Selection of a Fluorinated Aptamer Targeting the Viral RNA Frameshift Element with Different Chiralities.

Abstract

The development of RNA aptamers with high specificity and affinity for target molecules is a critical advancement in the field of therapeutic and diagnostic applications. This study presents the selection of a 2'-fluoro-modified mirror-image RNA aptamer through the in vitro SELEX process. Using a random RNA library, we performed iterative rounds of selection and amplification to enrich aptamers that bind specifically to the viral attenuator hairpin RNA containing the opposite chirality, which is an important part of the frameshift element. The unnatural chirality of the aptamer improved its enzymatic stability, and the incorporation of 2'-fluoro modifications was crucial in enhancing the binding affinity of the aptamers. After nine rounds of SELEX, the enriched RNA pool was sequenced and analyzed, revealing the dominant aptamer sequences. The selected 2'-fluoro-modified mirror-image RNA aptamer demonstrated a dissociation constant of approximately 1.6 μM, indicating moderate binding affinity with the target and exceptional stability against nuclease degradation. Our findings highlight the potential of 2'-fluoro-modified mirror-image RNA aptamers in enhancing the stability and utility of RNA-based therapeutics and diagnostics, paving the way for future applications in diverse biomedical fields.