A Short,Efficient Synthesis of 2′-Deoxypseudoisocytidine Based on Heck-Chemistry

Abstract

A novel synthesis of 2′-deoxypseudoisocytidine as well as of its phosphoramidite building block for oligonucleotide synthesis is presented. The synthesis is based on Heck-coupling between N-protected pseudoisocytosine and a silyl protected furanoid glycal. With this procedure the corresponding phosphoramidite building block is obtained in 5 steps and an overall yield of 28%.     

New synthesis of 5-carboxy-2'-deoxyuridine and its incorporation into synthetic oligonucleotides

5-Carboxy-2'-deoxyuridine is a methyl oxidation product of thymidine. It can be formed by the menadione-mediated photosensitization of thymidine in aerated aqueous solution. Here in we present a new four-step synthesis of the 5-carboxy-2'-deoxyuridine phosphoramidite building block based on the alkaline hydrolysis of 5-trifluoromethyl-2'-deoxyuridine. The phosphoramidite derivative has been incorporated at defined sites into oligonucleotides using the solid phase synthesis approach.     

Synthesis of an oligonucleotide analogue of ethenoadenosine

A phosphoramidite building block derived from 11-carboxy-1,N6-ethenoadenosine has been prepared to be used in a solid supported oligonucleotide synthesis.     

New Synthesis of 5-Carboxy-2′-deoxyuridine and Its Incorporation into Synthetic Oligonucleotides

Abstract

5-Carboxy-2′-deoxyuridine is a methyl oxidation product of thymidine. It can be formed by the menadione-mediated photosensitization of thymidine in aerated aqueous solution. Here in we present a new four-step synthesis of the 5-carboxy-2′-deoxyuridine phosphoramidite building block based on the alkaline hydrolysis of 5-trifluoromethyl-2′-deoxyuridine. The phosphoramidite derivative has been incorporated at defined sites into oligonucleotides using the solid phase synthesis approach.     

Synthesis and properties of phosphorylated 3'-O-beta-D-ribofuranosyl-2'-deoxythymidine

A strategy was developed for the synthesis of 3'-O-beta-D-ribofuranosyl 2'-deoxythymidine derivatives using three different protecting groups, which allows the synthesis of a phosphoramidite building block for oligonucleotide synthesis. Likewise the 5'-O- and 5'-O-phosphorylated analogues were synthesized and their conformation was determined using NMR spectroscopy.     

Aldehydo-oligonucleotides for Bioconjugation

Abstract

The synthesis of a new phosphoramidite building block with a masked aldehyde function is described. It was incorporated in an oligodeoxyribonucleotide. Coupling to amino derivatives by reductive animation was performed in high yield after unmasking the aldehyde function.     

Conjugation of oligonucleotides via an electrophilic tether: N-chloroacetamidohexyl phosphoramidite reagent

A novel method for the preparation of oligonucleotides conjugated with nucleophilic ligands is described. A new phosphoramidite building block derived from N-chloroacetyl-6-aminohexanol is attached at the 5′-terminus on the last step of oligonucleotide synthesis. Postsynthetic treatment of support-bound modified oligonucleotide with a variety of amines and mercaptans affords conjugates in high yield.     

Synthesis of an α-kojibiosyl substituted glycerol teichoic acid hexamer

In this paper the synthesis of an Enterococcus Faecalis teichoic acid (TA) hexamer is presented. The key kojibiosyl-glycerol phosphoramidite building block was obtained by condensation of thioglucose donors, provided with various protecting groups at the C2 hydroxyl function with an orthogonally protected glycerol acceptor. After selective deprotection, the resulting 1,2-cis-linked pseudodisaccharide acceptor was coupled to an α-directing thioglucose donor, giving the corresponding pseudotrisaccharide, which is then transformed to a phosphoramidite synthon. The kojibiosyl-glycerol phosphoramidite in combination with a glycerolphosphoramidite, an aminohexylphosphoramidite and dibenzylglycerol were coupled to a fully protected glycerol TA hexamer, using chemistry that can be amended for future automated synthesis. Global deprotection afforded the target hexamer kojibiosyl-glycerol containing TA (1).     

Synthesis of an Oligonucleotide Analogue of Ethenoadenosine

Abstract

A phosphoramidite building block derived from 11-carboxy-1,N ⁶-ethenoadenosine has been prepared to be used in a solid supported oligonucleotide synthesis.     

Use of [4,6-Di-14C]-5′-DMT-thymidine Phosphoramidite Reagent for the Radiolabeling of Synthetic Oligonucleotides

Abstract

A novel synthesis of 5′-radiolabeled oligonucleotides is described. The labeling is carried out by the phosphoramidite method with the aid of building block 1. The feasibility of the method is demonstrated by preparation of 5′-radiolabeled 3′-phosphorylated dodecathymidylate phosphorothioate.     

Synthesis of Oligonucleotide Conjugates with the Aid of N-Chloroacetamidohexyl Phosphoramidite Reagent

Abstract

A novel phosphoramidite building block derived from N-chloroacetyl-6-aminohexanol is attached at the 5′-terminus on the last step of oligonucleotide synthesis. Postsynthetic treatment of support-bound modified oligonucleotides with a variety of amines and mercaptanes affords oligonucleotide conjugates in a high yield.     

Synthesis and DNA binding properties of a new benzo[f]imidazo[1,5b]-isoquinoline-butan-1,2-diol derivative

A benzo[f]imidazo[1,5b]-isoquinoline derivative 4 with a 1,2-butandiol linker was prepared by reaction of a trimethylsilylated 5-naphthylidenehydantoin 3 with a 2,3-dideoxy-D-glycero-pentafuranoside 2 in 22% yield. After deprotection, the resulting compound 5 was converted to a DMT protected phosphoramidite building block 7 for standard DNA synthesis. DNA/DNA, DNA/RNA duplexes with 5 inserted as bulges were destabilized, except when the new amidite was used for the synthesis of a zipping duplex.     

