development of a single vector system that enhances trans-splicing of smn2 transcripts发展一个向量系统提高trans-splicing smn2成绩单.pdfVIP

Development of a Single Vector System that Enhances Trans-Splicing of SMN2 Transcripts 1 2 1 3 1,2 Tristan H. Coady , Travis D. Baughan , Monir Shababi , Marco A. Passini , Christian L. Lorson * 1 Department of Veterinary Pathobiology, Bond Life Sciences Center, University of Missouri, Columbia, Missouri, United States of America, 2 Department of Molecular Microbiology and Immunology, Bond Life Sciences Center, University of Missouri, Columbia, Missouri, United States of America, 3 Neuroscience, Genzyme Corporation, Framingham, Massachusetts, United States of America Abstract RNA modalities are developing as a powerful means to re-direct pathogenic pre-mRNA splicing events. Improving the efficiency of these molecules in vivo is critical as they move towards clinical applications. Spinal muscular atrophy (SMA) is caused by loss of SMN1. A nearly identical copy gene called SMN2 produces low levels of functional protein due to alternative splicing. We previously reported a trans-splicing RNA (tsRNA) that re-directed SMN2 splicing. Now we show that reducing the competition between endogenous splices sites enhanced the efficiency of trans-splicing. A single vector system was developed that expressed the SMN tsRNA and a splice-site blocking antisense (ASO-tsRNA). The ASO-tsRNA vector significantly elevated SMN levels in primary SMA patient fibroblasts, within the central nervous system of SMA mice and increased SMN-dependent in vitro snRNP assembly. These results demonstrate that the ASO-tsRNA strategy provides insight into the trans-splicing mechanism and a means of significantly enhancing trans-splicing activity in vivo. Citati