Bortezomib (Bor), a proteasome inhibitor, has marked therapeutic effects in multiple myeloma (MM), and its synergistic effects with other anticancer agents have been widely investigated. In the present study, endoplasmic reticulum (ER) stress was the target of the treatment strategy; anacardic acid (AA) and Bor induce ER stress, resulting in apoptosis of multiple myeloma cells. AA/Bor combination therapy exhibited overt cytotoxicity in MM cells, by synergistically reducing cell growth and promoting cell death. Notably, expression levels of the stress‑associated molecules binding protein, phosphorylated eukaryotic initiation factor 2α, activating transcription factor 4 (ATF4) and CCAAT‑enhancer binding protein homologous protein (CHOP) were increased following treatment. AA/Bor combination therapy‑induced U266 cell cytotoxicity was partially reversed by ATF4 gene silencing and slightly enhanced by CHOP knockdown. The results of the present study suggest that AA/Bor combination may be a potential therapeutic strategy for MM treatment.