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Biomedical subjects

M Weller

Publications and source records attributed to M Weller.

At least 163 records · Page 9Linked to original sources
PubMed1998

Potentiation of treosulfan toxicity by the glutathione-depleting agent buthionine sulfoximine in human malignant glioma cells: the role of bcl-2.

Median survival of human malignant glioma patients is less than one year even with cytoreductive surgery and postoperative radiotherapy. Adjuvant chemotherapy has been rather ineffective. Here, we studied the potentiation by L-buthionine-[S,R]-sulfoximine (BSO), a glutathione-depleting agent, of anticancer drug actions on two human malignant glioma cell lines, LN-229 and T98G. LN-229 has wild-type p53 status, T98G is mutant for p53. Glutathione levels were depleted by BSO with similar kinetics in both cell lines. Only LN-229 cells were growth-inhibited by BSO. BSO had minor effects on the toxicity of doxorubicin, ACNU (1-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-3-(2-chloroethyl)-3-nitrosou rea, nimustine) and vincristine. BSO failed to alter teniposide or cytarabine toxicity. BSO induced prominent sensitization to the alkylating agent, treosulfan, in both cell lines, as assessed by viability assays, in situ DNA end labeling and quantitative DNA fragmentation. Treosulfan is thought to mediate toxicity via formation of reactive epoxides. In the absence of BSO, treosulfan had little acute cytotoxic and moderate antiproliferative effects. Synergistic glioma cell cytotoxicity induced by treosulfan and BSO was not associated with reactive oxygen species formation. Ectopic expression of bcl-2 did not alter basal glutathione levels but attenuated glutathione depletion induced by BSO. Bcl-2 provided only moderate protection from synergistic induction of glioma cell death by treosulfan and BSO. Glutathione depletion may play a role in BSO-mediated chemosensitization, but other mechanisms are probably involved as well. BSO may be a useful agent for glioma cell sensitization to specific chemotherapeutic drugs such as treosulfan.

Antineoplastic Agents, Alkylating↗
PubMed1998

Synergy of CD95 ligand and teniposide: no role of cleavable complex formation and enhanced CD95 expression.

Teniposide (VM26) enhanced the anti-glioma activity of the cytotoxic cytokine, CD95 ligand. Synergy was observed at concentrations of teniposide that were insufficient for cleavable DNA topoisomerase II complex formation. CD95 ligand did not modulate the formation or removal of such complexes after teniposide treatment. These processes were also unaffected by ectopic expression of bcl-2. Teniposide enhanced CD95 expression in a glioma cell line with wild-type p53 (LN-229) but not in two p53 mutant cell lines (T98G, LN-308). Forced expression of a transdominant negative p53 mutant prevented the teniposide induced augmentation of CD95 expression in LN-229 cells but did not prevent the synergy of CD95 ligand and teniposide. Teniposide did not alter CD95 ligand expression, and forced expression of CD95 did not modulate sensitivity to VM26. Thus, teniposide-induced DNA lesions and alterations in CD95 or CD95 ligand are not necessary for teniposide-induced sensitization of human malignant glioma cells to CD95-mediated apoptosis.

Antineoplastic Agents, Phytogenic↗
PubMed1998

Increased serum levels of soluble CD95 (APO-1/Fas) in relapsing-remitting multiple sclerosis.

CD95/CD95 ligand interactions are critically involved in the negative regulation of peripheral T-cell responses. Here, we report that serum levels of soluble CD95 are significantly elevated in patients with relapsing remitting multiple sclerosis. In a transectional study, CD95 levels did not correlate with clinical disability or lesion formation on magnetic resonance imaging. Longitudinally, Expanded Disability Status Scale changes were associated with high CD95 levels. Interferon-beta (IFNbeta) treatment led to an initial increase and subsequent decline of serum CD95 levels. Interestingly, patients generating neutralizing antibodies to the drug had significantly higher baseline CD95 levels before IFNbeta treatment than those without neutralizing antibodies.

Adjuvants, Immunologic↗
PubMed1998

The topoisomerase I inhibitors, camptothecin and beta-lapachone, induce apoptosis of human retinal pigment epithelial cells.

