Bicyclic Nucleoside Research Articles

Synthesis of a Novel Bicyclic Nucleoside with a 3,7-Anhydrooctofuranosyl Skeleton

We have developed an efficient route for the synthesis of a novel bicyclic nucleoside using a key intermediate 13, which was prepared from 1,2:5,6-di-O-isopropylidene-α-d-glucose. 1,2-Nucleophilic addition of aldehyde 8 with allyltrimethylsilane and intramolecular Williamson reaction were successfully achieved to synthesize an intermediate with a 3,7-anhydrooctofuranosyl skeleton.

Synthesis of Conformationally Constrained 2’-N,4’-C-Ethylene-Bridged Adenosine (aza-ENA-A)

This thesis is based on six original research publications describing synthesis, structure and physicochemical and biochemical analysis of chemically modified oligonucleotides (ONs) in terms of their potential diagnostic and therapeutic applications. Synthesis of two types of bicyclic conformationally constrained nucleosides, North-East locked 1',2'-azetidine and North locked 2',4'-aza-ENA, is described. Study of the molecular structures and dynamics of bicyclic nucleosides showed that depending upon the type of fused system they fall into two distinct categories with their respective internal dynamics and type of sugar conformation. The physicochemical properties of the nucleobases in the conformationally constrained nucleosides found to be depended on the site and ring-size of the fused system. The incorporation of azetidine modified nucleotide units into 15mer ONs lowered the affinity toward the complementary RNA. However, they performed better than previously reported isosequential 1',2'-oxetane modified analogues. Whereas aza-ENA-T modification incorporated into ONs significantly enhanced affinity to the complementary RNA. To evaluate the antisense potential of azetidine-T and aza-ENA-T modified ONs, they were subjected to RNase H promoted cleavage as well as tested towards nucleolytic degradation. Kinetic experiments showed that modified ONs recruit RNase H, however with lower enzyme efficiency due to decreased enzyme-substrate binding affinity, but with enhanced turnover number. Both, azetidine-T and aza-ENA-T modified ONs demonstrated improved 3'-exonuclease stability in the presence of snake venom phosphodiesterase and human serum compared to the unmodified sequence. Oligodeoxynucleotides (ODNs) containing pyrene-functionalized azetidine-T (Aze-pyr X) and aza-ENA-T (Aza-ENA-pyr Y) modifications showed different fluorescence properties. The X modified ODNs hybridized to the complementary DNA and RNA showed variable increase in the fluorescence intensity depending upon the nearest-neighbor at the 3'-end to X modification (dA > dG > dT > dC) with high fluorescence quantum yield. However, the Y modified ODNs showed a sensible enhancement of the fluorescence intensity only with complementary DNA. Also, the X modified ODN showed decrease (~37-fold) in the fluorescence intensity upon duplex formation with RNA containing a G nucleobase mismatch opposite to the modification site, whereas a ~3-fold increase was observed for the Y modified probe.

Selective biocatalytic deacylation studies on furanose triesters: a novel and efficient approach towards bicyclonucleosides

Lipozyme TL IM catalyses the deacylation of 4-C-acyloxymethyl-3,5-di-O-acyl-1,2-O-(1-methylethylidene)-beta-L-threo-pentofuranose to form 3,5-di-O-acyl-4-C-hydroxymethyl-1,2-O-(1-methylethylidene)-alpha-D-xylo-pentofuranose in a highly selective and efficient manner. The rate of lipase-catalyzed deacylation of tributanoyl furanose is 2.3 times faster than the rate of deacylation of the triacetyl furanose derivative. In order to confirm the structure of the lipase-catalyzed deacylated product, it was converted to a bicyclic sugar derivative, which can be used for the synthesis of bicyclic nucleosides of importance in the development of novel antisense and antigene oligonucleotides. Further, it has been established that the monohydroxy product of the lipase-catalyzed reaction is the result of selective deacylation of the 4-C-acyloxymethyl function in the substrate and not of any acyl migration process.

