Low doses of NVP-BEZ235 are sufficient to inhibit proliferation of PEL cells in vitro, by inhibiting the downstream targets of PI3K/mTOR pathway. (A) Inhibition of proliferation of the indicated PEL cell lines on treatment with NVP-BEZ235 as measured by the MTS assay. Shown in each panel is the absorbance at 490 nm (y-axis) in the absence of drug (gray diamond) or increasing, indicated doses of NVP-BEZ235. Treatment time is represented on the x-axis. Each data point is the average of triplicate or quadruplicate measurements. Error bars represent the SD and, in most cases, are smaller than the symbol. (B) The IC₅₀ curve for NVP-BEZ235 showing an IC₅₀ value of 5.68 ± 1.76nM for BC-1 PEL cells. (C) Immunoblot analysis of extracts harvested from PEL cell line treated with DMSO (vehicle) or increasing doses of NVP-BEZ235. Membranes were probed with antibodies raised specifically against the phosphorylated forms of Akt (Ser473), mTOR, S6K, pS6, FOXO1, and GSK3β. Membranes were also probed with anti-actin antibody, as a loading control.