Syntheses of 2′-C-amidoalkyl and 2′-C-cyanoalkyl containing oligodeoxyribonucleotides and assessment of their hybridisation affinity for complementary DNA and RNA

Oligodeoxynucleotides containing 2′-C-branched nucleosides with an amide or nitrile appended to either a one or two carbon alkyl chain have been synthesised. The phosphoramidites of the 2′-C-modified nucleosides were prepared and incorporated into the oligonucleotides using automated DNA synthesis. The duplex stability with complementary RNA and DNA was measured by UV melting experiments, in order to assess whether the amide/nitrile function could induce any duplex stability without the presence of the 2′-oxygen. The duplex stabilities of the oligonucleotides containing the 2′-C-modifications were decreased in the absence of the 2′-oxygen.     

Regioselective 2′-O-debenzoylation of 2',3'-di-O-benzoyl threose nucleosides

A simple and convenient procedure for the regioselective 2′-O-debenzoylation of 2',3'-di-O-benzoyl threose nucleosides has been achieved successfully affording 3′-O-benzoyl threose nucleosides, which are useful starting material synthons for the synthesis of modified threose nucleosides for different purposes.     

New synthesis of 3′-deoxypurine nucleosides using samarium(III) iodide complex

Regio- and stereoselective iodinative cleavage of 2′,3′-anhydropurine nucleosides was achieved with samarium diiodide and ethyl bromoacetate to produce the corresponding 3′-iodopurine nucleosides, which were then converted to 3′-deoxypurine nucleosides including the natural product cordycepin.     

Synthesis of dinucleotides with 2′-C to phosphate connections by ring-closing metathesis

Four different nucleosides with olefinic 2′-modifications were prepared; 2′-C-methylene, 2′-C-(propen-1-yl), 2′-C-allyl and 2′-O-allyl uridines, respectively. These were incorporated into dinucleotides with allyl phosphate or vinyl phosphonate linkages. Hence, six different dinucleotides were studied as substrates for RCM reactions, and from four of these, cyclic dinucleotides with connections between 2′-C and phosphorus of 3-6 atoms were obtained.     

Efficient synthesis of 5′-O(N)-carbamyl and -polycarbamyl nucleosides

A simple and efficient method for the preparation of 5′-O(N)-carbamyl and 5′-O(N)-polycarbamyl nucleoside derivatives is reported. The method consisted of treatment of 2′,3′-O-protected purine (Ado, Ino) or pyrimidine nucleosides (Thd, Urd) with trichloroacetylisocyanate, followed by cleavage of the trichloroacetyl moiety by silica-gel promoted methanolysis during column chromatography. Iterative application of this method gave mono, di, and tricarbamyl derivatives in good to excellent yields (ave = 80%).     

Synthesis and anti-HIV activities of bis-(cycloSaligenyl) pronucleotides derivatives of 3′-fluoro-3′-deoxythymidine and 3′-azido-3′-deoxythymidine

Anti-HIV nucleoside monophosphates have limited cellular uptake due to the presence of negatively-charged phosphate group. Bis-(cycloSaligenyl) derivatives containing two anti-HIV nucleosides, 3′-fluoro-3′-deoxythymidine (FLT) and 3′-azido-3′-deoxythymidine (AZT) were synthesized to increase intracellular delivery of nucleoside monophosphates. 2,5-Bis(hydroxymethylene)benzene-1,4-diol was selected as a monocyclic bidentate scaffold and synthesized by three different methods from bis(hydroxymethylene)cyclohexan-1,4-diene-1,4-diol, or diethyl 2,5-dihydroxyterephthalate. The reaction of the tetraol with diisopropylphosphoramidous dichloride in the presence of 2,6-lutidine, followed by conjugation reactions with nucleosides (i.e., FLT and AZT) and oxidation afforded symmetrical and unsymmetrical bis-(cycloSaligenyl) diphosphate triester products, AZT-AZT, FLT-FLT, and FLT-AZT conjugates, in 63-74% overall yields and modest anti-HIV activities (IC₅₀ = 2.8-69.6 μM).     

5′-Epimeric 3′-deoxy-3′,4′-didehydronucleoside-5′-C-phosphonates: synthesis and structural assignment by NMR and X-ray analyses

Epimeric 5′-(RS) dialkyl 3′-deoxy-3′,4′-didehydro-5′-C-phosphonates were prepared by nucleophilic addition of various dialkyl phosphites to 3′-deoxy-3′,4′-didehydronucleoside-5′-aldehydes. Whereas direct NMR configuration assignment for the C5′ atom bearing the phosphoryl and hydroxy groups using the J (P,H4′) and J (H5′,H4′) coupling constants is impossible due to the absence of the H4′ atom, successful separation, crystallisation and X-ray crystallographic analysis of a pair of epimeric 5′-C-phosphonates, followed by correlation with a series of NMR parameters, led to efficacious configuration assignment of individual epimers in the mixtures.     

