Effect of Sugar 2′,4′-Modifications on Gene Silencing Activity of siRNA Duplexes

Malek-Adamian, Elise; Fakhoury, Johans; Arnold, Amy E.; Martı́nez-Montero, Saúl; Shoichet, Molly S.; Damha, Masad J.

doi:10.1089/nat.2019.0792

Cited by 18 publications

(22 citation statements)

References 25 publications

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“…The preferred sugar puckering was found in the northern region (C3′‐ endo ) for 2′‐OEt modification and unmodified 2′‐OH RNA, as expected. The introduction of 4′‐ C ‐aminoalkyl modifications favors the sugar puckering toward C2′‐ endo has also been observed in the previous studies [21,22,24,54,55] …”

Section: Resultssupporting

confidence: 70%

“…Moreover, Ueno Y. and coworkers have also obtained similar results for analogous compounds using NMR measurements [22,24] . The calculated percentages of sugar conformations for 4′‐ C ‐azidoethyl‐2′‐ O ‐methyluridine and 4′‐ C ‐azidopropyl‐2′‐ O ‐methyluridine compounds, observed ∼69 % South conformations for both the compounds [22,24] . These results confirm that the 4′‐modification helps to alter the sugar conformation exclusively towards the South conformation.…”

Section: Resultssupporting

confidence: 62%

“…Based on NMR studies, it was observed that 4′‐AzM‐2′‐OEt and 4′‐AM‐2′‐OEt‐uridine nucleosides predominantly adopt South conformation, and 2′‐ O ‐ethyl‐uridine nucleoside adopts exclusively North conformation, in solution. Moreover, Ueno Y. and coworkers have also obtained similar results for analogous compounds using NMR measurements [22,24] . The calculated percentages of sugar conformations for 4′‐ C ‐azidoethyl‐2′‐ O ‐methyluridine and 4′‐ C ‐azidopropyl‐2′‐ O ‐methyluridine compounds, observed ∼69 % South conformations for both the compounds [22,24] .…”

Section: Resultsmentioning

confidence: 54%

“…2′‐modifications such as 2′‐F, 2′‐OMe, and 2′‐MOE have attracted attention and have been explored in clinically approved AONs and siRNAs drugs [19,20] . Also, dual modifications at 2′‐ and 4′‐ sugar positions such as 4′‐ C ‐aminomethyl‐2′‐ O ‐methyl (4′‐AM‐2′‐OMe), 4′‐ C ‐aminoethyl‐2′‐ O ‐methyl (4′‐AMEt‐2′‐OMe), and 4′‐ C ‐aminopropyl‐2′‐ O ‐methyl (4′‐AMPr‐2′‐OMe) analogs have adopted C2′‐ endo (South , DNA ‐type) sugar conformation [21,22] . In contrast, other bi‐functional sugar modifications such as 4′‐ C ‐aminomethyl‐2′‐deoxy‐2′‐fluoro (4′‐AM‐2′‐F), 4′‐ C ‐aminoethyl‐2′‐deoxy‐2′‐fluoro (4′‐AMEt‐2′‐F), 2′,4′‐diF, 2′‐OMe‐4′‐F, 2′‐F‐4′‐OMe, 2′,4′‐diOMe, 2′‐F‐4′‐OMe‐araU, 2′,4′‐diF‐araU tuned the sugar conformation predominantly toward C3′‐ endo ( North , RNA ‐type), which has been attributed to the small size, high electronegativity of fluorine, and additional C−H⋅⋅⋅F interactions [23–30] .…”

Section: Introductionmentioning

confidence: 99%

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Synthesis, Structural, and Conformational Analysis of 4′‐C‐Alkyl‐2′‐O‐Ethyl‐Uridine Modified Nucleosides

Harikrishna

Gore

2021

Eur J Org Chem

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Sugar modifications have attracted much attention due to their potential structural and functional influence on therapeutic nucleic acids. Herein, we report the synthesis of dual modified 4′‐C‐azidomethyl‐2′‐O‐ethyl‐uridine (4′‐AzM‐2′‐OEt−U) and 4′‐C‐aminomethyl‐2′‐O‐ethyl‐uridine (4′‐AM‐2′‐OEt−U) nucleosides using linear multi‐step synthesis (16 linear steps). Additionally, we report an alternative route for the synthesis of 2′‐O‐ethyl‐uridine nucleoside which has been achieved in three steps with an overall yield of 40 %. X‐ray structure of 4′‐AzM‐2′‐OEt−U illustrates that the nucleoside adopts C2′‐endo (South) conformation having a DNA‐type glycosidic bond (χ) angle of −116.01°. Computational studies revealed the C2′‐endo and C4′‐exo conformations for 4′‐AzM‐2′‐OEt−U and 4′‐AM‐2′‐OEt−U free nucleosides, respectively. The C4′‐exo conformation in 4′‐AM‐2′‐OEt−U free nucleoside is collectively stabilized by various non‐covalent interactions between positively charged aminomethyl and 2′,3′‐hydroxyl groups. Insights into the structural and conformational analysis of dual sugar modified nucleosides and oligonucleotides will be helpful in the rational design of modified nucleosides and therapeutic oligonucleotides.

