Endowing RNase H-inactive antisense with catalytic activity: 2-5A-morphants

Longhu Zhou, Edgar R. Civitello, Nidhi Gupta, Robert H. Silverman, Ross J. Molinaro, David E. Anderson, Paul F. Torrence

Research output: Contribution to journalArticle

6 Scopus citations

Abstract

A convergent synthetic approach was used to conjugate 2′,5′- oligoadenylate (2-5A, p5′A2′ [p5′A2′] np5′A) to phosphorodiamidate morpholino oligomers (morphants). To provide requisite quantities of 2-5A starting material, commercially and readily available synthons for solid-phase synthesis were adapted for larger scale solution synthesis. Thus, the tetranucleotide 5′- phosphoryladenylyl(2′→5′)adenylyl-(2′→5′) adenylyl(2′→5′)adenosine (p5′A2′p5′A2′] 2p5′A2′, tetramer 2-5A, 9) was synthesized starting with 2′,3′-O-dibenzoyl-N6,N6-dibenzoyl adenosine prepared from commercially available 5′-O-(4-monomethoxytrityl) adenosine. Coupling with N6-benzoyl-5′-O-(4,4′-dimethoxytrityl)- 3′-O-(tert-butyldimethylsilyl) adenosine-2′-(N,N-diisopropyl-2- cyanoethyl)phosphoramidite, followed by oxidization and deprotection, generated 5′-deprotected dimer 2-5A. Similar procedures lengthened the chain to form protected tetramer 2-5 A. The title product 9 p5′A(2′p5′A) 3 (tetramer 2-5A) was obtained through phosphorylation of the terminal 5′-hydroxy of the protected tetramer and removal of remaining protecting groups using concentrated ammonium hydroxide-ethanol (3:1, v/v) at 55°C and tetrabutylammonium fluoride (TBAF) in THF at room temperature, respectively. The 2-5A-phosphorodiamidate morpholino antisense chimera 11 (2-5A-morphant) was synthesized by covalently linking an aminolinker- functionalized phosphorodiamidate morpholino oligomer with periodate oxidized 2-5A tetramer (p5′A2′[p5′A2′]2p5′A). The resulting Schiff base was reduced with cyanoborohydride thereby transforming the ribose of the 2′-terminal nucleotide of 2-5A N-substituted morpholine. RNase L assays demonstrated that this novel 2-5A-antisense chimera had significant biological activity, thereby providing another potential tool for RNA ablation.

Original languageEnglish (US)
Pages (from-to)383-390
Number of pages8
JournalBioconjugate Chemistry
Volume16
Issue number2
DOIs
StatePublished - Mar 1 2005

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Biomedical Engineering
  • Pharmacology
  • Pharmaceutical Science
  • Organic Chemistry

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    Zhou, L., Civitello, E. R., Gupta, N., Silverman, R. H., Molinaro, R. J., Anderson, D. E., & Torrence, P. F. (2005). Endowing RNase H-inactive antisense with catalytic activity: 2-5A-morphants. Bioconjugate Chemistry, 16(2), 383-390. https://doi.org/10.1021/bc049778q