Compound Shape Effects in Minor Groove Binding Affinity and Specificity for Mixed Sequence DNA

Abstract

AT specific heterocyclic cations that bind in the DNA duplex minor groove have had major successes as cell and nuclear stains and as therapeutic agents which can effectively enter human cells. Expanding the DNA sequence recognition capability of the minor groove compounds could also expand their therapeutic targets and have an impact in many areas, such as modulation of transcription factor biological activity. Success in the design of mixed sequence binding compounds has been achieved with N-methylbenzimidazole (N-MeBI) thiophenes which are preorganized to fit the shape of the DNA minor groove and H-bond to the −NH of G·C base pairs that project into the minor groove. Initial compounds bind strongly to a single G·C base pair in an AT context with a specificity ratio of 50 (KD AT-GC/KD AT) or less and this is somewhat low for biological use. We felt that modifications of compound shape could be used to probe local DNA microstructure in target mixed base pair sequences of DNA and potentially improve the compound binding selectivity. Modifications were made by increasing the size of the benzimidazole N-substituent, for example, by using N-isobutyl instead of N-Me, and by changing the molecular twist by introducing substitutions at specific positions on the aromatic core of the compounds. In both cases, we have been able to achieve a dramatic increase in binding specificity, including no detectible binding to pure AT sequences, without a significant loss in affinity to mixed base pair target sequences.