Triazole- and Tetrazole-Bridged Nucleic Acids: Synthesis, Duplex Stability, Nuclease Resistance, and in Vitro and in Vivo Antisense Potency

Antisense
oligonucleotides are attractive therapeutic agents for
several types of disease. One of the most promising modifications
of antisense oligonucleotides is the introduction of bridged nucleic
acids. As we report here, we designed novel bridged nucleic acids,
triazole-bridged nucleic acid (TrNA), and tetrazole-bridged nucleic
acid (TeNA), whose sugar conformations are restricted to N-type by
heteroaromatic ring-bridged structures. We then successfully synthesized
TrNA and TeNA and introduced these monomers into oligonucleotides.
In UV-melting experiments, TrNA-modified oligonucleotides exhibited
increased binding affinity toward complementary RNA and decreased
binding affinity toward complementary DNA, although TeNA-modified
oligonucleotides were decomposed under the annealing conditions. Enzymatic
degradation experiments demonstrated that introduction of TrNA at
the 3′-terminus rendered oligonucleotides resistant to nuclease
digestion. Furthermore, we tested the silencing potencies of TrNA-modified
antisense oligonucleotides using in vitro and in vivo assays. These
experiments revealed that TrNA-modified antisense oligonucleotides
induced potent downregulation of gene expression in liver. In addition,
TrNA-modified antisense oligonucleotides showed a tendency for increased
liver biodistribution. Taken together, our findings indicate that
TrNA is a good candidate for practical application in antisense methodology.