Vol. 296, Issue 1, 31-40, January 2001

Pyrrolo-1,5-benzoxazepines Induce Apoptosis in Chronic Myelogenous Leukemia (CML) Cells by Bypassing the Apoptotic Suppressor Bcr-Abl

Margaret M. Mc Gee, Giuseppe Campiani, Anna Ramunno, Caterina Fattorusso, Vito Nacci, Mark Lawler, D. Clive Williams and Daniela M. Zisterer

Department of Biochemistry, Trinity College, Dublin, Ireland (M.M.M.G., D.C.W., D.M.Z.); Dipartimento Farmaco Chimico Tecnologico, Universita' degli Studi di Siena, Siena, Italy (G.C., V.N.); Dipartimento di Scienze Farmaceutiche, Universita' degli Studi di Salerno, Fisciano, Italy (A.R., C.F.); and Department of Haematology, Sir Patrick Dun Research Labs, St. James's Hospital and Trinity College, Dublin, Ireland (M.L.)

Expression of the transforming oncogene bcr-abl in chronic myelogenous leukemia (CML) cells is reported to confer resistance against apoptosis induced by many chemotherapeutic agents such as etoposide, ara-C, and staurosporine. In the present study some members of a series of novel pyrrolo-1,5-benzoxazepines potently induce apoptosis, as shown by cell shrinkage, chromatin condensation, DNA fragmentation, and poly(ADP-ribose) polymerase (PARP) cleavage, in three CML cell lines, K562, KYO.1, and LAMA 84. Induction of apoptosis by a representative member of this series, PBOX-6, was not accompanied by either the down-regulation of Bcr-Abl or by the attenuation of its protein tyrosine kinase activity up to 24 h after treatment, when approximately 50% of the cells had undergone apoptosis. These results suggest that down-regulation of Bcr-Abl is not part of the upstream apoptotic death program activated by PBOX-6. By characterizing the mechanism in which this novel agent executes apoptosis, this study has revealed that PBOX-6 caused activation of caspase 3-like proteases in only two of the three CML cell lines. In addition, inhibition of caspase 3-like protease activity using the inhibitor z-DEVD-fmk blocked caspase 3-like protease activity but did not prevent the induction of apoptosis, suggesting that caspase 3-like proteases are not essential in the mechanism by which PBOX-6 induces apoptosis in CML cells. In conclusion, this study demonstrates that PBOX-6 can bypass Bcr-Abl-mediated suppression of apoptosis, suggesting an important potential use of these compounds in the treatment of CML.