Perturbation of proteasome function by bortezomib leading to ER stress-induced apoptotic cell death in cholangiocarcinoma.

Objective: Cholangiocarcinoma (CCA) or cancer of the biliary tract is heterogeneous; however, chronic inflammatory-related features are unique in CCA. Moreover, the genes involved in proteasome functions are evidently increased in CCA. Hence, CCA might be vulnerable to endoplasmic reticulum (ER) stressors, particularly a proteasome inhibitor. Therefore, bortezomib (BTZ), a specific 26S proteasome inhibitor, was selected, and its antitumor effects against CCA were investigated.

Methods: Liver fluke-associated CCA cell lines were used. Cell proliferation and apoptosis detection were determined by a tetrazolium-based assay, caspase detection and annexin V binding assay. The accumulations of proteasome substrates, the inductions of ER stress and unfolded protein response (UPR) proteins were demonstrated by western blot and reporter systems. The in vivo anti-proliferative effect was accessed in a subcutaneous transplantation mouse model.

Results: BTZ inhibited CCA proliferation and induced caspase-dependent apoptosis, independently of the NF-κB pathway. Inhibition of protein degradation by BTZ led to the induction of UPR; induction of XBP1 splicing, ATF6 proteolysis and nuclear ATF4 as well as BiP and CHOP expressions were evident. Nevertheless, ER stress-induced UPR was overwhelming, leading to the activation of apoptosis demonstrated by proteolytic cleavages of ER-related caspase 4 and 12 as well as classical caspase 8, 9 and 3. The growth inhibitory effect of BTZ was supported by an in vivo model.

Conclusions: BTZ treatment could be a promising therapeutic approach for CCA treatment.