Fludarabine induces apoptosis in chronic lymphocytic leukemia--the role of P53, Bcl-2, Bax, Mcl-1, and Bag-1 proteins.

The expression of P53, Bcl-2, Bax, Bag-1, and Mcl-1 proteins in CD5/CD20-positive B-chronic lymphocytic leukemia (B-CLL) cells from 30 typical CLL patients was evaluated before and after 48 h of incubation with 10(-6) M fludarabine using multiparametric flow cytometric analysis. Protein expression was correlated with annexin V expression, Rai modified clinical staging, lymphocyte doubling time, and previous treatment. Our main goal was to determine the predictive value of these proteins in CLL cells in terms of disease evolution. Bcl-2 expression decreased from a median fluorescence index (MFI) of 331.71 +/- 42.2 to 245.81 +/- 52.2 (P < 0.001) after fludarabine treatment, but there was no difference between viable cells (331.57 +/- 44.6 MFI) and apoptotic cells (331.71 +/- 42.2 MFI) before incubation (P = 0.859). Bax expression was higher in viable cells (156.24 +/- 32.2 MFI) than in apoptotic cells (133.56 +/- 35.7 MFI) before incubation, probably reflecting defective apoptosis in CLL (P = 0.001). Mcl-1 expression was increased in fludarabine-resistant cells and seemed to be a remarkable protein for the inhibition of the apoptotic process in CLL (from 233.59 +/- 29.8 to 252.04 +/- 35.5; P = 0.033). After fludarabine treatment, Bag-1 expression was increased in fludarabine-resistant cells (from 425.55 +/- 39.3 to 447.49 +/- 34.5 MFI, P = 0.012), and interestingly, this higher expression occurred in patients who had a short lymphocyte doubling time (P = 0.022). Therefore, we could assume that Bag-1 expression in such situation might identify CLL patients who will need treatment earlier.