Synthesis and Biological Evaluation of Bicyclic Nucleosides as Inhibitors of M. tuberculosis Thymidylate Kinase

Herein we describe the synthesis and conformational analysis of a series of bicyclic thymidine derivatives and their evaluation as inhibitors of thymidine monophosphate kinase from Mycobacterium tuberculosis (TMPKmt), based on previously discovered bicyclic sugar nucleosides. With a K(i) value of 2.3 microm, 1-[3-aminomethyl-3,5-dideoxy-2-O,6-N-(thiocarbonyl)-beta-D-ribofuranosyl]thymine emerged as the most potent TMPK inhibitor of this series. Moreover, this promising compound displays inhibitory potency against Mycobacteria cultures with an IC(99) value of 100 microg mL(-1), thus promoting TMPKmt for the first time as a validated target for further inhibitory design. Attempts to rationalise the observed structure-activity relationship (SAR) involving molecular modelling and conformational analysis are described.

Analogues of a Locked Nucleic Acid with Three-Carbon 2‘,4‘-Linkages: Synthesis by Ring-Closing Metathesis and Influence on Nucleic Acid Duplex Stability and Structure

Two bicyclic 2'-deoxynucleoside analogues containing a saturated and an unsaturated three-carbon 2',4'-linkage, respectively, have been synthesized using a ring-closing metathesis-based linear strategy starting from uridine. Both analogues have been incorporated into oligodeoxynucleotide sequences and increased the stability of DNA:RNA hybrid duplexes (DeltaT(m) approximately 2.5-5.0 degrees C per modification) and decreased the stability of dsDNA duplexes (DeltaT(m) approximately 2.5-1.0 degrees C per modification). CD spectroscopy revealed that the bicyclic nucleosides induced formation of A-type-like duplexes albeit to a lesser degree than found for locked nucleic acid (LNA) monomers. From the CD data and UV melting analysis, we propose that the 2'-oxygen atom of the bicyclic moiety is essential for the formation of stabilized A-type-like dsDNA but not for the formation of a stabilized A-type DNA:RNA hybrid.

Synthesis and Hybridization Studies of 2‘-Amino-α-L-LNA and Tetracyclic “Locked LNA”

A convergent route to a new class of locked nucleic acids, i.e., 2'-amino-alpha-L-LNA, has been developed. The optimized synthetic route to the corresponding phosphoramidite building block of thymine proceeds in 4% overall yield over 15 steps from the starting diol. Crucial synthetic steps include (a) introduction of a C2-azido group prior to nucleobase coupling, (b) Vorbrüggen glycosylation primarily affording the desired alpha-anomer, (c) separation of alpha-L-ribo- and beta-L-ribo-configured bicyclic nucleosides, and (d) selection of a suitable protecting group to avoid intramolecular Michael addition of the C2'-amino group onto the C6-position. Incorporation of a 2'-amino-alpha-L-LNA monomer into oligodeoxyribonucleotides results in modest changes in thermal stability with complementary DNA, whereas significant increases in thermal stability are observed with RNA complements along with excellent Watson-Crick discrimination. These results, along with the flexibility of the synthetic strategy allowing chemoselective N2'-functionalization at a late stage, render 2'-amino-alpha-L-LNA a promising building block for nucleic acid based nanobiotechnology and therapeutics. A slight modification in strategy facilitated the synthesis of the corresponding phosphoramidite building blocks of Michael adducts, which due to their tetracyclic skeletons exhibit a conformationally restricted furanose ring and glycosidic torsion angle (anti-range). Incorporation of such a "locked LNA" monomer into oligodeoxyribonucleotides results in large decreases in thermal affinity toward DNA/RNA complements.

