Nucleolytic cleavage of the mixed lineage leukemia breakpoint cluster region during apoptosis

VP-16 (etoposide) has recently been shown to induce topoisomerase II (TOP2) -mediated DNA cleavage within the mixed lineage leukemia (MLL) breakpoint c luster region (ber), suggesting a role of TOP2 in MLL gene rearrangement. I n our current studies, we have compared the induction of DNA cleavage withi n the MLL bcr in different cell lines after treatment with various anticanc er drugs. All anticancer drugs tested including VP-16 (a TOP2-directed drug ), camptothecin (a topoisomerase I-directed drug), 5-fluorouracil and metho trexate (antimetabolites), and vinblastine (a microtubule inhibitor) induce d the same site-specific cleavage within the MLL her. This cleavage was sho wn to nuclease-mediated but not TOP2-mediated by the following observations : 1) drug- induced cleavage within the MLL her was not protein-linked; 2) u nlike TOP2-mediated cleavage, drug-induced DNA cleavage with the MLL her wa s kinetically slow and coincided with the formation of the apoptotic nucleo somal DNA ladder; 3) drug-induced cleavage within the I ML ber was unaffect ed in cells with reduced nuclear TOP2; and 4) drug-induced cleavage within the MLL ber was abolished by the caspase inhibitor, Z-Asp(OCH3)-Glu(OCH3)-V al-Asp(OCH3)-FMK. The possibility that an apoptotic, nuclease may be involv ed in cleavage of the MLL her and MLL gene translocation is discussed.