Synthesis of 2‘-C-Difluoromethylribonucleosides and Their Enzymatic Incorporation into Oligonucleotides
2005-09-30T00:00:00Z (GMT) by
Nucleosides bearing a branched ribose have significant promise as therapeutic agents and biotechnological and biochemical tools. Here we describe synthetic entry into a new subclass of these analogues, 2‘-C-β-difluoromethylribonucleosides. We constructed the glycosylating agent 4 in three steps from 1,3,5-tri-O-benzoyl-α-d-ribofuranose 1. The key steps included nucleophilic addition of difluoromethyl phenyl sulfone to 2-ketoribose 2 followed by mild and efficient reductive desulfonation. Ribofuranose 4 glycosylated bis(trimethylsilyl)uracil directly, giving difluoromethyluridine 7 efficiently after deprotection. Conversion of 4 to the corresponding ribofuranosyl bromide allowed efficient access to C, A, and G analogues. A related approach starting from methyl d-ribofuranose offered synthetic entry into the diastereomeric manifold, 2‘-C-α-difluoromethyl-arabino-α-pyrimidine. To incorporate 2‘-C-β-difluoromethyluridine into an oligodeoxynucleotide we converted 7 to the bisphosphate and carried out successive ligation reactions using T4 RNA ligase and T4 DNA ligase. Analogous to natural RNA linkages, the resulting oligonucleotide undergoes hydroxide-catalyzed backbone scission at the difluoromethyluridine residue via internal transphosphorylation.