RESUMO
We recently argued for a major role of p53 in staurosporine(ST)-induced apoptosis of immortalized epithelial cells, depending on their p53 status. Here, we studied the effects of PRIMA-1 (p53 reactivation and induction of massive apoptosis) and Pifithrin-alpha (p fifty-three inhibitor) in combination with ST to reinforce our previous results by respectively restoring or inhibiting the p53 transcriptional activity in different cell lines.PRIMA-1 does modify neither expression of apoptosis-related proteins nor the percentage of wild-type p53 HeLa and CaSki cells with [symbol: see text]delta psi m and DNA cleavage, whilst it increases by 45% Bax expression and apoptosis of mutated p53 C33A cells. Pifithrin-alpha, does modify neither Bax expression nor apoptosis level of C33A cells, but readily inhibits both [symbol: see text]delta psi m and DNA fragmentation of p53wt cells with decreasing Bax expression. These data support the evidence that PRIMA-1 could be a good candidate, as an anti-cancer drug targeting mutant p53, in order to increase ST efficiency. Moreover, Pifithrin-alpha could be used in combination with ST and PRIMA-1 to prevent side effects of anti-tumor therapies in cells expressing mutant P53.
Assuntos
Apoptose/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Proteínas de Membrana/farmacologia , Proteínas do Tecido Nervoso/farmacologia , Tiazóis/farmacologia , Tolueno/análogos & derivados , Proteína Supressora de Tumor p53/genética , Benzotiazóis , Técnicas de Cultura de Células , Linhagem Celular Transformada , Linhagem Celular Tumoral , Transformação Celular Viral , DNA de Neoplasias/análise , DNA de Neoplasias/metabolismo , Combinação de Medicamentos , Inibidores Enzimáticos/farmacologia , Feminino , Células HeLa , Humanos , Potenciais da Membrana/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mutação , Papillomaviridae/fisiologia , Estaurosporina/farmacologia , Tolueno/farmacologia , Transcrição Gênica/efeitos dos fármacos , Proteína X Associada a bcl-2/metabolismoRESUMO
The mitochondrial localization of p53 is an important event in p53-dependent apoptosis. Some p53 mutants defective for transcription also facilitate apoptosis through changes of the mitochondria. Here, apoptosis of HeLa and CaSki cells (p53(wt)), C33A and HaCat cells (p53(mt)) and SaOs-2 cells (p53 deficient) was induced by 300 nM staurosporine. We showed that wild-type p53, as well as p53 mutants, were transiently located to the mitochondria with changes in the mitochondrial membrane potential (Delta Psi m). However, in C33A cells harboring a p53 mutated on its DNA binding domain, Delta Psi m collapse and Sub-G1 DNA content were reduced compared to p53(wt) cells, whereas no significant difference was observed in HaCat cells with a p53 mutated on UV hot spots. In addition, inhibition of the mitochondrial permeability transition pores by cyclosporine A significantly reduced the Delta Psi m loss and the sub-G1 DNA content in p53 positive cells. These results indicate that Delta Psi m collapse is an early and necessary event, which plays an important role in apoptosis of immortal mammalian cells.