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)

Multiple myeloma (MM) is an invariably fatal plasma cell malignancy, primarily due to the therapeutic resistance which ultimately arises. Much of the resistance results from the expression of various survival factors. Despite this, the ribonucleoside analogue, 8-chloro-adenosine (8-Cl-Ado), is cytotoxic to a number of MM cell lines. Previously, we established that the analogue incorporates into the RNA and inhibits mRNA synthesis. Because 8-Cl-Ado is able to overcome survival signals present in MM cells and inhibits mRNA synthesis, it is likely that the drug induces cytotoxicity by depleting the expression of critical MM survival genes. We investigated this question using gene array analysis, real-time reverse transcription-PCR, and immunoblot analysis on 8-Cl-Ado-treated MM.1S cells and found that the mRNA and protein levels of the receptor tyrosine kinase MET decrease prior to apoptosis. To determine MET's role in 8-Cl-Ado cytotoxicity, we generated MM.1S clones stably expressing a MET ribozyme. None of the clones expressed <25% of the basal levels of MET mRNA, suggesting that a threshold level of MET is necessary for their survival. Additionally, the ribozyme knockdown lines were more sensitive to the cytotoxic actions of 8-Cl-Ado as caspase-3 activation and the induction of poly-ADP-ribose polymerase (PARP) cleavage were more pronounced and evident 12 h earlier than in the parental cells. We further established MET's role in MM cell survival by demonstrating that a retroviral MET RNA interference construct induces PARP cleavage in MM.1S cells. These results show that MET provides a survival mechanism for MM cells. 8-Cl-Ado overcomes MM cell survival by a mechanism that involves the depletion of MET.
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PMID:Multiple myeloma cell killing by depletion of the MET receptor tyrosine kinase. 1794 23

Anthracyclines and anthracenediones are well-known cancer chemotherapeutic agents but their uses are limited with cardiotoxicity and drug resistance. Several l- and d-form amino acids were introduced into the anthraquinone skeleton and numerous derivatives were synthesized for the evaluation of anticancer activity. The screening tests showed that WRC-213, an l-methionine conjugation, was the most effective derivative to inhibit proliferative effect of human androgen-independent prostate cancer PC-3 cells (IC50=50 nM). In an extension evaluation, WRC-213 displayed a potent anti-proliferative activity in various cancer cell lines, including non-small cell lung cancer A549, androgen-independent prostate cancer DU145, colorectal cancer HT-29, breast cancer MCF-7 and hepatocellular carcinoma Hep3B and HepG2. It induced cell-cycle arrest at S and G2, but not mitotic phase, in PC-3 cells. The comet assay revealed that induction of DNA damage and inhibition of topoisomerase II were the primary insults. After the checkpoint arrest of the cell-cycle, WRC-213 induced the mitochondria-mediated intrinsic apoptotic pathway, including Mcl-1 cleavage, Bcl-2 down-regulation and activation of caspase-9/caspase-3 cascades. Survivin degradation and caspase-2 activation also contributed to WRC-213-induced apoptosis. Moreover, the assessment of cytotoxicity in H9c2 cardiomyocytes and drug resistance in NCI/ADR-RES cells demonstrated that WRC-213 showed much lower cardiotoxicity and P-glycoprotein-related resistance than those of mitoxantrone, etoposide and doxorubicin. In conclusion, it is suggested that WRC-213 is a potential topoisomerase II inhibitor with reduced cardiotoxicity and drug resistance. It inhibits topoisomerase II activity and induces chromosomal DNA strand breaks, leading to S and G2 arrest of the cell-cycle and activation of mitochondria-mediated apoptotic pathways.
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PMID:WRC-213, an l-methionine-conjugated mitoxantrone derivative, displays anticancer activity with reduced cardiotoxicity and drug resistance: identification of topoisomerase II inhibition and apoptotic machinery in prostate cancers. 1803 33

