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January 22nd, 2025
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Hiroshima University
genomics
biorxiv

Risk Prediction of RNA Off-Targets of CRISPR Base Editors in Tissue-Specific Transcriptomes Using Language Models

Nakamae, K.Open in Google Scholar•Suzuki, T.Open in Google Scholar•Yonezawa, S.Open in Google Scholar•Yamamoto, K.Open in Google Scholar•Kakuzaki, T.Open in Google Scholar•Ono, H.Open in Google Scholar•Naito, Y.Open in Google Scholar•Bono, H.Open in Google Scholar

Base-editing technologies, particularly cytosine base editors (CBEs), allow precise gene modification without introducing double-strand breaks; however, unintended RNA off-target effects remain a critical concern and are understudied. To address this gap, we developed PiCTURE, a standardized computational pipeline for detecting and quantifying transcriptome-wide CBE-induced RNA off-target events. PiCTURE identifies both canonical ACW (W = A or T/U) motif-dependent and non-canonical RNA off-targets, revealing a broader WCW motif that underlies many unanticipated edits. Additionally, we developed two machine learning models based on the DNABERT-2 language model, termed STL and SNL, which outperformed motif-only approaches in terms of accuracy, precision, recall, and F1 score. To demonstrate the practical application of our predictive model for CBE-induced RNA off-target risk, we integrated PiCTURE outputs with the PROTECTiO pipeline and estimated RNA off-target risk for each transcript showing tissue-specific expression. The analysis revealed differences among tissues: while the brain and ovaries exhibited relatively low off-target burden, the colon and lungs displayed relatively high risks. Our study provides a comprehensive framework for RNA off-target profiling, emphasizing the importance of advanced machine learning-based classifiers in CBE safety evaluations and offering valuable insights for the development of safer genome-editing therapies.

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