title: BH3 mimetic-elicited Ca2+signals in pancreatic acinar cells are dependent on Bax and can be reduced by Ca2+-like peptides.
abstract: BH3 mimetics are small-molecule inhibitors of B-cell lymphoma-2 (Bcl-2) and Bcl-xL, which disrupt the heterodimerisation of anti- and pro-apoptotic Bcl-2 family members sensitising cells to apoptotic death. These compounds have been developed as anti-cancer agents to counteract increased levels of Bcl-2 proteins often present in cancer cells. Application of a chemotherapeutic drug supported with a BH3 mimetic has the potential to overcome drug resistance in cancers overexpressing anti-apoptotic Bcl-2 proteins and thus increase the success rate of the treatment. We have previously shown that the BH3 mimetics, BH3I-2' and HA14-1, induce Ca2+release from intracellular stores followed by a sustained elevation of the cytosolic Ca2+concentration. Here we demonstrate that loss of Bax, but not Bcl-2 or Bak, inhibits this sustained Ca2+elevation. What is more, in the absence of Bax, thapsigargin-elicited responses were decreased; and in two-photon-permeabilised bax-/-cells, Ca2+loss from the ER was reduced compared to WT cells. The Ca2+-like peptides, CALP-1 and CALP-3, which activate EF hand motifs of Ca2+-binding proteins, significantly reduced excessive Ca2+signals and necrosis caused by two BH3 mimetics: BH3I-2' and gossypol. In the presence of CALP-1, cell death was shifted from necrotic towards apoptotic, whereas CALP-3 increased the proportion of live cells. Importantly, neither of the CALPs markedly affected physiological Ca2+signals elicited by ACh, or cholecystokinin. In conclusion, the reduction in passive ER Ca2+leak in bax-/-cells as well as the fact that BH3 mimetics trigger substantial Ca2+signals by liberating Bax, indicate that Bax may regulate Ca2+leak channels in the ER. This study also demonstrates proof-of-principle that pre-activation of EF hand Ca2+-binding sites by CALPs can be used to ameliorate excessive Ca2+signals caused by BH3 mimetics and shift necrotic death towards apoptosis.
authors.affiliation: Medical Research Council Group, Cardiff School of Biosciences, Cardiff University, Cardiff CF10 3AX, UK.
authors.forename: Pawel E
authors.initials: PE
authors.lastname: Ferdek
authors.affiliation: Medical Research Council Group, Cardiff School of Biosciences, Cardiff University, Cardiff CF10 3AX, UK.
authors.forename: Monika A
authors.initials: MA
authors.lastname: Jakubowska
authors.affiliation: Medical Research Council Group, Cardiff School of Biosciences, Cardiff University, Cardiff CF10 3AX, UK.
authors.forename: Polina
authors.initials: P
authors.lastname: Nicolaou
authors.affiliation: Medical Research Council Group, Cardiff School of Biosciences, Cardiff University, Cardiff CF10 3AX, UK.
authors.forename: Julia V
authors.initials: JV
authors.lastname: Gerasimenko
authors.affiliation: Medical Research Council Group, Cardiff School of Biosciences, Cardiff University, Cardiff CF10 3AX, UK.
authors.forename: Oleg V
authors.initials: OV
authors.lastname: Gerasimenko
authors.affiliation: Medical Research Council Group, Cardiff School of Biosciences, Cardiff University, Cardiff CF10 3AX, UK.|Systems Immunity Research Institute, Cardiff University, Cardiff CF14 4XN, UK.
authors.forename: Ole H
authors.initials: OH
authors.lastname: Petersen
mesh_terms: D061354:Acinar Cells / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D001549:Benzamides / Q000494:pharmacology; D001578:Benzopyrans / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009570:Nitriles / Q000494:pharmacology; D009842:Oligopeptides / Q000378:metabolism; D010179:Pancreas / Q000187:drug effects / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D019284:Thapsigargin / Q000378:metabolism; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism*
chemical_list: C474172:3-iodo-5-chloro-N-(2-chloro-5-((4-chlorophenyl)sulphonyl)phenyl)-2-hydroxybenzamide; D000970:Antineoplastic Agents; C474007:Bax protein (53-86); D001549:Benzamides; D001578:Benzopyrans; D009570:Nitriles; D009842:Oligopeptides; D010446:Peptide Fragments; D010455:Peptides; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein; C412708:calcium-like peptide CALP1; C412709:calcium-like peptide CALP3; C456800:ethyl 2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate; D019284:Thapsigargin; D002118:Calcium