title: Practical synthesis of cytidine-5-carboxamide-modified nucleotide reagents.
abstract: Chemically-modified derivatives of cytidine, bearing a 5-(N-substituted-carboxamide) functional group, are new reagents for use in aptamer discovery via the SELEX process (Systematic Evolution of Ligands by EXponential enrichment). Herein, we disclose a practical synthesis of 5-(N-benzylcarboxamide)-2'-deoxycytidine, and the corresponding 5-(N-1-naphthylmethylcarboxamide)- and 5-(N-3-phenylpropylcarboxamide)-2'-deoxycytidine analogs, as both the suitably-protected 3'-O-cyanoethylphosphoramidite reagents (CEP; gram scale) and the 5'-O-triphosphate reagents (TPP; milligram-scale). The key step in the syntheses is a mild, palladium(0)-catalyzed carboxyamidation of an unprotected 5-iodo-cytidine. Use of the CEP reagents for solid-phase oligonucleotide synthesis was demonstrated and incorporation of the TPP reagents by KOD polymerase in a primer extension assay confirmed the utility of these reagents for SELEX. Finally, the carboxyamidation reaction was also used to prepare the nuclease-resistant sugar-variants: 5-(N-benzylcarboxamide)-2'-O-methyl-cytidine and 5-(N-3-phenylpropylcarboxamide)-2'-deoxy-2'-fluoro-cytidine.
authors.affiliation: a SomaLogic Inc. , Boulder , Colorado , USA.
authors.forename: John C
authors.initials: JC
authors.lastname: Rohloff
authors.forename: Catherine
authors.initials: C
authors.lastname: Fowler
authors.forename: Brian
authors.initials: B
authors.lastname: Ream
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Carter
authors.forename: Greg
authors.initials: G
authors.lastname: Wardle
authors.forename: Tim
authors.initials: T
authors.lastname: Fitzwater
mesh_terms: D003562:Cytidine / Q000031:analogs & derivatives / Q000138:chemical synthesis* / Q000737:chemistry; D009841:Oligonucleotides / Q000138:chemical synthesis* / Q000737:chemistry; D060327:Solid-Phase Synthesis Techniques
chemical_list: D009841:Oligonucleotides; D003562:Cytidine
keywords: Aptamer; SELEX; cytidine-5-carboxamide; modified nucleotide