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

Retinoic acid and hydrocortisone (HC) have been shown to regulate the drug sensitivity of the blast cells of acute myeloblastic leukemia (AML). We asked if the proto-oncogene bcl-2 played a role in this regulation. As target cells we used the continuous lines, OCI/AML-1, OCI/AML-2 or OCI/AML-5; expression of bcl-2 can be detected by Northern analysis of RNA from OCI/AML-2 or OCI/AML-5 cells; bcl-2 expression can be found in OCI/AML-1 cells only by using RT-PCR. Exposure of OCI/AML-2 or OCI/AML-5 cells to retinoic acid (all-trans retinoic acid, ATRA) led to a down-regulation of bcl-2 expression that was first seen after 2 h of exposure and was complete after a day. The down-regulation could be prevented by exposing the cells to ara-C either before or after ATRA; decrease in bcl-2 protein was moderate and only obvious after 36 h of ATRA treatment. Nuclear run-on experiments provided evidence that bcl-2 down-regulation was occurring at transcriptional and post-translational levels. Since bcl-2 is considered to have anti-oxidant activity, we tested the sensitivity of the three cell lines to H2O2; we found that OCI/AML-1, the line with very low bcl-2 expression, was a 100-fold more H2O2-sensitive than OCI/AML-2 or OCI/AML-5, where bcl-2 expression can be detected readily. We then asked if H2O2 sensitivity could be regulated. We found that exposure of cells to HC before H2O2 was protective while ATRA after peroxide treatment increased killing; this is the same pattern of regulation observed when AML blasts are exposed to HC before, or ATRA after ara-C. Finally, we asked whether N-acetylcysteine (NAC), a known radical scavenger would protect cells against ara-C killing. Significant protection was observed when NAC was given before drug, but not if given after drug. NAC protection against ara-C killing was seen for OCI/AML-1 and 2 cells, but not for OCI/AML-5 cells. We interpret the results as follows: ara-C kills cells in two ways: first, directly, by incorporation into DNA and chain termination; second, indirectly, by inducing the production of toxic radicals. Bcl-2 reduces the oxidant activity of such radicals, and is protective. ATRA regulates ara-C toxicity by its action on bcl-2. Left unexplained are the action of HC, which does not affect bcl-2 expression and the mechanism by which ara-C prevents down-regulation of bcl-2 by ATRA.
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PMID:Mechanism of cytosine arabinoside toxicity to the blast cells of acute myeloblastic leukemia: involvement of free radicals. 776 41

The bcl-2 gene is expressed in many types of human tumours and becomes transcriptionally deregulated in the majority of non-Hodgkin's lymphomas as the result of t(14;18) chromosomal translocations. The 26-kDa Bcl-2 protein has been shown to block programmed cell death (apoptosis) induced by many types of stimuli, including a wide variety of chemotherapeutic drugs and radiation. The presence of bcl-2 in tumor cells has been correlated with poor responses to therapy in patients with some types of cancer. To explore further the relevance of bcl-2 to drug resistance, we used antisense (As) approaches to achieve reductions in the levels of steady state Bcl-2 protein levels in t(14;18)-containing human lymphoma cell lines. Both synthetic bcl-2-As oligonucleotides and inducible expression plasmids that produce bcl-2-As transcripts induced reductions in bcl-2 expression, resulting in a marked enhancement in the sensitivity of neoplastic cells to conventional chemotherapeutic drugs such as cytosine arabinoside (ara-C) and methotrexate (MTX). These results suggest that novel therapeutics targeted against bcl-2 could provide the means for improved treatment of cancer by affecting physiological pathways distal to the targets of cytotoxic drugs.
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PMID:Reversal of chemoresistance of lymphoma cells by antisense-mediated reduction of bcl-2 gene expression. 795 Mar 2

