CYTOTOXICITY OF ANTHRACYCLINES - CORRELATION WITH CELLULAR UPTAKE, INTRACELLULAR-DISTRIBUTION AND DNA-BINDING

In order to interact with topoisomerase II and induce genotoxicity, an thracyclines have to cross the outer cell membrane and the cytoplasm, enter the nucleus, and bind to the DNA. We incubated sensitive and res istant hematopoietic cells from cell lines and patient cells with daun orubicin, idarubicin, and its active derivative idarubicinol, extracte d the anthracyclines from whole cells and nuclei, and determined their concentration fluorimetrically. Additionally, the DNA binding of the drugs was evaluated in the same cells by determining fluorescence reso nance energy transfer between the anthracyclines and DNA-bound Hoechst dye 33342. We found a several thousand-fold accumulation of anthracyc lines in sensitive and resistant hematopoietic cells; 30-60% of the dr ugs are found in the nucleus, resulting in 200- to 300-fold difference s in concentration between the nucleus and outer fluids. A small propo rtion of the intracellular or intranuclear anthracyclines is bound to the DNA. The amount of DNA-bound anthracyclines correlates directly to cell death. It takes an additional 10 min for idarubicin and 30 min f or daunorubicin to satisfy DNA binding sites after the drugs have arri ved in the nucleus. The described methods provide the means to perform ex vivo studies on clinical material.