Eeyarestatin causes cervical cancer cell sensitization to bortezomib treatment by augmenting ER stress and CHOP expression

ObjectiveThe proteasome inhibitor bortezomib is currently being tested in clinical trials against refractory cervical cancer. However, high doses of bortezomib are associated with adverse effects, which may lead to treatment abrogation or to the use of lower, ineffective doses. We investigated combination drug treatments that could enhance the efficacy of low bortezomib concentrations on cervical cancer cells.MethodsThe cervical cancer cell lines CaSki, HeLa and SW756 were treated with various combinations of bortezomib and eeyarestatin. Treated cells were analyzed for cell viability by clonal assays and the MTT assay, and for expression of pro-apoptotic proteins and cell stress markers by immunofluorescence, immunoblots and RT-PCR analysis.ResultsCotreatment of bortezomib with eeyarestatin markedly enhanced cell death in cervical cancer cells, allowing reduction of the bortezomib concentration necessary for efficient cell death to as low as 5 ng/ml. Combination of bortezomib with eeyarestatin resulted in a massive induction of the endoplasmic reticulum stress reaction, small and large heat shock protein activation, autophagy, and upregulation of pro-apoptotic CHOP.ConclusionEeyarestatin is a small molecule recently shown to cause endoplasmic reticulum stress by inhibiting the endoplasmic reticulum-associated degradation pathway, which directs misfolded cytotoxic proteins to proteasomal degradation. Concomitant inhibition of both pathways markedly enhances the efficacy of bortezomib against cervical cancer cells and thus may be applied to reduce the bortezomib dosage required for efficient cervical cancer treatment.

► Eeyarestatin enhances the cytotoxic effect of bortezomib in cervical cancer cells.
► Bortezomib and eeyarestatin induce a mitochondria-independent cell death mechanism.
► Bortezomib and eeyarestatin cause upregulation of the pro-apoptotic protein CHOP.