Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.22 (cdc2)
 8,319 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Responses to the CDK inhibitor flavopiridol (FP) have been examined in U937 leukemia cells ectopically expressing full-length Bcl-2 or an N-terminal phosphorylation loop-deleted protein (Bcl-2). A 3-fold increase in full-length Bcl-2 protein conferred substantial resistance to ara-C-associated lethality, but not to FP-mediated cytochrome c release, caspase-3 and-9 activation and apoptosis. In a second clonal line, a 6-fold increase in Bcl-2 expression delayed but did not ultimately prevent FP-associated apoptosis. In marked contrast, cells ectopically expressing the Bcl-2 loop-deleted protein (32-80) were highly resistant to FP-mediated cytochrome c release, caspase-3, -8, and -9 activation, Bid and PARP cleavage, and apoptosis despite relatively low levels of protein expression. The loop-deleted Bcl-2, but not full-length Bcl-2 protein also protected clonogenic cells from FP-mediated lethality. Finally, in Bcl-2-overexpressing cells, FP lethality was not attenuated by the caspase-8 inhibitor IETD-FMK, arguing against a role for the extrinsic, receptor-mediated pathway in circumventing Bcl-2-associated resistance. Collectively, these findings indicate that FP induces cytochrome c release in leukemic cells despite overexpression of Bcl-2, and suggest that this event may be modulated by negative regulatory factors residing within the N-terminal phosphorylation loop region.
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PMID:Loss of the Bcl-2 phosphorylation loop domain is required to protect human myeloid leukemia cells from flavopiridol-mediated mitochondrial damage and apoptosis. 1217 Jul 73

We have previously reported that combretastatin-A4 prodrug (CA4P), anantitubulin/antiangiogenic agent isolated from the South African willow tree Combretum caffrum, induced cell death primarily through mitotic catastrophe in a panel of human B-lymphoid tumors. In this study, we investigated the molecular aspects of the mitotic catastrophe and whether or not it shares the same pathways of apoptosis. For this we studied the effect of CA4P on selected markers of apoptosis [caspases 9 and 3, poly(ADP-ribose) polymerase (PARP), bcl-2, and bax] and G2-M protein regulators (p53, MDM2, 14-3-3sigma, GADD45, cdc2, cdc25, chk1, wee1, p21, and cyclin B1). The chronic lymphocytic leukemia cell line WSU-CLL was used for this purpose. Western blot analysis showed that 24 h of CA4P (5 nM) exposure induces caspase 9 activation and PARP cleavage. However, the addition of Z-Val-Ala-Asp-fluoromethylketone (a general caspase inhibitor) or Z-Leu-Glu(OMe)-His-Asp(OMe)-CH2F (a caspase 9 inhibitor) before CA4P treatment did not block cell death. No change in bcl-2 or bax protein expression was observed. Exposure of WSU-CLL cells to 4 and 5 nM CA4P was associated with overproduction of total p53 and no dramatic change in MDM2, 14-3-3sigma, GADD45, the cyclin-dependent kinase cdc2, its inhibitory phosphorylation, the cdc2-inhibitory kinase (wee1), chk1, or cdc25 hyperphosphorylation. The overaccumulation of p21 and cyclin B1 protein was obvious at 24 h. Furthermore, CA4P treatment showed an increase in the expression of a marker of mitosis (mitotic protein monoclonal-2 antibody) and an overaccumulation of the cyclin B in the nucleus. Our findings suggest that CA4P induces mitotic catastrophe and arrest of WSU-CLL cells mostly in the M phase independent of p53 and independent of chk1 and cdc2 phosphorylation pathways. Apoptosis is a secondary mechanism of death in a small proportion of cells through activation of caspase 9 and PARP cleavage. The two mechanisms of cell death, i.e., mitotic catastrophe and apoptosis, are independent of each other in our model.
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PMID:Combretastatin-A4 prodrug induces mitotic catastrophe in chronic lymphocytic leukemia cell line independent of caspase activation and poly(ADP-ribose) polymerase cleavage. 1217 7

