Fig. 6.
eIF4E is amplified and overexpressed in HMECres and confers resistance to BEZ235. (A) SNP array analysis of HMECres vs. parental HMECs indicating a region of amplification at chromosome 4 present in the BEZ235-resistant HMECres. (B) Genomic QPCR confirming eIF4E amplification in HMECres. (C) QPCR analysis comparing eIF4E mRNA expression levels normalized to ACTB in HMECres with parental HMEC. (D) Western blot assay confirming eIF4E overexpression in HMECres. (E) Western blot analysis of HMECs transduced with vector control or eIF4E. (F) Growth assay in the presence of 50 nM BEZ235 comparing HMECres with HMECs transduced with vector control or eIF4E. (G) Western analysis of HMECs transduced with shLUC (control) or two sheIF4E constructs (1426 and 1774). (H) Growth curves of the cell lines in G in the absence or presence of 50 nM BEZ235. (I) Western analysis of HMECs overexpressing MYC transduced with shLUC (control) or two sheIF4E constructs (1426 and 1774). (J) Growth curves of the cell lines in I in the absence or presence of 50 nM BEZ235. (Data are represented as mean ± SEM.)