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Section: Resultssupporting

confidence: 70%

Section: Resultssupporting

confidence: 62%

Section: Resultsmentioning

confidence: 54%

Section: Introductionmentioning

confidence: 99%

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Synthesis, Structural, and Conformational Analysis of 4′‐C‐Alkyl‐2′‐O‐Ethyl‐Uridine Modified Nucleosides

Harikrishna

Gore

2021

Eur J Org Chem

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“… 24 Furthermore, the efficacy of the allele-specific siRNAs in this study may be improved by alternating the chemical modification of the siRNAs. 25 Also, more SNP targets could be tested to increase the percentage of the targeted patient population.…”

Section: Discussionmentioning

confidence: 99%

Ex vivo Improvement of a von Willebrand Disease Type 2A Phenotype Using an Allele-Specific Small-Interfering RNA

et al. 2020

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Von Willebrand disease (VWD) is the most common inherited bleeding disorder and is mainly caused by dominant-negative mutations in the multimeric protein von Willebrand factor (VWF). These mutations may either result in quantitative or qualitative defects in VWF. VWF is an endothelial protein that is secreted to the circulation upon endothelial activation. Once secreted, VWF multimers bind platelets and chaperone coagulation factor VIII in the circulation. Treatment of VWD focuses on increasing VWF plasma levels, but production and secretion of mutant VWF remain uninterrupted. Presence of circulating mutant VWF might, however, still affect normal hemostasis or functionalities of VWF beyond hemostasis. We hypothesized that inhibition of the production of mutant VWF improves the function of VWF overall and ameliorates VWD phenotypes. We previously proposed the use of allele-specific small-interfering RNAs (siRNAs) that target frequent VWF single nucleotide polymorphisms to inhibit mutant VWF. The aim of this study is to prove the functionality of these allele-specific siRNAs in endothelial colony-forming cells (ECFCs). We isolated ECFCs from a VWD type 2A patient with an intracellular multimerization defect, reduced VWF collagen binding, and a defective processing of proVWF to VWF. After transfection of an allele-specific siRNA that specifically inhibited expression of mutant VWF, we showed amelioration of the laboratory phenotype, with normalization of the VWF collagen binding, improvement in VWF multimers, and enhanced VWF processing. Altogether, we prove that allele-specific inhibition of the production of mutant VWF by siRNAs is a promising therapeutic strategy to improve VWD phenotypes.

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Atom‐Modified gDNA Enhances Cleavage Activity of TtAgo Enabling Ultra‐Sensitive Nucleic Acid Testing

Zhang,

Chen,

Jiang

et al. 2024

Advanced Science

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The DNA‐guided (gDNA) Argonaute from Thermus thermophilus (TtAgo) has little potential for nucleic acid detection and gene editing due to its poor dsDNA cleavage activity at relatively low temperature. Herein, the dsDNA cleavage activity of TtAgo is enhanced by using 2′‐fluorine (2′F)‐modified gDNA and developes a novel nucleic acid testing strategy. This study finds that the gDNA with 2′F‐nucleotides at the 3′‐end (2′F‐gDNA) can promote the assembly of the TtAgo‐guide‐target ternary complex significantly by increasing its intermolecular force to target DNA and TtAgo, thereby providing ≈40‐fold activity enhancement and decreasing minimum reaction temperature from 65 to 60°C. Based on this outstanding advance, a novel nucleic acid testing strategy is proposed, termed FAST, which is performed by using the 2′F‐gDNA/TtAgo for target recognition and combining it with Bst DNA polymerase for nucleic acid amplification. By integrating G‐quadruplex and Thioflavin T, the FAST assay achieves one‐pot real‐time fluorescence analysis with ultra‐sensitivity, providing a limit of detection up to 5 copies (20 µL reaction mixture) for miR‐21 detection. In summary, an atom‐modification‐based strategy has been developed for enhancing the cleavage activity of TtAgo efficiently, thereby improving its practicability and establishing a TtAgo‐based nucleic acid testing technology with ultra‐sensitivity and high‐specificity.

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Effect of Sugar 2′,4′-Modifications on Gene Silencing Activity of siRNA Duplexes

Cited by 18 publications

References 25 publications

Synthesis, Structural, and Conformational Analysis of 4′‐C‐Alkyl‐2′‐O‐Ethyl‐Uridine Modified Nucleosides

Synthesis, Structural, and Conformational Analysis of 4′‐C‐Alkyl‐2′‐O‐Ethyl‐Uridine Modified Nucleosides

Ex vivo Improvement of a von Willebrand Disease Type 2A Phenotype Using an Allele-Specific Small-Interfering RNA

Atom‐Modified gDNA Enhances Cleavage Activity of TtAgo Enabling Ultra‐Sensitive Nucleic Acid Testing

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