Effects of rapamycin and p27^kip1.

(A) Rapamycin and forced expression of p27^kip1 by VP22/p27^kip1 fusion protein prevent IL-7–mediated increase of T-ALL cells in the S+G2M phase and DNA synthesis. T-ALL cells were cultured with IL-7 either alone or in the presence of rapamycin, VP22/p27kip1 fusion protein, VP22 control protein, or the indicated combinations. Cultures were examined for percentage of cells at the S+G2M phases of the cell cycle and for DNA synthesis by thymidine incorporation as described in Materials and methods. Representative results from 1 among 3 patients studied are shown. (B) VP22/p27^kip1 prevents IL-7–mediated activation of cdk2. T-ALL cells were cultured for 72 hours under the indicated conditions, and activation of cdk2 was determined in cell lysates by in vitro kinase reactions by using Histone H1 as exogenous substrate. Reactions were analyzed by SDS-PAGE, transferred to PVDF membrane, and exposed to x-ray film. (C) Rapamycin, VP22/p27^kip1, and their combination prevent IL-7–mediated phosphorylation of Rb. T-ALL cells were cultured for 72 hours under the indicated conditions, analyzed by 6% SDS-PAGE, transferred on nitrocellulose membrane, and immunoblotted with mAb specific for Rb.