- Towards Zwitterionic Oligonucleotides with Improved Properties: the NAA/LNA-Gapmer Approach
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Oligonucleotides (ON) are promising therapeutic candidates, for instance by blocking endogenous mRNA (antisense mechanism). However, ON usually require structural modifications of the native nucleic acid backbone to ensure satisfying pharmacokinetic properties. One such strategy to design novel antisense oligonucleotides is to replace native phosphate diester units by positively charged artificial linkages, thus leading to (partially) zwitterionic backbone structures. Herein, we report a “gapmer” architecture comprised of one zwitterionic central segment (“gap”) containing nucleosyl amino acid (NAA) modifications and two outer segments of locked nucleic acid (LNA). This NAA/LNA-gapmer approach furnished a partially zwitterionic ON with optimised properties: i) the formation of stable ON-RNA duplexes with base-pairing fidelity and superior target selectivity at 37 °C; and ii) excellent stability in complex biological media. Overall, the NAA/LNA-gapmer approach is thus established as a strategy to design partially zwitterionic ON for the future development of novel antisense agents.
- Wojtyniak, Melissa,Schmidtgall, Boris,Kirsch, Philine,Ducho, Christian
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p. 3234 - 3243
(2020/09/01)
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- A new and convenient approach for the preparation of β-cyanoethyl protected trinucleotide phosphoramidites
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Herein we report a convenient approach for the preparation of fully protected trinucleotide synthons to be used for the synthesis of gene libraries. The trinucleotide synthons bear β-cyanoethyl groups at the phosphate residues, and thus can be used in standard oligonucleotide synthesis without additional steps for deprotection and work-up.
- Janczyk, Matthaeus,Appel, Bettina,Springstubbe, Danilo,Fritz, Hans-Joachim,Mueller, Sabine
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supporting information; experimental part
p. 1510 - 1513
(2012/03/22)
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- Pyrimidine-purine and pyrimidine heterodinucleosides synthesis containing a triazole linkage
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This article describes a synthetic route to generate two purine-pyrimidine and pyrimidine heterodinucleosides. Both microwave activated regioselective alkylation using hydride and copper-catalyzed-azide-alkyne-cycloaddition (CuAAC) were used in order to perform the synthesis.
- Lucas,Elchinger,Faugeras,Zerrouki
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scheme or table
p. 168 - 177
(2011/08/02)
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- Chelation-controlled regioselective alkylation of pyrimidine 2′-deoxynucleosides
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Protection-deprotection steps, which are usually needed for regioselective alkylation of pyrimidine deoxynucleosides, can be avoided by choosing the appropriate solvent. The combined effects of low dielectric constant and possible sodium chelation by pyrimidine nucleosides may account for the unexpected regioselectivity observed in THF.
- Lucas,Teste,Zerrouki,Champavier,Guilloton
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scheme or table
p. 199 - 207
(2010/03/23)
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- α,β-Difluoromethylene deoxynucleoside 5′-triphosphates: A convenient synthesis of useful probes for DNA polymerase β structure and function
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αβ-Difluoromethylene deoxynucleoside 5′-triphosphates (dNTPs, N = A or C) are advantageously obtained via phosphorylation of corresponding dNDP analogues using catalytic ATP, PEP, nucleoside diphosphate kinase, and pyruvate kinase. DNA pol β Kdd values for the α, β-CF2 and unmodified dNTPs, α, β-NH dUTP, and the α, β-CH2 analogues of dATP and dGTP are discussed in relation to the conformations of α, β-CF2 dTTP versus α, β-NH dUTP bound into the enzyme active site.2009 American Chemical Society.
- Upton, Thomas G.,Kashemirov, Boris A.,McKenna, Charles E.,Goodman, Myron F.,Prakash, G. K. Surya,Kultyshev, Roman,Batra, Vinod K.,Shock, David D.,Pedersen, Lars C.,Beard, William A.,Wilson, Samuel H.
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supporting information; experimental part
p. 1883 - 1886
(2009/10/10)
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- DNG cytidine: Synthesis and binding properties of octameric guanidinium-linked deoxycytidine oligomer
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The synthesis of guanidinium-linked cytidyl oligomer (DNG-C8), a cationic DNA analog, and the corresponding cytidine monomers is described. The DNG monomer synthesis was streamlined to produce a shorter route to the final monomer than previously reported for thymidine and subsequent solid-phase synthesis produced an octameric cytidyl DNG strand. Because octameric deoxyguanosine would be used as the complementary strand in our studies, it was necessary to investigate guanosine self-association. Singular value decomposition was used to mathematically deconvolve the spectral data and confirm the presence of transitions due to DNA-G8 self-association. Job plots show the binding stoichiometry of DNG-C8 with DNA-G 8 to be 1:1. Thermal denaturation studies of the DNG-C 8·DNA-G8 duplex established a T m≥90°C and a ΔG°=-13.3kcalmol-1, indicating the DNG-C8·DNA-G8 duplex is over 1000 times more stable than that of DNA-C8·DNA-G8.