The effects of combined exposure to the checkpoint abrogator UCN-01 and pharmacologic MEK1/2 inhibitors were examined in human multiple myeloma (MM) cell lines. Treatment of RPMI8226, NCI-H929, and U266 MM cells with a minimally toxic concentration of UCN-01 (150 nM) for 24 hours resulted in mitogen-activated protein (MAP) kinase activation, an effect that was blocked by coadministration of the MEK1/2 inhibitor PD184352. These events were accompanied by enhanced activation of p34(cdc2) and a marked increase in mitochondrial damage (loss of DeltaPsim; cytochrome c and Smac/DIABLO (direct IAP binding protein with low pI) release), poly(ADP-ribose) polymerase (PARP) cleavage, and apoptosis. PD184352/UCN-01 also dramatically reduced clonogenic survival in each of the MM cell lines. In contrast to As(2)0(3), apoptosis induced by PD184352/UCN-01 was not blocked by the free-radical scavenger N-acetyl-L-cysteine. Whereas exogenous interleukin 6 substantially prevented dexamethasone-induced lethality in MM cells, it was unable to protect them from PD184352/UCN-01-induced apoptosis despite enhancing Akt activation. Insulinlike growth factor 1 (IGF-1) also failed to diminish apoptosis induced by this drug regimen. MM cell lines selected for a high degree of resistance to doxorubicin, melphalan, or dexamethasone, or displaying resistance secondary to fibronectin-mediated adherence, remained fully sensitive to PD184352/UCN-01-induced cell death. Finally, primary CD138(+) MM cells were also susceptible to UCN-01/MEK inhibitor-mediated apoptosis. Together, these findings suggest that simultaneous disruption of cell cycle and MEK/MAP kinase signaling pathways provides a potent stimulus for mitochondrial damage and apoptosis in MM cells, and also indicate that this strategy bypasses the block to cell death conferred by several other well-described resistance mechanisms.
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PMID:Combined treatment with the checkpoint abrogator UCN-01 and MEK1/2 inhibitors potently induces apoptosis in drug-sensitive and -resistant myeloma cells through an IL-6-independent mechanism. 1238 35

Until recently, the ability of ARF (human p14(ARF), murine p19(ARF)) tumour-suppressor protein, encoded by the INK4A/ARF locus, to inhibit cell growth in response to various stimuli was related to its ability to stabilize p53 through the so-called ARF/MDM2/p53 pathway. However, recent data have demonstrated that ARF is not implicated in this unique p53-dependent pathway. By use of transient and stable expression, we show here that human p14(ARF) inhibits the growth of human tumoral cells lacking functional p53 by inducing a transient G(2) arrest and subsequently apoptosis. This p14(ARF)-induced G(2) arrest was correlated with inhibition of CDC2 activity, inactivation of CDC25C phosphatase and induction of the CDK inhibitor p21(WAFI). Apoptosis was demonstrated using Hoechst 33352 staining, proteolytic activation of caspase-3 and PARP cleavage. Similar results were obtained in experiments with cells synchronized by hydroxyurea block. Importantly, we were able to reproduce these effects 'in vivo' by showing that p14(ARF) inhibits the growth of p53 nullizygous human tumours in nude mice and induces the regression of p53 -/- established tumours. In these experiments, tumoral regression was associated with inhibition of cell proliferation as well as induction of apoptosis confirming the data obtained in cell lines.
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PMID:p14ARF induces G2 arrest and apoptosis independently of p53 leading to regression of tumours established in nude mice. 1266 Aug 18

