UNIPROT:P10415 - FACTA Search

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Query: UNIPROT:P10415 (Bcl-2)
33,771 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have investigated the effects of transient Bcl-2 down-regulation induced by the Bcl-2 antisense oligodeoxynucleotide (ODN) G3139 (Genta Incorporated) in high Bcl-2 protein expressing, estrogen receptor (ER) positive MCF-7 and low Bcl-2 expressing, ER negative MDA435/LCC6 human breast cancer cells. Treatment with Bcl-2 antisense ODN in vitro caused > 80% reduction of Bcl-2 protein levels in a sequence specific manner for both cell lines. Maximum mRNA reduction was achieved within 24 h of the first antisense ODN exposure whereas full protein down-regulation required antisense exposure over 48 h. This Bcl-2 reduction was associated with 80-95% loss of viable cells compared to untreated cells. Similar cytotoxic effects were observed in both cell lines despite a nine-fold intrinsic difference in Bcl-2 protein expression suggesting that the relative degree of down-regulation of Bcl-2 is more important than the absolute reduction. Cell death associated with G3139 exposure exhibited properties indicative of apoptosis such as mitochondrial membrane depolarization and caspase activation. Combined treatment with G3139 and cytotoxic agents resulted in additive cytotoxicity in both cell lines. However, under most conditions studied, the direct cytotoxic activity of G3139 antisense was not synergistic with the cytotoxic agents. These results suggest that while Bcl-2 clearly constitutes an attractive therapeutic target due to its role in regulating apoptosis in breast cancer cells, additional mechanisms are important in the control of apoptosis arising from exposure to anticancer agents in vitro.
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PMID:Effects of Bcl-2 modulation with G3139 antisense oligonucleotide on human breast cancer cells are independent of inherent Bcl-2 protein expression. 1111 54

Apoptosis induction by the pure antiestrogen faslodex, also known as ICI 182780 (ICI), is associated with an effective down-regulation of Bcl-2 expression in the human breast cancer cell line MCF-7. Recent observations point out that beside members of the Bcl-2 family also the TNFR1 signaling pathway may be involved in apoptosis induction by antiestrogens. In this report we have analyzed the expression of members of the TNFR1 signaling pathway during the apoptotic process induced by the pure antiestrogen faslodex and by tamoxifen (Tam) in MCF-7 breast cancer cells. Treatment with 10(-7) M ICI or 10(-7) M Tam leads to a time dependent increase of TNFR1 and TRADD steady-state mRNA levels in MCF-7 cells. In contrast, Bcl-2 expression was strongly decreased following administration of ICI but only weakly after administration of Tam. Western blot analysis and studies by the use of fluorescence microscopy and flow cytometry revealed a time dependent induction of TNFR1 protein and cell surface expression in MCF-7 cells in response to treatment with ICI. To investigate if TNFR1 is functionally involved in apoptosis induction by antiestrogens, we tested whether TNFR1 blocking antibodies can counteract the growth inhibitory action of Tam and ICI. Coincubation of MCF-7 cells with antiestrogens (ICI or Tam) and blocking TNFR1 antibodies lead to an increase in cell viability. Our results provide evidence for a cross talk between the TNFR1 signaling pathway and antiestrogens during the process of apoptosis induction in MCF-7 breast cancer cells. The superiority of the pure antiestrogen ICI to induce apoptosis in MCF-7 cells may result from its capability to modulate the induction of apoptosis via Bcl-2 as well as TNF-associated signal transduction pathways.
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PMID:Treatment with the pure antiestrogen faslodex (ICI 182780) induces tumor necrosis factor receptor 1 (TNFR1) expression in MCF-7 breast cancer cells. 1111 59

We have compared the anti-proliferative effects of ursodeoxycholic acid (UDCA), chenodeoxycholic acid (CDCA) and their derivatives, HS-1183, HS-1199 and HS-1200, on MCF-7 (wild-type p53) and MDA-MB-231 (mutant p53) cells. While UDCA and CDCA exhibited no significant effect, their novel derivatives inhibited the proliferation of both cell lines in a concentration-dependent manner, concomitant with apoptotic nuclear changes and the increase of a sub-G1 population and DNA fragmentation. Furthermore, we also observed an increase in the ratio of pro-apoptotic protein Bax to anti-apoptotic protein Bcl-2 and cleavages of lamin B and poly(ADP-ribose) polymerase (PARP) in MCF-7 and MDA-MB-231 cells. Cell cycle related proteins, cyclin D1 and D3, as well as retinoblastoma protein (pRb) were down-regulated, while the level of cyclin-dependent kinase inhibitor p21(WAF1/CIP1) was increased in both cancer cells after treatment with novel bile acids. These findings suggest that these cytotoxic effects of novel bile acid derivatives on human breast carcinoma cells were mediated via apoptosis through a p53-independent pathway.
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PMID:Novel bile acid derivatives induce apoptosis via a p53-independent pathway in human breast carcinoma cells. 1116 11

