To confirm the effect of Baf on the de-acidification of endosomes, acridine orange staining was performed. AO is a fluorescent weak base that is frequently used as a probe for monitoring the acidification of organelles. In neutral or alkali environments this dye emits green fluorescence, but when exposed to acidic compartments, it is ionized and its emission undergoes a red shift. In the untreated cells, red LBH-589 citations fluorescence was observed inside discrete cytoplasm organelles, indicating that the AO had accumulated in acidic organelles. However, the red fluorescence dramatically decreased after 6 hours of Baf treatment, indicating that the Baf had increased the pH of the endosomes in the MiaPaca-2 cells. These results suggest that the release of dye from the liposomes is dependent on low pH conditions in intracellular organelles. GGTI release and efficacy to inhibit protein geranylgeranylation inside cancer cells were investigated. Fig 4A shows that the delivery of GGTI by the liposomes results in the inhibition of protein geranylgeranylation inside the cell. In this experiment, MiaPaCa-2 cells were treated with liposomal-GGTI for 3 hours, and proteins were collected from cell lysate for Western analysis. As shown, treatment of cells with either GGTI P61-A6 alone or GGTI-loaded liposome led to the appearance of unprenylated Rap1 band, indicating that liposomes deliver and release GGTI compound inside of cells to function and inhibited protein geranylgeranylation. Since the pH-liposomes exhibit significant destabilization below pH 6, which corresponds to pH of endolysosome interior, the pH-liposomes were likely to be destabilized or fused with endosomal membranes resulting in the release of the drug cargo into cytosol, inhibiting GGTase-I and blocking Rap1 prenylation. To examine the pH-sensitivity of GGTI release, the same experiment was repeated after treating the cells with AZD6738 pH-altering compound Bafilomycin A1. As shown in Fig 4A, treatment with Bafilomycin A1 abolished the effect of liposomal- GGTI on Rap1 prenylation. This is in line with the idea that increased pH of lysosomes altered by Bafilomycin A1 could not destabil