Mutations Distal to the Substrate Site Can Affect Varicella Zoster Virus Thymidine Kinase Activity: Implications for Drug Design

Varicella zoster virus encodes a thymidine kinase responsible for the activation of antiherpetic nucleoside prodrugs such as acyclovir. In addition, herpes virus thymidine kinases are being explored in gene/chemotherapy strategies aimed at developing novel antitumor therapies. To investigate and improve compound selectivity, we report here structure-based site-directed mutagenesis studies of varicella zoster virus thymidine kinase (VZVTK). Earlier reports showed that mutating residues at the core of the VZVTK active site invariably destroyed activity; hence, we targeted more distal residues. Based on the VZVTK crystal structure, we constructed six mutants (E59S, R84V, H97Y/A, and Y21H/E) and tested substrate activity and competitive inhibition for several compound series. All VZVTK mutants tested retained significant phosphorylation activity with dThd as substrate, apart from Y21E (350-fold diminution in the k(cat)/K(m)). Some mutations give slightly improved affinities: bicyclic nucleoside analogs (BCNAs) with a p-alkyl-substituted phenyl group seem to require aromatic ring stacking interactions with residue 97 for optimal inhibitory effect. Mutation Y21E decreased the IC(50) value for the BCNA 3-(2'-deoxy-beta-D-ribofuranosyl)-6-octyl-2,3-dihydrofuro[2,3-d]pyrimidin-2-one (Cf1368) 4-fold, whereas mutation Y21H increased the IC(50) value by more than 15-fold. These results suggest that residue 21 is important for BCNA selectivity and might explain why HSV1TK is unable to bind BCNAs. Other mutants, such as the E59S and R84V thymidine kinases, which in wild-type VZVTK stabilize the dimer interface, give opposite results regarding the level of sensitivity to BCNAs. The work described here shows that distal mutations that affect the VZVTK active-site may help in the design of more selective substrates for gene suicide therapy or as anti-varicella zoster virus drugs.

Bicyclic Nucleoside Inhibitors of Varicella—Zoster Virus: The Effect of Branching in the p‐Alkylphenyl Side Chain.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Efficient Metathesis Route to the B‐Ring of Eleutherobin and Other Medium‐Sized Cyclic Ethers.

A short and efficient RCM route is reported for the synthesis of the B-ring of eleutherobin and other medium-sized cyclic ethers from readily available 1,2,5,6-diisopropylidene-D-glucose. This strategy is successfully extended to the synthesis of a few bicyclic ethers, which may find applications in the synthesis of novel bicyclic nucleosides.

Bicyclic nucleoside inhibitors of Varicella–Zoster virus: The effect of branching in the p-alkylphenyl side chain

Further to the discovery of bicyclic furanopyrimidine nucleoside analogues (BCNAs) as potent anti-VZV agents, a branched series of this family of compounds was synthesised. The aim was to study the impact of the geometry and steric hindrance in the side chain as well as lipophilic role of this moiety on biological activity. The results showed a detrimental effect of branching on antiviral activity, with a different magnitude depending on the position of branching in the side chain. This study again showed the importance of this moiety for biological activity, as well as the limited efficacy of the C log P value as a tool for predicting the potency of BCNAs, while suggesting an alternative rationale behind the design of future series.

Efficient metathesis route to the B-ring of eleutherobin and other medium-sized cyclic ethers

A short and efficient RCM route is reported for the synthesis of the B-ring of eleutherobin and other medium-sized cyclic ethers from readily available 1,2,5,6-diisopropylidene- d-glucose. This strategy is successfully extended to the synthesis of a few bicyclic ethers, which may find applications in the synthesis of novel bicyclic nucleosides.

