Molecular and cellular pharmacology of novel photoactive psoralen and coumarin conjugates of pyrrole- and imidazole-containing analogues of netropsin.

The molecular and cellular pharmacology of novel sequence-directed photoactive agents, in which either psoralen or coumarin is conjugated to minor groove-binding AT-selective pyrrole-, or more GC-selective imidazole-containing analogues of netropsin, is described. The compounds were relatively non-toxic in the dark and showed marked photoinduced cytotoxicities when irradiated at 366 nm UV. The psoralen-containing pyrrole (1) and imidazole (2) compounds gave the largest photoinduced effect, were more active than 8-methoxypsoralen (8-MP) itself by 333- and 22-fold, respectively, and were more potent than the corresponding coumarin-containing analogues 3 and 4. Following irradiation, 1 and 2 were > 300- and > 10-fold more efficient at producing interstrand cross-links in naked DNA, respectively, than 8-MP. 1 was at least 10-fold more efficient at producing cross-links in cells than 2, reflecting the difference in their IC50 values. No cross-links were observed with the coumarin analogues, but these compounds were more potent than 8-MP. AT and GC sequence recognition was confirmed by DNA footprinting, and sites of covalent modification mapped by a polymerase stop assay. All compounds produced blocks at thymine base sites following irradiation. 1 was more efficient than 8-MP and produced a different pattern of covalent modification, whereas 2 was more selective than either 1 or 8-MP. A 1H-NMR study on a 1:1 complex of 2 with the hexamer (5'-dA1T2G3C4A5T6-3')2 indicated that the imidazole carboxamide moieties of 2 reside in the minor groove of the sequence 5'-GCAT-3' of the hexamer with the C-terminus located on the 3'-TA site, and the psoralen group intercalated between the 5'-A1T2-3' base pairs.