1-beta-D-Arabinofuranosylcytosine (ara-C) is an anti-leukemic agent that incorporates into cellular DNA leading to inhibition of DNA synthesis and loss of clonogenic survival. In contrast, ionizing radiation induces DNA damage through the generation of reactive oxygen intermediates. Although little is known of the specific determinants of ara-C and ionizing radiation-induced cytotoxicity, recent work has shown that both are capable of inducing internucleosomal DNA fragmentation in a pattern consistent with programmed cell death (apoptosis). In order to assess the importance of apoptosis in drug and ionizing radiation-induced cytotoxicity in the U-937 myelomonocytic cell line, we created cell lines that constitutively express a transfected bcl-2 gene. Bcl-2 was capable of inhibiting 40-50% of the ara-C and ionizing radiation-induced internucleosomal DNA fragmentation at all tested concentrations. However, cell survival following exposure to these agents was only increased in the bcl-2 transfectants at relatively low doses of ara-C and ionizing radiation. These data demonstrate that although bcl-2 is capable of inhibiting ara-C and ionizing radiation-induced DNA fragmentation in myeloid cells, it increases cell survival only at low doses of these agents. This suggests that apoptosis may be a less important mechanism of cytotoxicity at higher doses of ara-C and ionizing radiation than it is at lower doses.
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PMID:Effect of Bcl-2 on ionizing radiation and 1-beta-D-arabinofuranosylcytosine-induced internucleosomal DNA fragmentation and cell survival in human myeloid leukemia cells. 826 Jul 51

Human chronic myelogenous leukemia-blast crisis K562 cells have been demonstrated to be relatively resistant to antileukemic drug-induced apoptosis. This has been attributed to the activity of p210bcr-abl tyrosine kinase present in the K562 cells, which is known to suppress drug-induced apoptosis. Recently, K562 cells have been shown to express the antiapoptosis Bcl-xL but not Bcl-2 proteins. To investigate the contribution of Bcl-xL toward resistance to drug-induced apoptosis, we created K562/Bcl-xS and K562/neo cells by electroporating the expression plasmids pSFFVneo-Bcl-xS and pSFFVneo, containing the bcl-xS and neomycin resistance genes, respectively, into K562 cells. K562/Bcl-xS but not K562/neo cells expressed the bcl-xS mRNA and p19Bcl-xS protein. In contrast, both cell types expressed equivalent levels of Bcl-xL, Bax, Bcl-2, Myc, retinoblastoma, p21cbor-abl, and p145abl proteins. A significant increase in the hemoglobin levels was observed in the K562/Bcl-xS compared with the K562/neo cells (P < 0.05). In addition, K562/Bcl-xS cells were significantly more sensitive than K562/neo cells to undergoing erythroid differentiation induced by low-dose 1-beta-D-arabinofuranosylcytosine (ara-C) and hexamethyl bisacetamide (P < 0.05), but not by all-trans-retinoic acid. Low-dose ara-C- or hexamethyl bisacetamide-induced differentiation was not associated with apoptosis of K562/Bcl-xS or K562/neo cells. Low-dose ara-C-induced erythroid differentiation was accompanied by conversion of the retinoblastoma protein to predominantly its underphosphorylated isoform as well as by down-regulation of Myc levels in K562/Bcl-xS and K562/neo cells. Importantly, exposure to high-dose ara-C (HIDAC; 100 microM ara-C for 4 h) caused internucleosomal DNA fragmentation and the morphological features of apoptosis in K562/Bcl-xS cells. These effects were modestly enhanced by cotreatment with HIDAC plus herbimycin A. In contrast, K562/neo cells were completely resistant to HIDAC- and herbimycin A-induced apoptosis. These results indicate that the expression of Bcl-xS induces erythroid differentiation and partially sensitizes chronic myelogenous leukemia-blast crisis-derived K562 cells to ara-C-induced differentiation and apoptosis.
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PMID:Enforced expression of Bcl-XS induces differentiation and sensitizes chronic myelogenous leukemia-blast crisis K562 cells to 1-beta-D-arabinofuranosylcytosine-mediated differentiation and apoptosis. 895 29