Hepatocyte growth factor (HGF) is crucial for the development and regeneration of the liver and offers a possible new therapeutic strategy for the treatment of canine liver disease. In this study, the in vitro and in vivo bioactivity of recombinant canine HGF (rcHGF) produced with a baculoviral expression system in insect cells was measured. In vitro rcHGF induced mitogenesis, motogenesis, and phosphorylated the HGF receptor c-MET and its downstream mediators PKB and ERK1/2 in two canine epithelial cell lines. After a partial hepatectomy (phx) in dogs, rcHGF increased phosphorylation of c-MET, PKB and ERK1/2. A moderate increase was seen with the cell cycle protein PCNA in rcHGF treated livers, but no HGF-induced increase in liver weight was seen 7 days after phx. Furthermore, rcHGF treated livers showed lower levels of the key mediator of apoptosis, caspase-3, at 7days after phx. It is concluded that rcHGF is a biologically active protein in vitro and in vivo and the baculoviral expression system supplies sufficient amounts of rcHGF for future clinical studies in dogs.
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PMID:In vitro and in vivo bioactivity of recombinant canine hepatocyte growth factor. 1831 58

Cobalamin-dependent methionine synthase, with a cofactor of vitamin B12, catalyzes the reaction of 5-methyltetrahydrofolate and homocysteine to form methionine and tetrahydrofolate, which takes a core position in folate cycle, one-carbon-unit transfer, and sulfur amino acid pathways. The 'methyl folate trap' hypothesis suggests that methionine synthase is a potential target for anticancer drug development. ZL031 and ZL033 are 5-methyltetrahydrofolate-like compounds that have been newly synthesized as potential inhibitors of the enzyme. To identify the effect of these two compounds on methionine synthase activity, a spectrophotometric assay was used and the results proved that ZL031 and ZL033 inactivated methionine synthase in HL-60 cells with an IC50 dose of 10.0 and 1.4 mumol/l, respectively. Moreover, obvious inhibitory effect on proliferation of HL-60 cells was observed, leading to our further investigation of the underlying anticancer mechanism. Under the circumstances of methionine synthase deficiency and subsequent folate depletion, cell cycle was arrested in G1/S phase and apoptosis was also observed. Analysis of cell cycle regulatory proteins demonstrated that cyclin E and cyclin-dependent kinase 2 were both increased. Furthermore, reduction of caspase-3, poly (ADP-ribose) polymerase, caspase-8, and caspase-9 protein levels were observed. In all the biological experiments we have performed, ZL033 has shown a better efficacy compared with ZL031. These results suggest that ZL031 and ZL033, as novel methionine synthase inhibitors, caused G1/S phase delay and apoptosis and eventually inhibit the proliferation of HL-60 cells in vitro. ZL033, with a carboxylic acid substituent, might have a better potential for drug development than ZL031 with an ester substituent.
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PMID:Two newly synthesized 5-methyltetrahydrofolate-like compounds inhibit methionine synthase activity accompanied by cell cycle arrest in G1/S phase and apoptosis in vitro. 1859 11

The higher expression of methionine cycle genes in melanoma cells than in normal melanocytes may be related with increased protein synthesis and transmethylation reactions and the subsequent need for high levels of methionine. 3-O-(3,4,5-trimethoxybenzoyl)-(-)-epicatechin (TMECG), a trimethoxy derivative of epicatechin-3-gallate (ECG), effectively suppressed proliferation of melanoma cells in cultures by inducing apoptosis. TMECG modulates the expression of genes involved in methionine metabolism, cellular methylation and glutathione synthesis in melanoma cells. TMECG treatment of melanoma cells resulted in the downregulation of antiapoptotic Bcl-2, the upregulation of proapoptotic Bax and the activation of caspase-3; however, it did not induce the expression of the apoptosis protease-activating factor-1 (Apaf-1). Having elucidated the effects of TMECG on the melanoma methionine cycle, we designed therapeuthical strategies to increase its effectiveness. Combinations of TMECG with S-adenosylmethionine or compounds that modulate the intracellular concentration of adenosine strongly increase the antiproliferative effects of TMECG. The ability of TMECG to target multiple aspects related with melanoma survival, with a high degree of potency, points to its clinical value in melanoma therapy.
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PMID:Targeting the methionine cycle for melanoma therapy with 3-O-(3,4,5-trimethoxybenzoyl)-(-)-epicatechin. 1872 82

