Low doses of paclitaxel potently induce apoptosis in human retinoblastoma Y79 cells by up-regulating E2F1.
Paclitaxel (PTX) is an anticancer drug currently in phase II clinical trials. This study shows for the first time that low doses of PTX (5 nM) potently induce apoptosis in human retinoblastoma Y79 cells. The effect of PTX is accompanied by a potent induction of E2F1 which appears to play a critical role in the effects induced by PTX. PTX induced a dose- and time-dependent effect, with G2/M arrest, cyclines A, E and B1 accumulation and a marked modification in the status of Cdc2-cyclin B1 complex, the major player of the G2/M checkpoint. Apoptosis followed G2/M arrest. An early and prolonged increase in p53 expression with its stabilization by phosphorylation and acetylation and its nuclear translocation occurred. Consistently, PTX increased p21WAF1, bax and MDM2 levels, suggesting that p53 is transcriptionally active. p53 accumulated following both E2F1 up-regulation and increase in the levels of p14ARF which interacts with MDM2 preventing ubiquitination and proteosomal degradation of p53. Both extrinsic (E2F1/Fas/JNK/caspase-2 activation) and intrinsic (Bcl-2 phosphorylation, Bid fragmentation and Bax increase) pathways seemed to be involved. Loss of mitochondrial potential and activation of apoptosome and executive caspase-3,-6 and-7 was shown. Incubation with either the irreversible pan-caspase inhibitors Z-VAD-FMK, or SP600125, a selective inhibitor of JNK, or pifithrin alpha, a potent p53 inhibitor, significantly inhibited the effects induced by PTX.
Drago-Ferrante
Rosa
R
Dipartimento di Scienze Biochimiche, Università degli Studi di Palermo, Policlinico, 90127 Palermo, Italy.
Santulli
Andrea
A
Di Fiore
Riccardo
R
Giuliano
Michela
M
Calvaruso
Giuseppe
G
Tesoriere
Giovanni
G
Vento
Renza
R
D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis; D002455:Cell Division; D045744:Cell Line, Tumor; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D050687:E2F1 Transcription Factor / Q000378:metabolism*; D016195:G2 Phase; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D053078:Membrane Potential, Mitochondrial; D017239:Paclitaxel / Q000494:pharmacology*; D010766:Phosphorylation; D012175:Retinoblastoma / Q000188:drug therapy* / Q000473:pathology*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D015854:Up-Regulation
D000972:Antineoplastic Agents, Phytogenic; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050687:E2F1 Transcription Factor; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D017239:Paclitaxel