Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0023418 (leukemia)
 93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We studied the role of proteases in apoptosis using a cell-free system prepared from a human leukemia cell line. HL60 cells are p53 null and extremely sensitive to a variety of apoptotic stimuli including DNA damage induced by the topoisomerase I inhibitor, camptothecin. We measured DNA fragmentation induced in isolated nuclei by cytosolic extracts using a filter elution assay. Cytosol from camptothecin-treated HL60 cells induced internucleosomal DNA fragmentation in nuclei from untreated cells. This fragmentation was suppressed by serine protease inhibitors. Serine proteases (trypsin, endoproteinase Glu-C, chymotrypsin A, and proteinase K) and papain by themselves induced DNA fragmentation in naive nuclei. This effect was enhanced in the presence of cytosol from untreated cells. Cysteine protease inhibitors (E-64, leupeptin, Ac-YVAD-CHO [ICE inhibitor]) did not affect camptothecin-induced DNA fragmentation. The apopain/Yama inhibitor, Ac-DEVD-CHO, and the proteasome inhibitor, MG-132, were also inactive both in the cell-free system and in whole cells. Interleukin-1 beta converting enzyme (ICE) or human immunodeficiency virus protease failed to induce DNA fragmentation in naive nuclei. Together, these results suggest that DNA damage activates serine protease(s) which in turn activate(s) nuclear endonuclease(s) during apoptosis in HL60 cells.
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PMID:DNA fragmentation induced by protease activation in p53-null human leukemia HL60 cells undergoing apoptosis following treatment with the topoisomerase I inhibitor camptothecin: cell-free system studies. 880 33

Some chemotherapeutic agents, as well as TNF and Fas, induce apoptotic cell death in tumor cells, but the cellular components involved in the process have not yet been identified. Interleukin 1 beta converting enzyme (ICE) is a mammalian homolog of CED-3, a protein required for programmed cell death in nematode Caenorhabditis elegans. We found that a selective inhibitor of ICE/ced 3 family proteases, benzyloxycarbonyl Asp CH2OC(O) 2 6,-dichlorobenzene (Z-Asp-CH2-DCB). completely blocked the apoptotic cell death of human leukemia cells caused by etoposide, camptothecin, 1-beta-D-arabinofuranosyl cytosine (Ara-C) and adriamycin. Moreover, in antitumor agent-treated U937 cells, an ICE-like (CPP32-like) protease was strongly activated. These results indicate that ICE/ ced 3 family proteases are involved in antitumor agent-induced apoptosis. Activation of ICE family proteases plays a key role in apoptosis. However, the subsequent mechanisms resulting in apoptosis are largely unknown. We identified actin as a substrate of ICE family proteases. Cleavage of actin and other substrate proteins by ICE family proteases could be critical in the ongoing process of antitumor agent-induced apoptosis in tumor cells.
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PMID:[Involvement of ICE/CED 3 family proteases in antitumor agent-induced apoptosis]. 903 Feb 33

Recent genetic and biochemical studies have implicated cysteine-dependent aspartate-directed proteases (caspases) in the active phase of apoptosis. In the present study, three complementary techniques were utilized to follow caspase activation during the course of etoposide-induced apoptosis in HL-60 human leukemia cells. Immunoblotting revealed that levels of procaspase-2 did not change during etoposide-induced apoptosis, whereas levels of procaspase-3 diminished markedly 2-3 h after etoposide addition. At the same time, cytosolic peptidase activities that cleaved DEVD-aminotrifluoromethylcoumarin and VEID-aminomethylcoumarin increased 100- and 20-fold, respectively; but there was only a 1. 5-fold increase in YVAD-aminotrifluoromethylcoumarin cleavage activity. Affinity labeling with N-(Nalpha-benzyloxycarbonylglutamyl-Nepsilon-biotin yllysyl)aspartic acid [(2,6-dimethylbenzoyl)oxy]methyl ketone indicated that multiple active caspase species sequentially appeared in the cytosol during the first 6 h after the addition of etoposide. Analysis on one- and two-dimensional gels revealed that two species comigrated with caspase-6 and three comigrated with active caspase-3 species, suggesting that several splice or modification variants of these enzymes are active during apoptosis. Polypeptides that comigrate with the cytosolic caspases were also labeled in nuclei of apoptotic HL-60 cells. These results not only indicate that etoposide-induced apoptosis in HL-60 cells is accompanied by the selective activation of multiple caspases in cytosol and nuclei, but also suggest that other caspase precursors such as procaspase-2 are present but not activated during apoptosis.
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PMID:Activation of multiple interleukin-1beta converting enzyme homologues in cytosol and nuclei of HL-60 cells during etoposide-induced apoptosis. 905 43

