Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P10415 (Bcl-2)
 33,771 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of the non-tumor-promoting protein kinase C (PKC) activator bryostatin 1 and the PKC inhibitors staurosporine and UCN-01 were examined with respect to modulation of 1-[beta-D-arabinofuranosyl]cytosine (ara-C)-induced apoptosis in human myeloid leukemia cells (HL-60) overexpressing the antiapoptotic protein Bcl-2. HL-60/Bcl-2 cells displayed a 5-fold increase in Bcl-2 protein compared with empty-vector counter-parts (HL-60/pCEP4) but comparable levels of Bax, Mcl-1, and Bcl-xL. After exposure to an equimolar concentration of ara-C (10 microM for 6 hr), HL-60/Bcl-2 cells were significantly less susceptible to apoptosis, DNA fragmentation, and loss of clonogenicity than HL-60/pCEP4 cells. The protective effect of increased Bcl-2 expression was manifested by a failure of ara-C to induce activation/cleavage of the Yama protease (CPP32; caspase-3) and degradation of one of its substrates, poly(ADP-ribose)polymerase to an 85-kDa cleavage product. When HL-60/Bcl-2 cells were preincubated with bryostatin 1 (10 nM; 24 hr) or coincubated with either staurosporine (50 nM; 6 hr) or UCN-01 (300 nM; 6 hr) after a 1-hr preincubation, exposures that exerted minimal effects alone, ara-C-induced apoptosis and DNA fragmentation were restored to levels equivalent to, or greater than, those observed in empty-vector controls. These events were accompanied by restoration of the ability of ara-C to induce CPP32 cleavage and activation, poly(ADP-ribose) polymerase degradation, and inhibition of colony formation. Western analysis of Bcl-2 protein obtained from overexpressing cells treated with bryostatin 1, staurosporine, or UCN-01 revealed the appearance of a slowly migrating species and a general broadening of the protein band, effects that were insensitive to the protein synthesis inhibitor cycloheximide. Alterations in Bcl-2 protein mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis were reversed by treatment of lysates with alkaline phosphatase or protein phosphatase 2A; actions of the latter were blocked by the specific phosphatase inhibitor okadaic acid. In vivo labeling studies of Bcl-2 protein demonstrated increased incorporation of [32PO4]orthophosphate in drug-treated cells. Last, phosphorylated Bcl-2 failed to display decreased binding to the proapoptotic protein Bax. Collectively, these findings indicate that bryostatin 1, which down-regulates PKC, and staurosporine and UCN-01, which directly inhibit the enzyme, circumvent resistance of Bcl-2-overexpressing leukemic cells to ara-C-induced apoptosis and activation of the protease cascade. They also raise the possibility that modulation of Bcl-2 phosphorylation status contributes to this effect.
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PMID:Agents that down-regulate or inhibit protein kinase C circumvent resistance to 1-beta-D-arabinofuranosylcytosine-induced apoptosis in human leukemia cells that overexpress Bcl-2. 939 80

Our goal was to determine the cytotoxicity of 7-OH-hydroxystaurosporine (UCN-01) as a single agent and in combination with cis-diamminedichloroplatinum(II) (CDDP) in a panel of ovarian carcinoma cells. We sought to examine the role of p53 gene function and alterations in cell cycle progression or other mechanisms of action of UCN-01 including perturbation of the apoptosis pathway mediated by NF-kappaB and Bcl-2/Bax. Cytotoxicity was determined from dose-response curves established by the Alamar blue vital dye indicator assay. Restoration of wild-type p53 in a p53 null cell line, SKOV 3, was achieved by transfection of a p53 expression vector. Cell cycle distribution was measured by fluorescence-activated cell sorting analysis of ethidium bromide-stained nuclei. Apoptosis was measured by quantitative fluorescence microscopy. NF-kappaB DNA binding activity was measured by electrophoretic mobility shift assay. Bcl-2 and Bax levels were determined by Western immunoblotting. UCN-01 was effective as a cytotoxic agent alone and in combination with CDDP in all cell lines studied, regardless of p53 status. The degree of sensitization to CDDP conferred by UCN-01, however, was found to correlate with p53 gene status. p53 wild-type cells seem to be more sensitive to the cytotoxic effects of the combination of UCN-01 + CDDP than the p53 mutant cells. This was confirmed in cells in which p53 wild-type function was restored by transfection of p53 cDNA, but these cells are also significantly more sensitive to CDDP alone. The effects of UCN-01 on cell cycle progression also appear to be p53 dependent but may not be the primary mechanism of action. The rate of apoptosis is increased 4-fold in UCN-01 + CDDP-treated cells compared to either agent alone. UCN-01 does not effect NF-kappaB DNA binding activity or Bcl-2 and Bax levels. UCN-01 enhances CDDP cytotoxicity and apoptosis in ovary cancer cells. This occurs regardless of p53 status, but wild-type p53 seems to increase the degree of sensitization.
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PMID:UCN-01 in ovary cancer cells: effective as a single agent and in combination with cis-diamminedichloroplatinum(II)independent of p53 status. 981 1

