Article Figures & Data
Figures
- Figure 1.
The effect of dasatinib on MPM cell downstream pathways and cytotoxicity. A, MPM cells were treated with 100 nmol/L dasatinib for the indicated times, lysed, and analyzed by Western blotting with the indicated antibodies. B, MPM cells were treated with dasatinib at the indicated concentrations for 72 h. Cell number was estimated using an MTT assay. Cells treated with vehicle alone were defined as 100%.
- Figure 2.
The effects of dasatinib on cell cycle and apoptosis. A, MPM cells (MSTO-211H, NCI-H28, NCI-H2052, NCI-H2452) were treated with 100 nmol/L dasatinib for 16 h before being stained with propidium iodide and analyzed with FACS to determine the proportion of cells in each phase of the cell cycle. B, MPM cells (MSTO-211H, NCI-H28, NCI-H2052, NCI-H2452) were treated with 100 nmol/L dasatinib for 6 h and stained with propidium iodide and Annexin V to estimate the number of necrotic cells (propidium iodide positive) and those that were undergoing early apoptosis (Annexin V positive). SFK inhibition in sensitive cell lines resulted in apoptosis and cell cycle arrest.
- Figure 3.
Effect of dasatinib on cell migration and invasion. A, MPM cells were plated to confluence on tissue culture plastic. A single scratch was made in the confluent monolayer. The scratch was monitored and photographed every 6 h for 24 h (representative data). B, cells that migrated into the scratch after 6 h were counted. The data represent two independent experiments. In each experiment, 20 fields were counted. Bars, SD. In all cases, when treated cells are compared with corresponding controls, P < 0.05. C, cells were plated onto Matrigel-coated filters in a modified Boyden chamber. Cells were allowed to attach and spread for 16 h before the addition of 25 nmol/L (MSTO-211H) or 100 nmol/L (all others) dasatinib to the upper chamber and conditioned medium to the lower chamber. After 24 h, cells still in the upper chamber were removed, and cells that had invaded through the Matrigel were stained and counted. In parallel, control cells treated identically were assessed for viability and cell number with trypan blue. The average number of cells per field is expressed as a percentage of the control after normalizing for cell number. In MSTO-211H, NCI-2052, and NCI-H2452 cells, dasatinib inhibited invasion compared with corresponding controls, P < 0.05.
- Figure 4.
Representative examples of immunohistochemical analysis of MPM cell lines and tissue specimens. A, c-Src expression in MPM cell line H28 and one tumor specimen with positive membrane and cytoplasmic staining. As a control, blocking peptide shows the absence of immunostaining in both specimens. B, examples of immunostaining for the three antibodies used on MPM tissue specimens showing positive membrane and cytoplasmic staining in tumor cells.
Tables
- Table 1.
Characteristics of four MPM cell lines and IC50 values to dasatinib therapy
Cell line IC50 (nmol/L) Histology Patient characteristics MSTO-211H 25 Biphasic 62-y-old male Caucasian NCI-H28 60 Mesothelioma 48-y-old male Caucasian smoker NCI-H2052 80 Mesothelioma 65-y-old male Caucasian smoker NCI-H2452 980 Epithelioid Male nonsmoker (age unknown) - Table 2.
Clinical and demographic patient data
Characteristic n Patients Percent of total (%) Sex Female 8 17 Male 38 83 Ethnicity African-American 1 2 Hispanic 1 2 Caucasian 44 96 Prior cancer None 39 85 Aerodigestive tract 0 0 Other (i.e., skin) 7 15 Family history of MPM No 44 96 Yes 2 4 Asbestos exposure No 14 31 Yes 31 67 Unknown 1 2 Smoking status Current 6 13 Former 24 52 Never 16 35 Surgery Extrapleural pneumonectomy 42 91 Pleurectomy 4 9 Side of resected tumor Right 26 57 Left 20 43 Pathologic stage II 1 2 III 29 63 IV 16 35 Pathologic T stage T2 10 22 T3 25 54 T4 11 24 Pathologic N stage N0 20 43 N1 6 13 N2 20 43 Pathologic M stage M0 39 85 M1 7 15 - Table 3.
P values for the associations between Src expression/activation and disease stage
Stage Src membrane Src cytoplasm Src Y 419 membrane Src Y 419 cytoplasm Src Y 530 membrane Src Y 530 cytoplasm Pathologic stage 0.93 0.22 0.05 0.19 0.36 0.18 T stage 0.6 0.23 0.57 0.09 0.59 0.43 N stage 0.45 0.96 0.74 0.09 0.02 0.07 M stage 0.62 0.99 0.03 0.04 0.79 0.28
Volume 6, Issue 7
