Lapatinib (Tykerb)

T47Dcells were pretreated with 100ng/ml EGF for 20 min and then treated with the indicated concentrations of Lapatinib for 24 hours.

Impact of the TKI erlotinib, lapatinib, dasatinib, and sorafenib on the viability of MDS/AML cells. MOLM-13 (A) and HL-60 (B) cells were incubated with the indicated doses (given in mM below the x-axis) of the 4 TKI, and cellular viability was assessed by MTT assay after 24, 48 and 72 h of incubation. Changes in viability are given as percentage of cells as compared to non-treated control samples. This experiment was repeated at least three times, yielding comparable results. Graphs show representative results of one experiment carried out in duplicates (mean  standard deviation).

Capacity of the TKI to overcome the AML-typical differentiation blockage. The myeloid cell lines MOLM-13 and HL-60 were incubated for 6 days with 0.01% DMSO (serving as a negative solvent control), 1 mM of ATRA (serving as a positive control), as well as with the indicated doses of the four TKI. (A) Representative May–Gruenwald– Giemsa staining of MOLM-13 cells, (B) quantitation of the percentage of MOLM-13 cells exhibiting at least two morphological signs of differentiation (that is a decrease in cytoplasmic basophilia, a reduction of the nucleo-cytoplasmic ratio, appearance of nuclear lobulation and/or cytoplasmic granules). Percentages were evaluated by examining at least 100 cells/condition; (C) representative FACS overlays of MOLM-13 cells depicting TKI-induced CD11b expression (black line) as compared to the isotype (shaded grey); (D) quantitation of TKI-induced CD11b-expression in MOLM-13 cells; (E) representative slides depicting morphology/staining of MOLM-13 cells assessed in the NBT-reduction assay; (F) respective quantitative assessment demonstrating the NBT-reducing capacity under the different drugs; (G) representative May–Gruenwald– Giemsa staining of HL-60 cells.

Inhibition of signaling pathway activation in lung tumor cell lines by kinase inhibitors. Lung tumor cells were cultured in 10% FBS until reaching ∼80% confluence and then the cells were starved in serum-free medium for overnight, followed by 4-hour treatment with the inhibitors. Cell lysates were then prepared and used for determination of the pathway activation signals by the CEER assay.

(B–C) LNCaP (B) and LNCaP-AI (C) cells were transiently transfected with sPLA2-IIa(-800)-Luc (0.5 lg). The cells were then treated with Erlotinib (20 lM), Gefitinib (20 lM), Lapatinib (20 lM), CI-1033 (8 lM), LY294002 (20 lM) and Bortezomib (20 lM) without or with EGF (100 ng/ml) for 24 h. Luciferase assay was performed according to a standard protocol with Renilla luciferase as an internal control. Data are presented as the mean (±SD) of duplicate values of a representative experiment that was independently repeated for five times.

LNCaP-AI cells were starved in 1% stripped medium for 24 h. The cells were then treated with Erlotinib (20 lM), Gefitinib (20 lM), Lapatinib (20 lM), CI-1033 (8 lM), LY294002 (20 lM) and Bortezomib (20 lM) for 24 h. Cell culture medium was collected from each sample and subjected to ELISA for sPLA2-IIa. The condition medium samples were diluted 10 times for ELISA. Average of duplicate samples was converted to nanogram per milliliter against standard curve. The data represent one of five repeated experiments.