title: Synthesis of 2'-O-guanidinopropyl-modified nucleoside phosphoramidites and their incorporation into siRNAs targeting hepatitis B virus.
abstract: Synthetic RNAi activators have shown considerable potential for therapeutic application to silencing of pathology-causing genes. Typically these exogenous RNAi activators comprise duplex RNA of approximately 21 bp with 2 nt overhangs at the 3' ends. To improve efficacy of siRNAs, chemical modification at the 2'-OH group of ribose has been employed. Enhanced stability, gene silencing and attenuated immunostimulation have been demonstrated using this approach. Although promising, efficient and controlled delivery of highly negatively charged nucleic acid gene silencers remains problematic. To assess the potential utility of introducing positively charged groups at the 2' position, our investigations aimed at assessing efficacy of novel siRNAs containing 2'-O-guanidinopropyl (GP) moieties. We describe the formation of all four GP-modified nucleosides using the synthesis sequence of Michael addition with acrylonitrile followed by Raney-Ni reduction and guanidinylation. These precursors were used successfully to generate antihepatitis B virus (HBV) siRNAs. Testing in a cell culture model of viral replication demonstrated that the GP modifications improved silencing. Moreover, thermodynamic stability was not affected by the GP moieties and their introduction into each position of the seed region of the siRNA guide strand did not alter the silencing efficacy of the intended HBV target. These results demonstrate that modification of siRNAs with GP groups confers properties that may be useful for advancing therapeutic application of synthetic RNAi activators.
authors.affiliation: Goethe-University, Institute of Organic Chemistry & Chemical Biology, Max-von-Laue-Str. 7, 60438 Frankfurt am Main, Germany.
authors.forename: Jolanta
authors.initials: J
authors.lastname: Brzezinska
authors.forename: Jennifer
authors.initials: J
authors.lastname: D'Onofrio
authors.forename: Maximilian C R
authors.initials: MC
authors.lastname: Buff
authors.forename: Justin
authors.initials: J
authors.lastname: Hean
authors.forename: Abdullah
authors.initials: A
authors.lastname: Ely
authors.forename: Musa
authors.initials: M
authors.lastname: Marimani
authors.forename: Patrick
authors.initials: P
authors.lastname: Arbuthnot
authors.forename: Joachim W
authors.initials: JW
authors.lastname: Engels
mesh_terms: D016503:Drug Delivery Systems; D004355:Drug Stability; D020868:Gene Silencing / Q000187:drug effects; D057809:HEK293 Cells; D006515:Hepatitis B virus / Q000187:drug effects; D006801:Humans; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000235:genetics; D009943:Organophosphorus Compounds / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D013386:Succinates / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology
chemical_list: D009841:Oligonucleotides; D009943:Organophosphorus Compounds; D034741:RNA, Small Interfering; D013386:Succinates; C434331:phosphoramidite; C022015:guanidinopropylsuccinic acid