TZT-1027, a newly synthesized dolastatin 10 derivative, is a potent antitumor agent which inhibits microtubule polymerization and perturbs microtubule dynamics. In this report, we investigated whether TZT-1027 inhibited the growth of various human cancer cells, and the cell death caused by TZT-1027 was due to apoptosis. In addition, we elucidated the apoptosis machinery induced by treatment with TZT-1027. The 50% growth-inhibitory concentrations (IC50 values) of TZT-1027 on cancer cells derived from various sources were not more than 5.9 ng/ml. TZT-1027 showed superior cytotoxicity than any other antitumor agents. Next, we evaluated morphological nuclear change, namely, chromatin condensation and DNA fragmentation. We used three cancer cell lines derived from different types in view of having apoptosis related protein, human leukemia HL-60 (in the presence of both Caspase-3 and Bcl-2), human breast cancer MCF-7 (in the absence of Caspase-3), and human prostate cancer DU145 (in the absence of Bcl-2). TZT-1027 induced DNA fragmentation in the presence but not absence of Caspase-3. Nevertheless, apoptic chromatin condensation was observed in all cancer cells even if there was no Caspase-3. Furthermore, we examined whether TZT-1027, microtubule-disrupting agent, influenced cell cycle progression. Flow cytometric analysis revealed the cells treated with TZT-1027, and with the other antimicrotubule agents, to be arrested at the G2/M phase and subsequently to show fragmented DNA smaller than that of G1 phase cells. Moreover, we tested TZT-1027 for its ability to induce Bcl-2 phosphorylation in human cancer cell lines. TZT-1027 and other agents which interacted with microtubules induced Bcl-2 phosphorylation, whereas DNA-damaging agents did not. The present results suggested an association of the growth-inhibitory effect of TZT-1027 with the induction of apoptosis and indicated that the apoptosis induced by TZT-1027 was followed by G2/M arrest even if there was no Caspase-3 or Bcl-2.
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PMID:Induction of apoptosis in human cancer cells by TZT-1027, an antimicrotubule agent. 1122 16

Most breast cancer patients treated with anti-oestrogens will eventually develop resistance towards treatment. Therefore it is important to find new therapeutic agents effective for treatment of patients relapsing on anti-oestrogen. The vitamin D analogue EB1089 (Seocalcitol(TM)) is a promising new agent for treatment of breast cancer patients with advanced disease, and in this study we show that two different anti-oestrogen-resistant human breast cancer cell lines are more sensitive towards treatment with EB1089, than the parent MCF-7 cell line. The two resistant cell lines both express a lower content of the anti-apoptotic protein Bcl-2, and we suggest that this may explain the higher sensitivity towards EB1089. The importance of Bcl-2 for response to EB1089 is supported by our observation that oestradiol abrogates the effect of EB1089 in cell lines which increase Bcl-2 in response to oestradiol treatment. Overall these results indicate that treatment with Seocalcitol(TM)may prove effective when patients become refractory to anti-oestrogen therapy, and that Bcl-2 may be used as a predictive marker.
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PMID:Anti-oestrogen resistant human breast cancer cell lines are more sensitive towards treatment with the vitamin D analogue EB1089 than parent MCF-7 cells. 1123 91

Transforming growth factor beta (TGF-beta) is a multifunctional cytokine capable of regulating diverse cellular processes. In this study we investigated the effect of autocrine TGF-beta signaling on tumor necrosis factor (TNF) alpha-induced cell death. We abrogated the TGF-beta autocrine loop by overexpression of a truncated TGF-beta type II receptor in MCF-7 breast carcinoma cells and found that this generated resistance to TNF-alpha-induced cytotoxicity. To elucidate the molecular basis of the influence of TGF-beta on TNF-alpha-induced cytotoxicity, we evaluated the expression levels or activities of proteins involved in TNF-alpha signal transduction or the regulation of apoptosis in general in TGF-beta-responsive and TGF-beta-nonresponsive MCF-7 cells. We observed no significant difference in the expression of TNF-alpha receptors or the TNF receptor-associated death domain protein. In addition, downstream activation of nuclear factor kappaB by TNF-alpha was not altered in cells that had lost TGF-beta responsiveness. Analysis of members of the Bcl-2 family of apoptosis-regulatory proteins revealed that Bcl-X(L) and Bax expression levels were not changed by disruption of TGF-beta signaling. In contrast, the TGF-beta-nonresponsive cells expressed much higher levels of Bcl-2 protein and mRNA than did cells with an intact TGF-beta autocrine loop. Furthermore, restoration of a TGF-beta signal to MCF-7 cells that had spontaneously acquired resistance to TGF-beta caused a reduction in Bcl-2 protein expression. Taken together, our data indicate that loss of autocrine TGF-beta signaling results in enhanced resistance to TNF-alpha-mediated cell death and that this is likely to be mediated by derepression of Bcl-2 expression.
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PMID:Inhibition of transforming growth factor beta signaling in MCF-7 cells results in resistance to tumor necrosis factor alpha: a role for Bcl-2. 1124 65

