PP2 and dasatinib inhibit constitutive phosphorylation of LYN and induce apoptosis of primary MCL cells. (A) Constitutive phosphorylation profiles of LYN in MCL patients’ samples. Phospho-Tyr397 LYN was detected using a pan phospho-src family antibody. The blots were stripped and re-probed for total LYN. (B) Total proteins from HBL-2 cells were immunoprecipitated with an anti LYN antibody (Ig-LYN) or with an irrelevant IgG control and immunobloted (IB) with either an anti-phosphotyrosine antibody (P-Tyr) or an anti-LYN antibody. (C) Primary MCL cells (UPN3, UPN14) were treated with variable concentrations of PP2 (2 to 20 μM) or dasatinib (1 to 200nM) for 2 h. Phospho-Tyr397 LYN and LYN total were analyzed by western-blot. (D) Primary MCL cells were treated with various concentrations of PP2 (UPN3, top panel) or 10 μM of PP2 (UPN 1–3-7–8-9–14, middle panel) for 24 h and apoptosis was measured by flow cytometry after gating on CD19+ cells. All measurements were done in duplicate and the mean is provided. Results are also shown as median ± quartile (box) ± SE (bars) bottom panel). Differences between groups were determined using the paired Student t test. (E) Primary MCL cells were treated with dasatinib for 24 h with various concentrations (top panel) or with 100nM (middle panel). Apoptosis was measured as described above. Results are also shown as median ± quartile (box) ± SE (bars) bottom panel).