Chemical synthesis of 2''-deoxyoligonucleotides containing 5-fluoro-2''-deoxycytidine.

2'-Deoxyoligonucleotides with 5-fluorocytosine residues incorporated at specific positions of the nucleotide sequence are tools of great potential in the study of the catalytic mechanism by which DNA cytosine methyltransferases methylate the 5-position of DNA cytosine residues in specific sequence contexts. Chemical synthesis of such oligonucleotides is described. Two alternative approaches have been developed, one of which proceeds via a fully protected phosphoramidite of 5-fluoro-4-methylmercapto-2'-deoxyuridine 2, the other via a fully protected phosphoramidite of 5-fluoro-2'-deoxycytidine 3. Either building block can be used in automated oligonucleotide synthesis applying standard elongation cycles and deprotection procedures exclusively. The methylmercapto function of 2 is replaced by an amino group in the final ammonia treatment used for cleavage from support and base deprotection.     

Synthesis and selective cleavage of an oligodeoxynucleotide containing a bridged internucleotide 5''-phosphorothioate linkage.

A self complementary oligodeoxynucleotide dodecamer containing an achiral bridged 5'-phosphorothioate linkage 3'-O-P-S-5' has been prepared using the solid phase phosphoramidite procedure. For the incorporation of the phosphorothioate link we have used a 5'-(S-trityl)-mercapto-5'-deoxythymidine-3'-phosphoramidite. After coupling this building block the S-trityl group was removed by silver ions. The free thiol moiety was then coupled with a standard phosphoramidite in the presence of tetrazole. After oxidation of the 5'-phosphorothioite with iodine/water the synthesis was continued with standard building blocks up to the desired dodecamer. This backbone modified dodecamer can be cleaved selectively and quantitatively at the P-S bond by silver or mercuric ions under very mild conditions.     

Preparation of a novel psoralen containing deoxyadenosine building block for the facile solid phase synthesis of psoralen-modified oligonucleotides for a sequence specific crosslink to a given target sequence.

4,5',8-Trimethylpsoralen was attached to the C8-position of deoxyadenosine via a sulfur atom and a five carbon atom linker. The modified deoxyadenosine was then converted to a protected phosphoramidite and used as unusual as a building block for solid phase oligodeoxyribonucleotide synthesis. The efficiency of the photoreaction of a psoralen-modified oligonucleotide to a complementary matrix strand reached more than 90% within a 1 hour irradiation time at a wavelength of 345 nm.     

Photochemically induced RNA and DNA abasic sites

Two phosphoramidite building blocks were synthesized that can easily be deprotected by UV light to reveal natural abasic sites in oligoribonucleotides as well as in oligodeoxyribonucleotides. Another building block which releases a 2'-O-methylated abasic site upon UV radiation is also described.     

Incorporation of an aldehyde function in oligonucleotides

A nucleotide-like phosphoramidite building block that has the nucleic base replaced by the tert-butyldimethylsilyl-protected styrene glycol was synthesized. After the automatic synthesis of an oligonucleotide incorporating this synthon, the benzaldehyde function was generated by fluoride deprotection and oxidation by sodium periodate. In a similar manner, an oligonucleotide where a nucleic base was replaced by the (CH2)8CH=O chain was synthesized and conjugated with biotin derivatives.     

Synthesis and matrix assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry study of phosphopeptide

The symmetry phosphoramidite reagents were synthesized via atwo-step route and used for the `one pot' synthesis of thephosphoserine building block Fmoc-Ser(PO(OBzl)OH)-OH. Theprocedure is simple and rapid, and product is obtained in highyield. When using this building block, the phosphopeptide(RKGS(PO₃H₂)SSNEPSSDSLSSPTLLAL) related to theC-terminal of c-Fos protein was synthesized manually byFmoc/t-Bu procedure and characterized by matrix assisted laserdesorption/ionization time of flight (MALDI-TOF) massspectrometry. In the fragment ion of phosphopeptide in theMALDI-TOF mass spectrometry with a curved field reflection, theacquired information can be used to identify the sequences andthe phosphorylation sites of the peptide.     

Synthesis and selective cleavage of oligodeoxyribonucleotides containing non-chiral internucleotide phosphoramidate linkages.

Oligodeoxynucleotides containing phosphoramidate internucleotide links 3'-OP(O)NH-5' have been prepared using standard solid phase phosphoramidite techniques. For the incorporation of the phosphoramidate linkages we have used monomer as well as dimer building blocks. With the monomer 3'-phosphoramidite building blocks, which are derived from 5'-amino-2',5'-dideoxynucleosides, it is possible to incorporate phosphoramidate links into specific positions within an oligodeoxynucleotide. Furthermore the synthesis of several dinucleoside phosphate derivatives which are linked by phosphoramidate bonds are described. The internucleotide phosphoramidate linkage was performed using the Staudinger reaction followed by a Michaelis-Arbuzov type transformation. After 3'-phosphitylation these dinucleosides are compatible with the current phosphoramidite methodology of oligodeoxynucleotide synthesis.     

Synthesis of a thymine glycol building block and its incorporation into oligonucleotides.

A phosphoramidite building block of thymine glycol, a major oxidative base damage in DNA, was synthesized in four steps from protected thymidine. The two hydroxyl functions of the oxidized base were protected with the tert-butyldimethylsilyl group. Using this building block, oligonucleotides containing this damage were synthesized.