Bcl-2 and Bax belong to a family of proteins involved in apoptosis regulation and are believed to reside in the cellular cytoplasm. The authors recently reported interphase nuclear localization of both proteins after immunofluorescence staining of formaldehyde- and methanol-fixed human and rodent cell monolayers. In addition, the authors' data confirmed earlier reports on Bcl-2 immunoreactivity of mitotic chromosomes in human cells. In their experience, nuclear or mitotic staining of Bcl-2, in contrast with cytoplasmic Bcl-2 immunoreactivity, is rarely observed in formaldehyde-fixed, paraffin-embedded breast cancer specimens. Therefore, the authors wondered if nuclear and mitotic Bcl-2 immunoreactivity could be irreversibly reduced by certain fixation procedures, including formaldehyde fixation. Here the authors investigated the effects of various routinely used fixation protocols and antigen retrieval techniques on Bcl-2 and Bax immunoreactivity in monolayers of MCF-7 human breast cancer cells. Whereas nuclear and mitotic immunoreactivity for Bcl-2 was clearly present after formaldehyde and methanol fixation, it was completely absent in cells fixed in acetone, methanol, or formaldehyde alone. In addition, it was found that in particular nuclear and mitotic Bcl-2, and to a lesser extent cytoplasmic Bcl-2 immunoreactivity, decreased after prolonged formaldehyde fixation, whereas Bax immunoreactivity diminished only slightly. Heat-mediated antigen retrieval after prolonged formaldehyde fixation elevated cytoplasmic, but not nuclear and mitotic, Bcl-2 immunoreactivity.
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PMID:Routine formaldehyde fixation irreversibly reduces immunoreactivity of Bcl-2 in the nuclear compartment of breast cancer cells, but not in the cytoplasm. 1127 19

There is ample evidence that deregulation of apoptosis results in the development, progression, and/or maintenance of cancer. Since many apoptotic regulatory genes (e.g. bcl-x) code for alternatively spliced protein variants with opposing functions, the manipulation of alternative splicing presents a unique way of regulating the apoptotic response. Here we have targeted oligonucleotides antisense to the 5'-splice site of bcl-x(L), an anti-apoptotic gene that is overexpressed in various cancers, and shifted the splicing pattern of Bcl-x pre-mRNA from Bcl-x(L) to Bcl-x(S), a pro-apoptotic splice variant. This approach induced significant apoptosis in PC-3 prostate cancer cells. In contrast, the same oligonucleotide treatment elicited a much weaker apoptotic response in MCF-7 breast cancer cells. Moreover, although the shift in Bcl-x pre-mRNA splicing inhibited colony formation in both cell lines, this effect was much less pronounced in MCF-7 cells. These differences in responses to oligonucleotide treatment were analyzed in the context of expression of Bcl-x(L), Bcl-x(S), and Bcl-2 proteins. The results indicate that despite the presence of Bcl-x pre-mRNA in a number of cell types, the effects of modification of its splicing by antisense oligonucleotides vary depending on the expression profile of the treated cells.
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PMID:Modification of alternative splicing of Bcl-x pre-mRNA in prostate and breast cancer cells. analysis of apoptosis and cell death. 1127 82

