Mechanistic analysis of the enhanced RNAi activity by 6-mCEPh-purine at the 5’ end of the siRNA guide strand
- 1 Institute for Quantitative Biosciences, The University of Tokyo;
- 2 R&D Division, Kyowa Kirin, Co., Ltd.;
- 3 Institut de Genetique Humaine, CNRS and Universite de Montpellier
- ↵* Corresponding author; email: tomari{at}iqb.u-tokyo.ac.jp
Abstract
A key approach for improving siRNA efficacy is chemical modifications. Through an in-silico screening of modifications at the 5′-end nucleobase of the guide strand, an adenine-derived compound called 6-(3-(2-carboxyethyl)phenyl)-purine (6-mCEPh-purine) was identified to improve the RNAi activity in cultured human cells and in vivo mouse models. Nevertheless, it remains unclear how this chemical modification enhances the siRNA potency. Here, we used a series of biochemical approaches to quantitatively evaluate the effect of the 6-mCEPh-purine modification at each step in the assembly of the RNAi effector complex called RISC. We found that the modification improves the formation of mature RISC at least in two different ways, by fixing the loading orientation of siRNA duplexes and increasing the stability of mature RISC after passenger strand ejection. Our data will provide a molecular platform for further development of chemically modified siRNA drugs.
Keywords
- Received October 25, 2019.
- Accepted October 15, 2020.
- Published by Cold Spring Harbor Laboratory Press for the RNA Society
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