In vivo comparison of 2′-O-methyl phosphorothioate and morpholino antisense oligonucleotides for Duchenne muscular dystrophy exon skipping
In vivo comparison of 2′-O-methyl phosphorothioate and morpholino antisense oligonucleotides for Duchenne muscular dystrophy exon skipping
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DOI：
10.1002/jgm.1288
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发表时间：
2009-03-01
影响因子：
3.5
通讯作者：
Aartsma-Rus, Annemieke
中科院分区：
文献类型：
--
作者：
Heemskerk, Hans A.;de Winter, Christa L.;Aartsma-Rus, Annemieke
背景反义介导的外显子跳跃是治疗杜氏肌营养不良症（DMD）的一种公认的方法。使用反义寡核苷酸（AON），破坏的DMD阅读框架被恢复，允许产生部分功能性肌营养不良蛋白，并将严重的杜氏肌营养不良症转化为较轻的贝克尔肌营养不良症表型。体内研究主要使用2 '-O-甲基硫代磷酸酯（2 OMePS）或吗啉代（PMO）AON进行。方法对mdx和人源化（h）DMD小鼠分别肌肉注射和静脉注射2 OMePS和PMO（小鼠外显子23和人外显子44、45、46和51）。结果肌肉注射2 OMePS AON的长度增加，人外显子45的跳跃效率增加，而小鼠外显子23的跳跃效率降低。尽管PMO诱导更多的小鼠外显子23跳跃，但PMO和2 OMePS对于人外显子更具有可比性。静脉给药后，两种化学物质均在心脏中显示外显子跳跃和新蛋白。此外，与2 OMePS相比，PMO显示出更低的肌内浓度和更高的外显子23跳跃水平，这可能是由于在细胞外基质中的隔离。最后，两个错配呈现2 OMePS，但不是PMO AONs几乎invalid.Conclusions在本研究中获得的结果表明，增加AON长度提高跳跃效率在某些情况下，但不是所有的情况下。在注射2 OMePS和未缀合的PMO后，在心脏中诱导外显子跳跃和肌营养不良蛋白恢复是可行的。此外，PMO和2 OMePS之间的效率差异似乎是序列而不是化学依赖性的。最后，结果表明PMO的序列特异性可能低于2 OMePS。版权所有（C）2009约翰威利父子有限公司
Background Antisense-mediated exon skipping is a putative treatment for Duchenne muscular dystrophy (DMD). Using antisense oligonucleotides (AONs), the disrupted DMD reading frame is restored, allowing generation of partially functional dystrophin and conversion of a severe Duchenne into a milder Becker muscular dystrophy phenotype. In vivo studies are mainly performed using 2'-O-methyl phosphorothioate (2OMePS) or morpholino (PMO) AONs. These compounds were never directly compared.Methods mdx and humanized (h)DMD mice were injected intramuscularly and intravenously with short versus long 2OMePS and PMO for mouse exon 23 and human exons 44, 45, 46 and 51.Results Intramuscular injection showed that increasing the length of 2OMePS AONs enhanced skipping efficiencies of human exon 45, but decreased efficiency for mouse exon 23. Although PMO induced more mouse exon 23 skipping, PMO and 2OMePS were more comparable for human exons. After intravenous administration, exon skipping and novel protein was shown in the heart with both chemistries. Furthermore, PMO showed lower intramuscular concentrations with higher exon 23 skipping levels compared to 2OMePS, which may be due to sequestration in the extracellular matrix. Finally, two mismatches rendered 2OMePS but not PMO AONs nearly ineffective.Conclusions The results obtained in the present study indicate that increasing AON length improves skipping efficiency in some but not all cases. It is feasible to induce exon skipping and dystrophin restoration in the heart after injection of 2OMePS and unconjugated PMO. Furthermore, differences in efficiency between PMO and 2OMePS appear to be sequence and not chemistry dependent. Finally, the results indicate that PMOs may be less sequence specific than 2OMePS. Copyright (C) 2009 John Wiley & Sons, Ltd.