A, HCC827 and H1975 cells were transfected with Shp2 siRNAs (siR), non-silencing control siRNA (NS), or mock-treated (P). Cell lysates were analyzed for Shp2 knockdown and active Erk1/2 by immunoblotting. B, Cells were plated in 96-well plates followed by transfection with Shp2 siRNAs or control siRNA as above. The relative number of viable cells was measured 6 days post transfection and normalized to untransfected cells. Data represent two (HCC827) and four (H1975) independent experiments performed in quintuple. *, p <0.05. C, HCC827 containing Dox-inducible Shp2 shRNAs [6] were treated with Dox or left untreated, cell lysates were analyzed with indicated antibodies. D, HCC827 cells were transfected with HA-Erk2 plus control vector (−), Gab1FF, or Shp2CSDA, and treated with EGF or left untreated. HA-Erk2 activation were analyzed by immunoblotting of pErk2 after immunoprecipitation of the transfected HA-Erk2. E, HCC827 and H1975 cells were treated with indicated concentrations of SPI-112Me in 96-well plates as described in Materials and Methods and viable cells were measured on Day 6. Data were from three independent experiments performed in triplicates (n = 9) (graph). Right panel, cells were treated with SPI-112Me overnight and cell lysates were analyzed by immunoblots with indicated antibodies.