6th ESACP Congress, Heidelberg, April 7-11, 1999

A027
CGP57148B AN INHIBITOR OF THE BCR-ABL TYROSINE KINASE COMMITS RESISTANT HL60 CELLS TO UNDERGO APOPTOSIS THROUGH BCL-XL DOWN-REGULATION
Belhoussine R 1, Morjani H 1, Palissot T 1, Belloc F 2, Dufer J 1, Manfait M 1

1 Unité Médian, UFR de Pharmacie, Université de Reims, Reims Cedex, France
2 Laboratoire d'Hématologie, Hôpital Haut Lêvèque, Pessac, France

Apoptosis is a genetically regulated cell death process which results in a variety of morphological changes like chromatin condensation and DNA fragmentation. We have examined cytosolic pH, mitochondrial membrane potential and reactive oxygen species (ROS) during daunorubicin (DNR) induced apoptosis in human leukemic HL60 cells. We showed that in sensitive cells apoptosis was accompanied by significant acidification of the cytosol and an increase of ROS level without mitochondrial membrane depolarization. The response to an apoptotic stimulus is determined and regulated in part by mitochondrial oncoproteins which include proteins of the bcl-2 family (bcl-2, bax, bcl-xL). Thus, we investigated the effect of these proteins on the induction of this phenomenon in the multidrug-resistant HL60 cells established with vincristine (HL60/VCR), DNR (HL60/DNR), C2-ceramide (HL60/C2), lonidamine (HL60/L2) and VP16 (HL60/V2) respectively. We showed that sensitive and HL60/VCR cells were able to undergo apoptosis when treated with DNR, while the other resistant cell lines do not. Western blotting analysis shows that bax is expressed at the same level in all cell lines whereas bcl-2 is overexpressed only in HL60/VCR. However, the other resistant cell lines displayed a higher expression of bclxL at the expense of the bcl-2 expression. Interestingly, bcr-abl, a dysregulated tyrosine kinase, was detected in cell lines overexpressing bcl-xL. Treatment with 1ÁM of the tyrosine kinase inhibitor CGP57148B commit these resistant cells to undergo apoptosis. Inhibition of the bcr-abl was accompanied by a decrease of bcl-xL expression. These data suggest that in addition of the multidrug-resistance phenotype, bcr-abl translocation and bclxL overexpression could also account for the development of resistance to cell death induced by anthracyclines in leukemic cells and that the CGP57148B could be useful for reversal of this phenotype.