Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.22.56 (caspase-3)
 35,750 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In monolayer cultures of P19 EC cells treated with both all-trans retinoic acid (RA) and bone morphogenetic protein (BMP)-4 (RA/BMP-4 treatment), many non-adherent apoptotic cells and activated caspase-3-positive cells were observed, but they were not observed in cells treated with RA or BMP-4 alone. Consistent with the appearance of activated caspase-3-positive cells, BMP-4 and RA together induced processing of caspase-9, Ac-DEVD-MCA cleavage activity and DNA fragmentation. These three activities were observed infrequently or not at all when cells were treated with RA or BMP-4 alone. In the RA/BMP-4 treatment-induced apoptosis, caspase-9 was upstream of caspase-3 in the enzyme cascade, and the caspase-9 to -3 step was key in the apoptotic pathway. Bcl-xL inhibited processing of caspase-9, Ac-DEVD-MCA cleavage activity and DNA fragmentation induced by RA/BMP-4 treatment. However, unlike staurosporine-induced apoptosis, cytochrome c, which activates caspase-9, was not detected in the cytosol of RA/BMP-4-treated cells. RA and BMP-4 may activate caspase-9 through an apoptotic pathway other than the Apaf-1/cytochrome c pathway. The prominent decrease of X-chromosome-linked inhibitory apoptosis protein (XIAP) in the cytosol may explain the activation of caspase-9 induced by RA and BMP-4 treatment.
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PMID:BMP-4 and retinoic acid synergistically induce activation of caspase-9 and cause apoptosis of P19 embryonal carcinoma cells cultured as a monolayer. 1057 80

Caspase activation plays a central role in the execution of apoptosis. The key components of the biochemical pathways of caspase activation have been recently elucidated. In this review, we focus on the two most well-studied pathways of caspase activation: the cell surface death receptor pathway and the mitochondria-initiated pathway. In the cell surface death receptor pathway, activation of caspase-8 following its recruitment to the death-inducing signaling complex (DISC) is the critical event that transmits the death signal. This event is regulated at several different levels by various viral and mammalian proteins. Activated caspase-8 can activate downstream caspases by direct cleavage or indirectly by cleaving Bid and inducing cytochrome c release from the mitochondria. In the mitochondrial-initiated pathway, caspase activation is triggered by the formation of a multimeric Apaf-1/cytochrome c complex that is fully functional in recruiting and activating procaspase-9. Activated caspase-9 will then cleave and activate downstream caspases such as caspase-3, -6, and -7. This pathway is regulated at several steps, including the release of cytochrome c from the mitochondria, the binding and hydrolysis of dATP/ATP by Apaf-1, and the inhibition of caspase activation by the proteins that belong to the inhibitors of apoptosis (IAP).
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PMID:Biochemical pathways of caspase activation during apoptosis. 1061 63

The tyrphostin AG957 (NSC 654705) inhibits p210bcr/abl, the transforming kinase responsible for most cases of chronic myelogenous leukemia (CML). The present studies were performed to determine the fate of AG957-treated cells and assess the selectivity of AG957 for CML myeloid progenitors. When K562 cells (derived from a patient with blast crisis CML) were treated with AG957, dose- and time-dependent p210bc/abl down-regulation was followed by mitochondrial release of cytochrome c, activation of caspase-9 and caspase-3, and apoptotic morphological changes. These apoptotic changes were inhibited by transfection with cDNA encoding dominant negative caspase-9 but not dominant-negative FADD or blocking anti-Fas antibodies. In additional experiments, a 24-h AG957 exposure caused dose-dependent inhibition of K562 colony formation in soft agar. To extend these studies to clinical samples of CML, peripheral blood mononuclear cells from 10 chronic phase CML patients and normal controls were assayed for the growth of hematopoietic colonies in vitro in the presence of increasing concentrations of AG957. These assays demonstrated selectivity of AG957 for CML progenitors, with median IC50s (CML versus normal) of 7.3 versus >20 microM AG957 in granulocyte colony-forming cells (P < 0.001), 5.3 versus >20 microM in granulocyte/macrophage colony-forming cells (P < 0.05), and 15.5 versus > 20 microM in erythroid colony-forming cells (P > 0.05). The adamantyl ester of AG957 (NSC 680410) down-regulated p210bcr/abl in K562 cells and inhibited granulocyte colony formation in CML specimens at lower concentrations without enhanced toxicity in normal progenitors. These observations not only demonstrate that AG957-induced p210bcr/abl down-regulation is followed by activation of the cytochrome c/Apaf-1/caspase-9 pathway but also indicate that this class of kinase inhibitor exhibits selectivity worthy of further evaluation.
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PMID:Effects of the bcr/abl kinase inhibitors AG957 and NSC 680410 on chronic myelogenous leukemia cells in vitro. 1065 55

