Given the great strides that have taken place over the past few decades in our understanding of nucleic acid’s role in cellular processes, it has become abundantly clear that DNA and RNA can provide a great tool and target for drug development. The human genome project has provided a major impetus in identifying human genes implicated in diseases and has opened the door to new possibilities with DNA-based therapeutics. Further developments in transcriptomics and proteomics will provide additional momentum for the advancement of therapeutics by supplying novel targets for drug design, screening, and selection. As new discoveries are made and our knowledge of nucleic acid’s role in life processes is expanding, the area of chemistry focused on learning how to target and exploit these nucleic acids for control of their relative processes is also expanding. New strategies to develop molecules that can both identify DNA or RNA targets and modulate their activity are of great interest to medicinal chemistry. The goal of this research was to delineate an efficient approach to targeting nucleic acids that yields cell permeable, biologically stable molecules that can be exploited in in vivo applications.

Herein describes our approach which utilizes cyclic peptide phage display for the evolution of novel cyclic peptide scaffolds that target a given oligonucleotide. Evolved scaffolds are then tested in vitro as discrete entities to assess their binding capabilities. Given that the phage display scaffolds employ a disulfide linker for cyclization, alternative redox stable macrocylic linkers were developed and synthesized. Generated analogues were subsequently assessed for the retention of the desired binding activity.

The details of this pragmatic approach were developed using the bTAR RNA oligo as a model system. Results indicated that not only could we evolve bTAR binders from a pool of 1.2 billion possible scaffolds in a relatively short time, but that these scaffolds bound with affinities in the low micromolar range when tested as discrete entities. We were also successful in developing an alternative dicarbon macrocyclic linker to yield redox stable analogues. Subsequent testing of the analogue scaffolds indicated the retention of the desired binding properties.