Figure 5. Grb7 silencing reduces cell viability and increases lapatinib efficacy.
A-E, SKBR3 or BT474 cells were transfected with Grb7 siRNA or non-targeting control siRNA (CNTR siRNA). A, 2×105 SKBR3 cells/well were seeded in 6-well plates, allowed to adhere for 48 h and subsequently used for protein lysates preparation. Grb7, phospho-Akt, total Akt and γ-tubulin levels were determined by immunoblotting. B, BT474 cells were transfected with Grb7 siRNA or CNTR siRNA and imaged by light microscopy five days later. C, D, 5×103 siRNA-transfected or untransfected SKBR3/well were seeded in 96-well plates, allowed to adhere for 24 h and subsequently treated with or without lapatinib at the indicated concentrations. Five days later, viability was determined and compared to that of untransfected SKBR3 (C); lapatinib-specific death was determined by normalizing to baseline cell death of CNTR siRNAs- or Grb7 siRNAs-transfected SKBR3 (D). E, SKBR3 cells were transfected with Grb7 siRNA or CNTR siRNA. mRNA was extracted 3 days after transfection and gene expression was detected by LDAs. A, B, One representative experiment out of three is shown. C, D, Results are means ± SD of four independent experiments. E, Results were calculated as means of two separate experiments. *: p<0.05.