An α-d-Configured Bicyclic Nucleoside Restricted in an E-type Conformation: Synthesis and Parallel RNA Recognition

An alpha-D-arabino configured bicyclic nucleoside strongly restricted in an E-type conformation by a 2'-3'-fused oxetane ring is synthesized. Several synthetic strategies toward the target compound are described, and the successful preparation from a D-xylose derivative is based on a ruthenium-mediated cleavage of a double bond, an S(N)2-inversion at the 2-position to give an arabino-configuration, nucleobase coupling, and finally ring closure to give the oxetane ring. The E-type conformation is confirmed by molecular modeling and NMR. The nucleoside is incorporated into short alpha-DNA sequences. In a mixed pyrimidine context, these recognize complementary parallel RNA-sequences with mainly increased affinity and complementary parallel DNA-sequences with decreased affinity. The present bicyclic analogue represents the first conformationally restricted alpha-DNA-analogue to improve nucleic acid recognition in mixmers with alpha-DNA monomers.

Bicyclic nucleoside analogues from d-glucose: synthesis of chiral as well as racemic 1,4-dioxepane ring-fused derivatives

The dioxepanofuranose derivatives 4 and 12, obtained through the cyclization of the 3-(2-hydroxyethyl) ether of a d- xylo-pentodialdose derivative, were appropriately functionalized and elaborated to the first examples of the new class of 3′- O and 5′- O-bicyclic nucleoside analogues 9, 10, and 14 with a fused seven-membered ring. Reactions carried out through the intermediacy of the d- xylo-pentodialdose derivative 5 yielded racemic products, while prior protection of the 4-formyl group (as in 7) before deprotection of the 1,2-hydroxyl groups led to optically active analogues.

Diastereoselective Reformatsky Reaction of Methyl 4‐Bromocrotonate with 1,2:5,6‐Di‐O‐isopropylidene‐α‐D‐ribo‐hexofuranose‐3‐ulose: Application to Novel Bicyclic Nucleosides.

ChemInformVolume 36, Issue 10 Natural Products Diastereoselective Reformatsky Reaction of Methyl 4-Bromocrotonate with 1,2:5,6-Di-O-isopropylidene-α-D-ribo-hexofuranose-3-ulose: Application to Novel Bicyclic Nucleosides. Mukund K. Gurjar, Mukund K. Gurjar Natl. Chem. Lab., Pune 411 008, IndiaSearch for more papers by this authorDandepally Srinivasa Reddy, Dandepally Srinivasa Reddy Natl. Chem. Lab., Pune 411 008, IndiaSearch for more papers by this authorMohan M. Bhadbhade, Mohan M. Bhadbhade Natl. Chem. Lab., Pune 411 008, IndiaSearch for more papers by this authorRajesh G. Gonnade, Rajesh G. Gonnade Natl. Chem. Lab., Pune 411 008, IndiaSearch for more papers by this author Mukund K. Gurjar, Mukund K. Gurjar Natl. Chem. Lab., Pune 411 008, IndiaSearch for more papers by this authorDandepally Srinivasa Reddy, Dandepally Srinivasa Reddy Natl. Chem. Lab., Pune 411 008, IndiaSearch for more papers by this authorMohan M. Bhadbhade, Mohan M. Bhadbhade Natl. Chem. Lab., Pune 411 008, IndiaSearch for more papers by this authorRajesh G. Gonnade, Rajesh G. Gonnade Natl. Chem. Lab., Pune 411 008, IndiaSearch for more papers by this author First published: 08 February 2005 https://doi.org/10.1002/chin.200510200Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat No abstract is available for this article. Volume36, Issue10March 8, 2005 RelatedInformation

NMR conformational analysis of p-tolyl furanopyrimidine 2′-deoxyribonucleoside and crystal structure of its 3′,5′-di- O-acetyl derivative

The conformation of a representative molecule of a new, potent class of antiviral-active modified nucleosides is determined. A bicyclic nucleoside, 3-(2′-deoxy-β- d-ribofuranosyl)-6-(4-methylphenyl)-2,3-dihydrofuro[2,3- d]pyrimidin-2-one, shows C2′- endo and C3′- endo ribose conformations in solution (63:37, 37 °C; DMSO- d 6), as determined by 1H NMR studies. The crystal structure of a 3′,5′-di- O-acetyl-protected derivative (monoclinic, P2 1, a/ b/ c = 6.666(1)/12.225(1)/24.676(2) Å, β = 90.24(1)°, Z = 4) shows exclusively C2′- endo deoxyribose puckering. The base is found in the anti position both in solution and in crystalline form.

