Evolved polymerases facilitate selection of fully 2'-OMe-modified aptamers

Area of Science:

• Biochemistry
• Molecular Biology
• Medicinal Chemistry

Background:

• 2 '-OMe modifications enhance nuclease resistance in RNA and DNA aptamers.
• However, these modifications hinder polymerase recognition, complicating aptamer identification.
• Novel thermostable DNA polymerases (SFM4-6 and SFM4-9) were previously developed to overcome this challenge.

Purpose of the Study:

• To report the first selection of fully 2 '-OMe modified aptamers.
• To identify aptamers that bind human neutrophil elastase (HNE).
• To evaluate the binding affinity and stability of these modified aptamers.

Main Methods:

• Selection of 2 '-OMe modified aptamers using evolved DNA polymerases SFM4-6 and SFM4-9.
• Systematic evolution of ligands by exponential enrichment (SELEX) for aptamer discovery.
• Characterization of aptamer binding affinity and stability under various conditions (salt, serum).

Main Results:

• Six fully 2 '-OMe modified aptamers (2mHNE-1 to 2mHNE-6) were successfully selected against HNE.
• All isolated aptamers demonstrated reasonable binding affinity, dependent on 2 '-OMe substituents.
• One aptamer (2mHNE-5) retained high affinity for HNE in high salt and serum concentrations, outperforming natural aptamers.

Conclusions:

• The developed DNA polymerases SFM4-6 and SFM4-9 enable efficient selection of 2 '-OMe modified aptamers.
• These modified aptamers exhibit robust binding to HNE, with potential for therapeutic applications.
• The findings pave the way for broader exploration and production of nuclease-resistant aptamers.