Synthesis of novel 4′-C-methyl-1′,3′-dioxolane pyrimidine nucleosides and evaluation of its anti-HIV-1 activity

The key glycosyl donor for the target molecule 12 was prepared by two-step sequences; (1) acetalization of tert-butyldimethylsilyloxyacetaldehyde with 3-bromopropanediol, (2) DBN-initiated β-elimination of the resulting 2-(tert-butyldimethylsilyloxy)methyl-4-bromomethyl-1,3-dioxolane 11. Electrophilic glycosidation between 12 and silylated pyrimidine nucleobase proceeded efficiently to provide a mixture of β- and α-anomers of the respective glycosides 14 and 15. Tin radical-mediated reduction of the bromomethyl functional group of 14 and 15 gave protected 4′-C-methyl-dioxorane uracil- 16 and thymine nucleoside 17. The respective cytosine nucleoside 18 was synthesized from 16. De-silylation of 4′-methyl-1′,3′-dioxolane pyrimidine nucleosides 16–18 gave the target molecules. Evaluation of the anti-HIV-1 activity of the β- and α-anomers of the novel 4′-C-methyl-1′,3′-dioxolane nucleosides 22β,α–24β,α revealed that none of the nucleoside derivatives possess anti-viral activity against HIV-1 and show cytotoxicity against MT-4 cells at 100 μM.     

3′-Selective modification of a 4′,5′-didehydro-5′-deoxy-2′,3′-epoxyuridine using nucleophiles

1-(2,3-Anhydro-5-deoxy-4,5-didehydro-α-l-erythro-pent-4-enofuranosyl)uracil 4 was obtained by the treatment of 5′-iodo-2′,3′-epoxyuridine 5 with LiHMDS in excellent yield. The pyrimidine nucleoside 4 possesses quite unique vinyl epoxide moiety within the molecules. The reactions of 4 with a variety of nucleophiles gave 3′-substituted pyrimidine nucleosides without the formation of the corresponding 2′-substituted isomers. In the case of NaN₃ or PhSH, the corresponding 5′-adduct was obtained as a minor product together with the expected 3′-adduct.     

A convenient approach towards boron cluster modifications with adenosine and 2′-deoxyadenosine

New 2′-deoxyadenosine and adenosoine modifications: 8-[(2-dimethylaminoethyl)amino]-2′-deoxyadenosine and 8-[(2-dimethylaminoethyl)amino]adenosine were prepared and their reactivity towards cyclic oxonium adducts of closo-dodecaborate and cobalt-bis-dicarbollide was studied. The cleavage reactions of clusters oxonium rings by N,N-dimethylanio group of modified nucleosides led to the first [B₁₂H₁₂]²⁻ and new [Co(C₂B₉H₁₁)₂]⁻ conjugates with adenosine and 2′-deoxyadenosine respectively. The proposed methodology provides a convenient route for the synthesis of libraries of boron cluster modified adenosine and 2′-deoxyadenosine derivatives for biological screening.     

First synthesis of 2′-oxabicyclo[3.1.0]hexyl nucleosides with a north conformation

The first synthesis of 2′-oxabicyclo[3.1.0]hexyl nucleosides, a novel class of bicyclonucleosides, with a north conformation was successfully accomplished starting from (S)-epichlorohydrin via a tandem alkylation-lactonization, a less steric hindrance-dependent silylation in equilibrium and a coupling reaction with nucleobases under Vorbruggen conditions. Addition of acetic acid prevented a benzoyl group from migrating during desilylation with TBAF. ¹H NMR and X-ray crystallographic analysis indicated that the anomeric effect worked on the β-2′-oxabicyclo[3.1.0]hexyl nucleosides.     

Dinucleotides of 4′-C-vinyl- and 5′-C-allylthymidine as substrates for ring-closing metathesis reactions

Thymidine derivatives containing a 4′-C-vinyl group or a 5′-C-allyl group are synthesized and used as building blocks for three different dinucleotides. These are evaluated as substrates for ring-closing metathesis cyclisations, and a protected 5′-C-allylthymidine homo-dimer is found to be the most reactive. A protected precursor for a conformationally restricted cyclic dinucleotide with a four carbon 5′-C to 5′-C connection is hereby efficiently obtained, whereas a corresponding three carbon 4′-C to 5′-C connection is obtained in a lower yield.     

Synthesis of 2′,3′-dideoxy-6′-fluorocarbocyclic nucleosides via Reformatskii-Claisen rearrangement

2′,3′-Dideoxy-6′-fluorocarbocyclic nucleosides, analogues of highly bioactive carbovir and abacavir were synthesized. The notable steps were the incorporation of fluoromethylene group by way of silicon-induced Reformatskii-Claisen rearrangement of allyl bromofluoroacetate, the construction of the carbocyclic ring via ring-closing metathesis (RCM) and the introduction of base by Mitsunobu reaction.     

