Piperlongumine and bortezomib synergically inhibit cholangiocarcinoma via ER stress-induced cell death.

Cholangiocarcinoma (CCA) is a lethal malignancy of the cholangiocytes lining the biliary tree. Only 25% of affected patients are eligible for resection due to late-stage diagnosis. Systemic chemotherapy is recommended for those inoperable patients; however, an inadequate response to such treatment remains a significant obstacle. Piperlongumine (PL) is a biologically active alkaloid that selectively kills various cancer cells through the induction of reactive oxygen species (ROS). The role of PL has been shown through its inhibiting the ubiquitin-proteasome system. The mechanism of PL-induced CCA cell death was investigated by inhibiting the UPS and testing the therapeutic potential of combining PL and the proteasome inhibitor bortezomib. A single treatment with PL or BTZ suppressed CCA cell growth. Combined treatment with PL with BTZ produced a synergistic interaction, evidenced by (1) a combination index of < 1 and (2) induction of cell cycle arrest and down-regulation of cell cycle markers. PL induced the accumulation of poly-ubiquitinated proteins in CCA cells but did not affect proteasome activity. PL, in combination with BTZ, amplified the accumulation of poly-ubiquitinated proteins in CCA cells, leading to an endoplasmic reticulum (ER) stress response through the induction of X-box binding protein mRNA splicing. Moreover, PL-combined BTZ promoted the activation of a proapoptotic unfolded protein response via the ATF4-CHOP axis. PL induced CCA cell death via increased accumulation of the poly-ubiquitinated proteins. PL also enhanced the anti-cancer activity of BTZ via ER stress-induced CCA cell death. Thus, the combination of PL and BTZ has potential as an alternative therapeutic option for CCA.