| 研究実績の概要 |
Rhabdomyosarcoma (RMS) is the most common pediatric sarcoma in the world arising from skeletal muscle progenitors. Malfunction of the Duchenne muscular dystrophy-related gene, dystrophin was implicated in the growth and metastasis of tumor in RMS.
In this project, the applicants attempted to study dystrophin intron retention in RMS to identify targets for RNA modulation by Antisense Oligonucleotides (AO).
This year, the applicants successfully confirmed the retention of intron 40 and 58 in a variety of RMS cell lines using primers to amplify exons 40 to 41 and 57 to 59 regions respectively. Next, they performed sequencing analysis to confirm the nucleotide arrangement and size of the sequence of intron 40 (851bp) and 58 (608bp), respectively. From the result, intron 40 retention was registered in 2 of 3 RMS cells in two patterns; retention of the whole of intron 40 and/or the 3’ end of intron 40 (exon 41E), respectively. Direct sequencing analysis also showed retention of whole intron 58 in RMS. In addition to introns 40 and 58, intron retention analysis also showed mild intron 70 retention in RMS cells which was confirmed by direct sequencing. Therefore, the identified retained introns might have arisen from malfunction of transcription regulation. From the results, intron retention could be a good focus of targeted therapy in RMS.
Next year, the applicants are going to search for candidate exonic/intronic splicing enhancers (ESE/ISE) by web based algorithms within the retained introns or nearby exons for AO-induced splicing analysis.
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