RESUMO
CD95 (Fas/Apo-1)-mediated apoptosis was shown to occur through two distinct pathways. One involves a direct activation of caspase-3 by large amounts of caspase-8 generated at the DISC (Type I cells). The other is related to the cleavage of Bid by low concentration of caspase-8, leading to the release of cytochrome c from mitochondria and the activation of caspase-3 by the cytochrome c/APAF-1/caspase-9 apoptosome (Type II cells). It is also known that the protein synthesis inhibitor cycloheximide (CHX) sensitizes Type I cells to CD95-mediated apoptosis, but it remains contradictory whether this effect also occurs in Type II cells. Here, we show that sub-lethal doses of CHX render both Type I and Type II cells sensitive to the apoptogenic effect of anti-CD95 antibodies but not to chemotherapeutic drugs. Moreover, Bcl-2-positive Type II cells become strongly sensitive to CD95-mediated apoptosis by the addition of CHX to the cell culture. This is not the result of a restraint of the anti-apoptotic effect of Bcl-2 at the mitochondrial level since CHX-treated Type II cells still retain their resistance to chemotherapeutic drugs. Therefore, CHX treatment is granting the CD95-mediated pathway the ability to bypass the mitochondria requirement to apoptosis, much alike to what is observed in Type I cells.
Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Apoptose/fisiologia , Mitocôndrias/metabolismo , Transdução de Sinais/fisiologia , Receptor fas/metabolismo , Anticorpos/farmacologia , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/efeitos dos fármacos , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/metabolismo , Caspases/efeitos dos fármacos , Caspases/metabolismo , Cicloeximida/farmacologia , Citocromos c/efeitos dos fármacos , Citocromos c/metabolismo , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/fisiologia , Sinergismo Farmacológico , Células HL-60 , Humanos , Mitocôndrias/efeitos dos fármacos , Inibidores da Síntese de Proteínas/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Receptor fas/antagonistas & inibidoresRESUMO
BACKGROUND: Apoptosis is an essential form of cell death, the failure of which can lead to cancer development. Cancer including leukemia is usually treated with chemotherapeutic drugs that can be effective, but frequently problems are encountered that impair the success of the treatment. Butyrate is a short-chain fatty acid that can have many effects on different cells, including apoptosis. METHODS: The effect of a combination treatment with butyrate and antineoplastic agents Ara-C, etoposide and vincristine is evaluate on the leukemic cell line THP-1. RESULTS: We show that butyrate increased apoptosis induced by the three agents as seen by measurement of DNA content, annexin exposure and morphological characteristics. We also demonstrate that the process of apoptosis induced by butyrate and chemotherapeutic drugs involves the participation of caspases and induced activation of caspase-3, -8 and -9. CONCLUSIONS: We believe that butyrate could be a promising therapeutic agent for the treatment of leukemia in combination with other antineoplastic drugs.
Assuntos
Antineoplásicos/farmacologia , Butiratos/farmacologia , Sinergismo Farmacológico , Leucemia Monocítica Aguda/tratamento farmacológico , Leucemia Monocítica Aguda/patologia , Clorometilcetonas de Aminoácidos/farmacologia , Clorometilcetonas de Aminoácidos/uso terapêutico , Antineoplásicos/classificação , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Western Blotting , Butiratos/química , Butiratos/uso terapêutico , Inibidores de Caspase , Caspases/metabolismo , Caspases/uso terapêutico , Linhagem Celular Tumoral , Citarabina/farmacologia , Citarabina/uso terapêutico , Replicação do DNA/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Quimioterapia Combinada , Etoposídeo/farmacologia , Etoposídeo/uso terapêutico , Humanos , Leucemia Monocítica Aguda/metabolismo , Vincristina/farmacologia , Vincristina/uso terapêuticoRESUMO
Bcr-Abl is one of the most potent antiapoptotic molecules and is the tyrosine-kinase implicated in Philadelphia (Ph) chromosome-positive leukemia. It is still obscure how Bcr-Abl provides the leukemic cell a strong resistance to chemotherapeutic drugs. A rational drug development produced a specific inhibitor of the catalytic activity of Bcr-Abl called STI571. This drug was shown to eliminate Bcr-Abl-positive cells both in vitro and in vivo, although resistant cells may appear in culture and relapse occurs in some patients. In the study described here, Bcr-Abl-positive cells treated with tyrosine-kinase inhibitors such as herbimycin A, genistein or STI571 lost their phosphotyrosine-containing proteins, but were still extremely resistant to apoptosis. Therefore, in the absence of tyrosine-kinase activity, Bcr-Abl-positive cells continue to signal biochemically to prevent apoptosis induced by chemotherapeutic drugs. We propose that secondary antiapoptotic signals are entirely responsible for the resistance of Bcr-Abl-positive cells. Precise determination of such signals and rational drug development against them should improve the means to combat Ph chromosome-positive leukemia.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas de Fusão bcr-abl/metabolismo , Piperazinas/farmacologia , Proteínas Tirosina Quinases/metabolismo , Pirimidinas/farmacologia , Benzamidas , Benzoquinonas , Western Blotting , Caspases/metabolismo , Resistencia a Medicamentos Antineoplásicos , Proteínas de Fusão bcr-abl/antagonistas & inibidores , Genisteína/farmacologia , Células HL-60/efeitos dos fármacos , Células HL-60/metabolismo , Humanos , Mesilato de Imatinib , Células K562/efeitos dos fármacos , Células K562/metabolismo , Lactamas Macrocíclicas , Proteínas Tirosina Quinases/antagonistas & inibidores , Quinonas/farmacologia , Rifabutina/análogos & derivados , Fatores de TempoRESUMO
Cyclobutane pyrimidine dimers (CPDs) are directly involved in signaling for UV-induced apoptosis in mammalian cells. Failure to remove these lesions, specially those located at actively expressing genes, is critical, as cells defective in transcription coupled repair have increased apoptotic levels. Thus, the blockage of RNA synthesis by lesions is an important candidate event triggering off active cell death. In this work, wild-type and XPB mutated Chinese hamster ovary (CHO) cells expressing a marsupial photolyase, that removes specifically CPDs from the damaged DNA, were generated, in order to investigate the importance of this lesion in both RNA transcription blockage and apoptotic induction. Photorepair strongly recovers RNA synthesis in wild-type CHO cell line, although the resumption of transcription is decreased in XPB deficient cells. This recovery is accompanied by the prevention of cells entering into apoptosis. These results demonstrate that marsupial photolyase has access to CPDs blocking RNA synthesis in vivo, and this may be affected by the presence of a mutated XPB protein.