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Site-Specifically Modified Circular Ribonucleic Acid Serves as Multitarget miRNA Sponge with Low Immunogenicity.

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Synthetic circular RNAs (circRNAs) modified with pseudouridine show reduced immunogenicity for improved miRNA-based therapies. This method enables precise synthesis of stable, effective circRNA therapeutics.

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Area of Science:

  • Biotechnology
  • RNA Therapeutics
  • Molecular Biology

Background:

  • MicroRNAs (miRNAs) regulate gene expression and are targets for therapy.
  • Linear miRNA sponges have limitations like low stability and high immunogenicity.
  • Artificial circular RNAs (circRNAs) offer better stability but still face immunogenicity challenges.

Purpose of the Study:

  • To develop a method for synthesizing site-specifically modified circRNAs with reduced immunogenicity.
  • To investigate the impact of pseudouridine modification on circRNA immunogenicity and function.
  • To enable long-term coregulation of multiple miRNAs using engineered circRNAs.

Main Methods:

  • Developed a DNA-templated RNA ligation method for synthesizing modified circRNAs.
  • Incorporated multiple miRNA binding sites into the circRNAs.
  • Introduced pseudouridine modifications (30%) into the synthetic circRNAs.

Main Results:

  • The developed method allows for high-yield, site-specific synthesis of modified circRNAs.
  • Pseudouridine modification (30%) effectively reduced circRNA immunogenicity, irrespective of modification site or pattern.
  • Engineered circRNAs demonstrated long-term coregulation of multiple miRNAs.

Conclusions:

  • Site-specifically modified circRNAs can be precisely synthesized using the developed DNA-templated ligation method.
  • Pseudouridine modification is a viable strategy to mitigate circRNA immunogenicity for therapeutic applications.
  • These advancements pave the way for developing next-generation circRNA-based therapeutics with balanced properties.