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Seven dicationic 2,5-diarylfurans have been synthesized, and their interactions with poly(dA-dT) and the duplex oligomer d(CGCCAATTCGCG)2 were evaluated by Tm measurements. The inhibition of topoisomerase II isolated from Giardia lamblia, the inhibition of growth of G. lamblia in cell culture by these furans, and the effectiveness of these compounds against Pneumocystis carinii in the immunosuppressed rat model have been assessed. Strong binding affinities to poly(dA-dT) and to the oligomer were observed for the dicationic furans, and the interaction strength is directly correlated to the biological activity of the compounds. An X-ray structure for the complex of the dicationic amidine derivative, 2,5-bis(4-guanylphenyl)furan (1), with the oligomer demonstrates the snug fit of these compounds with the AATT minor-groove binding site and hydrogen bonds to AT base pairs at the floor of the minor groove. The stronger DNA binding molecules are the most effective inhibitors of topoisomerase II and G. lamblia in cell culture, and there is a correlation for both DNA interaction and topoisomerase II inhibition with the biological activity of these compounds against G. lamblia. Compound 1 is the most effective against P. carinii, it is more active and less toxic than pentamidine on intravenous administration and it is also effective by oral dosage. The results presented here suggest a model for the biological action of these compounds in which the dication first binds in the minor groove of DNA and forms a complex that results in the inhibition of the microbial topoisomerase II enzyme.

Phenylamidine cationic groups linked by a furan ring (furamidine) and related compounds bind as monomers to AT sequences of DNA. An unsymmetric derivative (DB293) with one of the phenyl rings of furamidine replaced with a benzimidazole has been found by quantitative footprinting analyses to bind to GC-containing sites on DNA more strongly than to pure AT sequences. NMR structural analysis and surface plasmon resonance binding results clearly demonstrate that DB293 binds in the minor groove at specific GC-containing sequences of DNA in a highly cooperative manner as a stacked dimer. Neither the symmetric bisphenyl nor bisbenzimidazole analogs of DB293 bind significantly to the GC containing sequences. DB293 provides a paradigm for design of compounds for specific recognition of mixed DNA sequences and extends the boundaries for small molecule-DNA recognition. O rganic cations that bind in the DNA minor groove have biological activities that range from anti-opportunistic infection to anticancer properties (1-5). Such compounds have also provided a wealth of fundamental information about nucleic acid recognition properties, and they continue to be important models in the study of nucleic acid complexes (1, 2, 6-8). Netropsin ( Fig. 1) was the first minor-groove-binding compound crystallized with a B-form DNA, and the structure of the complex provided clear suggestions about the molecular basis for AT base pair sequence-specific recognition (9). The structure also lead to proposals by the Dickerson and Lown groups for minor-groove binding netropsin analogs, lexitropsins, that could specifically recognize GC base pairs and could, thus, have extended sequence recognition capability (10-12). Initial efforts in the design of analogs of netropsin that could recognize GC base pairs, however, yielded compounds of limited specificity. A breakthrough in this area occurred with the discovery that the monocationic relative of netropsin, distamycin ( Fig. 1), could bind into the minor groove of some AT sequences of DNA as a stacked, antiparallel dimer (7,13,14). Replacement of pyrrole groups in distamycin by imidazole provided lexitropsins with improved GC recognition specificity through dimer complexes, and current design efforts in the pyrrole-imidazole polyamide system have reached a high level of success (15-18). With recent developments by the Dervan group, AT and TA as well as GC and CG base pairs can now be effectively distinguished in DNA sequences by polyamides (15-18).Interestingly, the polyamide system is the only one of the well known minor-groove binding motifs that has yielded conclusive evidence for formation of the stacked-dimer recognition unit, although recent evidence indicates that some monocationic cyanine dyes can form an array of stacked dimers in the DNA minor groove (19). No dications have been found to form the stacked-dimer recognition motif. Netropsin, the first minorgroove-binding agent to be characterized in detail and a dicationic relative of the monocation distamycin (Fig. 1), for example, does n...

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