title: Molecular mechanisms of nutlin-induced apoptosis in multiple myeloma: evidence for p53-transcription-dependent and -independent pathways.
abstract: Multiple myeloma (MM) is an incurable plasma cell malignancy in which p53 is rarely mutated. Thus, activation of the p53 pathway by a small molecule inhibitor of the p53-MDM2 interaction, nutlin, in MM cells retaining wild type p53 is an attractive therapeutic strategy. Recently we reported that nutlin plus velcade (a proteasome inhibitor) displayed a synergistic response in MM. However, the mechanism of the p53-mediated apoptosis in MM has not been fully understood. Our data show that nutlin-induced apoptosis correlated with reduction in cell viability, upregulation of p53, p21 and MDM2 protein levels with a simultaneous increase in pro-apoptotic targets PUMA, Bax and Bak and downregulation of anti-apoptotic targets Bcl2 and survivin and activation of caspase in MM cells harboring wild type p53. Nutlin-induced apoptosis was inhibited when activation of caspase was blocked by the caspase inhibitor. Nutlin caused mitochondrial translocation of p53 where it binds with Bcl2, leading to cytochrome C release. Moreover, blocking the transcriptional arm of p53 by the p53-specific transcriptional inhibitor, pifithrin-α, not only inhibited nutlin-induced upregulation of p53-transcriptional targets but also augmented apoptosis in MM cells, suggesting an association of transcription-independent pathway of apoptosis. However, inhibitor of mitochondrial translocation of p53, PFT-μ, did not prevent nutlin-induced apoptosis, suggesting that the p53 transcription-dependent pathway was also operational in nutlin-induced apoptosis in MM. Our study provides the evidence that nutlin-induced apoptosis in MM cells is mediated by transcription-dependent and -independent pathways and supports further clinical evaluation of nutlin as a novel therapeutic agent in MM.
authors.affiliation: Division of Molecular and Cellular Biology, Toronto General Hospital Research Institute, Dept of Laboratory Medicine and Pathobiology, University of Toronto, Ontario, Canada.
authors.forename: Manujendra N
authors.initials: MN
authors.lastname: Saha
authors.forename: Hua
authors.initials: H
authors.lastname: Jiang
authors.forename: Hong
authors.initials: H
authors.lastname: Chang
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism; D052160:Benzothiazoles / Q000494:pharmacology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology*; D015151:Immunoblotting; D009101:Multiple Myeloma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D010879:Piperazines / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2 / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides / Q000494:pharmacology; D014050:Toluene / Q000031:analogs & derivatives / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
chemical_list: C545747:2-phenylacetylenesulfonamide; D051017:Apoptosis Regulatory Proteins; D052160:Benzothiazoles; D050759:Cyclin-Dependent Kinase Inhibitor p21; D007093:Imidazoles; D010879:Piperazines; D019253:Proto-Oncogene Proteins c-bcl-2; D013449:Sulfonamides; D016159:Tumor Suppressor Protein p53; D014050:Toluene; C482205:nutlin 3; C121565:pifithrin; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2