POTENTIATION OF CHLORAMBUCIL CYTOTOXICITY IN B-CELL CHRONIC LYMPHOCYTIC-LEUKEMIA BY INHIBITION OF DNA-DEPENDENT PROTEIN-KINASE ACTIVITY USING WORTMANNIN

In this study, we examined the ability of wortmannin to modulate chlor ambucil (CLB) cytotoxicity in lymphocyte samples from patients with B- cell chronic lymphocytic leukemia (B-CLL). It has been suggested previ ously that enhanced cross-link repair is a primary mechanism of resist ance to nitrogen mustards (NMs) in B-CLL. DNA-dependent protein kinase (DNA-PK) is involved in the repair of double-strand breaks and in rej oining steps in recombination mechanisms. Mutants defective in this pr ocess are hypersensitive to alkylating agents. We have recently demons trated that the activity of DNA-PK is a determinant in the cellular re sponse of B-CLL to CLB. The DNA-PK gene has homology to the P110 phosp hatidylinositol 3-kinase (PI 3-K). Wortmannin, an inhibitor of P110 PI 3-K, also inhibits DNA-PK activity in vino. We investigated the effec t of wortmannin on DNA-PK activity and CLB toxicity in the lymphocytes from 11 patients with B-CLL. Our results demonstrate that DNA-PK acti vity is decreased after exposure to wortmannin in a dose-dependent man ner. Wortmannin, at nontoxic concentrations, synergistically sensitize d B-CLL lymphocytes to the effects of CLB. Moreover, we observed a sig nificant correlation when me compared the fold decrease in DNA-PK acti vity and the synergistic value (I), obtained when wortmannin was used at 0.1 mu M in the resistant B-CLL lymphocyte samples, there was a hig hly significant correlation between the ability of wortmannin at 0.1 a nd 0.25 mu M to decrease the level of DNA-PK activity and to increase CLB sensitivity. In a model of primary human tumor cells, our findings suggest that the inhibition of DNA-PK activity may be a powerful way to overcome resistance to NMs such as CLB and point to new possibiliti es to improve the effectiveness of NM therapy.