The aim of the study was to determine whether the topoisomerase I inhibitors, camptothecin and beta-lapachone, are suitable agents for the adjuvant pharmacotherapy of proliferative vitreoretinopathy (PVR). The effects of the drugs on cultured human retinal pigment epithelial (RPE) cells were examined using growth assays, cytotoxicity assays, single cell agarose gel electrophoresis, in situ DNA end labeling and immunoblot analysis for apoptosis-regulatory proteins. Both agents killed RPE cells in a concentration-and time-dependent manner. Cell death was apoptotic as assessed by single cell agarose gel electrophoresis and in situ DNA end labeling. Camptothecin, but not beta-lapachone, induced accumulation of p53 and the major growth arrest-associated p53 response protein, p21. Both drugs enhanced expression of the proapoptotic BAX protein. Camptothecin, but not beta-lapachone, synergistically enhanced RPE cell apoptosis induced by the cytotoxic cytokine, CD95 ligand (CD95L). This effect was linked to camptothecin-induced inhibition of RNA synthesis. Atypical topoisomerase I inhibitors may be promising agents for the adjuvant pharmacotherapy of PVR. Experimental studies to assess possible ocular toxicity upon local administration and to confirm its therapeutic efficacy in an animal model of PVR are required.

Apoptosis↗
PubMed1998

Bcl-2, Bax and Bcl-x expression in neuronal apoptosis: a study of mutant weaver and lurcher mice.

We investigated the expression of the apoptosis modulating proteins Bcl-2, Bax and Bcl-x in the cerebellum of mutant lurcher and weaver mice. Lurcher Purkinje cells and weaver germinal (granule neuron progenitor) cells both die via apoptosis during the postnatal cerebellar development. No significant changes in the expression patterns were detected prior to the actual cell death process. Instead apoptotic lurcher Purkinje cells exhibited increased Bax and Bcl-x expression, while surviving cells had an expression pattern similar to that of healthy littermates. Increased Bax expression was also found in apoptotic weaver germinal cells, while no change of Bcl-x expression was detected. Bcl-2 was expressed at low levels in cerebellar neurons and no loss of Bcl-2 was evident. The observed expression patterns of Bcl-2, Bax and Bcl-x protein in apoptotic lurcher and weaver neurons support the hypothesis that the execution of neuronal apoptosis involves increased expression of Bax, which could represent a general mechanism in diverse neurodegenerative processes.

Animals↗
PubMed1998

Humoral immune response to p53 in malignant glioma.

p53 immunoreactivity and humoral immune response to p53 were examined in 14 patients with malignant glioma, including 4 patients with leptomeningeal glioma cell dissemination. Twelve patients expressed p53 protein within the tumour tissue. p53 antibodies were detected in the serum in 2 of 14 patients but never in the cerebrospinal fluid (CSF). Soluble p53 protein was detected neither in serum nor in CSF of the glioma patients. CSF levels of the immunosuppressive cytokine, transforming growth factor (TGF)-beta, were elevated in the glioma patients, including those with a humoral response to p53. These preliminary findings raise the possibility of systemic humoral immune responses to antigens, including mutant p53, expressed by glioma cells in the central nervous system.

Adult↗
PubMed1998

Cellular stimulation via CD95 involves activation of phospho-inositide-3-kinase.

Several distinct intracellular pathways have been recently shown to be activated during CD95/Fas/APO-1-mediated apoptosis. Here, we demonstrate that CD95 ligation induces a rapid and transient tyrosine phosphorylation and activation of phosphoinositide-3-kinase (PI-3-K) in Jurkat T lymphocytes or CD95-sensitive glioma cells. Experiments using p56lck-deficient or p56lck-reconstituted Jurkat clones and the tyrosine kinase inhibitor herbimycin A revealed that tyrosine phosphorylation and activation of PI-3-K by CD95 depends on expression of Src-like tyrosine kinases, in particular p56lck. PI-3-K stimulation seems to be critical for CD95 receptor signalling since, first, inhibition of PI-3-K prevents CD95-mediated apoptosis and, second, CD95 receptor ligation fails to induce tyrosine phosphorylation or activation of PI-3-K in CD95-resistant glioma cells. Thus, PI-3-K activation may be an early signalling event during CD95-induced apoptosis, and failure to stimulate PI-3-K may predict tumor cell resistance to CD95-triggered apoptosis.

