Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.22.56 (caspase-3)
35,750 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Treatment with the proteasome inhibitor, PS-341 resulted in concentration- and time-dependent effects on Bcl-2 phosphorylation and cleavage in H460 cells that coincided with the PS-341-induced G2-M phase arrest. The observed Bcl-2 cleavage paralleled the degree of PS-341-induced apoptosis but was detected to a similar extent with comparable concentrations of two other proteasome inhibitors (MG-132 and PSI). Calpain inhibitors, ALLM and ALLN, and the caspase inhibitors, Z-VAD and AC-YVAD did not induce BcI-2 phosphorylation and cleavage. Exposure to PS-341 resulted in an additional Mr 25,000 cleavage fragment of Bcl-2, whereas only a Mr 23,000 fragment was observed with other anticancer agents. The formation of the Mr 25,000 fragment was not prevented by caspase inhibitors unlike the Mr 23,000 fragment, which suggests mediation by a caspase-independent pathway. Cell fractionation studies revealed that the Bcl-2 cleaved fragments localize within membrane structures and was an early event (at approximately 12 h, posttreatment), and before the observed cleavage of poly(ADP-ribose) polymerase (PARP), beta-catenin, and DNA fragmentation (at approximately 36 h posttreatment). The Mr 23,000 Bcl-2 cleavage product was inhibited by the pan-caspase inhibitor and the inhibitors of capase-3, -8, -9; but the PARP cleavage was prevented only by the pan-caspase and caspase-3 inhibitors, which suggests that the Mr 23,000 Bcl-2 cleavage occurred at both the initiation and execution stages of apoptosis. The inhibition of the ubiquitin/proteasome pathway by PS-341 leads, at an early stage of apoptosis, to Bcl-2 phosphorylation and a unique proteolytic cleavage product, which are associated with G2-M phase arrest and the induction of apoptosis.
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PMID:PS-341, a novel proteasome inhibitor, induces Bcl-2 phosphorylation and cleavage in association with G2-M phase arrest and apoptosis. 2207 12

Although the participation of the ubiquitin-dependent pathway and of the proteasome in apoptosis has been proposed, its role in this process is not yet clearly defined. In previous studies, we have shown that in the central nervous system of the rat, programmed cell death and the ubiquitin-dependent proteolytic pathway are closely related to each other and that different types of neurons and of glial cells, shown different types of correlation between the two phenomena. In this work, we have used lactacystin, a highly specific inhibitor of the proteasome, to explore in Schwann cell cultures the relationship between the activity of the Ub-dependent pathway and apoptosis. Apoptosis was explored analyzing changes in nuclear morphology, using the Annexin V assay and by flow cytometry. Activity of caspase-3 was also measured. Changes in the levels of ubiquitin-protein conjugates and of the ubiquitin activating enzymes, E1, as well as expression of proteins that instruct the cells to apoptosis (p53, NFkappaB-IkappaB, Bcl2), or that participate in the control and regulation of the cell cycle, were also examined. Our results indicate that the decrease in the activity of the proteasome induced by lactacystin in Schwann cells, induces apoptotic cell death through changes in the concentration of certain key proteins that are involved in the apoptosis-signaling pathways.
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PMID:Apoptosis in Schwann cell cultures is closely interrelated with the activity of the ubiquitin-proteasome proteolytic pathway. 1251 44

