Dose-response transition from cell cycle arrest to apoptosis with selective degradation of Mdm2 and p21WAF1/CIP1 in response to the novel anticancer agent, aminoflavone (NSC 686,288).
Aminoflavone (AF, NSC 686,288) is beginning clinical trials. It induces replication-mediated histone H2AX phosphorylation, DNA-protein crosslinks and activates p53. Here, we studied p21(CIP1/WAF1) and Mdm2 responses to AF. Although p53 stabilization and phosphorylation at serine 15 increased with dose and time of exposure, Mdm2 and p21(CIP1/WAF1) protein levels displayed a biphasic response, as they accumulated at submicromolar doses and then decreased with increasing AF. As both Mdm2 and p21(CIP1/WAF1) mRNA levels increased with AF concentration without reduction at higher concentrations, we measured the half-lives of Mdm2 and p21(CIP1/WAF1) proteins. Mdm2 and p21(CIP1/WAF1) half-lives were shortened with increasing AF concentrations. Proteasomal degradation appears responsible for the decrease of both Mdm2 and p21(CIP1/WAF1), as MG-132 prevented their degradation and revealed AF-induced Mdm2 polyubiquitylation. AF also induced protein kinase B (Akt) activation, which was reduced with increasing AF concentrations. Suppression of Akt by small interfering RNA was associated with downregulation of Mdm2 and p21(CIP1/WAF1) and with enhanced apoptosis. These results suggest that the cellular responses to AF are determined at least in part by Mdm2 and p21(CIP1/WAF1) protein levels, as well as by Akt activity, leading either to cell cycle arrest when Mdm2 and p21(CIP1/WAF1) are elevated, or to apoptosis when Mdm2 and p21(CIP1/WAF1) are degraded by the proteasome and Akt insufficiently activated to protect against apoptosis.
Meng
L-H
LH
Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4255, USA.
Kohn
K W
KW
Pommier
Y
Y
D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D015153:Blotting, Western; D002453:Cell Cycle / Q000187:drug effects*; D045744:Cell Line, Tumor; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism*; D003513:Cycloheximide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005419:Flavonoids / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D008954:Models, Biological; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2 / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D005419:Flavonoids; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D025801:Ubiquitin; C413760:aminoflavone; D003513:Cycloheximide; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D051057:Proto-Oncogene Proteins c-akt; D046988:Proteasome Endopeptidase Complex