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5′‐( E )‐Vinylphosphonate: A Stable Phosphate Mimic Can Improve the RNAi Activity of siRNA–GalNAc Conjugates
Author(s) -
Parmar Rubina,
Willoughby Jennifer L. S.,
Liu Jingxuan,
Foster Donald J.,
Brigham Benjamin,
Theile Christopher S.,
Charisse Klaus,
Akinc Akin,
Guidry Erin,
Pei Yi,
Strapps Walter,
Cancilla Mark,
Stanton Matthew G.,
Rajeev Kallanthottathil G.,
SeppLorenzino Laura,
Manoharan Muthiah,
Meyers Rachel,
Maier Martin A.,
Jadhav Vasant
Publication year - 2016
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201600130
Subject(s) - small interfering rna , gene silencing , rna interference , rna , microbiology and biotechnology , rna silencing , biology , phosphorylation , in vitro , kinase , in vivo , rna induced silencing complex , conjugate , biochemistry , chemistry , gene , genetics , mathematical analysis , mathematics
Small interfering RNA (siRNA)‐mediated silencing requires siRNA loading into the RNA‐induced silencing complex (RISC). Presence of 5′‐phosphate (5′‐P) is reported to be critical for efficient RISC loading of the antisense strand (AS) by anchoring it to the mid‐domain of the Argonaute2 (Ago2) protein. Phosphorylation of exogenous duplex siRNAs is thought to be accomplished by cytosolic Clp1 kinase. However, although extensive chemical modifications are essential for siRNA–GalNAc conjugate activity, they can significantly impair Clp1 kinase activity. Here, we further elucidated the effect of 5′‐P on the activity of siRNA–GalNAc conjugates. Our results demonstrate that a subset of sequences benefit from the presence of exogenous 5′‐P. For those that do, incorporation of 5′‐( E )‐vinylphosphonate (5′‐VP), a metabolically stable phosphate mimic, results in up to 20‐fold improved in vitro potency and up to a threefold benefit in in vivo activity by promoting Ago2 loading and enhancing metabolic stability.