  
Heme Oxygenase ◽  
Kidney Injury ◽  
Heme Oxygenase 1 ◽  
Zinc Protoporphyrin ◽  
Mrna Synthesis ◽  
Proximal Tubule Cells ◽  
Tnf Α ◽  
Tubule Cells

Acute kidney injury (AKI) due to renal ischemia-reperfusion (I/R) is a major clinical problem without effective therapy. Ginger is one of the most widely consumed spices in the world, and 6-shogaol, a major ginger metabolite, has anti-inflammatory effects in neuronal and epithelial cells. Here, we demonstrate our novel findings that 6-shogaol treatment protected against renal I/R injury with decreased plasma creatinine, blood urea nitrogen, and kidney neutrophil gelatinase-associated lipocalin mRNA synthesis compared with vehicle-treated mice subjected to renal I/R. Additionally, 6-shogaol treatment reduced kidney inflammation (decreased proinflammatory cytokine and chemokine synthesis as well as neutrophil infiltration) and apoptosis (decreased TUNEL-positive renal tubular cells) compared with vehicle-treated mice subjected to renal I/R. In cultured human and mouse kidney proximal tubule cells, 6-shogaol significantly attenuated TNF-α-induced inflammatory cytokine and chemokine mRNA synthesis. Mechanistically, 6-shogaol significantly attenuated TNF-α-induced NF-κB activation in human renal proximal tubule cells by reducing IKKαβ/IκBα phosphorylation. Furthermore, 6-shogaol induced a cytoprotective chaperone heme oxygenase (HO)-1 via p38 MAPK activation in vitro and in vivo. Consistent with these findings, pretreatment with the HO-1 inhibitor zinc protoporphyrin IX completely prevented 6-shogaol-mediated protection against ischemic AKI in mice. Taken together, our study showed that 6-shogaol protects against ischemic AKI by attenuating NF-κB activation and inducing HO-1 expression. 6-Shogaol may provide a potential therapy for ischemic AKI during the perioperative period.

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