BH3 mimetic-elicited Ca signals in pancreatic acinar cells are dependent on Bax and can be reduced by Ca-like peptides.

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 Ca release from intracellular stores followed by a sustained elevation of the cytosolic Ca concentration. Here we demonstrate that loss of Bax, but not Bcl-2 or Bak, inhibits this sustained Ca elevation. What is more, in the absence of Bax, thapsigargin-elicited responses were decreased; and in two-photon-permeabilised bax cells, Ca loss from the ER was reduced compared to WT cells. The Ca-like peptides, CALP-1 and CALP-3, which activate EF hand motifs of Ca-binding proteins, significantly reduced excessive Ca 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 Ca signals elicited by ACh, or cholecystokinin. In conclusion, the reduction in passive ER Ca leak in bax cells as well as the fact that BH3 mimetics trigger substantial Ca signals by liberating Bax, indicate that Bax may regulate Ca leak channels in the ER. This study also demonstrates proof-of-principle that pre-activation of EF hand Ca-binding sites by CALPs can be used to ameliorate excessive Ca signals caused by BH3 mimetics and shift necrotic death towards apoptosis.