Derivatives of camptothecins, topoisomerase I inhibitors and 7-hydroxystaurosporine (UCN-01), a protein kinase C (PKC) inhibitor and cell cycle checkpoint abrogator, are promising anticancer drugs. We characterized the apoptotic response to camptothecin and UCN-01 for the 8 human breast carcinoma cell lines (MCF-7, MCF-7/ADR, T47D, HS578T, BT549, MDA-N, MDA MB231, MDA435) from the National Cancer Institute (NCI) Anticancer Drug Screen. MCF-7 and T47D cells exhibited marked resistance to apoptosis, whereas MCF-7/ADR (NCI/ADR-RES) and HS578T cells exhibited the most pronounced apoptotic response. Apoptotic response was not correlated with growth inhibition measured by sulforhodamine B (SRB) assay, indicating that apoptosis is not the only mechanism of drug-induced cell death. Measurements of topoisomerase I levels and cleavage complexes and of PKC isoforms demonstrated that primary target inhibition was not correlated with apoptotic response. Several key apoptotic pathways were evaluated. Only MCF-7 cells had wild-type p53, indicating that p53 is not required for drug-induced apoptosis. MCF-7 cells also showed the highest MDM-2 expression (along with T47D cells, which were also resistant to apoptosis). Bcl-2, Mcl-1 and caspases 2 and 3 protein levels varied widely, whereas Bax expression was comparable among cell lines. Interestingly, Bcl-2, Mcl-1 and Bcl-X(L) cumulative expressions were inversely correlated with apoptotic response. Our results provide a comparative molecular characterization for the breast cancer cell lines of the NCI Anticancer Drug Screen and demonstrate the diversity of cellular responses to drugs (apoptosis vs. cell cycle arrest) and the importance of multifactorial analyses for modulating/predicting the apoptotic response to chemotherapy.
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PMID:Apoptotic response to camptothecin and 7-hydroxystaurosporine (UCN-01) in the 8 human breast cancer cell lines of the NCI Anticancer Drug Screen: multifactorial relationships with topoisomerase I, protein kinase C, Bcl-2, p53, MDM-2 and caspase pathways. 1039 57

Compounds that inhibit protein kinases are currently undergoing clinical evaluation for the treatment of a variety of malignancies. The kinase inhibitors flavopiridol and 7 hydroxy-staurosporine (UCN-01) were examined for their effects on B-cell chronic lymphocytic leukemia (B-CLL) cells in vitro (n = 49). Flavopiridol and UCN-01 induced concentration-dependent apoptosis of most B-CLL samples tested, with greater than 50% cell killing occurring at concentrations of less than 1 mcmol/L, and with flavopiridol displaying more potent activity than UCN-01. Flavopiridol (0.1 mcmol/L) and UCN-01 (1 mcmol/L) also induced striking decreases in the levels of the antiapoptosis proteins Mcl-1, X-linked inhibitor of apoptosis (XIAP), and BAG-1 in nearly all cases of B-CLL and of Bcl-2 in approximately half of B-CLL specimens evaluated. In contrast, expression of the proapoptotic proteins Bax and Bak was not significantly influenced by these kinase inhibitors. Flavopiridol-induced decreases in the levels of antiapoptosis proteins Mcl-1 and XIAP preceded apoptosis and were not substantially affected by the addition of caspase inhibitors to cultures. In contrast, UCN-01-stimulated decreases in antiapoptosis proteins were slower, occurred concurrently with apoptosis, and were partially prevented by caspase inhibitors. The findings suggest that flavopiridol and UCN-01 induce apoptosis of B-CLL cells through different mechanisms. The potent apoptotic activities of flavopiridol and UCN-01 against cultured B-CLL cells suggest that they may be effective as single agents in the treatment of B-CLL or for sensitizing B-CLL cells to conventional cytotoxic drugs. (Blood. 2000;96:393-397)
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PMID:Protein kinase inhibitors flavopiridol and 7-hydroxy-staurosporine down-regulate antiapoptosis proteins in B-cell chronic lymphocytic leukemia. 1088 97

