Synthesis and Biomedical Applications of Gamma Peptide Nucleic Acid

Nucleic acid is a promising material for biomedical applications due to its programmability and predictability of sequence binding. However, native nucleic acid itself could not be widely exploited in in vivo applications because of enzymatic degradation, weak and off-targeting binding and cellular delivery. Such issues could be potentially addressed by a novel synthetic nucleic acid analogue called Gamma Peptide Nucleic Acid (γPNA), in which backbone is replaced with a gamma modified N-(2- aminoethyl)-glycine. Nonetheless, the biomedical applications of γPNA will impede until the synthesis and binding properties are better investigated. The presented work in this Dissertation first offers a robust synthetic strategy for preparing optically pure γPNA monomers. The strategy utilizes a bulky temporary
protecting group to shield the alpha proton of the aminoaldehyde intermediate and performs reductive amination to build the γPNA backbone. As a result, the final monomers have enantiomeric excess more than 99.5% confirmed by HPLC and 19F NMR. This methodology will facilitate the production of γPNA for a wider range of applications. Earlier, γPNA is demonstrated to be preorganized into either right-handed (RH) or left-handed (LH) motifs by simply inverting the stereochemistry at γ position. The RH motif can bind to RH-γPNA, unmodified PNA and native nucleic acid with exquisite binding affinity and sequence specificity, while the LH motif can only bind to LH-γPNA and unmodified PNA. The second work presented in this Dissertation demonstrates that the sequence information embedded in conformationally orthogonal γPNA can be interconverted by toehold-mediated strand displacement reaction. This work opens up the opportunities for developing orthogonal molecular circuits to rapidly and sensitively detect genetic materials. Designing a tight binding hybridization probe with high sequence specificity is
challenging in that the probes are too stable to be sensitive to a base mispairing. Inspired by Nature’s template-directed assembly, the third work presented in this Dissertation
demonstrates a relatively weak binding MPγPNA probe can be concatenated in the presence of disease-relevant RNA repeated expansions and becomes a tight binding and
potentially selective hybridization probe. This work may benefit the development of future nucleic acid therapeutics.
Together, this Dissertation offers a robust synthetic strategy for preparing optically pure γPNA monomers, systematically understands the binding relationships of γPNA and
provides a novel strategy to in-situ assemble tight binding probe with high sequence selectivity. This work envisions γPNA as a versatile biomolecular self-assembly platform
with control over sequence and helical sense and will hopefully facilitate the potential applications of γPNA in diagnostics and therapeutics.