Alex Azhayev

1-Alkylthioalkylation of nucleoside hydroxyl functions and its synthetic applications : a new versatile method in nucleoside chemistry

Abstract Treatment of appropriately protected nucleosides with a mixture of acetic acid, acetic anhydride and dialkylsulfoxide was shown to give O-(1-alkylthioalkylated) nucleosides that were oxidized to the corresponding sulfoxides and sulfones, or converted via O-halogenomethyl derivatives to various O-substituted nucleosides.

Amide group assisted 3′-dephosphorylation of oligonucleotides synthesized on universal A-supports

Abstract (±)-3-Amino-1-(4,4′-dimethoxytriphenylmethyl)-2-propanediol was attached to succinylated alkylamino-controlled pore glass via the second amide bond. The resulting solid phase was acylated to give seven new universal solid supports, compatible with the preparation of all common types of oligodeoxyribonucleotides. These resins allow for fast elimination of the 3′-terminal phosphodiester or phosphorothioate function by ammonia in methanol at room temperature.

Novel solid supports for the preparation of 3′-derivatized oligonucleotides: Introduction of 3′-alkylphosphate tether groups bearing amino, carboxy, carboxamido, and mercapto functionalities

Abstract Syntheses of 5 non-nucleosidic solid supports (1–5) that enable preparation of oligonucleotides bearing a carboxy,-amino,-carboxamido,- or mercaptoalkyl spacer arm at their 3′-terminus are described. They all contain an ester bond of moderate susceptibility toward nucleophiles. Upon the completion of oligonucleotide chain assembly, this bond may be cleaved by a variety of nucleophiles. These release the oligonucleotide from the support and simultaneously introduce the desired functionality. Differences in the reactivity between the supports prepared are discussed.

A new universal solid support for oligonucleotide synthesis

Starting from 2,3:5,6-di-O-isopropylidene-α-D-mannofuranose, benzyl 5,6-ditrifluoroacetamido-5,6-dideoxy-2-O-(4,4′-dimethoxytrityl)-α-D-mannofuranose was prepared and attached to a controlled pore glass to generate a new universal solid support for oligonucleotide synthesis. This support is applicable for the preparation of all types of oligonucleotides as well as oligomers with unusual baselabile nucleosides.

Towards a general method for the stepwise solid-phase synthesis of peptide–oligonucleotide conjugates

Abstract Three peptide–oligonucleotide phosphorothioate hybrids were synthesized using a new approach for stepwise solid-phase synthesis of conjugates. This method utilizes commercially available N α -Fmoc amino acids for peptide synthesis and a new solid support. Three specific modifications of the solid-phase were made after the peptide and before the oligonucleotide assembly.

Stepwise solid-phase synthesis of peptide–oligonucleotide phosphorothioate conjugates employing Fmoc peptide chemistry

Abstract A straightforward stepwise method for the preparation of peptide–oligonucleotide phosphorothioate conjugates, was developed, based on the highly efficient Fmoc peptide solid phase synthesis, followed by oligonucleotide phosphorothiate chain assembly. The three conjugates synthesized contained 15- or 17-mer oligonucleotide phosphorothioates and 10- or 16-mer peptides, incorporating two or three arginine residues.

Oligonucleotides Containing 9-(2-Deoxy-2-Fluoro-β-D-Arabinofuranosyl)-Adenine and -Guanine: Synthesis, Hybridization and Antisense Properties

Abstract Synthesis of 9-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)-adenine (7, ara-A2′F) and -guanine (12 ara-G2′F) was accomplished via the condensation of 2, 6-dichloropurine (1) with 2-deoxy-2-fluoro-1, 3, 5-tri-O-benzoyl-α-D-arabinofuranose (2) as a key chemical step. Condensation of silylated N 6-benzoyladenine (6) with 2 gave, after deblocking and chromatographic separation, ara-A2,F (7) (14%), its α-anomer 8 (14%) and N 7-α-isomer 9 (25%). The PSEUROT analysis of N 9-β-D-arabinosides 7 and 12 manifested slight preference for the S rotamer (64%) for the former, and an equal population of the N and S rotamers for the latter. The arabinosides 7 and 12 were used for the preparation of the respective phosphoamidite building blocks 13 and 14 for automated oligonucleotide synthesis. Four 15-mer oligonucleotides (ONs) complementary to the initiation codon region of firefly luciferase mRNA were prepared: unmodified 2′-deoxy-ON (AS1) and containing (i) ara-A2,F instead of the only A (AS2), (ii) ara-G2,F vs. 3-G from the 5′-terminus (AS3), and (iii) both arabinosides at the same positions (AS4). All these ONs display practically the same (i) affinity to both complementary DNA and RNA, and (ii) ability to inhibit a luciferase gene expression in a cell-free transcription-translation system.

Synthesis of 3'-functionalized oligonucleotides on a single solid support

Abstract Oligodeoxyribonucleotides tethered with an amino-, carboxy,- amidocarbonyl-, or mercaptoalkyl spacer arm at 3′-terminus were obtained by assembling the chains on a modified aminoalkyl-CPG ( 1 ) and releasing them with appropriate reagents.

Improved Synthesis of Trinucleotide Phosphoramidites and Generation of Randomized Oligonucleotide Libraries

A new method to produce a set of 20 high quality trinucleotide phosphoramidites on a 5–10 g scale each was developed. The procedure starts with condensation reactions of P-components with N-acyl nucleosides, bearing the 3 ′-hydroxyl function protected with 2-azidomethylbenzoyl, to give fully protected dinucleoside phosphates 13. Upon cleavage of dimethoxytrityl group from 13, dinucleoside phosphates 16 are initially transformed into trinucleoside diphosphates 19 and then the 2-azidomethylbenzoyl is selectively removed under neutral conditions to generate trinucleoside diphosphates 5 in excellent yield. Subsequent 3 ′-phosphitylation affords target trinucleotide phosphoramidites 7. When mutagenic oligonucleotides are synthesized employing mixtures of building blocks 7 as well as following the new synthetic protocol, representative oligonucleotide libraries are generated in good yields.

Synthesis of 3′-O-(ω-aminoalkoxymethyl)thymidine 5′-triphosphates, terminators of DNA synthesis that enable 3′-labelling

Treatment of 5′-O-benzoylthymidine 1 with a mixture of acetic anhydride, acetic acid and dimethyl sulfoxide yielded 5′-O-benzoyl-3′-O-methylthiomethylthymidine 2, which was converted via the 3′O-bromomethyl derivative into 3′-O-(ω-aminoalkoxymethyl)thymidines 7 bearing a 6, 8 or 10 methylene groups long hydrocarbon chain, and finally to their 5′-triphosphates 10. The latter compounds were shown to be terminators of DNA synthesis catalysed by thermostable Tet/z DNA-polymerase, and may be labelled at the aliphatic amino group with fluorescent probes.