DOI: https://doi.org/10.21203/rs.3.rs-2762001/v1
Off-target interactions between antisense oligonucleotides (ASOs) with state-of-the-art modifications and biological components still pose clinical safety liabilities. To prevent both hybridization-dependent and -independent off-target interactions, a novel nanoarchitecture called BRace On a THERapeutic aSo (BROTHERS or BRO) consisting of typical gapmer ASO and partially complementary peptide nucleic acid (PNA) strand was devised. This non-canonical ASO/PNA hybrids have been shown to have non-specific protein-binding repellent characteristics. Feasible optimization of the structural and thermodynamic characteristics of this duplex system was also demonstrated to elicit in vivo operative toehold-mediated strand displacement (TMSD) reaction, which can reduce hybridization to RNA off-targets. The optimized BROs dramatically mitigated hepatotoxicity while maintaining the on-target knockdown activity of their parent ASOs in vivo. This new technique will not only bring a novel BRO class of drugs that would have a transformative impact on the extrahepatic delivery of ASOs, but also help uncover the toxicity mechanism of ASOs.