Methionine adenosyltransferase II (MAT II) is a key enzyme in cellular metabolism and catalyzes the formation of S-adenosylmethionine (SAMe) from L-methionine and ATP. Normal resting T lymphocytes have minimal MAT II activity, whereas activated proliferating T lymphocytes and transformed T leukemic cells show significantly enhanced MAT II activity. This work was carried out to examine the role of MAT II activity and SAMe biosynthesis in the survival of leukemic T cells. Inhibition of MAT II and the resultant decrease in SAMe levels enhanced expression of FasL mRNA and protein, and induced DISC (Death Inducing Signaling Complex) formation with FADD (Fas-associated Death Domain) and procaspase-8 recruitment, as well as concomitant increase in caspase-8 activation and decrease in c-FLIP(s) levels. Fas-initiated signaling induced by MAT II inhibition was observed to link to the mitochondrial pathway via Bid cleavage and to ultimately lead to increased caspase-3 activation and DNA fragmentation in these cells. Furthermore, blocking MAT 2A mRNA expression, which encodes the catalytic subunits of MAT II, using a small-interfering RNA approach enhanced FasL expression and cell death, validating the essential nature of MAT II activity in the survival of T leukemic cells.
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PMID:Inhibition of methionine adenosyltransferase II induces FasL expression, Fas-DISC formation and caspase-8-dependent apoptotic death in T leukemic cells. 1904 23

Puerarin, a major isoflavonoid derived from the Chinese medical herb Radix puerariae (kudzu root), has been reported to be useful in the treatment of various cardiovascular diseases. In the present study, we examined the detailed mechanisms underlying the inhibitory effects of puerarin on inflammatory and apoptotic responses induced by middle cerebral artery occlusion (MCAO) in rats. Treatment of puerarin (25 and 50 mg/kg; intraperitoneally) 10 min before MCAO dose-dependently attenuated focal cerebral ischemia in rats. Administration of puerarin at 50 mg/kg, showed marked reduction in infarct size compared with that of control rats. MCAO-induced focal cerebral ischemia was associated with increases in hypoxia-inducible factor-1alpha (HIF-1alpha), inducible nitric oxide synthase (iNOS), and active caspase-3 protein expressions as well as the mRNA expression of tumor necrosis factor-alpha (TNF-alpha) in ischemic regions. These expressions were markedly inhibited by the treatment of puerarin (50 mg/kg). In addition, puerarin (10-50 microM) concentration-dependently inhibited respiratory bursts in human neutrophils stimulated by formyl-Met-Leu-Phe. On the other hand, puerarin (20-500 microM) did not significantly inhibit the thiobarbituric acid-reactive substance reaction in rat brain homogenates. An electron spin resonance (ESR) method was conducted on the scavenging activity of puerarin on the free radicals formed. Puerarin (200 and 500 microM) did not reduce the ESR signal intensity of hydroxyl radical formation. In conclusion, we demonstrate that puerarin is a potent neuroprotective agent on MCAO-induced focal cerebral ischemia in vivo. This effect may be mediated, at least in part, by the inhibition of both HIF-1alpha and TNF-alpha activation, followed by the inhibition of inflammatory responses (i.e., iNOS expression), apoptosis formation (active caspase-3), and neutrophil activation, resulting in a reduction in the infarct volume in ischemia-reperfusion brain injury. Thus, puerarin treatment may represent a novel approach to lowering the risk of or improving function in ischemia-reperfusion brain injury-related disorders.
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PMID:Neuroprotective mechanisms of puerarin in middle cerebral artery occlusion-induced brain infarction in rats. 1927 72

Osteosarcoma (OSA), the most common malignant bone tumor in dogs and children, exhibits a similar clinical presentation and molecular biology in both species. Unfortunately, 30-40% of children and 90% of dogs still die of disease despite aggressive therapy. The purpose of this study was to test the biologic activity of a novel heat shock protein 90 (HSP90) inhibitor, STA-1474, against OSA. Canine and human OSA cell lines and normal canine osteoblasts were treated with STA-1474 and evaluated for effects on proliferation (CyQuant), apoptosis (Annexin V, PARP cleavage, caspase 3/7 activation) and known HSP90 client proteins. HSP90 was immunoprecipitated from normal and malignant osteoblasts and Western blotting for co-chaperones was performed. Mice bearing canine OSA xenografts were treated with STA-1474, and tumors samples were evaluated for caspase-3 activation and loss of p-Akt/Akt. Treatment with STA-1474 promoted loss of cell viability, inhibition of cell proliferation and induction of apoptosis in OSA cell lines. STA-1474 and its active metabolite STA-9090 also demonstrated increased potency compared to 17-AAG. STA-1474 exhibited selectivity for OSA cells versus normal canine osteoblasts, and HSP90 co-precipitated with co-chaperones p23 and Hop in canine OSA cells but not in normal canine osteoblasts. Furthermore, STA-1474 downregulated the expression of p-Met/Met, p-Akt/Akt and p-STAT3. Finally, STA-1474 induced tumor regression, caspase-3 activation and downregulation of p-Met/Met and p-Akt/Akt in OSA xenografts. Together, these data suggest that HSP90 represents a relevant target for therapeutic intervention in OSA.
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PMID:The novel HSP90 inhibitor STA-1474 exhibits biologic activity against osteosarcoma cell lines. 1954 63