Apoptosis↗
PubMed1998

Predicting response to cancer chemotherapy: the role of p53.

Loss of wild-type p53 activity is thought to be a major predictor of failure to respond to radiotherapy and chemotherapy in various human cancers. This assumption is largely based on some cell-death studies in p53-knockout mice and on correlations of p53 status assessed by immunochemistry or single-strand conformational polymorphism (SSCP) analysis, and responses to therapy in human cancers in vivo. In principle, p53 may enhance chemosensitivity by promoting apoptosis via transcription-independent mechanisms as well as transcriptional activation of proapoptotic genes such as bax and transcriptional repression of antiapoptotic genes such as bcl-2. Drug-induced suicide mediated by the CD95/CD95 ligand system may also involve a p53-controlled pathway. Yet, p53 may decrease chemosensitivity by promoting p21-mediated and p21-independent growth arrest, DNA repair, and differentiation, and by enhancing the transcription of antiapoptotic genes such as bcl-x. Cell-culture work indicates that the effects of altering the p53 status on chemosensitivity depend very much on the cellular context. Disruption of p53 function in otherwise normal, nonneoplastic cells may enhance rather than decrease chemosensitivity. However, targeted p53 gene disruption in some cell types obtained from p53-knockout mice results in enhanced rather than decreased sensitivity, e.g., to irradiation. Transformed cells that have retained wild-type p53 function tend to acquire chemoresistance when p53 function is disabled, with few exceptions. Thus, preexisting molecular alterations or consecutive accumulation of molecular alterations after loss of p53 rather than the loss of wild-type p53 activity per se may confer chemoresistence to tumor cells. Moreover, p53 accumulation resulting from the increased half-life of mutant p53 proteins can act as a gain-of-function mutation, presumably as a consequence of multiple protein-protein interactions. Finally, significant tumor cell-type- and drug-specific patterns of modulation of chemosensitivity by p53 are beginning to emerge. Transfer of wild-type p53 genes into tumor cells commonly induces growth arrest but may render these cells relatively more resistant to most chemotherapeutic drugs. Therefore, careful experimental in vitro and in vivo studies are required before chemotherapy-supported p53 gene therapy for human cancer is introduced into clinical practice.

Animals↗
PubMed1998

CD95 expression and CD95-mediated apoptosis of T cells in multiple sclerosis. No differences from normal individuals and no relation to HLA-DR2.

CD95-mediated apoptosis is a potent endogenous pathway of T cell elimination that has been suggested to be altered in multiple sclerosis (MS). MS is associated with the HLA-DR2, Dw2, DQ6 HLA class II haplotype. We have previously reported that T cell lines from HLA-DR2-positive individuals show enhanced production of tumor necrosis factor (TNF), a cytokine homologous to CD95 ligand, in response to specific antigen. Here we have studied CD95 expression and susceptibility to CD95-mediated apoptosis in peripheral blood mononuclear cells (PBMC) and activated T cells of 20 healthy individuals and 20 MS patients, half of whom were HLA-DR2-positive. MS patients did not differ from healthy individuals in either parameter. There was also no difference in CD95 expression or CD95-mediated apoptosis when MS patients and healthy individuals were grouped and compared according to HLA-DR status. These data reveal no differential regulation of PBMC/T cell apoptosis induced by CD95 receptor ligation in MS and show no impact of HLA-DR2 status on PBMC/T cell susceptibility to the same apoptotic stimulus. However, to assess the contribution of T cell apoptosis to the pathogenesis of MS further studies on other details of the complex system leading to T cell apoptosis are required.

Apoptosis↗
PubMed1998

Anticonvulsant drugs fail to modulate chemotherapy-induced cytotoxicity and growth inhibition of human malignant glioma cells.