Modulation of ara-C-induced apoptosis in human leukemia cells by the macrocyclic lactone PKC activator bryostatin 1 occurs at multiple levels, and involves a variety of oncogenes and signalling pathways. Under some circumstances, bryostatin 1 may lead to enhanced conversion of ara-C to its lethal metabolite, ara-CTP. However, bryostatin 1 is able to potentiate ara-C-mediated cytotoxicity in the absence of metabolic perturbations, presumably by modulating the cell death pathway itself. For example, chronic exposure of cells to bryostatin 1 leads to PKC down-regulation, which may alter the balance between survival (e.g., ERK) versus stress (e.g., SAPK/JNK)-related pathways. The ability of bryostatin 1 to enhance ara-C-mediated apoptosis is inversely related to its capacity to induce leukemic cell maturation and may involve the failure to down-regulate expression of the cell cycle progression-related proto-oncogene, c-myc. Finally, recent evidence suggests that bryostatin 1 may act, through modification of Bcl-2 phosphorylation status, at a distal site in the cell death pathway. These studies could provide a paradigm important for understanding the mechanism(s) by which agents acting through signal transduction pathways modulate cytotoxic drug-induced cell death
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PMID:Modulation of ara-C induced apoptosis in leukemia by the PKC activator bryostatin 1. 919 93

We selected an apoptosis-resistant subline (VC-33) in a human promyelocytic leukemia cell line, HL-60, by alternating exposure to camptothecin (CPT) and etoposide (VP-16). When wild-type (WT) and VC-33 cells were incubated with various concentrations of either CPT or VP-16 for 4 h, VC-33 showed several-fold resistance to apoptosis induced by these agents in comparison with WT cells. VC-33 cells also exhibited cross-resistance to apoptosis induced by 1-beta-d-arabinofuranosylcytosine, hydroxyurea, a calcium ionophore (A23187), cycloheximide, or UV irradiation. The levels of protein-DNA cross-linking induced by CPT or VP-16, and the amounts of ara-CTP generation, tended to be smaller in VC-33 cells, but the difference was not sufficient to explain the difference in the sensitivity to apoptosis. The initial rise of intracellular calcium ions with A23187 and the expression of P-glycoprotein, Bcl-2, and Bcl-Xl were comparable between WT and VC-33 cells. This mutant may represent a new phenotype of resistance to apoptosis induced by a variety of agents, and may thus be useful in the study of the mechanisms of apoptosis.
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PMID:Apoptosis-resistant phenotype selected by alternating exposure to camptothecin and etoposide. 928 62

1-beta-D-Arabinofuranosylcytosine (ara-C) stimulates the formation of both diglyceride and ceramide in the acute myelogenous leukemia cell line HL-60 (Strum, J. C., Small, G. W., Pauig, S. B., and Daniel, L. W. (1994) J. Biol. Chem 269, 15493-15497). ara-C also causes apoptosis in HL-60 cells which can be mimicked by exogenous ceramide. However, the signaling role for ara-C-induced diacylglycerol (DAG) is not defined. We found that Bcl-2 levels were increased by treatment of HL-60 cells with exogenous DAG or 12-O-tetradecanoylphorbol-13-acetate (TPA). In contrast, exogenous ceramide treatment caused a decrease in cellular Bcl-2 levels. Thus, ara-C stimulates the synthesis of two second messengers with opposing effects on Bcl-2. Since the effects of ara-C-induced DAG could be due to protein kinase C (PKC) activation, we determined the effects of ara-C on PKC isozymes. ara-C caused an increase in membrane-bound PKCbetaII (but not PKCalpha or PKCdelta). ara-C or TPA-induced translocation of PKCbetaII was inhibited by 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3), and ara-C-induced apoptosis was stimulated by pretreatment of the cells with ET-18-OCH3. ET-18-OCH3 also inhibited stimulation of Bcl-2 by TPA and enhanced the decrease in Bcl-2 observed in ara-C-treated cells. These data indicate that ara-C-induced apoptosis is limited by ara-C-stimulated PKCbetaII through effects on Bcl-2. To further determine the role of PKC, we used antisense oligonucleotides directed toward PKCbetaII. The antisense, but not the sense, oligonucleotide inhibited PKCbetaII activation and enhanced ara-C-induced apoptosis. These data demonstrate that the stimulation of apoptosis by ara-C is self-limiting and can be enhanced by inhibition of PKC.
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PMID:Protein kinase CbetaII activation by 1-beta-D-arabinofuranosylcytosine is antagonistic to stimulation of apoptosis and Bcl-2alpha down-regulation. 929 81

