The NEDD8 pathway is a known activator of the ubiquitin–protease system, a complex that is partially responsible for the degradation of proteins involved in cell-cycle regulation and neoplastic growth. In this study, we evaluated the antitumor potential of MLN4924 (pevonedistat), a potent NEDD8 inhibitor. We hypothesized that MLN4924 treatment induces apoptosis in human melanoma cells. A375 and Mel39 BRAF V600E mutant melanoma cell lines were treated in vitro with MLN4924 alone or in combination with interferon-α (IFN-α) or vemurafenib – therapeutic agents utilized on melanoma patients. Annexin/propidium iodine flow cytometry analysis showed that treatment with MLN4924 for 72 h induced apoptosis in A375 and Mel39 melanoma cells with an IC50 of 1200 and 143 nmol/l, respectively. Combination therapy of A375 cells with 104 U/ml IFN-α and 1200 nmol/l MLN4924 led to a significant increase in cell death (78.2±3.7%) compared with single-agent treatment by IFN-α (17.5±2.5%) or MLN4924 (50.7±1.0%; P<0.005). Treatment of A375 cells with 1 μmol/l vemurafenib had a notable effect on cell viability. However, the addition of MLN4924 to vemurafenib had an inhibitory effect on apoptosis. Results from MTS proliferation assays indicate that MLN4924 has antiproliferative effects on melanoma cells in vitro, with the addition of IFN-α further inhibiting proliferation. Pretreatment with MLN4924 led to A375 cell sensitization to vemurafenib treatment and immunoblot analysis of MLN4924-treated cells revealed cleavage of caspase-3, caspase-7, caspase-9, and poly-ADP-ribose polymerase. These results show that MLN4924 does have an efficacy in treating melanoma in vitro alone or in combination with IFN-α, and thus it may have potential use in patients with advanced melanoma.