Facile synthesis and fundamental properties of an N-methylguanidine-bridged nucleic acid (GuNA[NMe])

Abstract

An N-methylguanidine-bridged nucleic acid (GuNA[NMe]), a guanidine-bridged nucleic acid (GuNA) bearing a methyl substituent at the bridge, was successfully synthesised and incorporated into oligonucleotides. By employing an acetyl protecting group, GuNA[NMe]-modified oligonucleotides bearing acid-sensitive purine nucleobases were successfully prepared. The obtained GuNA[NMe]-modified oligonucleotides exhibit excellent binding affinity towards the complementary single-stranded RNA and DNA. Furthermore, even a single GuNA[NMe] modification provides robust enzymatic stability, similar to that achieved by the well-established phosphorothioate backbone modification. These data indicate that such a GuNA[NMe] represents a valuable modification for the development of therapeutic oligonucleotides.

Polyamine‐Functionalized 2′‐Amino‐LNA in Oligonucleotides: Facile Synthesis of New Monomers and High‐Affinity Binding towards ssDNA and dsDNA

“…By combining these two strategies, nucleotide modifications such as N ‐methylguanidino‐, guanidino‐, piperazino‐ and glycyl‐2′‐amino‐LNA nucleosides have been studied and shown to equip the resulting ONs with 3′‐nucleolytic stability and high affinity toward single‐stranded DNA (ssDNA) and ssRNA targets (Figure 1). [27, 31, 32, 35, 36] Significant triplex stability was also observed for ONs carrying such 2′‐amino‐LNA monomers [27, 32] . Moreover, after being introduced into so‐called bisLNAs [a triplex‐forming ON (TFO) linked to a Watson–Crick interacting ON, both targeting the same ssDNA], piperazino‐ and glycyl‐modified 2′‐amino‐LNA nucleotides showed a remarkable ability to invade double‐stranded DNA (dsDNA) targets in vitro [37] .…”

Synthesis and properties of oligonucleotides modified with an N-methylguanidine-bridged nucleic acid (GuNA[Me]) bearing adenine, guanine, or 5-methylcytosine nucleobases

“…The preparation of the GuNA[Me]-A, -G, and -m C phosphoramidites 3a-c needed for the synthesis of the GuNA[Me]-modified oligonucleotides is detailed in Scheme 1. The acetyl group was selected as a protecting group for the guanidine moiety because it can be easily removed under the basic conditions (ammonia/methylamine solution) used for the DNA synthesis [20]. The phosphoramidite synthesis was started from 2'-amino-LNAs 1a-c, which were rapidly prepared via the transglycosylations of 2'-amino-LNA-T [25].…”

“…Notably, since the nucleobases were introduced at the late stage of the synthesis, we had no difficulty preparing these phosphoramidites. [20]. 5-(Ethylthio)-1H-tetrazole (ETT) was used as an activator for the coupling, and the coupling time was extended from 40 s to 20 min for the GuNA[Me] phosphoramidites.…”

“…This compound was synthesized in a similar manner as described in reference [20]. N 2 -isobutyrylguanine-9-yl)…”

“…The modification of oligonucleotides with GuNA[H]-T improved the nuclease resistance, cell membrane permeability, and binding affinity toward complementary single-stranded DNAs (ssDNAs) and RNAs (ssRNAs). We also synthesized and evaluated a GuNA[H]-T analog bearing a methyl group in the guanidine moiety (GuNA[Me]-T; Figure 1) [20]. The GuNA[Me]-T exhibited a similar duplexforming ability and nuclease resistance as GuNA[H]-T. Since a subtle change in the structure of the 2',4'-BNA/LNA modulated its biophysical and pharmacological properties, in vivo experiments with GuNA[H] and GuNA[Me] are expected to provide further mechanistic insights into how small substituents affect the efficacy and safety of therapeutic oligonucleotides.…”