- Szabo, Istvan E.,Bruice, Thomas C.
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p. 4233 - 4244
(2007/10/03)
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- An improved transient method for the synthesis of N-benzoylated nucleosides
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The Jones' transient method for the synthesis of N-benzoylated nucleosides is improved by reducing the amounts of chlorotrimethylsilane (TMSCl) and benzoyl chloride to nearly equivalent quantities. The easy work-up and high yields of products are the major advantages of this approach. Jones' method is further simplified by omitting the addition of ammonium hydroxide. The utility of this modification for the preparation of some useful protected nucleosides is also presented.
- Zhu, Xue-Feng,Williams Jr., Howard J.,Scott, A. Ian
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p. 1233 - 1243
(2007/10/03)
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- Microwave assisted high yielding preparation of N-protected 2′-deoxyribonucleosides useful for oligonucleotide synthesis
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A rapid and high yielding method for the synthesis of precursors of synthons for DNA synthesis, N-protected 2′-deoxyribonucleosides is described, which occur under mild conditions using microwave irradiation. The desired material, N-protected nucleosides, was obtained in 93-96% yield in few minutes. The final products were then characterized by H-NMR and MALDI-TOF and compared with the standard samples. The method is amenable to small to moderate scale of synthesis.
- Rao,Kumar,Chauhan,Garg,Gupta
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p. 393 - 400
(2007/10/03)
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- Building blocks for the solution phase synthesis of oligonucleotides: Regioselective hydrolysis of 3′,5′-di-O-levulinylnucleosides using an enzymatic approach
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A short and convenient synthesis of 3′- and 5′-O-levulinyl-2′-deoxynucleosides has been developed from the corresponding 3′,5′-di-O-levulinyl derivatives by regioselective enzymatic hydrolysis, avoiding several tedious chemical protection/deprotection steps. Thus, Candida antartica lipase B (CAL-B) was found to selectively hydrolyze the 5′-levulinate esters, furnishing 3′-O-levulinyl-2′-deoxynucleosides 3 in >80% isolated yields. On the other hand, immobilized Pseudomonas cepacia lipase (PSL-C) and Candida antarctica lipase A (CAL-A) exhibit the opposite selectivity toward the hydrolysis at the 3′-position, affording 5′-O-levulinyl derivatives 4 in >70% yields. A similar hydrolysis procedure was successfully extended to the synthesis of 3′- and 5′-O-levulinyl-protected 2′-O-alkylribonucleosides 7 and 8. This work demonstrates for the first time application of commercial CAL-B and PSL-C toward regioselective hydrolysis of levulinyl esters with excellent selectivity and yields. It is noteworthy that protected cytidine and adenosine base derivatives were not adequate substrates for the enzymatic hydrolysis with CAL-B, whereas PSL-C was able to accommodate protected bases during selective hydrolysis. In addition, we report an improved synthesis of dilevulinyl esters using a polymer-bound carbodiimide as a replacement for dicyclohexylcarbodiimide (DCC), thus considerably simplifying the workup for esterification reactions.
- Garcia, Javier,Fernandez, Susana,Ferrero, Miguel,Sanghvi, Yogesh S.,Gotor, Vicente
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p. 4513 - 4519
(2007/10/03)
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- Selective cleavage of O-(dimethoxytrityl) protecting group with sodium periodate
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Sodium periodate in aqueous organic solvents selectively removes, under mild reaction conditions, the O-(dimethoxytrityl) protecting group. Selectivity of the cleavage was studied using the nucleoside derivatives protected by various types of groups commonly used in nucleoside and nucleotide chemistry.
- Rejman, Dominik,Kralikova, Sarka,Tocik, Zdenek,Liboska, Radek,Rosenberg, Ivan
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p. 502 - 508
(2007/10/03)
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- Unexpected shift to a 4-imino tautomer upon N4-acylation of 5- methylcytosin-1-yl nucleoside analogues
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In contrast with the behaviour of 5-unsubstituted cytosine nucleoside analogues, 5-methylcytosine derivatives show upon N4-benzoylation, commonly used as base protection in oligonucleotide synthesis, a tautomeric change of the base moiety from a 4-amino- into a 4-imino isomer. In the latter form, which is easily diagnosticized by 13C NMR and confirmed by X-ray data, the compounds seem to be hydrolytically less stable.
