2‘-C-Branched Ribonucleosides:  Synthesis of the Phosphoramidite Derivatives of 2‘-C-β-Methylcytidine and Their Incorporation into Oligonucleotides

Abstract

We describe a strategy for the incorporation of a 2'-C-branched ribonucleoside, 2'-C-beta-methylcytidine, into oligonucleotides via solid-phase synthesis using phosphoramidite derivatives. 4-N-Benzoyl-2'-C-beta-methylcytidine (2b) was synthesized by coupling persilylated 4-N-benzoylcytosine with 1,2,3,5-tetra-O-benzoyl-2-C-beta-methyl-alpha-(and beta)-D-ribofuranose (1) in the presence of SnCl(4) in acetonitrile, followed by selective deprotection with NaOH in pyridine/methanol. The 3'- and 5'-hydroxyl groups were blocked as a cyclic di-tert-butylsilanediyl ether 3 by treatment with di-tert-butyldichlorosilane/AgNO(3) in DMF. The 2'-hydroxyl group was then protected as a tert-butyldimethylsilyl ether 4a by treatment with tert-butylmagnesium chloride followed by addition of tert-butyldimethylsilyl trifluoromethanesulfonate in THF. As an alternative to 2'-silyl protection, the corresponding 2'-O-tetrahydropyranyl ether 4b was prepared by treatment of 3 with 4,5-dihydro-2H-pyran in the presence of a catalytic amount of 10-camphorsulfonic acid in methylene chloride. The di-tert-butylsilanediyl groups of 4a and 4b were removed by treatment with pyridinium poly(hydrogen fluoride) to afford 5a and 5b, respectively. Protection of the 5'-hydroxyl group as a dimethoxytrityl ether and phosphitylation of the 3'-hydroxyl group by the standard procedure gave the phosphoramidite derivatives 7a and 7b. Both 7a and 7b could be used to incorporate 2'-C-beta-methylcytidine into oligonucleotides efficiently via standard solid-phase synthesis, but the tetrahydropyranyl group of 7b was more readily removed from oligonucleotides than the tert-butyldimethylsilyl group of 7a. Oligonucleotides containing 2'-C-beta-methylcytidine undergo base-catalyzed degradation analogous to natural RNA.

Aspartic acid based nucleoside phosphoramidate prodrugs as potent inhibitors of hepatitis C virus replication

“…28 The synthesis of 2′-C-Me containing nucleosides 1 and 2 was accomplished via the Vorbrüggen glycosylation method starting from the commercially available benzoyl-protected 2′-C-Me ribose sugar 12, followed by the deprotection of the benzoyl group in a methanolic ammonia solution at room temperature (Scheme 2). 29 To assist in both the 5′-regioselectivity of phosphorylation and the general organic solubility of the nucleoside, isopropylidene protection was introduced for the 2′,3′-diol unit of the sugar moiety (Scheme 2) to obtain protected nucleosides 15 and 16. They have been used for the synthesis of phosphoramidates 1b-d and 2a-i where the isopropylidene group was deprotected using aqueous 80% TFA at room temperature without affecting the amino acid ester part (Scheme 6).…”

2′-C-Methyl Branched Pyrimidine Ribonucleoside Analogues: Potent Inhibitors of RNA Virus Replication

“…This precursor sugar is easily accessible from commercially available material, and 2′-C-methyl-uridine (4) was prepared as described in the literature (Harry-O'kuru et al, 1997b;Tang et al, 1999), introducing only a slight modification that permitted easy isolation. For practical considerations, we prepared the cytosine derivative (5) converting the uracil nucleoside (Figure 1) via the transient generation of the 4-nitrophenoxy-derivative (Legorburu et al, 1999).…”

“…The 2′-C-methylribonucleosides 3-5 derived from the three naturally occurring pyrimidine bases were synthesized using the strategy reported by Harry-O'kuru et al (Wolfe & Harry-O'kuru 1995;Harry-O'kuru et al, 1997a;Harry-O'kuru et al, 1997b), which allows the use of a common peracylated 2′-C-methylribofuranose (1) which can be condensed with the appropriate heterocyclic base (Figure 1). This precursor sugar is easily accessible from commercially available material, and 2′-C-methyl-uridine (4) was prepared as described in the literature (Harry-O'kuru et al, 1997b;Tang et al, 1999), introducing only a slight modification that permitted easy isolation. For practical considerations, we prepared the cytosine derivative (5) converting the uracil nucleoside (Figure 1) via the transient generation of the 4-nitrophenoxy-derivative (Legorburu et al, 1999).…”

Partial acylation of cytidine and its 2'-C-methyl analogue as a tool to functionalize the ribonucleosidic 2’,3’-cis-diol system

“…2'-C-β-Methylcytidine (2) was obtained according to Ref. 16 Melting points were determined on a Laboratory Devices Mel-Temp II micromelting points apparatus and are uncorrected. UV spectra were measured on a Beckman DU-65 spectrophotometer.…”