Folates are essential for DNA synthesis and methylation reactions. The antifolate methotrexate (MTX) is a widely used chemotherapeutic drug which inhibits DNA synthesis and induces apoptosis. Changes in activity of a critical folate-metabolizing enzyme, methylenetetrahydrofolate reductase (MTHFR), might alter the chemosensitivity to MTX, as the MTHFR substrate is required for nucleotide synthesis and its product is used in homocysteine remethylation to methionine. Mild MTHFR deficiency is common in many populations due to a polymorphism at bp 677. We previously showed that altered expression of MTHFR enhanced MTX-induced myelosuppression in mice. To determine the cause of the impaired hematopoietic profile in mice with decreased or increased MTHFR expression, we evaluated MTX-induced apoptosis in the major hemolytic organ, spleen, using the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL) staining and caspase-3/7 activity assays, in MTHFR-deficient mice and in MTHFR-overexpressing mice after MTX administration. Decreased or increased expression of MTHFR in mice significantly increased TUNEL-positive cells and caspase-3/7 activities in MTX-treated spleen, compared with that of wild-type littermates. Plasma homocysteine levels correlated with apoptotic index in MTX-treated MTHFR-deficient mice and dUTP/dTTP ratios correlated with apoptotic index in MTX-treated MTHFR-overexpressing mice. The increased apoptosis may therefore relate to hyperhomocysteinemia and deoxyribonucleotide pool imbalances, respectively. Our results suggest that MTHFR underexpression and overexpression enhances MTX-induced apoptosis and myelosuppression, and that genotyping for the MTHFR polymorphism may have therapeutic implications.
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PMID:Methotrexate-induced apoptosis is enhanced by altered expression of methylenetetrahydrofolate reductase. 1959 6

Under physiological conditions, astrocytes take up L-glutamate from the synaptic gap, metabolize it to L-glutamine and return it to neurons, where L-glutamine is metabolized to L-glutamate and stored in neurotransmitter vesicles. However, under pathological conditions, such as hepatic failure, L-glutamine and ammonium are elevated globally in the brain. The Trojan horse hypothesis of L-glutamine toxicity assumes that intramitochondrial hydrolysis of L-glutamine enhances ammonium locally and leads to mitochondrial dysfunction. In the present study, we show that exposure of murine primary microglia as well as of the microglial cell-line BV-2 to L-glutamine promotes chromatin condensation and formation of crescent-like structures in the nucleus. Furthermore, L-glutamine induced an increase in annexin-V labelling, cell shrinkage (apoptotic volume decrease), cell fragmentation and formation of apoptotic bodies. Inhibition of the phosphate-activated glutaminase with 6-diazo-5-oxo-L-norleucine suppressed chromatin condensation and annexin-V labelling in L-glutamine-exposed cells. In addition, inhibition of the glutamine synthetase with L-methionine sulfoximine suppressed chromatin condensation and annexin-V labelling in ammonium-exposed cells. L-glutamine and ammonium enhanced production of reactive oxygen species, as detected with CM-H(2)DCFDA. Apoptosis, induced by L-glutamine, was inhibited either by the radical scavenger alpha-tocopherol or by the nitric oxide synthase blocker N (G)-methyl-L-arginine. Cyclosporin A, a ligand of the permeability transition pore complex component cyclophilin D, prevented L-glutamine-triggered apoptosis. Furthermore, blockade of caspase-9 activity with Z-LEHD-FMK prevented L-glutamine-triggered apoptosis. Taken together, our results indicate that hydrolysis of l-glutamine and, accordingly, accumulation of ammonium in mitochondria induce the intrinsic pathway of apoptosis, characterized by mitochondrial dysfunction and activation of caspase-9, which activates caspase-3.
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PMID:L-Glutamine-induced apoptosis in microglia is mediated by mitochondrial dysfunction. 1961 80


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