METHOD FOR QUANTIFYING EXPRESSION OF FUNCTIONALLY ACTIVE TOPOISOMERASE-II IN PATIENTS WITH LEUKEMIA

Aims-To produce a method to measure and quantify enzymatically active topoisomerase II in normal and neoplastic human cells. Methods-A crude cell lysate from density separated mononuclear cells from either peri pherial blood or bone marrow was prepared as a source of topoisomerase s. Using the lysate, minicircles from the Crithedia kinetoplast DNA co mplex were decatenated before being separated by agarose gel electroph oresis and visualised using ethidium bromide/ultraviolet fluorescence. Results-Cell number, sample volume and drug inhibition concentration required to produce reliable and reproducible assay conditions were es tablished. Intra- and interassay standards were included which permitt ed the quantification of active topoisomerase II in matched peripheral blood, bone marrow, presentation, and relapse samples from patients w ith acute lymphoblastic leukaemia. Active topoisomerase II has been co nverted to a unit scale which has been used to compare topoisomerase I I activities in cells from patients with normal blood and bone marrow samples. Conclusions-There was no change in topoisomerase II activitie s between samples taken at presentation and those taken during a recur rence. However, topoisomerase II activity in leukaemic blast populatio ns was increased compared with topoisomerase II active in normal cells .