Cancer chemopreventive effects of inositol hexaphosphate (IP6), a dietary constituent, have been demonstrated against a variety of experimental tumors, however, limited studies have been done against prostate cancer (PCA), and molecular mechanisms are not well defined. In the present study, we investigated the growth inhibitory effect and associated mechanisms of IP6 in advanced human PCA cells. Advanced human prostate carcinoma DU145 cells were used to study the anticancer effect of IP6. Flow cytometric analysis was performed for cell cycle progression and apoptosis studies. Western immunoblotting, immunoprecipitation and kinase assay were performed to investigate the involvement of G1 cell cycle regulators and their interplay, and end point markers of apoptosis. A significant dose- as well as time-dependent growth inhibition was observed in IP6-treated cells, which was associated with an increase in G1 arrest. IP6 strongly increased the expression of CDKIs (cyclin-dependent kinase inhibitors), Cip1/p21 and Kip1/p27, without any noticeable changes in G1 CDKs and cyclins, except a slight increase in cyclin D2. IP6 inhibited kinase activities associated with CDK2, 4 and 6, and cyclin E and D1. Further studies showed the increased binding of Kip1/p27 and Cip1/p21 with cyclin D1 and E. In down-stream of CDKI-CDK/cyclin cascade, IP6 increased hypophosphorylated levels of Rb-related proteins, pRb/p107 and pRb2/p130, and moderately decreased E2F4 but increased its binding to both pRb/p107 and pRb2/p130. At higher doses and longer treatment times, IP6 caused a marked increase in apoptosis, which was accompanied by increased levels of cleaved PARP and active caspase 3. IP6 modulates CDKI-CDK-cyclin complex, and decreases CDK-cyclin kinase activity, possibly leading to hypophosphorylation of Rb-related proteins and an increased sequestration of E2F4. Higher doses of IP6 could induce apoptosis and that might involve caspases activation. These molecular alterations provide an insight into IP6-caused growth inhibition, G1 arrest and apoptotic death of human prostate carcinoma DU145 cells.
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PMID:Inositol hexaphosphate inhibits growth, and induces G1 arrest and apoptotic death of prostate carcinoma DU145 cells: modulation of CDKI-CDK-cyclin and pRb-related protein-E2F complexes. 1266 18

In several recent studies, we have shown that silibinin inhibits the growth of human prostate cancer cells (PCA) both in vitro and in vivo. Here, we investigated the effect of silibinin in combination with cisplatin and carboplatin on human PCA DU145 cell growth and apoptosis. Cisplatin alone at 2 microg/ml dose produced 48% cell growth inhibition, whereas a combination with 50-100 microM silibinin resulted in 63-80% (p<0.05-0.001) growth inhibition. Similarly, compared to 68% growth inhibition at 20 microg/ml carboplatin, addition of 50-100 microM doses of silibinin caused 80-90% inhibition (p<0.005-0.001). In the studies assessing the effect of these combinations on cell cycle progression, a combination of cisplatin or carboplatin with silibinin resulted in a stronger G2-M arrest, compared to these agents alone showing a moderate G2-M and G1 arrests in case of cisplatin and silibinin, and a complete S phase arrest with carboplatin, respectively. A stronger G2-M arrest by these combinations was accompanied by a substantial decrease in the levels of cdc2, cyclin B1 and cdc25C. Silibinin/platinum compound combinations were also effective in inducing apoptosis where cisplatin and carboplatin when combined with silibinin enhanced apoptosis from 8 to 15% and from 20 to 40%, respectively. Apoptosis induction was further confirmed by PARP and caspases 3, 9 and 7 whose cleaved levels were also enhanced by combination treatment. In addition, there was a significant increase in cytochrome c release in the cytosol following treatment of DU145 cells with these combinations. Together, these results show a substantial increase in the efficacy of platinum compounds on human PCA cells, when combined with silibinin, which provide a rationale for further investigations with these combinations.
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PMID:Silibinin sensitizes human prostate carcinoma DU145 cells to cisplatin- and carboplatin-induced growth inhibition and apoptotic death. 1286 29

