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Multivalent Aptamer-Functionalized Single-Strand RNA Origami as Effective, Target-Specific Anticoagulants with Corresponding Reversal Agents

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DOI

  • Abhichart Krissanaprasit, North Carolina State Univ, University of North Carolina, North Carolina State University, Dept Entomol & Plant Pathol
  • ,
  • Carson M Key, North Carolina State Univ, University of North Carolina, North Carolina State University, Dept Entomol & Plant Pathol
  • ,
  • Kristen Froehlich
  • ,
  • Sahil Pontula
  • ,
  • Emily Mihalko
  • ,
  • Daniel M Dupont
  • Ebbe S Andersen
  • Jørgen Kjems
  • Ashley C Brown
  • ,
  • Thomas H LaBean, North Carolina State Univ, University of North Carolina, North Carolina State University, Dept Entomol & Plant Pathol

Anticoagulants are commonly utilized during surgeries and to treat thrombotic diseases like stroke and deep vein thrombosis. However, conventional anticoagulants have serious side-effects, narrow therapeutic windows, and lack safe reversal agents (antidotes). Here, an alternative RNA origami displaying RNA aptamers as target-specific anticoagulant is described. Improved design and construction techniques for self-folding, single-molecule RNA origami as a platform for displaying pre-selected RNA aptamers with precise orientational and spatial control are reported. Nuclease resistance is added using 2'-fluoro-modified pyrimidines during in vitro transcription. When four aptamers are displayed on the RNA origami platform, the measured thrombin inhibition and anticoagulation activity is higher than observed for free aptamers, ssRNA-linked RNA aptamers, and RNA origami displaying fewer aptamers. Importantly, thrombin inhibition is immediately switched off by addition of specific reversal agents. Results for single-stranded DNA (ssDNA) and single-stranded peptide nucleic acid (PNA) antidotes show restoration of 63% and 95% coagulation activity, respectively. To demonstrate potential for practical, long-term storage for clinical use, RNA origami is freeze-dried, and stored at room temperature. Freshly produced and freeze-dried RNA show identical levels of activity in coagulation assays. Compared to current commercial intravenous anticoagulants, RNA origami-based molecules show promise as safer alternatives with rapid activity switching for future therapeutic applications.

OriginalsprogEngelsk
TidsskriftAdvanced Healthcare Materials
Sider (fra-til)e2001826
ISSN2192-2640
DOI
StatusUdgivet - 9 jun. 2021

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