- Busson,Kerremans,Van Aerschot,Peeters,Blaton,Herdewijn
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p. 1079 - 1082
(2007/10/03)
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- The S-pixyl group: An efficient photocleavable protecting group for the 5' hydroxy function of deoxyribonucleosides
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The 9-phenylthioxanthyl (S-pixyl or S-Px) group has been investigated as a photocleavable protecting group for primary alcohols, and specifically as a 5' hydroxy protecting group for deoxyribonucleosides. Several alcohols, including the four nucleosides with protected exocyclic amino functions, were protected in very good to excellent yield by treatment of 9-chloro-9-phenylthioxanthene 3 in dry pyridine to reveal the derivatized compounds. Irradiation of the protected substrates in neutral, aqueous solution regenerated the starting alcohols in excellent yield.
- Coleman, Michael P.,Boyd, Mary K.
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p. 7911 - 7915
(2007/10/03)
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- Deuterated nucleosides
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Deuterated nucleotide and nucleoside units are used to synthesize strands of RNA and DNA wherein one unit is non-deuterated, with adjacent units deuterated. The deuteration is sufficiently high in order that resonance from the deuterated units do not overlap with resonances from non-deuterated units in NMR-experiments. Thereby a "NMR"-window is created in the DNA or RNA strands, which can be beneficially used for advanced studies of structure versus biological activity in DNA and RNA.
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- Solid-phase synthesis of branched RNA and branched DNA/RNA chimeras
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An effective method for synthesising branched oligonucleotides on solid phase in the 5' to 3' direction has been developed. Special branch-point monomers enable the synthesis of branched oligonucleotides which can have sequences of different length and base composition attached to the 2'- and 3'-hydroxyl groups of the branch point ribonucleoside. The branched oligonucleotides are assembled on commercial DNA synthesisers, the crude products are readily purified by reversed phase HPLC and the fully deprotected products are conveniently analysed by mass spectrometry.
- Grotli, Morten,Eritja, Ramon,Sproat, Brian
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p. 11317 - 11346
(2007/10/03)
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- Nucleoside 5'-methylene phosphonates
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Novel oligonucleotides analogs and nucleoside analogs as well as methods for their synthesis are described. The oligonucleotides are useful in diagnostic and therapeutic applications. The oligonucleotides are stable to nuclease degradation.
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- 1,1,1,3,3,3-Hexafluoro-2-propanol for the Removal of the 4,4'-Dimethoxytrityl Protecting Group from the 5'-Hydroxyl of Acid-Sensitive Nucleosides and Nucleotides
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1,1,1,3,3,3-Hexafluoro-2-propanol is introduced as a suitable reagent and solvent for the detritylation of 5'-O-(4,4'-dimethoxytrityl)-nucleosides and -deoxy- nucleosides, especially those that are susceptible to N-glycosyl cleavage under more strongly acidic conditions.
- Leonard, Nelson J.,Neelima
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p. 7833 - 7836
(2007/10/02)
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- General Approach to the Synthesis of Specifically Deuterium-Labeled Nucleosides
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Starting from D-(-)-ribose, a set of synthetic routes sharing common intermediates has been developed and exemplified in -, -, -, (5'R)--, and (5'S)--N4-benzoylcytidine and, by deoxygenation, their corresponding 2'-deoxynucleosides.These syntheses provide convenient access to millimolar quantities of deuterium/tritium-labeled natural or unnatural nucleosides for direct use or automated oligonucleotide synthesis.
- Voss, James J. De,Hangeland, Jon J.,Townsend, Craig A.
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p. 2715 - 2723
(2007/10/02)
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- Synthesis of oligonucleotide phosphorodithioates
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The synthesis of DNA containing sulfur at the two nonbonding internucleotide valencies is reported. Several different routes using either tervalent or pentavalent mononucleotide synthons are described.
