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Related Experiment Video

Updated: May 24, 2025

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Mitigating Cellular Dysfunction Through Contaminant Reduction in Synthetic circRNA for High-Efficiency mRNA-Based

Ziwei Zhang1, Weiyu Li1,2, Xiangyu Ren1,3

  • 1CAS Key Laboratory for Biological Effects of Nanomaterials and Nanosafety, National Center for Nanoscience and Technology, Chinese Academy of Sciences, Haidian District, Beijing, 100190, China.

Advanced Science (Weinheim, Baden-Wurttemberg, Germany)
|March 5, 2025
PubMed
Summary

Synthetic circular RNA (circRNA) impurities trigger immune responses. A novel purification method removes these contaminants, enhancing circRNA efficacy for regenerative medicine and cancer immunotherapy applications.

Keywords:
cell reprogrammingcircular RNAiPSCmRNA therapypurification strategy

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Area of Science:

  • Biotechnology
  • RNA Therapeutics
  • Immunology

Background:

  • Synthetic circular RNA (circRNA) shows therapeutic potential but is hindered by impurities.
  • Immunogenic byproducts like double-stranded RNA and 5' triphosphates activate innate immune responses, reducing circRNA efficacy.
  • RNA-sensing pathways are implicated in the immune response to these contaminants.

Purpose of the Study:

  • To identify key immunogenic byproducts in synthetic circRNA.
  • To develop and validate a purification strategy for removing these contaminants.
  • To evaluate the performance of purified circRNA in cellular reprogramming and immunotherapy.

Main Methods:

  • Identification of immunogenic byproducts in synthetic circRNA.
  • Development of a multi-step purification process involving enzymatic treatments and cellulose-based filtration.
  • Assessment of purified circRNA in induced pluripotent stem cell (iPSC) reprogramming and chimeric antigen receptor (CAR) T-cell applications.

Main Results:

  • The purification process effectively removed key immunogenic byproducts, significantly reducing immune activation.
  • Manufacturing yields of circRNA were increased by over 10-fold.
  • Purified circRNA achieved over 300% efficiency in iPSC reprogramming with minimal transfections.
  • Purified circRNA demonstrated high and persistent CAR expression in T cells, leading to potent anti-tumor activity.

Conclusions:

  • A scalable and reliable purification strategy for therapeutic-grade circRNA has been established.
  • The purified circRNA shows significant promise for applications in regenerative medicine and cancer immunotherapy.
  • This purification approach overcomes major hurdles in the clinical translation of synthetic RNA therapeutics.