Zongyi Yi , Liang Qu , Huixian Tang, Zhiheng Liu, Ying Liu, Feng Tian , Chunhui Wang, Xiaoxue Zhang, Ziqi Feng, Ying Yu, Pengfei Yuan, Zexuan Yi, Yanxia Zhao and Wensheng Wei
Abstract
Current methods for programmed RNA editing using endogenous ADAR enzymes and engineered ADAR-recruiting RNAs
(arRNAs) suffer from low efficiency and bystander off-target editing. Here, we describe LEAPER 2.0, an updated version of
LEAPER that uses covalently closed circular arRNAs, termed circ-arRNAs. We demonstrate on average ~3.1-fold higher editing efficiency than their linear counterparts when expressed in cells or delivered as in vitro-transcribed circular RNA oligonucleotides. To lower off-target editing we deleted pairings of uridines with off-target adenosines, which almost completely
eliminated bystander off-target adenosine editing. Engineered circ-arRNAs enhanced the efficiency and fidelity of editing
endogenous CTNNB1 and mutant TP53 transcripts in cell culture. Delivery of circ-arRNAs using adeno-associated virus in a
mouse model of Hurler syndrome corrected the pathogenic point mutation and restored α-L-iduronidase catalytic activity, lowering glycosaminoglycan accumulation in the liver. LEAPER 2.0 provides a new design of arRNA that enables more precise,
efficient RNA editing with broad applicability for therapy and basic research.
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