Synthesis of Nucleotides Bearing the 2′-O-Trifluoromethyl Group and Their Application in RNA Analogs Preparation

The integration of fluorine atoms into biologically active organic compounds has proved to be a vital technique in small molecule drugs. This technique can substantially enhance crucial properties, including metabolic stability, lipophilicity, and bioavailability, often with a mere addition of a single fluorine atom or a trifluoromethyl group. Over the past few decades, this concept has also been applied in nucleic acid chemistry. A commonly employed 2′-OH substitution is the introduction of a 2′-deoxy-2′-fluoro (2′-F) group. The strong electronegativity of fluorine prompts the modified siRNA to readily adopt a C3′- endo conformation, resulting in significant advantages in terms of binding affinity. To enrich the toolbox of chemical modification of oligonucleotides, the replacement of the 2′-OH with the 2′- O -trifluoromethyl group has been developed in RNA analog synthesis. Oligodeoxynucleotides containing the 2′- O -trifluoromethyl group can greatly increase the thermal stability of DNA/RNA duplexes depending on the position and amount of the modification. Moreover, 2′- O -trifluoromethylated oligodeoxynucleotide also exhibited a slightly higher resistance to snake venom phosphodiesterase than the unmodified oligodeoxynucleotide. The 2′- O -trifluoromethylated oligonucleotides can emerge as a label to study RNA structure and function as well, or to develop DNA/RNA-based diagnostics. Hence, it is necessary to report an effective method for the synthesis, deprotection, purification, and characterization of oligonucleotides bearing a 2′ -O -trifluoromethyl group. © 2024 Wiley Periodicals LLC. Basic Protocol 1 : Preparation of 6- N -benzoyl-5′- O -dimethoxytrityl-2′- O -trifluoromethyl adenosine 3′-(2-cyanoethyl N,N -diisopropyl)phosphoramidite Basic Protocol 2 : Preparation of 4- N -acetyl-5′- O -dimethoxytrityl-2′- O -trifluoromethyl cytidine 3′-(2-cyanoethyl N,N -diisopropyl)phosphoramidite Basic Protocol 3 : Preparation of 2- N -isobutyryl-5′- O -dimethoxytrityl-2′- O -trifluoromethyl guanine 3′-(2-cyanoethyl N,N -diisopropyl)phosphoramidite Basic Protocol 4 : Preparation of 5′- O -dimethoxytrityl-2′- O -2-trifluoromethyl uridine 3′-(2-cyanoethyl N,N -diisopropyl) phosphoramidite Basic Protocol 5 : Solid-phase synthesis of 2′- O -trifluoromethylated RNA analogs Basic Protocol 6 : Deprotection and purification of 2′- O -trifluoromethyl-RNAs