Effects of OSI-027 and rapamycin on phosphorylation of mTORC1 and mTORC2 substrates in malignant human lymphoid lines. (A,D) The MCL line Jeko (left) and T-cell ALL line Jurkat (right) were treated for 8 hours with diluent (lanes 1,8); OSI-127 at 1.25, 2.5, 5, 10 or 20μM (lanes 2-6 and 9-13, respectively); or 10nM rapamycin (lanes 7,14). Whole cell lysates²⁸  were then subjected to SDS-PAGE followed by immunoblotting with antibody that recognizes the indicated antigen. (B,E) Jeko cells were treated for 24 hours with diluent (lanes 1,5); OSI-127 at 0.5, 2, or 5μM (lanes 2-4); or rapamycin at 1, 10 and 100nM (lanes 6-8). Cell lysates²⁶  were then subjected to SDS-PAGE followed by immunoblotting with antibody that recognizes the indicated antigen. (C,F) Samples from 3 MCL patients were treated for 24 hours with diluent or OSI-127 at 2 and 5μM. After SDS-PAGE, cell lysates were probed as indicated. (G) After Jeko cells were treated with the indicated OSI-027 concentration for 24 hours, mTOR immunoprecipitates (top) or cell lysates (bottom) were probed as indicated.