Eugeny M. Zubin

Oligonucleotide-peptide conjugates as potential antisense agents

Oligonucleotide‐peptide conjugates have several applications, including their potential use as improved antisense agents for interfering with the RNA function within cells. In order to provide robust and generally applicable conjugation chemistry, we developed a novel approach of fragment coupling of pre‐synthesized peptides to the 2′‐position of a selected nucleotide within an otherwise protected oligonucleotide chain attached to a solid support.

1,2-Diol and Hydrazide Phosphoramidites for Solid-Phase Synthesis and Chemoselective Ligation of 2′-Modified Oligonucleotides

Abstract The preparation of two novel 2′-O-alkyl phosphoramidites bearing 1,2-diol and hydrazide functions for a chemoselective ligation is described. The former amidite was used to obtain 2′-modified oligodeoxyribonucleotides, which can be later oxidised by NaIO4 to generate 2′-aldehyde oligonucleotides. These were successfully conjugated to acceptor molecules. The latter amidite also showed good coupling yields, but the hydrazide function was demonstrated to be labile under basic deprotection conditions.

DNA Duplexes Containing 2′-Deoxy-2′-Iodoacetamidouridine as Reagents for Affinity Modification of Proteins

DNA duplexes containing the iodoacetamido group at position 2′ of the ribose moiety were proposed for affinity modification of Cys in DNA-binding proteins. Reactive DNA derivatives were obtained with iodoacetic anhydride and synthetic oligodeoxyribonucleotides containing 2′-amino-2′-deoxyuridine in place of thymidine at various positions. The derivatives were tested for reaction with amino acids and peptides and shown to specifically interact with Cys-containing proteins. The possibility of using the modified DNA duplexes to probe the protein SH group close to the DNA sugar-phosphate backbone in DNA–protein complexes was demonstrated with the example of subunit p50 of human transcription factor NF-κB.

2′-aldehyde oligonucleotides: Synthesis and use for affinity modification of DNA-recognizing proteins

Oligonucleotides with 1,2-diol group were prepared from 2′-O-[2-(2,3-dihydroxypropyl)amino-2-oxoethyl]uridine 3′-phosphoramidite. The thermal stability of modified DNA duplexes and their ability to form complexes with the p50 subunit of the NF-ϰB transcription factor and (cytosine-5)-DNA methyltransferase M.SsoII were studied. The periodate oxidation of the 1,2-diol group of the oligonucleotides resulted in reactive 2′-aldehyde derivatives. The opportunity of their use for the affinity modification of DNA-recognizing proteins was studied.

A convenient solid-phase method for the synthesis of novel oligonucleotide-folate conjugates.

We describe the preparation of two batches of a polymer support for the incorporation of folic acid into oligonucleotides. The method permits the regioselective attachment of a target nucleic acid sequence through its 3′-end to either the α-or γ-carboxyl group of L-glutamic acid, respectively. The supports have been tested in solid-phase synthesis of oligonucleotide-folate conjugates for cell delivery studies.

[2'-Modified oligoribonucleotides, containing 1,2-diol and aldehyde groups. Synthesis and properties].

Abstract1,2-Diol-oligoribonucleotides were prepared using fully protected 2′-O-[2-(2,3-dihydroxypropyl)amino-2-oxoethyl]uridine 3′-phosphoramidite. Incorporation of the modified uridine residue into oligonucleotide chains was not shown to significantly affect the thermal stability of RNA-RNA and RNA-DNA duplexes. Periodate oxidation of the 1,2-diol group resulted in reactive 2′-aldehyde oligoribonucleotides. These oligonucleotides were studied for their application in the affinity modification of RNA recognizing proteins with an example of bacterial ribonuclease P.

DNA or RNA Oligonucleotide 2′-Hydrazides for Chemoselective Click-Type Ligation with Carbonyl Compounds

An efficient method for the synthesis of DNA or RNA oligonucleotide 2′-hydrazides is described. Fully deprotected oligonucleotides containing a hydrazide group at the 2′-position of a uridine residue were obtained by a novel two-step procedure: periodate cleavage of an oligonucleotide with 1,2-diol group followed by conversion of the aldehyde to hydrazide with an extended linker arm using a homobifunctional reagent succinic dihydrazide and NaBH3CN. The resulting oligonucleotide 2′-hydrazides were efficiently conjugated by a click-type reaction at acidic pH to aliphatic or aromatic aldehydes with or without NaBH3CN reduction to afford novel 2′-conjugates.