Recent studies have shown that the transcription factor, nuclear factor kappaB (NF-kappaB), regulates critical survival pathways in a variety of different cell types, including human pancreatic cancer cells. The activation of NF-kappaB is controlled by proteasome-mediated degradation of its endogenous polypeptide inhibitor, inhibitor of nuclear factor kappaBalpha. We investigated the effects of PS-341, a peptide boronate inhibitor of the proteasome in human pancreatic cancer cells in vitro and in vivo. Comparison of PS-341's effects on the growth of eight different human pancreatic cancer cell lines revealed marked heterogeneity in drug responsiveness, ranging from highly resistant (IC50 > 10 microM; Panc-48, HS766T, and Mia-PaCa-2) to extremely sensitive (IC50 < 40 nM; L3.6pl, Hpaf2, and BxPC3). However, these effects did not correlate with differential inhibition of NF-kappaB activation. Direct quantification of apoptosis revealed that PS-341's effects on cell growth largely correlated with sensitivity to programmed cell death. Evaluation of PS-341's effects on established orthotopic tumor xenografts demonstrated that biweekly intravenous administration of the maximum-tolerated dose of the drug (1 mg/kg) led to significant reductions in the volumes of L3.6pl tumors but not Mia-PaCa-2 tumors. Laser scanning cytometer-mediated quantification of drug-induced apoptosis in the xenografts confirmed that PS-341 induced DNA fragmentation and activation of caspase-3 in L3.6pl tumors but not in Mia-PaCa-2 tumors. However, histological examination of drug-treated tumors revealed extensive central necrosis and reductions in microvessel density and VEGF expression in both tumor types. Taken together, our results demonstrate that PS-341 inhibits the growth of human pancreatic tumors via direct effects on tumor cells and indirect effects on the tumor vasculature.
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PMID:Effects of the proteasome inhibitor PS-341 on apoptosis and angiogenesis in orthotopic human pancreatic tumor xenografts. 1251 57

The proteasome is a multi-subunit protease complex that is involved in intracellular protein degradation in eukaryotes. Previously, we have reported that selective, synthetic chymotryptic proteasome inhibitors inhibit A-NK cell-mediated cytotoxicity by approximately 50%; however, the exact role of the proteasome in NK cell-mediated cytotoxicity remains unknown. Herein, we report that proteasome inhibitors, MG115 and MG132, decreased the proteasome chymotrypsin-like activity in the rat natural killer cell line RNK16 by 85% at a concentration of 5 microM. The viability of RNK16 cells was also reduced in the presence of these inhibitors. Both inhibitors induced the apoptosis of RNK16 cells, as shown by DNA fragmentation, caspase-3 activation and the appearance of sub-G-cell populations. An increase in the fraction of apoptotic cells was observed in a dose- and time-dependent manner in our studies. In addition, the activity of caspase-1, -2, -6, -7, -8, and -9, was increased following the treatment of RNK16 cells with these inhibitors. Further investigation revealed that the expression of Fas (CD95) protein on the RNK16 cell surface was increased after the treatment by MG115 or MG132, indicating that apoptosis induced by proteasome inhibitors in RNK16 cells might be mediated through the Fas (CD95)-mediated death pathway as well. Our studies indicate, for the first time, that proteasomal chymotryptic inhibitors can reduce natural killer cell viability and therefore indirectly inhibit cell-mediated cytotoxicity via the apoptosis-inducing properties of these agents.
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PMID:Activation of multiple caspases and modification of cell surface fas (CD95) in proteasome inhibitor-induced apoptosis of rat natural killer cells. 1253 25

The naturally occurring cyclic tetrapeptide chlamydocin is a very potent inhibitor of cell proliferation. Here we show that chlamydocin is a highly potent histone deacetylase (HDAC) inhibitor, inhibiting HDAC activity in vitro with an IC(50) of 1.3 nM. Like other HDAC inhibitors, chlamydocin induces the accumulation of hyperacetylated histones H3 and H4 in A2780 ovarian cancer cells, increases the expression of p21(cip1/waf1), and causes an accumulation of cells in G(2)/M phase of the cell cycle. In addition, chlamydocin induces apoptosis by activating caspase-3, which in turn leads to the cleavage of p21(cip1/waf1) into a 15-kDa breakdown product and drives cells from growth arrest into apoptosis. Concomitant with the activation of caspase-3 and cleavage of p21(cip1/waf1), chlamydocin decreases the protein level of survivin, a member of the inhibitor of apoptosis protein family that is selectively expressed in tumors. Although our data indicate a potential link between degradation of survivin and activation of the apoptotic pathway induced by HDAC inhibitors, stable overexpression of survivin does not suppress the activation of caspase-3 or cleavage of p21(cip1/waf1) induced by chlamydocin treatment. The decrease of survivin protein level is mediated by degradation via proteasomes since it can be inhibited by specific proteasome inhibitors. Taken together, our results show that induction of apoptosis by chlamydocin involves caspase-dependent cleavage of p21(cip1/waf1), which is strikingly associated with proteasome-mediated degradation of survivin.
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PMID:Inhibition of histone deacetylases by chlamydocin induces apoptosis and proteasome-mediated degradation of survivin. 1253 46