Synthesis and thermal denaturation studies of novel 2′-O,3′-C-linked bicyclic oligonucleotides with a methoxy or a piperazino group facing the major groove of nucleic acid duplexes

With the aim of evaluating duplex stabilities of oligonucleotides (ONs) with major groove facing functionalities, two novel 2'-O,3'-C-linked bicyclic nucleoside phosphoramidite building blocks were synthesized by routes involving regioselective O-methylation or piperazine attachment using carbonyldiimidazole coupling chemistry. The novel monomers were incorporated into 9-mer mixed base ONs and the thermal stability toward complementary single stranded DNA and RNA was evaluated by thermal denaturation experiments. O-methylated ONs confirmed the applicability of the functionalized bicylic sugar unit for attachment of groups facing the major groove and satisfactory binding properties towards the RNA complement were observed. For the piperazino modified ONs, experiments were performed in aqueous buffers with low (40 mM) and medium (110 mM) salt concentrations, at pH 5 and pH 7. A change from a medium to a low salt concentration induced a significant relative increase in the thermal stability of modified duplexes toward both DNA and RNA complements, which suggests protonation of the piperazino group under the experimental conditions applied.

Bicyclic Nucleoside Inhibitors of Varicella-Zoster Virus Modified on the Sugar Moiety: 3′ and 5′ Derivatives

Bicyclic furanopyrimidine nucleoside analogues (BCNAs) have been previously reported as potent and selective anti-varicella-zoster virus (VZV) agents. Few modifications on the sugar moiety have been considered so far but some of them have shown interesting activity against human cytomegalovirus (HCMV) while losing activity against VZV. In addition, recent work has led to an entirely new series of anti-HCMV bicyclic furopyrimidine agents, acting through a non-nucleoside mechanism. In order to further investigate structure-activity relationship studies on the sugar moiety, some 3'- and 5'-chloro derivatives and 5'-deoxygenated derivatives have been synthesized. The lack of anti-VZV activity of the 5'-modified derivatives is further proof of a mechanism of action involving VZV thymidine kinase (TK)-mediated phosphorylation. Similarly, the replacement of the 3'-OH with chlorine showed a decrease of antiviral activity, which can be correlated to the lack of interaction with VZV TK as demonstrated by enzyme assays. These results confirm free 5'-OH and 3'-OH as necessary requirements for efficient recognition by VZV TK and for potent anti-VZV activity in cell culture.

New Ruthenium‐Based Protocol for Cleavage of Terminal Olefins to Primary Alcohols: Improved Synthesis of a Bicyclic Nucleoside.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Discovery of Bicyclic Thymidine Analogues as Selective and High-Affinity Inhibitors of Mycobacterium tuberculosis Thymidine Monophosphate Kinase

Thymidine monophosphate kinase of Mycobacterium tuberculosis (TMPKmt) represents an attractive target for selectively blocking bacterial DNA synthesis. Hereby, we report on the discovery of a novel class of bicyclic nucleosides (10 and 11) and one dinucleoside (12), belonging to the most selective inhibitors of TMPKmt discovered so far.

Diastereoselective Reformatsky reaction of methyl 4-bromocrotonate with 1,2:5,6-di- O-isopropylidene-α- d- ribo-hexofuranos-3-ulose: application to novel bicyclic nucleosides

This paper describes an efficient synthetic route for novel bicyclic nucleosides. The stereochemistry of the targeted bicyclic nucleosides was successfully achieved by vinylogous Reformatsky reaction and ring closing metathesis reaction on a carbohydrate backbone.