CD5+ B-chronic lymphocytic leukaemia (B-CLL) and mantle cell lymphoma (MCL) in leukaemic phase are characterized by defects in cell death induction that primarily involves the Bcl-2 family of genes. Fludarabine (9-beta-D-arabinofuranosyl-2-fluoradenine, F-ara-A) is a potent inducer of apoptosis in CLL cells. This study aimed to determine whether F-ara-A-induced apoptosis might be related to Bcl-2 modifications and to evaluate in vitro/in vivo correlations. Peripheral blood lymphocytes from eight B-CLL and four leukaemic MCL were cultured in the presence of different concentrations of F-ara-A +/- methylprednisolone (MP). F-ara-A down-regulated the expression of Bcl-2 in 5/12 cases. mRNA down-regulation was maximal at 48 h; protein down-regulation was prominent after 48 h. Both events were dose-dependent. The amount of apoptosis was significantly higher in the samples treated with F-ara-A than in those exposed to MP alone. In the seven remaining cases, no Bcl-2 down-regulation was observed after exposure to F-ara-A and the degree of F-ara-A-induced apoptosis overlapped that induced by MP. The in vivo outcome after treatment with three to six courses of F-ara-A was evaluable in 10 patients: 4/5 cases, whose cells had shown in vitro Bcl-2 down-regulation and prominent apoptosis after exposure to F-ara-A, had a complete response (CR) and a partial response (PR) was observed in the remaining patient. Of the five patients whose cells had shown no in vitro Bcl-2 modulation after exposure to F-ara-A, two had a PR, but the other three did not show any in vivo clinical response.
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PMID:Fludarabine ability to down-regulate Bcl-2 gene product in CD5+ leukaemic B cells: in vitro/in vivo correlations. 935 15

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

Bax and Bcl-2 are a pair of important genes that control programmed cell death, or apoptosis, with Bax being the apoptosis promoter and Bcl-2 the apoptosis protector. Although the detailed mechanism is unknown, the protein products of these two genes form protein dimers with each other and the relative ratio of the two proteins is believed to be a determinant of the balance between life and death. In our preliminary study, we found that K562 erythroleukemia cells have an extremely low level of endogenous Bcl-2 expression and a fairly high level of endogenous Bax expression. We constructed Bax and Bcl-2 expression vectors and transfected them into K562 cells. We found that transfection of Bax vector increased the expression of Bax protein; a shortened form of Bax also appeared. Cell death analysis using the Annexin V assay showed that the Bax vector caused significantly more apoptotic cells that the Bcl-2 or pCI-neo vector did. After selection with G418, Bax, Bcl-2 and pCI-neo stably transfected cells were established. These three cell lines were examined for their response to the chemotherapeutic agents ara-C, doxorubicin, etoposide and SN-38. Bax-K562 cells showed significantly higher fractions of apoptotic cells than pCI-neo-K562 cells when treated with ara-C, doxorubicin or SN-38. No sensitization effect was seen when etoposide was used. In contrast, Bcl-2-K562 cells had fewer apoptotic cells than pCI-neo-K562 cells after treatment with all these agents. Therefore, Bax may sensitize K562 cells to apoptosis induced by a wide range of, but not all, chemotherapeutic agents.
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PMID:Overexpression of Bax gene sensitizes K562 erythroleukemia cells to apoptosis induced by selective chemotherapeutic agents. 956 26


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