Ectopic overexpression of Apaf-1 (2.5-fold) in human acute myelogenous leukemia HL-60 cells (HL-60/Apaf-1 cells) induced apoptosis and sensitized HL-60/Apaf-1 cells to etoposide- and paclitaxel-induced apoptosis (C. Perkins et al., Cancer Res., 58: 4561-4566, 1998). In this report, we demonstrate that in HL-60/Apaf-1 cells, the activity of caspase-9 and -3 induced by Apaf-1 overexpression was associated with a significant increase (5-fold) in the cytosolic accumulation of cytochrome c (cyt c), loss of mitochondrial membrane potential (deltapsim), and an increase in the reactive oxygen species. These were also associated with the processing of procaspase-8 and Bid (cytosolic, proapoptotic BH3 domain containing protein). Transient transfection of Apaf-1 into the Apaf-1-containing mouse embryogenic fibroblasts (MEFs; Apaf-1+/- MEFs) or Apaf-1-/- MEFs also induced the processing of procaspase-9 and procaspase-8, Bid cleavage, and apoptosis. These events were secondary to the activity of the downstream caspases induced by Apaf-1. This conclusion is supported by the observation that in HL-60/Apaf-1 cells, ectopic expression of dominant negative caspase-9, its inhibitory short isoform caspase-9b, or XIAP or treatment with the caspase inhibitor zVAD (50 microM) inhibited Apaf-1-induced caspase-8 and Bid cleavage, mitochondrial deltapsim, release of cyt c, and apoptosis. In contrast, a transient transfection of dominant negative caspase-8 or CrmA or exposure to caspase-8 inhibitor zIETD-fmk inhibited the processing of procaspase-8 and Bid but did not inhibit the cytosolic accumulation of cyt c in either the untreated HL-60/Apaf-1 cells or the etoposide-treated HL-60/Apaf-1 and HL-60/neo cells. These results indicate that Apaf-1 overexpression lowers the apoptotic threshold by activating caspase-9 and caspase-3. This triggers the mitochondrial deltapsim and cyt c release into the cytosol through a predominant mechanism other than cleavage of caspase-8 and/or Bid. This mechanism may involve a cytosolic mitochondrial permeability transition factor, which may be processed and activated by the downstream effector caspases, thereby completing an amplifying feedback loop, which triggers the mitochondrial events during apoptosis.
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PMID:The role of Apaf-1, caspase-9, and bid proteins in etoposide- or paclitaxel-induced mitochondrial events during apoptosis. 1074 35

Bcl-2 and related proteins are key regulators of apoptosis or programmed cell death implicated in human disease including cancer. We recently showed that cell-permeable Bcl-2 binding peptides could induce apoptosis of human myeloid leukemia in vitro and suppress its growth in severe combined immunodeficient mice. Here we report the discovery of HA14-1, a small molecule (molecular weight = 409) and nonpeptidic ligand of a Bcl-2 surface pocket, by using a computer screening strategy based on the predicted structure of Bcl-2 protein. In vitro binding studies demonstrated the interaction of HA14-1 with this Bcl-2 surface pocket that is essential for Bcl-2 biological function. HA14-1 effectively induced apoptosis of human acute myeloid leukemia (HL-60) cells overexpressing Bcl-2 protein that was associated with the decrease in mitochondrial membrane potential and activation of caspase-9 followed by caspase-3. Cytokine response modifier A, a potent inhibitor of Fas-mediated apoptosis, did not block apoptosis induced by HA14-1. Whereas HA14-1 strongly induced the death of NIH 3T3 (Apaf-1(+/+)) cells, it had little apoptotic effect on Apaf-1-deficient (Apaf-1(-/-)) mouse embryonic fibroblast cells. These data are consistent with a mechanism by which HA14-1 induces the activation of Apaf-1 and caspases, possibly by binding to Bcl-2 protein and inhibiting its function. The discovery of this cell-permeable molecule provides a chemical probe to study Bcl-2-regulated apoptotic pathways in vivo and could lead to the development of new therapeutic agents.
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PMID:Structure-based discovery of an organic compound that binds Bcl-2 protein and induces apoptosis of tumor cells. 1086 Sep 79

