Synthesis, Affinity for Complementary RNA and DNA, and Enzymatic Stability of Triazole-Linked Locked Nucleic Acids (t-LNAs)

Abstract

Dinucleoside phosphoramidites containing a triazole internucleotide linkage flanked by locked nucleic acid (LNA) were synthesized and incorporated into oligonucleotides (ONs). ONs bearing both LNA and triazole at multiple sites were obtained and their biophysical properties including enzymatic stability and binding affinity for RNA and DNA targets were studied. t-LNAs with four incorporations of a dinucleoside monomer having LNA on either side of the triazole linkage bind to their RNA target with significantly higher affinity and greater specificity than unmodified oligonucleotides, and are remarkably stable to nuclease degradation. A similar but reduced effect on enzymatic stability and binding affinity was noted for LNA only on the 3′-side of the triazole linkage. Thus, by combining unnatural triazole linkages and LNA in one unit (t-LNA), we produced a promising class of ONs with reduced anionic charge and potential for antisense applications.

Strict conformational demands of RNA cleavage in bulge-loops created by peptidyl-oligonucleotide conjugates

“…Although the peptide provides the oligonucleotide with some level of protection from cellular nucleases, ( 17 , 20 ) yet more biologically stable variants can be produced by harnessing recent advances in nuclease-resistant chemical modification, ( 60 , 61 ) which may also enhance binding to the target RNA. Indeed, the newly discovered triazole-LNA and carbamate-LNA modified oligonucleotides ( 62 , 63 ) show a much reduced level of degradation in vivo and significantly higher affinity toward RNA/DNA, which is re-enforced by locked nucleic acid sugars, thus providing us with new opportunities. This might allow the use of shorter oligonucleotides with improved cellular uptake properties, especially when conjugated to the arginine-rich catalytic peptides, which have been previously shown to facilitate transport of large hydrophilic molecules across biological barriers ( 64–68 ).…”

Modified internucleoside linkages for nuclease-resistant oligonucleotides

“…19). 351 Hybridization studies have been undertaken on ODN containing one to four blocks of the dimers described above (Fig. 19).…”

Searching for the ideal triazole: Investigating the 1,5-triazole as a charge neutral DNA backbone mimic

“…Several strategies have been developed to improve target affinity of triazole containing oligonucleotides, including flanking the triazole with LNA [27][28][29] and adding a G-clamp to the neighbouring nucleobase. 25 Whilst these are valid approaches, it is clear that the precise nature of the TL can have a significant effect on the hybridisation properties of the parent oligonucleotides and that optimization of the linkage should be considered carefully.…”