Paclitaxel and beta-lapachone synergistically induce apoptosis in human retinoblastoma Y79 cells by downregulating the levels of phospho-Akt.
Paclitaxel (PTX) and beta-lapachone (LPC) are naturally occurring compounds that have shown a large spectrum of anticancer activity. In this article we show for the first time that PTX/LPC combination induces potent synergistic apoptotic effects in human retinoblastoma Y79 cells. Combination of suboptimal doses of PTX (0.3 nM) and LPC (1.5 microM) caused biochemical and morphological signs of apoptosis at 48 h of treatment. These effects were accompanied by potent lowering in inhibitor of apoptosis proteins and by activation of Bid and caspases 3 and 6 with lamin B and PARP breakdown. PTX/LPC combination acted by favoring p53 stabilization through a lowering in p-Akt levels and in ps166-MDM2, the phosphorylated-MDM2 form that enters the nucleus and induces p53 export and degradation. Treatment with wortmannin or transfection with a dominant negative form of Akt anticipated at 24 h the effects induced by PTX/LPC, suggesting a protective role against apoptosis played by Akt in Y79 cells. In line with these results, we demonstrated that Y79 cells contain constitutively active Akt, which forms a cytosolic complex with p53 and MDM2 driving p53 degradation. PTX/LPC treatment induced a weakness of Akt-MDM2-p53 complex and increased nuclear p53 levels. Our results suggest that phospho-Akt lowering is at the root of the apoptotic action exerted by PTX/LPC combination and provide strong validation for a treatment approach that targets survival signals represented by phospho-Akt and inhibitor of apoptosis proteins.
D'Anneo
Antonella
A
Dipartimento di Scienze Biochimiche, Università degli Studi di Palermo, Policlinico, Palermo, Italy.
Augello
Giuseppa
G
Santulli
Andrea
A
Giuliano
Michela
M
di Fiore
Riccardo
R
Messina
Concetta
C
Tesoriere
Giovanni
G
Vento
Renza
R
D021581:Active Transport, Cell Nucleus; D000730:Androstadienes / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D051023:BH3 Interacting Domain Death Agonist Protein / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D053178:Caspase 6 / Q000378:metabolism; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004357:Drug Synergism; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000378:metabolism; D034921:Lamin Type B / Q000378:metabolism; D009285:Naphthoquinones / Q000494:pharmacology; D017239:Paclitaxel / Q000494:pharmacology; D010766:Phosphorylation; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D055550:Protein Stability; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism; D012175:Retinoblastoma / Q000201:enzymology / Q000473:pathology*; D013997:Time Factors; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D000077191:Wortmannin
D000730:Androstadienes; D000972:Antineoplastic Agents, Phytogenic; D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; D051034:Inhibitor of Apoptosis Proteins; D034921:Lamin Type B; D009285:Naphthoquinones; D047428:Protein Kinase Inhibitors; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; C014638:beta-lapachone; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D011065:Poly(ADP-ribose) Polymerases; D051057:Proto-Oncogene Proteins c-akt; C505407:CASP3 protein, human; C505514:CASP6 protein, human; D053148:Caspase 3; D053178:Caspase 6; D017239:Paclitaxel; D000077191:Wortmannin