in and these fibers appeared disorganized. In HUVEC and H28 MM cells, treatment with 0.5, 1 and 5 mM lovastatin for 24 hrs induced a dramatic up-regulation of both rhoA and cdc42 protein levels. Cyclin D1 is a regulator of cell cycle progression and is up-regulated by a wide variety of cellular signaling pathways including rhoA activation. The significant increase of rhoA protein levels did not result in up-regulation cyclinD1 protein levels but were reduced with lovastatin treatment of HUVEC and H28 cells. Furthermore, employing a colorimetric rhoA activation assay, we determined the effect of lovastatin on VEGF165 induced rhoA activation in HUVEC and H28 cells. Serum starved cell extract represent inactive levels of rhoA while 0.2M GTP loaded extract represents fully active rhoA. As expected VEGF stimulation induced rhoA activity to approximately 60 of the GTP loaded activity. Lovastatin inhibited VEGF165 induced rhoA activation in both HUVEC and H28 cells while co-administration of mevalonate and GGPP reversed the inhibitory 1381289-58-2 effects of lovastatin. These results demonstrate that lovastatininduced rhoA is inactive likely due to the lack of GGPP modification. Our previous studies have demonstrated that the combination of lovastatin and EGFR-TKI have resulted in synergistic cytotoxicity in a variety of human cancer derived cell lines. Other studies have demonstrated the utility of combining EGFRTKI with downstream inhibitors of the AKT pathway including rapamycin. Mammalian target of rapamycin plays a central role in regulating AKT driven translation initiation by regulating S6K1 and 4EBP1 activity. Rapamycin has limited clinical activity due to a feedback loop that activates AKT and acquired resistance suggesting that lovastatin may represent a novel therapeutic approach to target this pathway and enhance RTK-TKI activity. In this study, we evaluated the ability of rapamycin or lovastatin to augment the effects of the VEGFR-2 inhibitor KRN633. The H28 MM cell line had a relatively weak response to lovastatin-induced AKT inhibition. H28 cells express both VEGF and VEGFR-2. By Western blot analysis of activated AKT and its downstream targets S6K1 and 4EBP1, KRN633 and rapamycin treatments alone had minimal effects on the activation of these proteins. The combination of these agents showed enhanced inhibition of this pathway. In contrast, lovastatin treatment alone inhibited AKT, S6K1 and 4EPB1 phosphorylation and the combination of lovastatin and KRN633 induced a dramatic inhibition of the AKT pathway in this MM derived cell line. We 9-Azido-Neu5DAz further evaluated the combination of lovastatin and VEGFR-2 TKI on tumor cell cytotoxicity in HUVEC and MM cells. Utilizing MTT analysis and propidium iodide flow cytometry, we investigated the effects of combining two dif
