Effects of EGFR and IGF1R inhibitors on crosstalk between EGFR and IGF1R signaling pathways at multiple levels.

<p><b>A</b>, Immunoprecipitation detecting EGFR/IGF1R interaction. DU145 and PC3 cells were irradiated at the dose of 2 Gy. Then cellular proteins were extracted, and subjected to immunoprecipitation assay detecting EGFR and IGF1R interaction. <b>B</b>, IGF-I activates EGFR in DU145 and PC3 cells. Cells were serum-starved overnight, and then subjected to IGF-I stimulation for 30 min. EGFR phosphorylation was detected by Western blot. <b>C</b>, EGF activates IRS1, the key factor of IGF1R signal pathway in DU145 and PC3 cells. Cells were serum-starved overnight, and then subjected to EGF stimulation for 30 min. IRS1 phosphorylation was detected by Western blot. <b>D</b>, MAPK or AKT activation after EGFR and IGF1R simulation of inhibition. DU145 and PC3 cells were treated with or without AG1024 (10 µM), Erlotinib (10 µM), AG1024 plus Erlotinib; IGF (50 ng/mL), EGF (50 ng/mL), or EGF plus IGF for 30 min. Then ERK1/2 expression and phosphrylation, AKT expression and phosphrylation, IRS1 expression and phosphrylation was determined by Western blotting. β-tubulin was used as a loading control.