CD40 binding produces multifaceted growth signals in normal and malignant B cells, whereas its physiological role is less well characterized in epithelial cancers. We examined the growth outcome of CD40 ligation in human breast cancer cells, using CD40+ (T47D and BT-20) and CD40-negative (MCF-7, ZR-75-1) cell lines as defined by flow cytometric analysis, immunohistochemistry, and reverse transcription-PCR. Treatment with the soluble recombinant CD40 ligand (CD40L) molecules gp39 or CD40L-trimer significantly reduced [3H]thymidine uptake in BT-20 and T47D cells by up to 40%, but did not affect the growth of CD40-negative MCF-7 or ZR-75-1 cells. Similarly, significant growth inhibition was observed after co-incubation with CD40L-transfected murine L cells (55.0 +/- 8.9%, P < 0.001) that express membrane CD40L constitutively, or with paraformaldehyde-fixed, CD3+ CD40L+ PBLs from three different HLA-mismatched donors (39.7 +/- 3.7%, P < 0.01). Untransfected L cells and non-CD40L-expressing lymphocytes did not produce significant growth inhibition. The in vivo antitumorigenic effects of CD40L were examined using a s.c. severe combined immunodeficient-hu xenograft model. Pretreatment with two different soluble recombinant CD40L constructs (CD40L and gp39) produced similar xenograft growth-inhibitory effects [67 +/- 24% (n = 4), and 65 +/- 14% (n = 8) inhibition, respectively], which were reversed by co-treatment with the CD40L-neutralizing antibody LL48. In vitro analysis indicated that CD40L-induced growth inhibition was accompanied by apoptotic events including cell shrinkage, rounding, and detachment from the adherent T47D culture monolayer. Thirty-one and 27% of gp39-treated T47D and BT-20 cells underwent apoptosis, respectively, as compared with 56 and 65% from the same cell lines after treatment with the Fas agonistic antibody CH-11. An up-regulation of the proapoptotic protein Bax in T47D and BT-20 cells was observed, which indicated that this Bcl-2 family member may contribute to this growth-inhibitory effect. To explore the clinical relevance of CD40L-CD40 interaction, retrospective immunohistochemical analysis was carried to characterize in situ CD40- and CD40L-expression in breast cancer patient biopsies. All of the infiltrating ductal (5 of 5 cases tested) and lobular (4 of 4 cases) breast carcinomas, carcinomas in situ (6 of 6 cases), and mucinous carcinoma tested (1 case) expressed CD40. Varying proportions of tumor cells also expressed CD40L in the majority of infiltrating ductal (3 of 5 cases) and lobular (3 of 4 cases) carcinomas, and carcinomas in situ (4 of 6 cases), as determined by immunohistochemistry and validated by RT-PCR detection of the CD40L message in only CD40L positive-staining cases. Tumor infiltrating mononuclear cells from infiltrating carcinomas and carcinomas in situ expressed CD40 (10 of 10 cases), but less commonly CD40L (1 case of infiltrating lobular carcinoma, 2 cases of carcinoma in situ). Our findings indicate that the CD40 signaling pathway is active in human breast carcinoma cells. However, tumor-infiltrating lymphocytes from primary tumor tissues may be limited in their capacity to directly modulate tumor growth through the CD40L-CD40 loop.
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PMID:Growth-inhibitory effects of CD40 ligand (CD154) and its endogenous expression in human breast cancer. 1129 66

The natural polyamines -putrescine, spermidine, and spermine- are essential for cell growth and differentiation. Polyamines are involved in several gene regulatory functions, although their mechanism(s) of action has not been elucidated. We investigated the role of polyamines in the function of NF-kappa B and estrogen receptor-alpha (ER alpha), two transcription factors implicated in breast cancer cell proliferation and cell survival, using MCF-7 breast cancer cells. We found that spermine facilitated the binding of ER alpha and NF-kappa B to estrogen response element (ERE)- and NF-kappa B response element (NRE), respectively, and enhanced ER alpha-mediated transcriptional activation in transient transfection experiments. We also found that the association of the co-regulatory protein CBP/p300 with ER alpha and NF-kappa B was increased by spermine treatment of MCF-7 cells. Spermine also increased the nuclear translocation of NF-kappa B compared to the control. In contrast, treatment of MCF-7 cells with polyamine analogs, BE-3-4-3 and BE-3-3-3, resulted in transcriptional inhibition of both ERE- and NRE-driven reporter plasmids. In addition, polyamine analogs inhibited the association of ER alpha and NF-kappa B with CBP/p300 and were unable to facilitate nuclear translocation of NF-kappa B. APO-BRDU assay demonstrated that polyamine analogs induced apoptosis, with a loss of the anti-apoptotic protein Bcl-2. These data show a gene regulatory function of polyamines involving transcriptional activation of ER alpha and NF-kappa B, potentially leading to the up-regulation of genes involved in breast cancer cell proliferation. Our results with BE-3-4-3 and BE-3-3-3 suggest that down-regulation of ER alpha- and NF-kappa B-regulated genes is a possible mechanism for the action of polyamine analogs in inducing apoptosis of breast cancer cells.
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PMID:Regulation of estrogenic and nuclear factor kappa B functions by polyamines and their role in polyamine analog-induced apoptosis of breast cancer cells. 1131 19

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