Structural basis for the synergy of 4'- and 2'-modifications on siRNA nuclease resistance, thermal stability and RNAi activity.
Structural basis for the synergy of 4'- and 2'-modifications on siRNA nuclease resistance, thermal stability and RNAi activity.
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DOI：
10.1093/nar/gky703
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发表时间：
2018-09-19
影响因子：
14.9
通讯作者：
Egli M
中科院分区：
文献类型：
--
作者：
Harp JM;Guenther DC;Bisbe A;Perkins L;Matsuda S;Bommineni GR;Zlatev I;Foster DJ;Taneja N;Charisse K;Maier MA;Rajeev KG;Manoharan M;Egli M
化学修饰是用于改善代谢稳定性、摄取和活性的寡核苷酸治疗剂的先决条件，无论其作用模式如何，即反义、RNAi或适体。磷酸基团和核糖C2′/O2′原子是最常见的修饰位点。与2′-O-取代基相比，核糖4′-C-取代基位于3′-和5′-相邻磷酸酯的附近。为了研究对核酸酶抗性的潜在有益作用，我们将2′-F和2′-OMe与4′-Cα-和4′-Cβ-OMe以及2′-F与4′-Cα-甲基修饰组合。合成4′-C-OMe-尿苷的α-和β-差向异构体和4′-C-Me-尿苷单体的α-差向异构体并掺入siRNA中。4′α-差向异构体对热稳定性的影响很小，无论2′-取代基（H、F、OMe）如何，核酸酶稳定性都有所增加。4′β-差向异构体具有强烈的去稳定性，但对具有磷酸或硫代磷酸骨架的核酸外切酶具有完全抗性。含有2′-F，4′-Cα-OMe-U，2′-F，4′-Cβ-OMe-U，2′-OMe，4′-Cα-OMe-U，2 ′-OMe，4 ′-Cβ-OMe-U或2′-F，4′-Cα-Me-U的RNA八聚体的晶体结构有助于合理化这些观察结果，并指出链反向4′β-U差向异构体所观察到的前所未有的核酸酶抗性的空间和静电起源。我们使用人Argonaute 2与在种子区的不同位点具有2′-F，4′-Cα-OMe-U或2′-F，4′-Cβ-OMe-U的指导siRNA复合的结构模型来解释具有相应2′-/4′-C修饰的siRNA的体外活性。
Chemical modification is a prerequisite of oligonucleotide therapeutics for improved metabolic stability, uptake and activity, irrespective of their mode of action, i.e. antisense, RNAi or aptamer. Phosphate moiety and ribose C2′/O2′ atoms are the most common sites for modification. Compared to 2′-O-substituents, ribose 4′-C-substituents lie in proximity of both the 3′- and 5′-adjacent phosphates. To investigate potentially beneficial effects on nuclease resistance we combined 2′-F and 2′-OMe with 4′-Cα- and 4′-Cβ-OMe, and 2′-F with 4′-Cα-methyl modification. The α- and β-epimers of 4′-C-OMe-uridine and the α-epimer of 4′-C-Me-uridine monomers were synthesized and incorporated into siRNAs. The 4′α-epimers affect thermal stability only minimally and show increased nuclease stability irrespective of the 2′-substituent (H, F, OMe). The 4′β-epimers are strongly destabilizing, but afford complete resistance against an exonuclease with the phosphate or phosphorothioate backbones. Crystal structures of RNA octamers containing 2′-F,4′-Cα-OMe-U, 2′-F,4′-Cβ-OMe-U, 2′-OMe,4′-Cα-OMe-U, 2′-OMe,4′-Cβ-OMe-U or 2′-F,4′-Cα-Me-U help rationalize these observations and point to steric and electrostatic origins of the unprecedented nuclease resistance seen with the chain-inverted 4′β-U epimer. We used structural models of human Argonaute 2 in complex with guide siRNA featuring 2′-F,4′-Cα-OMe-U or 2′-F,4′-Cβ-OMe-U at various sites in the seed region to interpret in vitro activities of siRNAs with the corresponding 2′-/4′-C-modifications.
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影响因子：
14.9
作者：
Pallan PS;Greene EM;Jicman PA;Pandey RK;Manoharan M;Rozners E;Egli M
通讯作者：
Egli M
DOI：
10.1021/jo502948t
发表时间：
2015-03-20
期刊：
The Journal of organic chemistry
影响因子：
--
作者：
Martínez-Montero S;Deleavey GF;Dierker-Viik A;Lindovska P;Ilina T;Portella G;Orozco M;Parniak MA;González C;Damha MJ
通讯作者：
Damha MJ
影响因子：
3.2
作者：
Rajeev, Kallanthottathil G.;Nair, Jayaprakash K.;Manoharan, Muthiah
通讯作者：
Manoharan, Muthiah
影响因子：
64.5
作者：
Elkayam E;Kuhn CD;Tocilj A;Haase AD;Greene EM;Hannon GJ;Joshua-Tor L
通讯作者：
Joshua-Tor L
DOI：
10.1107/s0907444909052925
发表时间：
2010-02
期刊：
Acta crystallographica. Section D, Biological crystallography
影响因子：
--
作者：
Adams PD;Afonine PV;Bunkóczi G;Chen VB;Davis IW;Echols N;Headd JJ;Hung LW;Kapral GJ;Grosse-Kunstleve RW;McCoy AJ;Moriarty NW;Oeffner R;Read RJ;Richardson DC;Richardson JS;Terwilliger TC;Zwart PH
通讯作者：
Zwart PH