Antileukemic interactions between the nucleoside analog 1-beta-D-arabinofuranosylcytosine (ara-C) and the kinase inhibitor 7-hydroxystaurosporine (UCN-01) have been examined in relation to Bcl-2 expression/phosphorylation, mitochondrial damage, caspase activation, and loss of clonogenic potential. Subsequent exposure of ara-C-pretreated U937 cells (1 microM; 6 hr) to UCN-01 (300 nM; 24 hr) resulted in marked potentiation of pro-caspase-3 and -9 cleavage/activation, poly(ADP-ribose)polymerase degradation, diminished mitochondrial membrane potential (Deltapsi(m)), enhanced cytochrome c release, reduction in the S-phase fraction, and induction of classic apoptotic morphologic features. Enforced expression of full-length Bcl-2 significantly protected cells (at 24 hr) from ara-C/UCN-01-induced caspase activation and apoptosis, but was ineffective in preventing loss of Deltapsi(m) and cytochrome c release. Ectopic expression of a Bcl-2 N-terminal phosphorylation loop-deleted protein (Bcl-2Delta(32-80)) was more potent than its full-length counterpart in blocking drug-induced loss of Deltapsi(m, ) caspase activation, and apoptotic morphology, but not cytochrome c release. Examination of cells at later intervals revealed that ectopic expression of Bcl-2 or Bcl-2Delta(32-80) could only delay, but not prevent, mitochondrial damage, caspase activation, and cell death induced by ara-C/UCN-01 treatment. Despite their initial ability to inhibit apoptosis, neither full-length nor truncated Bcl-2 protein restored clonogenic potential to drug-treated cells. These findings indicate that subsequent exposure of ara-C-pretreated human leukemia cells to UCN-01 potently triggers mitochondrial damage and apoptosis, and that these events are postponed but not prevented by ectopic expression of Bcl-2 or its phosphorylation loop-deleted counterpart.
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PMID:Potentiation of 1-beta-D-arabinofuranosylcytosine-mediated mitochondrial damage and apoptosis in human leukemia cells (U937) overexpressing bcl-2 by the kinase inhibitor 7-hydroxystaurosporine (UCN-01). 1102 Apr 46

Interactions between the purine analogue 2-fluoroadenine 9-beta-D-arabinofuranoside (F-ara-A) and the kinase inhibitor UCN-01 have been examined in human leukemia cells (U937 and HL-60) with respect to induction of mitochondrial damage, caspase activation, apoptosis, and loss of clonogenic survival. Simultaneous or subsequent exposure of F-ara-A-treated cells (2 microM) to UCN-01 (100 nM) resulted in a marked potentiation of apoptosis, manifested by loss of mitochondrial membrane potential (delta psi(m)), cleavage/activation of procaspase-9 and procaspase-3, DNA fragmentation, and degradation of poly-ADP(ribosyl) polymerase. Coadministration of UCN-01 with F-ara-A was also associated with diminished phosphorylation of the cdc25 phosphatase. In contrast, exposure of cells to the sequence UCN-01, followed by F-ara-A, resulted in only a modest increase in apoptotic cells. The ability of UCN-01 to potentiate F-ara-A-mediated lethality was not mimicked by the selective PKC inhibitor bisindolylmaleimide, nor did treatment of cells with UCN-01 enhance formation of F-ara-ATP or increase incorporation of [3H]F-ara-A into DNA. Enhanced apoptosis in cells exposed sequentially or simultaneously to F-ara-A and UCN-01 was accompanied by a substantial reduction in colony formation (e.g., to 0.01% of control values). Cotreatment with UCN-01 also increased F-ara-A-mediated apoptosis and loss of delta psi(m) in U937 cells ectopically expressing Bcl-2, although not to the same extent as that observed in empty-vector controls. Finally, simultaneous exposure (24 h) of malignant B lymphocytes from the pleural effusion of a patient with indolent non-Hodgkin's lymphoma to F-ara-A and UCN-01 ex vivo resulted in a striking increase in apoptosis, as determined by terminal deoxynucleotidyltransferase-mediated nick end labeling assay. These findings indicate that UCN-01 increases F-ara-A-induced mitochondrial damage and apoptosis in human leukemia cells in a sequence-dependent manner, and that these events occur in at least some primary human lymphoma cells.
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PMID:Interactions between 2-fluoroadenine 9-beta-D-arabinofuranoside and the kinase inhibitor UCN-01 in human leukemia and lymphoma cells. 1123 87

