2′,4′‐BNA/LNA with 9‐(2‐Aminoethoxy)‐1,3‐diaza‐2‐oxophenoxazine Efficiently Forms Duplexes and Has Enhanced Enzymatic Resistance

Artificial nucleic acids are widely used in various technologies, such as nucleic acid therapeutics and DNA nanotechnologies requiring excellent duplex‐forming abilities and enhanced nuclease resistance. 2′‐O,4′‐C‐Methylene‐bridged nucleic acid/locked nucleic acid (2′,4′‐BNA/LNA) with 1,3‐diaza‐2‐oxophenoxazine (BNAP (B(H))) was previously reported. Herein, a novel B(H) analogue, 2′,4′‐BNA/LNA with 9‐(2‐aminoethoxy)‐1,3‐diaza‐2‐oxophenoxazine (G‐clamp), named BNAP‐AEO (B(AEO)), was designed. The B(AEO) nucleoside was successfully synthesized and incorporated into oligodeoxynucleotides (ODNs). ODNs containing B(AEO) possessed up to 10(4)‐, 152‐, and 11‐fold higher binding affinities for complementary (c) RNA than those of ODNs containing 2′‐deoxycytidine (C), 2′,4′‐BNA/LNA with 5‐methylcytosine (L), or 2′‐deoxyribonucleoside with G‐clamp (P(AEO)), respectively. Moreover, duplexes formed by ODN bearing B(AEO) with cDNA and cRNA were thermally stable, even under molecular crowding conditions induced by the addition of polyethylene glycol. Furthermore, ODN bearing B(AEO) was more resistant to 3′‐exonuclease than ODNs with phosphorothioate linkages.