Immunohistochemical analysis of the apoptosis-effector protease CPP32 (Caspase-3) in normal lymph nodes, tonsils, and nodes affected with reactive hyperplasia (n = 22) showed strong immunoreactivity in the apoptosis-prone germinal center B-lymphocytes of secondary follicles, but little or no reactivity in the surrounding long-lived mantle zone lymphocytes. Immunoblot analysis of fluorescence-activated cell sorted germinal center and mantle zone B cells supported the immunohistochemical results. In 22 of 27 (81%) follicular small cleaved cell non-Hodgkin's B-cell lymphomas, the CPP32-immunopositive germinal center lymphocytes were replaced by CPP32-negative tumor cells. In contrast, the large cell component of follicular mixed cells (FMs) and follicular large cell lymphomas (FLCLs) was strongly CPP32 immunopositive in 12 of 17 (71%) and in 8 of 14 (57%) cases, respectively, whereas the residual small-cleaved cells were poorly stained for CPP32 in all FLCLs and in 12 of 17 (71%) FMs, suggesting that an upregulation of CPP32 immunoreactivity occurred during progression. Similarly, cytosolic immunostaining for CPP32 was present in 10 of 12 (83%) diffuse large cell lymphomas (DLCLs) and 2 of 3 diffuse mixed B-cell lymphomas (DMs). Immunopositivity for CPP32 was also found in the majority of other types of non-Hodgkin's lymphomas studied. Plasmacytomas were CPP32 immunonegative in 4 of 12 (33%) cases, in contrast to normal plasma cells, which uniformly contained intense CPP32 immunoreactivity, implying downregulation of CPP32 in a subset of these malignancies. All 12 peripheral blood B-cell chronic lymphocyte leukemia specimens examined were CPP32 immunopositive, whereas 3 of 3 small lymphocytic lymphomas were CPP32 negative, suggesting that CPP32 expression may vary depending on the tissue compartment in which these neoplastic B cells reside. The results show dynamic regulation of CPP32 expression in normal and malignant lymphocytes.
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PMID:Immunolocalization of the ICE/Ced-3-family protease, CPP32 (Caspase-3), in non-Hodgkin's lymphomas, chronic lymphocytic leukemias, and reactive lymph nodes. 916 Jun 89

In a previous study, we showed that geranylgeraniol (GGO) is a potent inducer of apoptosis in human leukemia cells. The present study describes the effects of GGO on the activity of cysteine-dependent aspartate-directed proteases (caspases) in human leukemia U937 cells. The caspase-3 (CPP32) activity was increased in a time-dependent manner by treatment with 50 microM GGO, whereas no activation of caspase-1 (interleukin-1beta converting enzyme (ICE)) was observed in any time period under the same experimental conditions. Other isoprenyl compounds such as geraniol, geranylgerany-lacetone, and vitamin K2 had no measurable effects on the activities of either caspase-3 or caspase-1. A inhibitor that preferentially inhibits the caspase-3 related caspases, Z-DEVD-FMK, strongly blocked the GGO-induced DNA fragmentation. These results suggest the involvement of caspase-3 in GGO-induced apoptosis in U937 human leukemia cells.
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PMID:Geranylgeraniol potently induces caspase-3-like activity during apoptosis in human leukemia U937 cells. 917 67