Syntheses of 1′,2′-cyclopentyl nucleosides as potential antiviral agents

To discover novel nucleosides as potential antiviral agents, 1′,2′-cyclopentyl nucleosides were designed as hybrids of sofosbuvir and GS-6620. An asymmetric aldol condensation reaction was used as the key transformation to prepare the versatile 1′,2′-cyclopentyl ribose 6, which is useful to explore diverse bases at 1′ and its utility was demonstrated via the syntheses of nucleosides 9 and 11. The 2′-β-methyl-1′,2′-cyclopentyl ribonucleoside scaffold was exemplified via a C-nucleoside which was prepared using a RCM reaction as the key step leading to novel nucleoside 35.     

Phosphoramidite building blocks for efficient incorporation of 2′-O-aminoethoxy(and propoxy)methyl nucleosides into oligonucleotides

A simple and efficient method for the preparation of eight phosphoramidite building blocks for incorporation of 2′-O-(2-aminoethoxymethyl)ribonucleosides and 2′-O-(3-aminopropoxymethyl)ribonucleosides into synthetic oligonucleotides has been developed. The synthetic routes are maximally convergent and provide sufficient amounts of phosphoramidites for several solid-phase synthesis coupling reactions. Using acyclic derivatives 17a,b the overall yields of phosphoramidites 2 and 3 were increased up to 50% for pyrimidine nucleosides and up to 30% for purine derivatives with substantial decrease of total reaction steps. The 2′-O-substituent was found to be stable during oligonucleotide synthesis. The resulting oligonucleotides are of particular interest for post-synthetic functionalization and conjugation.     

Synthesis of cyclic bis(3′-5′)diguanylic acid (c-di-GMP) analogs

This paper reports the synthesis of cyclic bis(3′-5′)diguanylic acid (c-di-GMP) analogs, including the monophosphorothioic acid of c-di-GMP (c-GpGps), cyclic bis(3′-5′)guanylic/adenylic acid (c-GpAp), and cyclic bis(3′-5′)guanylic/inosinic acid (c-GpIp). These compounds are expected to be important, both in elucidating the mechanism of bioactive c-di-GMP and in designing and creating new bioactive c-di-GMP-related artificial derivatives.     

Scalable synthesis of substituted 2,7-dimethyl-9-phenylxanthen-9-ol (DMPx-OH): useful for the preparation of crystalline 5′-O-DMPx-protected nucleosides

A convenient single-step synthesis of several 2,7-dimethyl-9-phenylxanthen-9-ol (DMPx-OH) analogs has been accomplished using a Friedel-Crafts reaction. Treatment of various DMPx-OH with unprotected 2′-O-methoxyethyl-ribonucleosides (MOE) in the presence of B(C₆F₅)_3, as a Lewis Acid catalyst, furnished 5′-O-protected derivatives of 2′-MOE-ribonucleosides in good yields. The deprotection of the DMPx groups was accomplished by acid hydrolysis under very mild conditions. Among the five DMPx analogs synthesized, the 2,7-dimethyl-9-(4-nitrophenyl)xanthene-9-yl group furnished crystalline products enabling non-chromatographic isolation of 5′-O-protetced nucleosides.     

Novel and efficient regioselective enzymatic approach to 3′-, 5′- and 3′,5′-di-O-crotonyl 2′-deoxynucleoside derivatives

Regioselective syntheses of several O-crotonyl 2′-deoxynucleoside derivatives have been efficiently achieved using a biocatalytic methodology. While Candida antarctica lipase B (CAL-B) afforded the 5′-O-acylated compounds, immobilized lipase from Pseudomonas cepacia (PSL-C) provided the 3′-O-crotonylated analogs. Since classical chemical approaches did not work appropriately due to side isomerization reactions, a mixture of both lipases was used to achieve a useful synthetic route toward diacylated nucleosides.     

Synthesis of 3′-deoxy-3′-C-methyl-β-d-ribonucleoside analogs

3′-Deoxy-3′-C-methyl-β-d-ribonucleoside analogs bearing the five canonical bases of nucleic acids have been synthesized. All these derivatives were prepared by glycosylation of the corresponding heterocyclic bases with a suitable peracylated 3-C-methyl sugar precursor. The synthesis of the 3-C-methyl sugar precursor is described following a new stereoselective synthetic pathway.     

Practical synthetic routes to carbon-substituted nucleosides

Robust practical routes to three different carbon-substituted nucleosides are described. Short synthetic procedures for preparing 1′-homonucleosides, 2′-C-methyl-carbanucleosides and 2′-C-methyl-cyclopropyl/cyclopentyl fused bicyclic ribose analogues are described starting from readily available building blocks.