Interactions between the checkpoint abrogator UCN-01 and several pharmacological inhibitors of the mitogen-activated protein kinase (MAPK) kinase (MEK)/MAPK pathway have been examined in a variety of human leukemia cell lines. Exposure of U937 monocytic leukemia cells to a marginally toxic concentration of UCN-01 (e.g., 150 nM) for 18 h resulted in phosphorylation/activation of p42/44 MAPK. Coadministration of the MEK inhibitor PD184352 (10 microM) blocked UCN-01-induced MAPK activation and was accompanied by marked mitochondrial damage (e.g., cytochrome c release and loss of DeltaPsi(m)), caspase activation, DNA fragmentation, and apoptosis. Similar interactions were noted in the case of other MEK inhibitors (e.g., PD98059; U0126) as well as in multiple other leukemia cell types (e.g., HL-60, Jurkat, CCRF-CEM, and Raji). Coadministration of PD184352 and UCN-01 resulted in reduced binding of the cdc25C phosphatase to 14-3-3 proteins, enhanced dephosphorylation/activation of p34(cdc2), and diminished phosphorylation of cyclic AMP-responsive element binding protein. The ability of UCN-01, when combined with PD184352, to antagonize cdc25C/14-3-3 protein binding, promote dephosphorylation of p34(cdc2), and potentiate apoptosis was mimicked by the ataxia telangectasia mutation inhibitor caffeine. In contrast, cotreatment of cells with UCN-01 and PD184352 did not substantially increase c-Jun-NH(2)-terminal kinase activation nor did it alter expression of Bcl-2, Bcl-x(L), Bax, or X-inhibitor of apoptosis. However, coexposure of U937 cells to UCN-01 and PD184352 induced a marked increase in p38 MAPK activation. Moreover, SB203580, which inhibits multiple kinases including p38 MAPK, partially antagonized cell death. Lastly, although UCN-01 +/- PD184352 did not induce p21(CIP1), stable expression of a p21(CIP1) antisense construct significantly increased susceptibility to this drug combination. Together, these findings indicate that exposure of leukemic cells to UCN-01 leads to activation of the MAPK cascade and that interruption of this process by MEK inhibition triggers perturbations in several signaling and cell cycle regulatory pathways that culminate in mitochondrial injury, caspase activation, and apoptosis. They also raise the possibility that disrupting multiple signaling pathways, e.g., by combining UCN-01 with MEK inhibitors, may represent a novel antileukemic strategy.
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PMID:Pharmacological inhibitors of the mitogen-activated protein kinase (MAPK) kinase/MAPK cascade interact synergistically with UCN-01 to induce mitochondrial dysfunction and apoptosis in human leukemia cells. 1143 48

7-Hydroxystaurosporine (UCN-01) is a selective protein kinase C (PKC) inhibitor and is being developed as a novel anticancer agent. Because of reports that PKC may be involved in the pathogenesis of some forms of thyroid cancers, we examined four thyroid carcinoma lines (FRO, KAT5, NPA, and WRO). These cells were found to have different susceptibility to UCN-01 treatment, and there appeared to be a correlation between UCN-01-induced death and expression levels of endogenous Bcl-2. KAT5 cells, which normally express a low amount of Bcl-2, exhibited significantly higher sensitivity to UCN-01-induced death than the other cell lines. Of interest, susceptibility did not relate to PKC activity or its inhibition by UCN-01. In order to investigate the role of Bcl-2 in UCN-01-induced death, KAT5 cells were transfected to overexpress Bcl-2. KAT5/Bcl-2 cells were capable of conferring resistance to UCN-01-induced death. Furthermore, upregulating of Bcl-2 by 1alpha,25-dihydroxyvitamin D3 (VD3) could protect primary thyroid cell from death induced by UCN-01. Both in situ TUNEL staining and the flow cytometric analysis of cytokeratin-18 (CK18) cleavage confirmed that UCN-01 was indeed inducing apoptosis, and that this effect was inhibited by increased expression of Bcl-2. These results suggest that the Bcl-2 can block the UCN-01-activated cell death pathway and that the expression of Bcl-2 is inversely related to thyroid carcinoma cell susceptibility to UCN-01. Therefore, the analysis of the expression of apoptosis suppressors provides a basis for the use of UCN-01 in the treatment of thyroid cancer.
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PMID:Susceptibility of thyroid cancer cells to 7-hydroxystaurosporine-induced apoptosis correlates with Bcl-2 protein level. 1152 64

