Syk activity, sensitivity to R406-induced apoptosis, and ZAP-70 expression in TCL1 leukemias treated with R788. (A) Leukemic cells were isolated from the spleens of Eμ-TCL1 transgenic mice 1-2 months after treatment with R788 had been discontinued. TCL1 leukemias that were used in the adoptive-transfer experiments (TCL1-002 and TCL1-551) and normal B-cell samples (NB1 and NB2) were loaded as controls. Syk activity was evaluated by immunoblotting analysis with phospho-Syk and phospho-BLNK antibodies. (B) Percentage of viable leukemic cells (annexin V/PI negative) after 48 hours in culture with R406. (C) Immunoblotting analysis of ZAP-70 expression in TCL1 leukemias. Leukemia cells were purified by negative selection, the percentage of remaining T cells after purification was less than 3%. Negative controls included 2 normal B-cell samples purified by positive selection from spleens of wild-type mice (NB4 and NB5) and 1 ZAP-70–negative CLL sample (G57). Positive controls were the mouse T-cell line B3Z and 1 ZAP-70–positive CLL sample (G90). The CLL samples were purified by negative selection, as described elsewhere.15 Actin served as a loading control.
