Proteasome inhibitor synergizes with histone deacetylase inhibitor to trigger ROS- and ER stress-induced apoptosis of nasopharyngeal carcinoma independent of aggresome disruption

Abstract

Proteasome inhibitor and histone deacetylase inhibitor (HDACi) can synergistically induce apoptosis of cancer cells through aggresome disruption or induction of endoplasmic reticulum (ER) stress. We previously reported that suberoylanilide hydroxamic acid (SAHA), a pan-HDACi, and bortezomib, a proteasome inhibitor can synergistically induce apoptosis of nasopharyngeal carcinoma (NPC) cells through reactive oxygen species (ROS)-driven caspase-dependent mechanism. Here, we wish to investigate the role of aggresome disruption or ER stress in the process. We first tested the effect of bortezomib combining with a more specific HDACi, including MS-275, apicidin or romidepsin, on proliferation of NPC cells. Bortezomib could synergize with each of the specific HDACis to induce killing of the NPC cells. Strong apoptosis was demonstrated by the proteolytic cleavage of PARP and caspase-3, -8 and -9 as well as the high percentage of annexin V/propidium iodide (AV/PI)-positive NPC cells upon treatment with combination of bortezomib at 7.5 to 15 nM and romidepsin at 2.5 to 5 nM. Consistently, the apoptosis was ROS- and caspase-dependent. Interestingly, -tubulin, a key substrate of histone deacetylase 6, was not acetylated, arguing against the involvement of aggresome disruption. In contrast, ER stress induction might be involved because ATF-4 and CHOP, which are markers of ER stress, were up-regulated by combined bortezomib/romidepsin and inhibition of caspase-4 could suppress the apoptosis. Furthermore, addition of ROS scavenger, N-acetyl-cysteine, suppressed the expression of ATF-4 and CHOP. We conclude that proteasome inhibitor can synergize with HDACi to trigger ROS- and ER stress-induced apoptosis of NPC cells, independent of aggresome disruption. This project is funded by NPC Area of Excellence (AoE/M 06/08 Center for Nasopharyngeal Carcinoma Research), CRCG (#10401264) and Epstein-Barr virus research (# 20004525) grants of A.K.S. Chiang.