c-Abl regulates STAT3 activation and Mcl-1 transcription in CLL cells. (A) Western blot analysis of the effect of imatinib on pSTAT3 and Mcl-1 protein expression. CLL cells were cultured overnight and then incubated in the presence or absence of imatinib (20μM) for the indicated times. (B) Graphical representation of the data in panel A, showing percentage inhibition of pSTAT3 and Mcl-1 expression after imatinib treatment (mean ± SEM). Results are representative of 3 different CLL patient samples and are expressed as the percentage inhibition relative to the levels of pSTAT3 and Mcl-1 in untreated CLL cells incubated overnight. (C) The effect of imatinib on Mcl-1 gene expression. CLL cells from 3 different patients were incubated overnight before treating with imatinib (20μM) for the indicated time points. Mcl-1 mRNA levels were quantified using quantitative RT-PCR. Results are expressed as the percentage inhibition of Mcl-1 mRNA expression relative to the levels in untreated CLL cells incubated overnight (mean ± SEM). (D) Western blot analysis of STAT3 phosphorylation in CLL cells transfected with control (c) or c-Abl–specific (−Abl) siRNA. Cell viability (% viable) of transfected cells, as measured by trypan blue, and percentage knockdown (% KD) of pS⁷²⁷- and pY⁷⁰⁵-STAT3 are indicated; % KD of c-Abl is presented in Figure 1C.