Bax : Bcl-2 ratio modulation by bryostatin 1 and novel antitubulin agents is important for susceptibility to drug induced apoptosis in the human early pre-B acute lymphoblastic leukemia cell line, Reh

The ratio of Bar to Bcl-2 protein can determine whether cells will die via apoptosis or be protected from it. Reh was found to express a high basal le vel of Bcl-2 but was lacking of Bar protein expression. Treatment with bryo statin 1 induced a down-regulation in Bcl-2 protein that was not accompanie d by an obvious Bar protein induction or apoptosis. These results suggest t hat a decreased level of Bcl-2 alone in this cell line is not sufficient fo r apoptosis induction. In an effort to identify the mechanism whereby apopt osis could be induced in this ALL model, we treated Reh cells with three mi crotubule inhibitors: dolastatin 10, auristatin PE and vincristine, in the presence and absence of bryostatin 1. When used alone, only dolastatin 10 i nduced apoptosis that was detected morphologically, and by flow cytometry. Western blots revealed that dolastatin 10-induced apoptosis was accompanied by the induction of Bar protein and the reduction in Bcl-2 protein. Aurist atin PE and vincristine induced both Bar and Bcl-2 protein, leaving the Bax :Bcl-2 ratio constant. Reh cells pretreated for 24 h with bryostatin 1 foll owed by dolastatin 10, auristatin PE or vincristine showed significant apop tosis which was accompanied by Bcl-2 protein down regulation and Bar protei n up regulation. We conclude that: (1) expression of bar is necessary for a poptosis-induction in this model; (2) a decrease in Bcl-2 level alone is no t sufficient and might not be necessary for apoptosis-induction; and (3) th e ratio of Bax:Bcl-2 plays a critical role in susceptibility to apoptosis i n Reh cells. The results from this study should prove useful in guiding the clinical application of these novel agents in the treatment of acute lymph oblastic leukemia. (C) 1999 Elsevier Science Ltd. All rights reserved.