Adjuvant chemotherapy after cytoreductive surgery and irradiation plays an increasingly important role in the management of human malignant glioma. Here we have examined the effect of three anticonvulsants most commonly administered to glioma patients, carbamazepine, phenytoin and valproic acid, on the cytotoxic and antiproliferative actions in vitro of several cancer chemotherapy drugs currently evaluated for human gliomas. We find that none of the anticonvulsants reduces glioma cell viability or proliferation or modulates glioma cell clonogenicity at clinically relevant concentrations when administered alone. Therapeutic concentrations of either drug fail to alter the effect of cancer chemotherapy drugs in acute cytotoxicity assays or modified clonogenicity assays. A lack of interactions of anticonvulsants and cytotoxic drugs is also observed when the glioma cells are preexposed to the anticonvulsants for prolonged times, suggesting that chronic exposure to anticonvulsants in vivo may not change intrinsic glioma cell sensitivity to cancer chemotherapy. Thus, changes in hepatic enzyme activity or immunological parameters, but not modulation of intrinsic chemotherapeutic drug sensitivity, may influence the choice of an anticonvulsant for seizure control in glioma patients receiving adjuvant chemotherapy.

Anticonvulsants↗
PubMed1998

Chemosensitivity of human malignant glioma: modulation by p53 gene transfer.

Loss of wild-type p53 activity is one of the most common molecular abnormalities in human cancers including malignant gliomas. The p53 status is also thought to modulate sensitivity to irradiation and chemotherapy. Here, we studied the effect of a p53 gene transfer on the chemosensitivity of three human glioma cell lines with different endogenous p53 status (LN-229, wild-type; LN-18, mutant; LN-308, deleted), using the murine temperature-sensitive p53 val135 mutant. Expression of mutant p53 enhanced proliferation of LN-308 cells but reduced proliferation in the other cell lines. Expression of wild-type p53 caused reversible growth arrest of all cell lines but failed to induce apoptosis. Growth arrest induced by wild-type p53 was associated with strong induction of p21 expression. Strong induction of BAX expression and loss of BCL-2 expression, which are associated with p53-dependent apoptosis rather than growth arrest, were not observed. Wild-type p53 failed to sensitize glioma cells to cytotoxic drugs including BCNU, cytarabine, doxorubicin, teniposide and vincristine. The combined effects of wild-type p53 gene transfer and drug treatment were less than additive rather than synergistic, suggesting that the intracellular cascades activated by p53 and chemotherapy are redundant. Unexpectedly, forced expression of mutant p53 modulated drug sensitivity in that it enhanced the toxicity of some drugs but attenuated the effects of others. These effects may represent a dominant negative effect of mutant p53 in LN-229 cells which have wild-type p53 activity but must be considered a gain of function-type effect in the other two cell lines which have no wild-type p53 activity. Importantly, no clear-cut pattern emerged among the three cell lines studied. We conclude that somatic gene therapy based on the reintroduction of p53 will limit the proliferation of human malignant glioma cells but is unlikely to induce clinically relevant sensitization to chemotherapy in these tumors.

Amino Acid Substitution↗
PubMed1998

Soluble CD95 (Fas/APO-1) in malignant glioma: (no) implications for CD95-based immunotherapy?

CD95 targeting is a novel approach of immunotherapy for malignant glioma that might be antagonized by the release of soluble CD95 by the tumor cells. An alternatively spliced CD95 mRNA that encodes a secreted CD95 variant has been detected in glioma cell lines in vitro and in human tumors in vivo. Here, we report that the levels of soluble CD95 in the serum of malignant glioma patients do not differ from those of lumbar disk disease patients. Soluble CD95 was detected in the CSF in 2 of 20 malignant glioma patients by ELISA. Bioassay studies indicate that these low levels of soluble CD95 in the CSF of some patients with malignant glioma cells are unlikely to interfere with CD95-based immunotherapy of malignant gliomas in vivo.

Apoptosis↗
PubMed1998

Taxol-mediated augmentation of CD95 ligand-induced apoptosis of human malignant glioma cells: association with bcl-2 phosphorylation but neither activation of p53 nor G2/M cell cycle arrest.