Apaf-1(-/-) or caspase-3(-/-) cells treated with a variety of apoptosis inducers manifest apoptosis-associated alterations including the translocation of apoptosis-inducing factor (AIF) from mitochondria to nuclei, large scale DNA fragmentation, and initial chromatin condensation (stage I). However, when compared with normal control cells, Apaf-1(-/-) or caspase-3(-/-) cells fail to exhibit oligonucleosomal chromatin digestion and a more advanced pattern of chromatin condensation (stage II). Microinjection of such cells with recombinant AIF only causes peripheral chromatin condensation (stage I), whereas microinjection with activated caspase-3 or its downstream target caspase-activated DNAse (CAD) causes a more pronounced type of chromatin condensation (stage II). Similarly, when added to purified HeLa nuclei, AIF causes stage I chromatin condensation and large-scale DNA fragmentation, whereas CAD induces stage II chromatin condensation and oligonucleosomal DNA degradation. Furthermore, in a cell-free system, concomitant neutralization of AIF and CAD is required to suppress the nuclear DNA loss caused by cytoplasmic extracts from apoptotic wild-type cells. In contrast, AIF depletion alone suffices to suppress the nuclear DNA loss contained in extracts from apoptotic Apaf-1(-/-) or caspase-3(-/-) cells. As a result, at least two redundant parallel pathways may lead to chromatin processing during apoptosis. One of these pathways involves Apaf-1 and caspases, as well as CAD, and leads to oligonucleosomal DNA fragmentation and advanced chromatin condensation. The other pathway, which is caspase-independent, involves AIF and leads to large-scale DNA fragmentation and peripheral chromatin condensation.
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PMID:Two distinct pathways leading to nuclear apoptosis. 1095 27

Caspases, which play crucial roles during apoptosis, are activated from their inactive proforms in a sequential cascade of cleavage by other members of the caspase family. Caspase-9 is autoprocessed by the Apaf-1/cytochrome c pathway and acts at an early point in this cascade, whereas Bcl-xL, an antiapoptotic member of the Bcl-2 family, prevents activation of caspases in vitro. Little is known, however, about the relation between caspase-9 and Bcl-xL during development of the mammalian nervous system. We used antisera against two cleavage sites in mouse caspase-9 that recognize only the activated form of mouse caspase-9, and we examined immunohistochemically the activation of mouse caspase-9 in the nervous system of Bcl-x-deficient mouse embryos. Mouse caspase-9 is processed at both D(353) and D(368), but it is processed preferentially at D(368) during apoptosis of cultured cells induced by various stimuli and in the nervous system of Bcl-x-deficient mouse embryos. We show that Bcl-xL protects against caspase-9- and/or caspase-3-dependent apoptosis in the caudal portion of the ventral hindbrain, anterior horn cells, and dorsal root ganglia neurons of the normal mouse embryos and against caspase-9/caspase-3-independent apoptosis in the dorsal region of the nervous system including the dorsal spinal cord. Furthermore, we demonstrate that Bcl-xL blocks cytochrome c release from mitochondria, causing activation of caspase-9 in anterior horn cells and dorsal root ganglia neurons in mouse embryos at embryonic day 11.5.
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PMID:Detection of caspase-9 activation in the cell death of the Bcl-x-deficient mouse embryo nervous system by cleavage sites-directed antisera. 1096 Jun 82