- Beaton,Brill,Grandas,Ma,Nielsen,Yau,Caruthers
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p. 2377 - 2388
(2007/10/02)
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- Method for producing an active protein
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An active protein can be easily, safely produced by a method comprising providing a first peptide fragment having a first amino acid sequence corresponding to part of an active protein and a second peptide fragment having a second amino acid sequence corresponding to the remaining part of the active protein, at least one of said first peptide fragment and said second peptide fragment being one which has been obtained by recombinant DNA technique or has been obtained by a method comprising producing a predetermined peptide fragment by recombinant DNA technique and deleting from or adding to said predetermined peptide fragment at its N-terminus at least one amino acid residue; and linking said first peptide fragment at its C-terminus to said second peptide fragment at its N-terminus. The method of the present invention may be practiced, with further advantages, by predeterming said first peptide fragment and said second peptide fragment so that a first occurring methionine residue subsequent to the N-terminal amino acid residue of the active protein constitutes the N-terminal amino acid of the amino acid sequence of said second peptide fragment, or so that an amino acid residue positioned near the first occurring methionine residue subsequent to the N-terminal amino acid residue of the desired protein on the side of the N-terminus of the desired protein constitutes the N-terminal amino acid residue of said second peptide fragment.
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- Transient Protection: Efficient One-Flask Syntheses of Protected Deoxynucleosides
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Application of the concept of transient protection to the synthesis of protected deoxynucleosides is described.The deoxynucleosides are first treated with trimethylchlorosilane in pyridine for protection of the hydroxyl groups, and then immediately reacted with an acylating agent - benzoyl chloride for 1a and 1b and isobutyric anhydride for 1c - to effect N-acylation.Hydrolysis of the trimethylsilyl groups takes a few hours in aqueous pyridine or a few minutes with dilute ammonia.The ammonia also effects selective hydrolysis of the initially formed N,N-dibenzoyldeoxyadenosine derivative (3a) to the desired N-benzoyldeoxyadenosine (4a).This one-flask procedure gives crystalline N-acyl deoxynucleosides 4a and 4b in 95percent yield and 4c in 75percent yield, in only a few hours.The 5'-O-dimethoxytrityl deoxynucleosides 8a and 8b are also obtained in a one-flask procedure by initial reaction of the deoxynucleosides with 4,4'-dimethoxytrityl chloride, followed by treatment with trimethylchlorosilane and then benzoyl chloride.Although with deoxycytidine some of the 4-N,5'-O-bis(dimethoxytrityl) derivative (5c) is formed, benzoyl chloride effects conversion to the 4-N-benzoyl derivative (7b).After simple purification by flash chromatography 8a and 8b are each obtained in 80-90percent overall yield from 1a or 1b.
- Ti, G. S.,Gaffney, B. L.,Jones, R. A.
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p. 1316 - 1319
(2007/10/02)
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- Simplified Liquid-Phase Preparation of Four Decadeoxyribonucleotides and their Preliminary Spectroscopic Characterization
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Four self-complementary decadeoxyribonucleotides, dApTpApTpApTpApTpApT,dApTpApTpCpGpApTpApT, dApTpApTpGpCpApTpApT and dApApApApApTpTpTpTpT, were chemically synthesized by the phosphotriester procedure.Efforts were concentrated on keeping the procedure as simple as possible, concomitant with obtaining high-purity products at each step of the process.The decamers were elaborated from the 3'-end, starting with a 3'-O-benzoylated monomer according to the scheme: monomer + monomer -> dimer + dimer -> tetramer + dimer -> hexamer + tetramer -> decamer.Putity of intermediates and of the fully blocked decamers were monitored by chromatography and by 300-MHz 1H-NMR. spectroscopy.The deblocked decadeoxyribonucleotides were characterized by their UV., CD., and 1H-NMR. spectra.
- Denny, William A.,Leupin, Werner,Kearns, David R.
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p. 2372 - 2393
(2007/10/02)
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- N-ACYL PROTECTING GROUPS FOR DEOXYNUCLEOTSIDES A QUANTITATIVE AND COMPARATIVE STUDY
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A detailed study in search for suitable N-acyldeoxy-nucleosides which could serve as building blocks for the stepwise synthesis of deoxyoligonucleotides was undertaken.Several acyl groups namely 4-t-butylphenylacetyl, 4-t-butylphenoxyacetyl, 4-chlorobenzoyl, 3,4-dichlorobenzoyl, 2-methyl-benzoyl, 2,4-dimethylbenzoyl, nicotinoyl, 4-t-butylbenzoyl and 4-phenylazobenzoyl have been used for the protection of the exocyclic amino groups of deoxynucleosides.Interesting data concerning the stability of N-acyl groups towards a potent deacylating system (0.2N NaOH/MeOH) are presented.
- Koester, Hubert,Kulikowski, Konrad,Liese, Thomas,Heikens, Wiebke,Kohli, Vipin
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p. 363 - 369
(2007/10/02)
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