The anti-tumour alkaloid taxol shows strong cytotoxic and antiproliferative activity in two human malignant glioma cell lines, T98G and LN-229. CD95 (Fas/APO-1) ligand is a novel cytotoxic cytokine of the tumour necrosis factor (TNF) family that exerts prominent antiglioma activity. At clinically relevant taxol concentrations of 5-100 nM, taxol and CD95 ligand showed significant synergistic cytotoxicity and growth inhibition. High concentrations of taxol induced G/M cell cycle arrest in both cell lines. The synergy of taxol and CD95 ligand was independent of cell cycle effects of taxol as synergy was achieved at much lower taxol concentrations than G2/M arrest and as cell cycle effects of taxol were unaffected by co-exposure to CD95 ligand. Similarly, high concentrations of taxol were required to induce p53 activity in the p53 wild-type cell line LN-229. This effect was not modulated by CD95 ligand, suggesting that synergy is also independent of p53 activation. However, taxol induced a mobility shift of the bcl-2 protein on immunoblot analysis, indicative of bcl-2 phosphorylation. Bcl-2 phosphorylation on serine was confirmed by immunoprecipitation and phosphoserine immunoblot analysis. Considering (1) that phosphorylation of bcl-2 interferes with its heterodimerization with bax and (2) the inhibition of CD95-mediated apoptosis by bcl-2, we propose that taxol sensitizes malignant glioma cells to CD95 ligand by increasing the functional bax/bcl-2 rheostat in favour of bax and thus cell death.

Antineoplastic Agents, Phytogenic↗
PubMed1998

Fas/CD95/Apo-I activates the acidic sphingomyelinase via caspases.

Fas/CD95/Apo-I has been shown to stimulate a variety of molecules including several members of the caspase family and the acidic sphingomyelinase (Martin and Green 1995; Gulbins et al, 1995). Here, we demonstrate that Fas receptor-triggered activation of the acidic sphingomyelinase, consumption of sphingomyelin, release of ceramide, and subsequent activation of JNK and p38-K are regulated by caspases. Inhibition of caspases by Ac-YVAD-chloromethylketone or transient CrmA transfection prevented stimulation of acidic sphingomyelinase, release of ceramide and activation of JNK and p38-K upon Fas-receptor crosslinking. Likewise, Fas triggered apoptosis was almost completely blocked by Ac-YVAD-chloromethylketone or CrmA mediated inhibition of caspases. The results suggest a new signalling cascade from the Fas receptor via caspases to acidic sphingomyelinase, ceramide and JNK/p38-K.

Acids↗
PubMed1998

Extended therapeutic window for caspase inhibition and synergy with MK-801 in the treatment of cerebral histotoxic hypoxia.

In rats, striatal histotoxic hypoxic lesions produced by the mitochondrial toxin malonate resemble those of focal cerebral ischemia. Intrastriatal injections of malonate induced cleavage of caspase-2 beginning at 6 h, and caspase-3-like activity as identified by DEVD biotin affinity-labeling within 12 h. DEVD affinity-labeling was prevented and lesion volume reduced in transgenic mice overexpressing BCL-2 in neuronal cells. Intrastriatal injection of the tripeptide, N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (zVAD-fmk), a caspase inhibitor, at 3 h, 6 h, or 9 h after malonate injections reduced the lesion volume produced by malonate. A combination of pretreatment with the NMDA antagonist, dizocilpine (MK-801), and delayed treatment with zVAD-fmk provided synergistic protection compared with either treatment alone and extended the therapeutic window for caspase inhibition to 12 h. Treatment with cycloheximide and zVAD-fmk, but not with MK-801, blocked the malonate-induced cleavage of caspase-2. NMDA injections alone resulted in a weak caspase-2 cleavage. These results suggest that malonate toxicity induces neuronal death by more than one pathway. They strongly implicate early excitotoxicity and delayed caspase activation in neuronal loss after focal ischemic lesions and offer a new strategy for the treatment of stroke.

Amino Acid Chloromethyl Ketones↗