title: The proteasome inhibitor carfilzomib functions independently of p53 to induce cytotoxicity and an atypical NF-κB response in chronic lymphocytic leukemia cells.
abstract: OBJECTIVE
  The proteasome consists of chymotrypsin-like (CT-L), trypsin-like, and caspase-like subunits that cleave substrates preferentially by amino acid sequence. Proteasomes mediate degradation of regulatory proteins of the p53, Bcl-2, and nuclear factor-κB (NF-κB) families that are aberrantly active in chronic lymphocytic leukemia (CLL). CLL remains an incurable disease, and new treatments are especially needed in the relapsed/refractory setting. We therefore investigated the effects of the proteasome inhibitor carfilzomib (CFZ) in CLL cells.
  
  
  METHODS
  Tumor cells from CLL patients were assayed in vitro using immunoblotting, real-time polymerase chain reaction, and electrophoretic mobility shift assays. In addition, a p53 dominant-negative construct was generated in a human B-cell line.
  
  
  RESULTS
  Unlike bortezomib, CFZ potently induces apoptosis in CLL patient cells in the presence of human serum. CLL cells have significantly lower basal CT-L activity compared to normal B and T cells, although activity is inhibited similarly in T cells versus CLL. Co-culture of CLL cells on stroma protected from CFZ-mediated cytotoxicity; however, PI3K inhibition significantly diminished this stromal protection. CFZ-mediated cytotoxicity in leukemic B cells is caspase-dependent and occurs irrespective of p53 status. In CLL cells, CFZ promotes atypical activation of NF-κB evidenced by loss of cytoplasmic IκBα, phosphorylation of IκBα, and increased p50/p65 DNA binding, without subsequent increases in canonical NF-κB target gene transcription.
  
  
  CONCLUSIONS
  Together, these data provide new mechanistic insights into the activity of CFZ in CLL and support phase I investigation of CFZ in this disease.
authors.affiliation: College of Pharmacy, The Ohio State University, Columbus, Ohio, USA.
authors.forename: Sneha V
authors.initials: SV
authors.lastname: Gupta
authors.forename: Erin
authors.initials: E
authors.lastname: Hertlein
authors.forename: Yanhui
authors.initials: Y
authors.lastname: Lu
authors.forename: Ellen J
authors.initials: EJ
authors.lastname: Sass
authors.forename: Rosa
authors.initials: R
authors.lastname: Lapalombella
authors.forename: Timothy L
authors.initials: TL
authors.lastname: Chen
authors.forename: Melanie E
authors.initials: ME
authors.lastname: Davis
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Woyach
authors.forename: Amy
authors.initials: A
authors.lastname: Lehman
authors.forename: David
authors.initials: D
authors.lastname: Jarjoura
authors.forename: John C
authors.initials: JC
authors.lastname: Byrd
authors.forename: David M
authors.initials: DM
authors.lastname: Lucas
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D001402:B-Lymphocytes / Q000187:drug effects / Q000201:enzymology; D001593:Benzyl Compounds / Q000494:pharmacology; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D061945:Caspase Inhibitors / Q000494:pharmacology; D020169:Caspases / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D003497:Cyclic N-Oxides; D004353:Drug Evaluation, Preclinical; D006801:Humans; D006845:Hydrocarbons, Fluorinated / Q000494:pharmacology; D007212:Indolizines; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000188:drug therapy*; D016328:NF-kappa B / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D061988:Proteasome Inhibitors / Q000494:pharmacology*; D021122:Protein Subunits / Q000037:antagonists & inhibitors / Q000378:metabolism; D011726:Pyridinium Compounds / Q000494:pharmacology; D013601:T-Lymphocytes / Q000187:drug effects / Q000201:enzymology; D014407:Tumor Cells, Cultured / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
chemical_list: D000970:Antineoplastic Agents; D001593:Benzyl Compounds; D019086:Bridged Bicyclo Compounds, Heterocyclic; D061945:Caspase Inhibitors; D020169:Caspases; D003497:Cyclic N-Oxides; D006845:Hydrocarbons, Fluorinated; D007212:Indolizines; D016328:NF-kappa B; D009842:Oligopeptides; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D046988:Proteasome Endopeptidase Complex; D061988:Proteasome Inhibitors; D021122:Protein Subunits; D011726:Pyridinium Compounds; D016159:Tumor Suppressor Protein p53; C476029:Boc-D-FMK; C495901:TP53 protein, human; C553669:dinaciclib; C524865:carfilzomib