The prevention of apoptosis by Zn2+ has generally been attributed to its inhibition of an endonuclease acting in the late phase of apoptosis. In this study we investigated the effect of Zn2+ on an earlier event in the apoptotic process, the proteolysis of the "death substrate" poly(ADP-ribose) polymerase (PARP). Pretreatment of intact Molt4 leukemia cells with micromolar concentrations of Zn2+ caused an inhibition of PARP proteolysis induced by the chemotherapeutic agent etoposide. Using a cell-free system consisting of purified bovine PARP as a substrate and an apoptotic extract or recombinant caspase-3 as the PARP protease, Zn2+ inhibited PARP proteolysis in the low micromolar range. To rule out an effect of Zn2+ on PARP, a protein with two zinc finger domains, we used recombinant caspase-3 and a chromogenic tetrapeptide substrate containing the caspase-3 cleavage site. In this system, Zn2+ inhibited caspase-3 with an IC50 of 0.1 microM. These results identify caspase-3 as a novel target of Zn2+ inhibition in apoptosis and suggest a regulatory role for Zn2+ in modulating the upstream apoptotic machinery.
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PMID:Zinc is a potent inhibitor of the apoptotic protease, caspase-3. A novel target for zinc in the inhibition of apoptosis. 922 15

The Fas/Fas ligand (FasL) pathway is widely involved in apoptotic cell death in lymphoid and nonlymphoid cells. It has recently been postulated that many chemotherapeutic agents also induce cell death by activating the Fas/FasL pathway. In the present study we compared apoptotic pathways induced by anti-Fas or chemotherapeutic agents in the Jurkat human T-cell leukemia line. Immunoblotting showed that treatment of wild-type Jurkat cells with anti-Fas or the topoisomerase II-directed agent etoposide resulted in proteolytic cleavage of precursors for the cysteine-dependent aspartate-directed proteases caspase-3 and caspase-7 and degradation of the caspase substrates poly(ADP-ribose) polymerase (PARP) and lamin B1. Likewise, affinity labeling with N-(N(alpha)-benzyloxycarbonylglutamyl-N(epsilon)-biotinyllysyl+ ++)aspartic acid [(2,6-dimethyl-benzoyl)oxy]methyl ketone [Z-EK(bio)D-amok] labeled the same five active caspase species after each treatment, suggesting that the same downstream apoptotic pathways have been activated by anti-Fas and etoposide. Treatment with ZB4, an antibody that inhibits Fas-mediated cell death, failed to block etoposide-induced apoptosis, raising the possibility that etoposide does not initiate apoptosis through Fas/FasL interactions. To further explore the relationship between Fas- and chemotherapy-induced apoptosis, Fas-resistant Jurkat cells were treated with various chemotherapeutic agents. Multiple independently derived Fas-resistant Jurkat lines underwent apoptosis that was indistinguishable from that of the Fas-sensitive parental cells after treatment with etoposide, doxorubicin, topotecan, cisplatin, methotrexate, staurosporine, or gamma-irradiation. These results indicate that antineoplastic treatments induce apoptosis through a Fas-independent pathway even though Fas- and chemotherapy-induced pathways converge on common downstream apoptotic effector molecules.
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PMID:Comparison of apoptosis in wild-type and Fas-resistant cells: chemotherapy-induced apoptosis is not dependent on Fas/Fas ligand interactions. 924 21

7-hydroxystaurosporine (UCN-01) is a more selective protein kinase C inhibitor than staurosporine. UCN-01 exhibits antitumor activity in experimental tumor models and is presently in clinical trials. Our study reveals that human myeloblastic leukemia HL60 and K562 and colon carcinoma HT29 cells undergo internucleosomal DNA fragmentation and morphological changes characteristic of apoptosis after UCN-01 treatment. These three cell lines lack functional p53, and K562 and HT29 cells are usually resistant to apoptosis. DNA fragmentation in HT29 and K562 cells occurred after 1 day of treatment while it took less than 4 h in HL60 cells. Cycloheximide prevented UCN-01-induced DNA fragmentation in HT-29 cells, but not in HL60 and K562 cells, suggesting that macromolecular synthesis is selectively required for apoptotic DNA fragmentation in HT29 cells. UCN-01-induced DNA fragmentation was preceded by activation of cyclin B1/cdc2 kinase. Further studies in HL60 cells showed that UCN-01-induced apoptosis was associated with degradation of CPP32, PARP, and lamin B and that the inhibitor of caspases (ICE/CED-3 cysteine proteases), Z-VAD-FMK, and the serine protease inhibitor, DCI, protected HL60 cells from UCN-01-induced DNA fragmentation. However, only DCI and TPCK, but not Z-VAD-FMK, inhibited DNA fragmentation in the HL60 cell-free system, suggesting that serine protease(s) may play a role in the execution phase of apoptosis in HL60 cells treated with UCN-01. Z-VAD-FMK and DCI also inhibited apoptosis in HT29 cells. These data demonstrate that the protein kinase C inhibitor and antitumor agent, UCN-01 is a potent apoptosis inducer in cell lines that are usually resistant to apoptosis and lack p53 and that caspases and probably serine proteases are activated during UCN-01-induced apoptosis.
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PMID:7-Hydroxystaurosporine (UCN-01) induces apoptosis in human colon carcinoma and leukemia cells independently of p53. 926 Sep 9