A clinically relevant dose (2.0 Gy) of ionizing radiation (IR) was employed to determine if subsequent exposure to the protein kinase C (PKC) and Chk 1 inhibitor UCN-01 for 24 h could abrogate IR-induced G2/M arrest and promote apoptosis in U937 leukemic cells ectopically expressing Bcl-2 (U937/Bcl-2). To this end, empty-vector control (U937/pCEP4) and U937/Bcl-2 cells were exposed to two UCN-01 concentrations following IR: i) a 50 nM concentration, which by itself was minimally toxic to both cell lines, and ii) a 150 nM concentration, which modestly induced apoptosis (e.g., ~19%) in control cells after 24 h. The effects of UCN-01 on IR responses were examined in relation to apoptosis induction, suspension culture growth inhibition, loss of clonogenic survival, and cell cycle perturbations. IR (2 Gy) alone minimally induced apoptosis in both U937 transfectant cell lines (e.g., <5% at 24 h in each case). Although UCN-01 failed to potentiate IR-mediated apoptosis at either early (e.g., 24 h) or late (e.g., 72 h) intervals, exposure to 50 or 150 nM UCN-01 resulted in a significant, albeit modest, reduction in proliferation and colony formation in irradiated U937/pCEP4 and U937/Bcl-2 cells. Despite failing to enhance apoptosis, UCN-01 treatment abrogated IR-induced G2/M arrest in both cell lines, an event associated with enhanced activation of cyclin-dependent kinase 1 (cdk1), promotion of G0/G1 arrest, and dephosphorylation of the retinoblastoma protein (pRb). Together, these findings indicate that exposure of U937 cells ectopically-expressing Bcl-2 to the combination of UCN-01 + IR leads to a further reduction in cell proliferation, and that this phenomenon appears to involve a non-apoptotic mechanism.
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PMID:7-hydroxystaurosporine (UCN-01) and ionizing radiation combine to inhibit the growth of Bcl-2-overexpressing U937 leukemia cells through a non-apoptotic mechanism. 1211 31

An increasing number of unique active new chemotherapeutic and biologic agents are currently available for clinical research studies. Nucleoside analogs in development for non-Hodgkin's lymphoma (NHL) include clofarabine, troxacitabine, and bendamustine, a hybrid of an alkylating nitrogen mustard group and a purine-like benzimidazole, with demonstrated activity in NHL. Drugs directed at the cell cycle include flavopiridol and UCN-01. The proteasome plays a pivotal role in cellular protein regulation and activation of NFkappaB, which maintains cell viability through the transcription of inhibitors of apoptosis. PS-341 is a specific, selective inhibitor of the 26S proteasome which induces apoptosis and has activity in cell types characterized by overexpression of Bcl-2. Response rates of 50%, including complete remissions, have been reported using this agent in patients with refractory multiple myeloma. Studies are ongoing in NHL and chronic lymphocytic leukemia. G3139, an antisense oligonucleotide, has shown promise in early studies. Rituximab has revolutionized the treatment of NHL. However, other active antibodies are now available, including alemtuzumab, epratuzumab, and Hu1D10. The radioimmunoconjugates (90)Y-ibritumomab tiuxetan and (131)I-tositumomab may also play an important role in the management of NHL. Future therapeutic strategies should involve rational combinations of new chemotherapy drugs, biologic agents, and antisense compounds to increase the cure rate in patients with lymphoma.
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PMID:Hematologic malignancies: new developments and future treatments. 1217 Apr 31


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