The effects of synthetic derivatives of ursodeoxycholic acid (UDCA), HS-1183, and chenodeoxycholic acid (CDCA), HS-1199 and HS-1200, on the proliferation of human prostate carcinoma PC-3 cells were investigated. Whereas CDCA and UDCA had no effects on the growth of cells in a concentration range we have tested, HS-1199 and HS-1200 completely inhibited the cell proliferation, and HS-1183 showed a weak inhibitory activity. This proliferation-inhibitory effect of the synthetic bile acid derivatives was due to the induction of apoptosis, which was confirmed by observing DNA fragmentation, chromatin condensation and cleavage of PARP. Flow cytometric analysis also revealed that the synthetic bile acid derivatives arrested the cell cycle progression at the G1 phase, which effects were associated with inhibition of phosphorylation of pRB and enhanced binding of pRB and E2F-1. They also suppressed Cdk2 and cyclin E-dependent kinase activities without changes of their expressions. Furthermore, the synthetic bile acids increased the levels of Cdk inhibitor, p21WAF1/CIP1, expression and activated the reporter construct of p21WAF1/CIP1 promoter in p53-independent manner, and p21WAF1/CIP1 proteins induced by the synthetic bile acid derivatives were associated with Cdk2 and proliferating cell nuclear antigen. These distinctive features suggest that it is possible to create the new drugs useful for cancer therapy from the synthetic bile acid derivatives as lead compounds.
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PMID:Apoptosis and modulation of cell cycle control by synthetic derivatives of ursodeoxycholic acid and chenodeoxycholic acid in human prostate cancer cells. 1296 88

Therapeutic options to inhibit growth of human NETs of the GEP system are limited. Since NSAIDs might provide an antiproliferative treatment alternative with acceptable toxicity, we examined the effects of different NSAIDs on growth and survival in a representative set of human GEP NET cell lines. Growth and apoptosis were determined based on cell numbers, cell-cycle analyses, kinase assays, DNA fragmentation and PARP cleavage. Expression of COX and cell cycle-regulatory molecules was examined by immunoblotting and reporter gene assays. Depending on the drug and cell line investigated, NSAID treatment resulted in profound growth inhibition of GEP NET cells. Growth-inhibitory effects were achieved with either COX-2 selective (NS398) or unselective (indomethacin, sulindac) compounds. Cell-cycle analyses documented a G1 arrest in NSAID-treated GEP NET populations. In addition, 100 microM sulindac or indomethacin induced apoptosis. All 3 COX inhibitors prevented CDK-2 activation. In parallel to the NSAID-mediated reduction of CDK-2 activity, p21(cip-1) promoter activity and cellular p21(cip-1) levels increased and p21(cip-1) was sequestered into CDK-2 complexes. Thus, the G1 arrest likely resulted from p21(cip-1)-dependent inhibition of CDK-2 activity. At therapeutically relevant concentrations, sulindac significantly reduced GEP NET cell numbers, whereas IFN-alpha and octreotide remained ineffective. The extent of growth inhibition in GEP NETs was comparable to the antiproliferative effects of sulindac in established NSAID-sensitive cell models. NSAIDs acted as potent antiproliferative agents in GEP NET cells via G1 cell-cycle arrest and might therefore offer a therapeutic alternative to current treatment modalities.
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PMID:Nonsteroidal anti-inflammatory drugs inhibit growth of human neuroendocrine tumor cells via G1 cell-cycle arrest. 1456 37

