An improved guanidine protecting group for the efficient synthesis of GuNA[t-Bu]-modified antisense oligonucleotides

Abstract

An improved protecting strategy for the N-(tert-butyl)guanidine-bridged nucleic acid (GuNA[t-Bu]) phosphoramidite was established with (2,7-di-tert-butyl-9-fluorenyl)methyloxycarbonyl (di-tert-butyl-Fmoc) as a robust protecting group for the guanidine moiety. This protecting group was stable under standard oligonucleotide synthesis conditions and can be removed together with nucleobase protecting groups during the cleavage and deprotection step without the need for special conditions. The improved phosphoramidite enables stable and scalable production of GuNA[t-Bu]-modified oligonucleotides, thereby facilitating broader application of this monomer in oligonucleotide therapeutics.