Chemical Modifications for a Next Generation of Nucleic Acid Aptamers

Abstract

In the past three decades, in vitro systematic evolution of ligands by exponential enrichment (SELEX) has yielded many aptamers for translational applications in both research and clinical settings. Despite their promise as an alternative to antibodies, the low success rate of SELEX (∼30 %) has been a major bottleneck that hampers the further development of aptamers. One hurdle is the lack of chemical diversity in nucleic acids. To address this, the aptamer chemical repertoire has been extended by introducing exotic chemical groups, which provide novel binding functionalities. This review will focus on how modified aptamers can be selected and evolved, with illustration of some successful examples. In particular, unique chemistries are exemplified. Various strategies of incorporating modified building blocks into the standard SELEX protocol are highlighted, with a comparison of the differences between pre‐SELEX and post‐SELEX modifications. Nucleic acid aptamers with extended functionality evolved from non‐natural chemistries will open up new vistas for function and application of nucleic acids.

Traceless enzymatic synthesis of monodispersed hypermodified oligodeoxyribonucleotide polymers from RNA templates

“…However, for many target proteins, the natural nucleic acids do not have suitable functional groups for effective interactions. Therefore, base-modified aptamers 5 bearing non-natural substituents have been pursued and several examples of DNA aptamers containing one 6,7 or two 8 modified nucleotides have been reported to bind more efficiently and selectively than natural nucleic acids. However, to target some undruggable hydrophobic proteins, it could be advantageous to have aptamers displaying even three or four functional groups.…”

Ferrocene as a potential electrochemical reporting surrogate of abasic sites in DNA

“…Similar results were obtained previously with nucleotide 2 and templates containing THFabasic sites. 48 Next, we turned out attention to nucleobase-modified nucleotides due to their relevance in various fields such as aptamer and DNAzyme selection experiments 18,70 or in the context of epigenetic modifications. 71 Since 2'-deoxyadenine is preferentially incorporated opposite Fc and THF-abasic sites, we evaluated whether the presence of functional groups interfered with n+1 product formation.…”

“…Open Lastly, we evaluated the possibility of incorporating modified dUTP analogs bearing various side-chains and functional groups at position C5 of the pyrimidine which is readily tolerated by numerous polymerases. 10,18,67,70,71,73,74 Therminator, Hemo Klem Taq, and Dpo4 efficiently incorporated the commercially available dU*TP analog 5 opposite an Fc site (Figure S10A), while additional polymerases also appended this nucleotide on primer P1 opposite a THFabasic site (Figure S10B). A slightly lower incorporation efficiency was observed when M1-Sp18 was used as template (Figure S10C).…”

Ferrocene as an electrochemical reporting surrogate of abasic sites in DNA

“…Similar results were obtained previously with nucleotide 2 and templates containing THFabasic sites. 48 Next, we turned out attention to nucleobase-modified nucleotides due to their relevance in various fields such as aptamer and DNAzyme selection experiments 18,68 or in the context of epigenetic modifications. 69 Since deoxyadenine is preferentially incorporated opposite Fc and THF-abasic sites, we evaluated whether the presence of functional groups interfered with n+1 product formation.…”

“…Lastly, we evaluated the possibility of incorporating modified dUTP analogs bearing various side-chains and functional groups at position C5 of the pyrimidine which is readily tolerated by numerous polymerases. 10,18,65,68,69,71,72 Therminator, Hemo Klem Taq, and Dpo4 efficiently incorporated the commercially available dU*TP analog 5 opposite an Fc site (Figure S10A), while additional polymerases also appended this nucleotide on primer P1 opposite a THFabasic site (Figure S10B). A slightly lower incorporation efficiency was observed when M1-C18 was used as template (Figure S10C).…”