The human leukemia cell line, HL60 is very sensitive to various apoptotic stimuli and p53-null. The death-related cysteine proteases of the caspases family play a central role in the execution phase of apoptosis, and we recently reported the importance of serine protease activation in camptothecin-induced apoptotic endonuclease activation in HL60 cells. In the present study, we investigated the role of caspases (ICE/CED-3-related cysteine proteases) and serine proteases in cell death induced by the topoisomerase I inhibitor, camptothecin, in HL60 cells and in a cell-free system. We found that CPP32 is activated during camptothecin-induced apoptosis, and that N-benzyloxycarbony-Val-Ala-Asp (O-methyl) -fluoromethyketone (Z-VAD-fmk), a cell permeable caspase inhibitor blocks all features of apoptosis: morphological changes, cleavage of caspase 3 (CPP32/Yama/Apopain) and poly(ADP-ribose) polymerase, lamin B degradation and DNA fragmentation. However, Z-VAD-fmk and two other ICE/CED-3 inhibitors, YVAD-CHO and DEVD-CHO, were inactive in a cell-free system reconstituted from nuclei of untreated HL60 cells and cytosol from camptothecin-treated cells, suggesting that caspases are not required for endonuclease activation or lamin B cleavage in the cell-free system. By contrast, the serine protease inhibitors, 3,4-dichloroisocoumarin (DCI) and L-1-chloro-3-(4-tosylamido)-4-phenyl-2-butanone tosyl-L-phenylalanine chloromethyl ketone (TPCK), abolished the apoptosis-associated biochemical changes induced by camptothecin both in whole cells and in a cell-free system. DCI also inhibited CPP32 cleavage. Taken together, these results suggest that in HL60 cells, both CPP32 and serine proteases are activated in camptothecin-induced apoptosis.
Leukemia 1997 Aug
PMID:Camptothecin-induced apoptosis in p53-null human leukemia HL60 cells and their isolated nuclei: effects of the protease inhibitors Z-VAD-fmk and dichloroisocoumarin suggest an involvement of both caspases and serine proteases. 926 76

Apoptotic signaling cascades converge in the activation of caspases (interleukin-1beta converting enzyme like proteases). Treatment of the human promyelocytic leukaemia cell line U937 with actinomycin D resulted in the activation of caspase-3 also known as CPP32. Protease activity was measured in cytosolic extracts by fluorometric analysis of the time-dependent cleavage of acetyl-Asp-Glu-Val-Asp-aminomethylcoumarin (DEVD-AMC), a caspase-3 substrate. Caspase activity was inhibited by thiol modifying agents such as N-ethylmaleimide or iodoacetamide and NO donors such as S-nitrosoglutathione (GSNO), BF4NO, and spermine-NO. NO-mediated enzyme inhibition was fully reversible upon the addition of DTT (dithiothreitol). NO. itself was not primarily responsible for downregulation of caspase-3, as we found no correlation between rates of NO* release and the magnitude of enzyme inhibition. It is likely that S-nitrosation accounts for enzyme inhibition by various NO donors. SIN-1 and peroxynitrite were inhibitory as well. In this case, however, enzyme activity was not restored upon DTT addition, suggesting oxidation as an additional thiol modification mechanism. Our studies provide evidence that caspases are targeted by NO via S-nitrosation and oxidation of critical thiol groups.
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PMID:Inhibition of caspase-3 by S-nitrosation and oxidation caused by nitric oxide. 929 18


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