Sulforaphane (SUL), an isothiocyanate found in broccoli and other cruciferous vegetables, has been shown to induce phase II detoxification enzymes, inhibit chemically induced mammary tumors in rats, and more recently to induce cell cycle arrest and apoptosis in cancer cells of the colon. Here, we provide evidence that SUL also acts as a breast cancer anti-proliferative agent. The BALB/c mouse mammary carcinoma cell line F3II was treated with SUL at concentrations up to 15 microM and examined for markers of cell cycle arrest and apoptosis. Treatment of asynchronous F3II cells with 15 microM SUL resulted in G2/M cell cycle arrest, elevated p34cdc2 (cdc2) kinase activity, Bcl-2 down-regulation, evidence of caspase activation, and aggregation of condensed nuclear chromatin. Subsequent exposure of synchronized cells to 15 microM SUL resulted in elevated numbers of prophase/prometaphase mitotic figures, indicating cell cycle progression beyond G2 and arrest early within mitosis. Moreover, cells treated with 15 microM SUL displayed aberrant mitotic spindles, and higher doses of SUL inhibited tubulin polymerization in vitro. In addition, BALB/c mice injected s.c. with F3II cells and subsequently injected daily i.v. with SUL (15 nmol/day for 13 days) developed significantly smaller tumors (approximately 60% less in mass) than vehicle-treated controls. Western blot analysis of tumor proteins demonstrated significantly (P<0.05) reduced PCNA and elevated PARP fragmentation in samples from animals dosed with SUL. Taken together, these results indicate that SUL has mammary cancer suppressive actions both in cell culture and in the whole animal. Inhibition of mammary carcinogenesis appears in part to involve perturbation of mitotic microtubules and early M-phase block associated with cdc2 kinase activation, indicating that cells arrest prior to metaphase exit.
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PMID:Sulforaphane: a naturally occurring mammary carcinoma mitotic inhibitor, which disrupts tubulin polymerization. 1457 57

Silymarin, a defined mixture of natural flavonoid, has recently been shown to have potent cancer chemopreventive efficacy against colon carcinogenesis in rat model; however, the mechanism of such efficacy is not elucidated. Here, using pure active agent in silymarin, namely silibinin, we show its antiproliferative and apoptotic effects, and associated molecular alterations in human colon carcinoma HT-29 cells. Silibinin treatment of cells at 50-100 microg/ml doses resulted in a moderate to very strong growth inhibition in a dose- and a time-dependent manner, which was largely due to a G0/G1 arrest in cell cycle progression; higher dose and longer treatment time also caused a G2/M arrest. In mechanistic studies related its effect on cell cycle progression, silibinin treatment resulted in an upregulation of Kip1/p27 and Cip1/p21 protein as well as mRNA levels, and decreased CDK2, CDK4, cyclin E and cyclin D1 protein levels together with an inhibition in CDK2 and CDK4 kinase activities. In other studies, we observed that G2/M arrest by silibinin was associated with a decrease in cdc25C, cdc2/p34 and cyclin B1 protein levels, as well as cdc2/p34 kinase activity. In the studies assessing biological fate of silibinin-treated cells, silibinin-induced cell cycle arrest and growth inhibition were not associated with cellular differentiation, but caused apoptotic death. The quantitative apoptosis analysis showed up to 15% apoptotic cell death after 48 h of silibinin treatment. Interestingly, silibinin-induced apoptosis in HT-29 cells was independent of caspases activation, as all caspases inhibitor did not reverse silibinin-induced apoptosis. This observation was further confirmed by the findings showing a lack in caspases activity increase and caspases and PARP cleavage as well as a lack in cytochrome c release in cytosol following silibinin treatment of HT-29 cells. Additional studies conducted in mice showed that silibinin doses found effective in HT-29 cells are achievable in plasma, which increases the significance of the present findings and their possible translation in in vivo anticancer efficacy of silibinin against colon cancer. Together, these results identify molecular mechanisms of silibinin efficacy as a cell cycle regulator and apoptosis inducer in human colon carcinoma HT-29 cells, and justify further studies to investigate potential usefulness of this nontoxic agent in colon cancer prevention and intervention.
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PMID:Silibinin upregulates the expression of cyclin-dependent kinase inhibitors and causes cell cycle arrest and apoptosis in human colon carcinoma HT-29 cells. 1461 51


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