An L1210 cell line (Y8) selected for resistance to deoxyadenosine does not express p53 mRNA or protein but expresses WAF1/p21 even under basal conditions. The Y8 cell line had been previously shown to have an increased apoptotic response to a variety of agents that included DNA damaging agents, kinase inhibitors and drugs directed at NFkappa B activation. In this study we show that lactacystin (LC, an inhibitor of proteasome activity) in combination with parthenolide (PA) caused a synergistic increase in the apoptotic fraction of the Y8 cells. LC (2.5 microM) alone and PA (5.0 microM) caused less than 20% of the Y8 cells to undergo apoptosis. However, the combination of LC (2.5 microM) plus PA (5.0 microM) caused 60% of the Y8 cells to undergo apoptosis. The combination of drugs had no effects on the parental wild-type L1210 cells. Pretreatment of the intact Y8 cells with the caspase-3 inhibitor, Ac-DEVD-CHO, resulted in a marked decrease in the apoptosis caused by the LC plus PA combination. Cell-free extracts prepared from the LC plus PA combination-treated cells had activated caspase activities in the caspase cascade: caspase-3 >> caspase-8 > caspase-6 and caspase-10. These results suggest that there are interacting pathways involving aspects of NFkappa B activation and proteasome activity that could be exploited in therapy directed at p53-deficient tumor cells that would lead to caspase-3 activation and apoptosis bypassing the p53-dependent chemotherapy insensitivity.
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PMID:Lactacystin, a proteasome inhibitor, potentiates the apoptotic effect of parthenolide, an inhibitor of NFkappaB activation, on drug-resistant mouse leukemia L1210 cells. 1255 98

There is substantial evidence that cytokines induce apoptosis of vascular smooth muscle cells (VSMCs) in atherosclerosis. Its regulation, however, is not completely defined. The aim of this study is to investigate whether proteasome activity is related with apoptosis in VSMCs by tumor necrosis factor-alpha (TNF-alpha). Rat aorta smooth muscle cells were treated with TNF-alpha and proteasome inhibitor MG132 and then cell death was determined by morphology, viability, and DNA fragmentation. MG132 or TNF-alpha alone did not induce cell death. In contrast, co-treatment of TNF-alpha and proteasome inhibitor induced death and DNA degradation in VSMCs, suggesting proteasome inhibitor enhanced death activity of TNF-alpha. The death was not blocked by ascorbic acid but by nitric oxide synthase inhibitor N(G)-monomethyl-L-arginine. Both caspase-3 and -8 were activated during the death by the proteasome inhibitor and TNF-alpha. The death was effectively blocked by the caspase-3 inhibitor z-DEVD-fmk, suggesting a role of caspase-3 in the death. Nonetheless, there were no significant alterations in the level of Bcl-2, Bcl-X(L), Bax and Bak by the proteasome inhibitor, nor any evidence of cytochrome (cyt) c release into cytosol from dying cells, suggesting that cyt c is not involved. These results suggest that proteasome inhibition potentiates TNF-mediated death in VSMCs in a cyt c-independent pathway. The present study proposes a new mechanism by which VSMCs undergo death by cytokines.
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PMID:Enhancement of TNF-alpha-mediated cell death in vascular smooth muscle cells through cytochrome c-independent pathway by the proteasome inhibitor. 1256 Jan 2

