P-31 NMR-STUDIES ON THE INTERACTION OF MORPHOLINYL ANTHRACYCLINES ANDRELATED-COMPOUNDS WITH D(CGTACG)2 - THERMODYNAMIC AND KINETIC-PARAMETERS

P-31 NMR spectroscopy has been used to study the intercalation of the anthracyclines doxorubicin 1, daunorubicin 2, 4-demethoxydaunorubicin 3, morpholinodoxorubicin 4, methoxymorpholinodoxorubicin 5 and 9-deoxy daunorubicin 6 with the DNA fragment d(CGTACG)2. The individual phosph ate resonances of the oligonucleotide were assigned in the free as wel l as in the intercalated species. The P-31 chemical shift variations a llowed us to identify the intercalation sites, which resulted to be th e same for all compounds i.e. between the terminal CG base-pairs of th e helix (two molecules of drug per duplex). The binding constants, the dissociation rate constants and DELTAG# values have been determined i n different conditions of ionic strength and temperature. The kinetic constant (k(off)) of the slow step of the anthracycline/duplex interca lation process has been directly measured by NOE exchange techniques. Binding constants depend on the ionic strength and on the self-associa tion process so greatly, that their use to study by NMR anthracycline/ DNA interactions is questionable. On the contrary, the k(off) are not affected by these phenomena and present an interesting trend for 1-6, thus showing that the average life-time of the drug in the intercalati on site appears to be important for determining the cytotoxicity and t he antimitotic activity.