title: Knockdown of BAG3 sensitizes bladder cancer cells to treatment with the BH3 mimetic ABT-737.
abstract: OBJECTIVE
  BAG3 is overexpressed in several malignancies and mediates a non-canonical, selective form of (macro)autophagy. By stabilizing pro-survival Bcl-2 proteins in complex with HSP70, BAG3 can also exert an apoptosis-antagonizing function. ABT-737 is a high affinity Bcl-2 inhibitor that fails to target Mcl-1. This failure may confer resistance in various cancers.
  
  
  METHODS
  Urothelial cancer cells were treated with the BH3 mimetics ABT-737 and (-)-gossypol, a pan-Bcl-2 inhibitor which inhibits also Mcl-1. To clarify the importance of the core autophagy regulator ATG5 and BAG3 in ABT-737 treatment, cell lines carrying a stable lentiviral knockdown of ATG5 and BAG3 were created. The synergistic effect of ABT-737 and pharmaceutical inhibition of BAG3 with the HSF1 inhibitor KRIBB11 or sorafenib was also evaluated. Total cell death and apoptosis were quantified by FACS analysis of propidium iodide, annexin. Target protein analysis was conducted by Western blotting.
  
  
  RESULTS
  Knockdown of BAG3 significantly downregulated Mcl-1 protein levels and sensitized urothelial cancer cells to apoptotic cell death induced by ABT-737, while inhibition of bulk autophagy through depletion of ATG5 had no discernible effect on cell death. Similar to knockdown of BAG3, pharmacological targeting of the BAG3/Mcl-1 pathway with KRIBB11 was capable to sensitize both cell lines to treatment with ABT-737.
  
  
  CONCLUSIONS
  Our results show that BAG3, but not bulk autophagy has a major role in the response of bladder cancer cells to BH3 mimetics. They also suggest that BAG3 is a suitable target for combined therapies aimed at synergistically inducing apoptosis in bladder cancer.
authors.affiliation: Department of Urology, Goethe University Hospital, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany. jens.mani@kgu.de.
authors.forename: Jens
authors.initials: J
authors.lastname: Mani
authors.affiliation: Experimental Neurosurgery, Neuroscience Center, Goethe University Hospital, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany.
authors.forename: Patrick
authors.initials: P
authors.lastname: Antonietti
authors.affiliation: Experimental Neurosurgery, Neuroscience Center, Goethe University Hospital, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Rakel
authors.affiliation: Department of Urology, Goethe University Hospital, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany.
authors.forename: Roman
authors.initials: R
authors.lastname: Blaheta
authors.affiliation: Department of Urology, Goethe University Hospital, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany.
authors.forename: Georg
authors.initials: G
authors.lastname: Bartsch
authors.affiliation: Department of Urology, Goethe University Hospital, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany.
authors.forename: Axel
authors.initials: A
authors.lastname: Haferkamp
authors.affiliation: Experimental Neurosurgery, Neuroscience Center, Goethe University Hospital, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany.
authors.forename: Donat
authors.initials: D
authors.lastname: Kögel
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis / Q000235:genetics*; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis / Q000235:genetics*; D001713:Biphenyl Compounds / Q000627:therapeutic use*; D015153:Blotting, Western; D002084:Butylated Hydroxytoluene / Q000031:analogs & derivatives; D002295:Carcinoma, Transitional Cell / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D045744:Cell Line, Tumor; D004273:DNA, Neoplasm / Q000235:genetics*; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D009596:Nitrophenols / Q000627:therapeutic use*; D010879:Piperazines / Q000627:therapeutic use; D019253:Proto-Oncogene Proteins c-bcl-2; D013449:Sulfonamides / Q000627:therapeutic use*; D001749:Urinary Bladder Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology
chemical_list: C501332:ABT-737; D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C414431:BAG3 protein, human; D001713:Biphenyl Compounds; D004273:DNA, Neoplasm; D009596:Nitrophenols; D010879:Piperazines; D019253:Proto-Oncogene Proteins c-bcl-2; D013449:Sulfonamides; D002084:Butylated Hydroxytoluene; C006008:BH 3
keywords: ABT-737; ATG5; BAG3; Bladder cancer