Figure 3.

Potency and mechanism of liposomal staurosporine in vitro. (A) In vitro MTT cell viability assay results for free and liposomal staurosporine versus empty liposomes (control) were determined with U87 glioblastoma cells. First, 2,000 cells were plated and after 18 hours free or liposomal staurosporine was added. The concentration of free or liposomal staurosporine added to cultures ranged between 200 nm and 0.1 nm, with two-fold decrements down the range. Free and liposomal staurosporine had very similar potency while empty liposomes caused essentially no tumor cell killing. (B) Western blot analysis for Akt, P-Akt, and cleaved poly(ADP-ribose) polymerase. U87 cells were incubated with 200 nM, 10 nM, and 0.1 nM of staurosporine encapsulated in liposomes and staurosporine for 32 hours. (C) Bar graph showing caspase-3/7 activity as fold change from control (which is 1). Free and liposomal staurosporine yielded similar changes in caspase activity. (D) Fluorescence-activated cell sorting analysis for U87 glioblastoma cells incubated with either empty liposomes, 200 nM of staurosporine encapsulated in liposomes, dimethylsulfoxide, or free staurosporine. Representative plots of two repeats per condition are shown, and reveal that both free and liposomal staurosporine induced cell death, indicated by a G₀/G₁ shift, compared with the dimethylsulfoxide and empty liposome control treatments.

Abbreviations: conc, concentration; MTT, (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; Lipo, liposomal; PARP, poly(ADP-ribose) polymerase; GADPH, Glyceraldehyde 3-phosphate dehydrogenase; Akt, protein kinase B; DMSO, Dimethyl sulfoxide; Veh, DMSO vehicle control.