Multiple myeloma (MM) is a fatal hematological malignancy associated with disruption of RAS-to-MAP kinase (MAPK/ERK) signaling. Prenylation inhibitors such as farnesyltransferase inhibitors (FTIs) and the competitive HMG-CoA reductase inhibitor lovastatin have been shown to block RAS post-translational modification and thus transforming ability. Targeting down-stream members of the RAS signaling cascade with compounds such as the MEK inhibitor U0126 is another strategy used to successfully disrupt oncogenic RAS signal transduction. In order to assess the effects of co-treating MM cells with U0126 and prenylation inhibitors (e.g. FTI L-744,832 and lovastatin), six MM cell lines were assayed for viability using the MTT assay, induction of apoptosis by TUNEL assay, and MEK activation by a phospho-MEK-specific FACS assay. Treated cells were also subjected to Western blotting to determine levels of phosphorylated MAP kinase as well as processed and unprocessed RAS and RAS-related proteins. Co-treating MM cells with U0126 and FTI L-744,832 or lovastatin synergistically inhibited MM cell proliferation. Incubation of NCI-H929 cells with U0126 caused a concentration dependent increase of cells in the G0/G1 phase of the cell cycle. Accordingly, 48 h treatment of NCI-H929 cells with 20 μM U0126 caused a reduction in activated MEK in both G0/G1 and G2/M cell cycle phases (41.9 and 39.5%, respectively) while 50 μM reduced activated MEK levels by 64.8% in G0/G1 and 74.5% in G2/M. Lovastatin treatment (48 h, 10 μM) of NCI-H929 led to an accumulation of cells at the G1/S boundary of the cell cycle and >90% loss of activated MEK in both G0/G1 and G2/M. Lovastatin-induced changes in cell cycle distribution and loss of MEK activation could not be rescued by farnesyl pyrophosphate, but were restored to levels similar to those measured in untreated cells by geranylgeranyl pyrophosphate or mevalonate. Co-treating OPM-2 cells 48 h with 10 μM lovastatin and 50 μM U0126 led to >96% decrease in activated MEK in G0/G1 and >76% reduction in G2/M. Lovastatin treatment also caused a loss of prenylated RAS and RAS-family proteins as demonstrated by Western blot analyses. Additionally, Western blotting showed that U0126 treatment (5 μM, 30 min) alone or in combination with lovastatin (5 μM, 30 min) blocked IL-6 stimulated activation of MAP kinase in OPM-2 cells but had no effect on pAKT levels. Our results support that inhibition of RAS processing and down-stream signaling are the major mechanisms of action through which U0126 and lovastatin synergistically inhibit MM cell growth. Furthermore, these data demonstrate the potential therapeutic usefulness of these compounds in the treatment of multiple myeloma.