7 research outputs found
Synthesis and properties of 2′-deoxy-2′,4′-difluoroarabinose-modified nucleic acids
© 2015 American Chemical Society. We report the synthesis, thermal stability, and RNase H substrate activity of 2′-deoxy-2′,4′-difluoroarabino-modified nucleic acids. 2′-Deoxy-2′,4′-difluoroarabinouridine (2,′4′-diF-araU) was prepared in a stereoselective way in six steps from 2′-deoxy-2′-fluoroarabinouridine (2′-F-araU). NMR analysis and quantum mechanical calculations at the nucleoside level reveal that introduction of 4′-fluorine introduces a strong bias toward the North conformation, despite the presence of the 2′-βF, which generally steers the sugar pucker toward the South/East conformation. Incorporation of the novel monomer into DNA results on a neutral to slightly stabilizing thermal effect on DNA-RNA hybrids. Insertion of 2′,4′-diF-araU nucleotides in the DNA strand of a DNA-RNA hybrid decreases the rate of both human and HIV reverse transcriptase-associated RNase H-mediated cleavage of the complement RNA strand compared to that for an all-DNA strand or a DNA strand containing the corresponding 2′-F-araU nucleotide units, consistent with the notion that a 4′-fluorine in 2′-F-araU switches the preferred sugar conformation from DNA-like (South/East) to RNA-like (North).Peer Reviewe
Cellular Uptake and Intracellular Trafficking of Oligonucleotides: Implications for Oligonucleotide Pharmacology
One of the major constraints on the therapeutic use of oligonucleotides is inefficient delivery to their sites of action in the cytosol or nucleus. Recently it has become evident that the pathways of cellular uptake and intracellular trafficking of oligonucleotides can strongly influence their pharmacological actions. Here we provide background information on the basic processes of endocytosis and trafficking and then review recent literature on targeted delivery and subcellular trafficking of oligonucleotides in that context. A variety of approaches including molecular scale ligand-oligonucleotide conjugates, ligand-targeted nanocarriers, and the use of small molecules to enhance oligonucleotide effects are discussed
Locked 2′-Deoxy-2′,4′-Difluororibo Modified Nucleic Acids: Thermal Stability, Structural Studies, and siRNA Activity
© 2015 American Chemical Society. 2′-Deoxy-2′,4′-difluorouridine (2′,4′-diF-rU) was readily incorporated into DNA and RNA oligonucleotides via standard solid phase synthesis protocols. NMR and thermal denaturation (Tm) data of duplexes was consistent with the 2′,4′-diF-rU nucleotides adopting a rigid North (RNA-like) sugar conformation, as previously observed for the nucleoside monomer. The impact of this modification on Tm is neutral when incorporated within RNA:RNA duplexes, mildly destabilizing when located in the RNA strand of a DNA:RNA duplex, and highly destabilizing when inserted in the DNA strand of DNA:RNA and DNA:DNA duplexes. Molecular dynamics calculations suggest that the destabilization effect in DNA:DNA and DNA:RNA duplexes is the result of structural distortions created by A/B junctions within the helical structures. Quantum mechanics calculations suggest that the neutral effect imparted to A-form duplexes is caused by alterations in charge distribution that compensate the stabilizing effect expected for a pure North-puckered furanose sugar. 2′,4′-diF-RNA modified siRNAs were able to trigger RNA interference with excellent efficiency. Of note, incorporation of a few 2′,4′-diF-rU residues in the middle of the guide (antisense) strand afforded siRNAs that were more potent than the corresponding siRNAs containing LNA and 2′-F-ANA modifications, and as active as the 2′-F-RNA modified siRNAs.Peer Reviewe
Backbone FC-H⋯O hydrogen bonds in 2′F-substituted nucleic acids
Polarizing C-H⋯O hydrogen bonds: The structure of oligonucleotides containing alternating and contiguous tracts of 2′F-RNA and 2′F-ANA nucleotides reveals that nonconventional FC-H⋯O hydrogen bonds have a strong stabilizing effect on 2′-fluorinated duplexes. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.Peer Reviewe
Dramatic effect of furanose C2′ substitution on structure and stability: Directing the folding of the human telomeric quadruplex with a single fluorine atom
Human telomeric DNA quadruplexes can adopt different conformations in solution. We have found that arabinose, 2′F-arabinose, and ribose substitutions stabilize the propeller parallel G-quadruplex form over competing conformers, allowing NMR structural determination of this particularly significant nucleic acid structure. 2′F-arabinose substitution provides the greatest stabilization as a result of electrostatic (F-CH - -O4′) and pseudo-hydrogen-bond (F - -H8) stabilizing interactions. In contrast, 2′F-rG substitution provokes a dramatic destabilization of the quadruplex structure due to unfavorable electrostatic repulsion between the phosphate and the 2′-F. © 2013 American Chemical Society.Peer Reviewe
Rigid 2′,4′-difluororibonucleosides: Synthesis, conformational analysis, and incorporation into nascent RNA by HCV polymerase
We report on the synthesis and conformational properties of 2′-deoxy-2′,4′-difluorouridine (2′,4′-diF-rU) and cytidine (2′,4′-diF-rC) nucleosides. NMR analysis and quantum mechanical calculations show that the strong stereoelectronic effects induced by the two fluorines essentially ́locḱ the conformation of the sugar in the North region of the pseudorotational cycle. Our studies also demonstrate that NS5B HCV RNA polymerase was able to accommodate 2′,4′-diF-rU 5′-triphosphate (2′,4′-diF-rUTP) and to link the monophosphate to the RNA primer strand. 2′,4′-diF-rUTP inhibited RNA synthesis in dinucleotide-primed reactions, although with relatively high half-maximal inhibitory concentrations (IC50 > 50 μM). 2′,4′- diF-rU/C represents rare examples of ́locked́ ribonucleoside mimics that lack a bicyclic ring structure. © 2014 American Chemical Society.Peer Reviewe
Synthesis and Properties of 2′-Deoxy-2′,4′-difluoroarabinose-Modified Nucleic Acids
We report the synthesis, thermal
stability, and RNase H substrate
activity of 2′-deoxy-2′,4′-difluoroarabino-modified
nucleic acids. 2′-Deoxy-2′,4′-difluoroarabinouridine
(2,′4′-diF-araU) was prepared in a stereoselective way
in six steps from 2′-deoxy-2′-fluoroarabinouridine (2′-F-araU).
NMR analysis and quantum mechanical calculations at the nucleoside
level reveal that introduction of 4′-fluorine introduces a
strong bias toward the North conformation, despite the presence of
the 2′-βF, which generally steers the sugar pucker toward
the South/East conformation. Incorporation of the novel monomer into
DNA results on a neutral to slightly stabilizing thermal effect on
DNA–RNA hybrids. Insertion of 2′,4′-diF-araU
nucleotides in the DNA strand of a DNA–RNA hybrid decreases
the rate of both human and HIV reverse transcriptase-associated RNase
H-mediated cleavage of the complement RNA strand compared to that
for an all-DNA strand or a DNA strand containing the corresponding
2′-F-araU nucleotide units, consistent with the notion that
a 4′-fluorine in 2′-F-araU switches the preferred sugar
conformation from DNA-like (South/East) to RNA-like (North)