The differentiation and apoptosis-sensitizing effects of the Bcr-Abl-specific tyrosine kinase inhibitor CGP57148B, also known as STI-571, were determined in human Bcr-Abl-positive HL-60/Bcr-Abl and K562 cells. First, the results demonstrate that the ectopic expression of the p185 Bcr-Abl fusion protein induced hemoglobin in the acute myeloid leukemia (AML) HL-60 cells. Exposure to low-dose cytosine arabinoside (Ara-C; 10 nmol/L) increased hemoglobin levels in HL-60/Bcr-Abl and in the chronic myeloid leukemia (CML) blast crisis K562 cells, which express the p210 Bcr-Abl protein. As compared with HL-60/neo, HL-60/Bcr-Abl and K562 cells were resistant to apoptosis induced by Ara-C, doxorubicin, or tumor necrosis factor-alpha (TNF-alpha), which was associated with reduced processing of caspase-8 and Bid protein and decreased cytosolic accumulation of cytochrome c (cyt c). Exposure to CGP57148B alone increased hemoglobin levels and CD11b expression and induced apoptosis of HL-60/Bcr-Abl and K562 cells. CGP57148B treatment down-regulated antiapoptotic XIAP, cIAP1, and Bcl-x(L), without affecting Bcl-2, Bax, Apaf-1, Fas (CD95), Fas ligand, Abl, and Bcr-Abl levels. CGP57148B also inhibited constitutively active Akt kinase and NFkappaB in Bcr-Abl-positive cells. Attenuation of NFkappaB activity by ectopic expression of transdominant repressor of IkappaB sensitized HL-60/Bcr-Abl and K562 cells to TNF-alpha but not to apoptosis induced by Ara-C or doxorubicin. Importantly, cotreatment with CGP57148B significantly increased Ara-C- or doxorubicin-induced apoptosis of HL-60/Bcr-Abl and K562 cells. This was associated with greater cytosolic accumulation of cyt c and PARP cleavage activity of caspase-3. These in vitro data indicate that combinations of CGP57148B and antileukemic drugs such as Ara-C may have improved in vivo efficacy against Bcr-Abl-positive acute leukemia.
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PMID:CGP57148B (STI-571) induces differentiation and apoptosis and sensitizes Bcr-Abl-positive human leukemia cells to apoptosis due to antileukemic drugs. 1097 73

A novel human inhibitor of apoptosis protein (IAP) family member termed Livin was identified, containing a single baculoviral IAP repeat (BIR) domain and a COOH-terminal RING finger domain. The mRNA for livin was not detectable by Northern blot in most normal adult tissues with the exception of the placenta, but was present in developmental tissues and in several cancer cell lines. Highest levels were observed in two melanoma-derived cell lines, G361 and SK-Mel29. Transfection of livin in HeLa cells resulted in protection from apoptosis induced by expression of FADD, Bax, RIP, RIP3, and DR6. Similar to other IAP family members, the anti-apoptotic activity of Livin was dependent on the BIR domain. Livin was also capable of inhibiting DEVD-like caspase activity triggered by tumor necrosis factor-alpha. In vitro binding studies demonstrated a direct interaction between Livin and the active form of the downstream caspases, caspase-3 and -7, that was dependent on the BIR domain of Livin. In addition, the unprocessed and cleaved forms of caspase-9 co-immunoprecipitated with Livin in vivo, and recombinant Livin could inhibit the activation of caspase-9 induced by Apaf-1, cytochrome c, and dATP. The subcellular distribution of the transfected Livin was analyzed by immunofluorescence. Both Livin and Survivin were expressed in the nucleus and in a filamentous pattern throughout the cytoplasm. In contrast to the apoptotic activity, the COOH-terminal RING domain mediated its subcellular localization patterning. Further studies found that transfection of an antisense construct against livin could trigger apoptosis specifically in cell lines expressing livin mRNA. This was associated with an increase in DNA fragmentation and in DEVD-like caspase activity. Thus, disruption of Livin may provide a strategy to induce apoptosis in certain cancer cells.
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PMID:Livin, a novel inhibitor of apoptosis protein family member. 1102 45

Caspase-8 plays an essential role in apoptosis triggered by death receptors. Through the cleavage of Bid, a proapoptotic Bcl-2 member, it further activates the mitochondrial cytochrome c/Apaf-1 pathway. Because caspase-8 can be processed also by anticancer drugs independently of death receptors, we investigated its exact role and order in the caspase cascade. We show that in Jurkat cells either deficient for caspase-8 or overexpressing its inhibitor c-FLIP apoptosis mediated by CD95, but not by anticancer drugs was inhibited. In the absence of active caspase-8, anticancer drugs still induced the processing of caspase-9, -3 and Bid, indicating that Bid cleavage does not require caspase-8. Overexpression of Bcl-x(L) prevented the processing of caspase-8 as well as caspase-9, -6 and Bid in response to drugs, but was less effective in CD95-induced apoptosis. Similar responses were observed by overexpression of a dominant-negative caspase-9 mutant. To further determine the order of caspase-8 activation, we employed MCF7 cells lacking caspase-3. In contrast to caspase-9 that was cleaved in these cells, anticancer drugs induced caspase-8 activation only in caspase-3 transfected MCF7 cells. Thus, our data indicate that, unlike its proximal role in receptor signaling, in the mitochondrial pathway caspase-8 rather functions as an amplifying executioner caspase.
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PMID:Caspase-8/FLICE functions as an executioner caspase in anticancer drug-induced apoptosis. 1103 Jan 45


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