The ubiquitin-proteasome pathway is the principal mechanism for the degradation of short-lived proteins in eukaryotic cells. Recently, proteasome inhibitors have been shown to induce apoptosis in many kinds of human malignant cells. In this study, the mechanism of apoptosis induced by proteasome inhibitor in leukemic cells was examined. Evaluated by MTT assay, treatment of leukemic cells with Z-LLL-CHO, a reversible proteasome inhibitor, induced cell death in a dose-dependent manner. Appearance of the sub G(0)/G(1) fraction of cell cycle observed in flow cytometry assay suggested the induction of apoptosis, which was further proved by typical DNA ladder and morphological study. Western blot displayed the cleavage of bcl-2 into a shortened 22 kD fragment and the decrease in the levels of caspase-3 precursor. A highly sensitive colorimetric assay was employed and the elevation of caspase-3 activity was detected in both cell lines after treatment with Z-LLL-CHO. By comparison, these results showed that the leukemic cell line M-07e and KG-1a, which both express bcl-2 at a relative high level, had different susceptibility to undergo apoptosis induced by Z-LLL-CHO, which possibly due to their different levels of expression and activation of caspase-3 precursor, as well as their different degree of bcl-2 cleavage after treated by Z-LLL-CHO.
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PMID:[Induction of Apoptosis in Leukemic Cells by Inhibiting the Ubiquitin-Proteasome Pathway and Its Possible Mechanism] 1257 13

Thymocytes undergo negative and positive selection during development in the thymus. During this selection process, the majority of thymocytes are eliminated by apoptosis through signaling via TCR or die by neglect, possibly mediated through glucocorticoids. In this study, we report that thymocytes require molecular oxygen to undergo apoptosis induced by dexamethasone (DEX), a synthetic glucocorticoid, and treatment with N-acetyl-L-cysteine (NAC), a thiol antioxidant, inhibits thymocyte apoptosis in vivo as well as ex vivo. We detected elevated intracellular levels of hydrogen peroxide (H(2)O(2)) during DEX-induced apoptosis, which is reduced by NAC treatment, indicating that the elevated levels of intracellular H(2)O(2) are proapoptotic. We also show that loss of mitochondrial membrane potential, cytochrome c release, as well as caspase-3 activation induced by DEX are attenuated by NAC treatment. We identified the production site for H(2)O(2) as the ubiquinone cycle at complex III of mitochondria by using various inhibitors of the mitochondrial electron transport chain, and we show that the cell death events mediated by mitochondria are also significantly reduced when the inhibitors were used. Through inhibition of the proteasome, we also show that the production of H(2)O(2) and the cell death events mediated by mitochondria are regulated by proteosomal activities in DEX-induced thymocyte apoptosis. We conclude that in DEX-treated thymocytes, the increased production of H(2)O(2) originates from mitochondria and is proapoptotic for cell death mediated by mitochondria. We also conclude that all the apoptotic events mediated by mitochondria are regulated by proteasomes.
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PMID:Glucocorticoid-induced apoptosis of thymocytes: requirement of proteasome-dependent mitochondrial activity. 1259 72

We investigate the death route induced by potassium depletion in cerebellar granule cells in 0-15 h time range and study whether and how mutual relationship occurs between the cell antioxidant and proteolytic system. To achieve this, we incubated cells in the absence or presence of inhibitors of the antioxidant system, including superoxide dismutase and catalase, and of the proteolytic system, consisting of proteasomes and caspases, and investigated whether and how (i) cell survival, (ii) reactive oxygen species (ROS) production and (iii) antioxidant enzyme and caspase-3 activity change as a function of time after the apoptotic stimulus. The involvement of both antioxidant and proteolytic system on cytochrome c release was also investigated. Cell survival was found to increase in the presence of either proteasome or caspase inhibitors. On the contrary, as a result of the antioxidant system impairment, shift from apoptosis to necrosis occurs. We show that the antioxidant system, which exhibits a huge activity increase up to 3 h after apoptosis induction, is subjected to the proteasome-dependent proteolysis and that the increase in the antioxidant system found in the absence of proteasome activity is accompanied by ROS production decrease. Consistently, the early ROS-dependent release of cytochrome c was found to be prevented when the activity of the antioxidant system increased. Finally, caspase-3 activation was prevented by the inhibitors of both antioxidant system and proteasome.
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PMID:The apoptosis/necrosis transition in cerebellar granule cells depends on the mutual relationship of the antioxidant and the proteolytic systems which regulate ROS production and cytochrome c release en route to death. 1260 21


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