RESUMO
Chimeric tumor suppressor-1 (CTS-1) is based on the sequence of p53 and was designed as a therapeutic tool resisting various mechanisms of p53 inactivation. We previously reported that an adenovirus expressing CTS-1 (Ad-CTS-1) has superior cell death-inducing activity in glioma cells compared with wild-type p53. Here, we used cDNA microarrays to detect changes in gene expression preferentially induced by Ad-CTS-1. The putative serine threonine kinase, PCTAIRE3, and the quinone oxireductase, PIG3, were strongly induced by Ad-CTS-1 compared with wild-type p53. An adenoviral vector encoding PCTAIRE3 (Ad-PCTAIRE3) induced growth arrest and killed a minor proportion of the glioma cells. Ad-PIG3 alone affected neither growth nor viability. However, coinfection with Ad-PCTAIRE3 and Ad-PIG3 resulted in enhanced growth inhibition compared with Ad-PCTAIRE3 infection alone. Ad-CTS1, Ad-PCTAIRE3 or Ad-PIG3 induced the formation of free reactive oxygen species (ROS). However, the prevention of ROS formation induced by Ad-PCTAIRE3 and Ad-CTS-1 did not block growth arrest and cell death, suggesting that ROS formation is not essential for these effects. Altogether, these data identify PCTAIRE3 as one novel growth-inhibitory and death-inducing p53 response gene and suggest that changes in the expression of specific target genes contribute to the superior anti-glioma activity of CTS-1.
Assuntos
Apoptose , Neoplasias Encefálicas/genética , Quinases Ciclina-Dependentes/genética , Glioma/genética , Proteínas Recombinantes de Fusão/metabolismo , Adenoviridae/genética , Apoptose/genética , Neoplasias Encefálicas/enzimologia , Neoplasias Encefálicas/patologia , Ciclo Celular/genética , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Técnicas de Transferência de Genes , Genes Neoplásicos/genética , Genes Supressores de Tumor , Glioma/enzimologia , Glioma/patologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas Proto-Oncogênicas/genética , Espécies Reativas de Oxigênio/metabolismo , Proteínas Recombinantes de Fusão/genética , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Alkylphosphocholines (APC) are candidate anticancer agents. We here report that APC induce the formation of large vacuoles and typical features of apoptosis in human glioma cell lines, but not in immortalized astrocytes. APC promote caspase activation, poly(ADP-ribose)-polymerase (PARP) processing and cytochrome c release from mitochondria. Adenoviral X-linked inhibitor of apoptosis (XIAP) gene transfer, or exposure to the caspase inhibitor, benzyloxycarbonyl-Val-Ala-DL-Asp-fluoro-methylketone zVAD-fmk, blocks caspase-7 and PARP processing, but not cell death, whereas BCL-X(L) blocks not only caspase-7 and PARP processing but also cell death. APC induce changes in Delta Psi m in sensitive glioma cells, but not in resistant astrocytes. The changes in Delta Psi m are unaffected by crm-A (cowpox serpin-cytokine response modifier protein A), XIAP or zVAD-fmk, but blocked by BCL-X(L), and are thus a strong predictor of cell death in response to APC. Free radicals are induced, but not responsible for cell death. APC thus induce a characteristic morphological, BCL-X(L)-sensitive, apparently caspase-independent cell death involving mitochondrial alterations selectively in neoplastic astrocytic cells.
Assuntos
Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Neoplasias Encefálicas/tratamento farmacológico , Caspases/metabolismo , Glioma/tratamento farmacológico , Fosforilcolina/análogos & derivados , Fosforilcolina/toxicidade , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Apoptose/fisiologia , Astrócitos/efeitos dos fármacos , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/ultraestrutura , Caspases/efeitos dos fármacos , Linhagem Celular Tumoral , Colágeno Tipo XI/efeitos dos fármacos , Colágeno Tipo XI/metabolismo , Citoplasma/efeitos dos fármacos , Citoplasma/patologia , Citoplasma/ultraestrutura , Radicais Livres/metabolismo , Glioma/patologia , Glioma/ultraestrutura , Humanos , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/genética , Microscopia Eletrônica de Transmissão , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Poli(ADP-Ribose) Polimerase-1 , Poli(ADP-Ribose) Polimerases , Proteínas/genética , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Estearatos/toxicidade , Vacúolos/efeitos dos fármacos , Vacúolos/patologia , Vacúolos/ultraestrutura , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X , Proteína bcl-XRESUMO
Diva is a novel proapoptotic member of the Bcl-2 protein family which binds apoptosis activating factor-1 (APAF-1). Diva is identical with Boo which was identified as a novel antiapoptotic Bcl-2 family protein. Here, we report that Diva promotes the cell cycle exit of human glioma cells in response to serum deprivation and inhibits apoptosis of these cells induced by CD95 ligand or chemotherapeutic drugs. In glioma cells, Diva interferes with apoptotic signaling downstream of cytochrome c release, but upstream of caspase activation, consistent with an inhibitory effect on the mitochondrial amplification step involving the apoptosome and APAF-1.
Assuntos
Neoplasias do Sistema Nervoso Central/patologia , Glioma/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fator Apoptótico 1 Ativador de Proteases , Caspase 3 , Caspase 8 , Caspase 9 , Caspases/metabolismo , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Neoplasias do Sistema Nervoso Central/tratamento farmacológico , Neoplasias do Sistema Nervoso Central/metabolismo , Cumarínicos/metabolismo , Meios de Cultura Livres de Soro/farmacologia , Grupo dos Citocromos c/metabolismo , Inibidores Enzimáticos/farmacologia , Proteína Ligante Fas , Genes myc/imunologia , Glioma/tratamento farmacológico , Glioma/metabolismo , Humanos , Glicoproteínas de Membrana/metabolismo , Compostos de Nitrosoureia/farmacologia , Oligopeptídeos/metabolismo , Proteínas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/genética , Teniposídeo/farmacologia , Transfecção , Células Tumorais CultivadasRESUMO
The cell cycle regulatory protein p27, an inhibitor of cyclin-dependent kinases (CDK), has been attributed a role in (i) prognosis in breast and colon cancer, (ii) induction of apoptosis in cancer cells, and (iii) resistance to cancer chemotherapy. Here we report that p27 is widely expressed in human malignant gliomas in vivo and in glioma cell lines in vitro. Serum deprivation or confluency promotes p27 protein accumulation in vitro. Neither baseline p27 levels nor p27 levels induced by confluency or serum deprivation correlate with p53 status or drug sensitivity of human glioma cell lines. Expression of antisense p27 mRNA increased the doubling times in T98G glioma cells, whereas sense p27 mRNA had no such effect. There was a density-dependent and drug-specific modulation of chemosensitivity by sense or antisense mRNA expression in T98G cells. Taken together, these data define a strong p27 response to altered growth conditions and suggest a role for p27 in modulating response to chemotherapy in human malignant glioma cells.