Figure 4. The combination effect of AUY922 and rapamycin on KIT protein degradation in GIST430 and GIST48 cells.

GIST430 (A) and GIST48 (B) cells were treated with the indicated dose of AUY922 alone or in a combination with 40 μM rapamycin for 8 h. Cells were lysed and analyzed by immunoblotting against KIT and MAP1LC3. Relative KIT expression was normalized to ACTIN and compared with that of the untreated control. Cells were treated with 20 nM AUY922 and 5 μM rapamycin, either alone or in combination, for 12 to 48 h and analyzed by Annexin V staining (C). Cells were treated with various concentrations of AUY922 combined with rapamycin (D). Cell growth was determined by the methylene blue dye assay as described in the Materials and Methods section. The interaction between AUY922 and rapamycin at the IC₅₀ point was analyzed using the isobologram method. The data are expressed as the mean ± S.E. of two or more independent experiments.