使用MYOD1转导的人尿液来源细胞作为杜氏肌营养不良症外显子跳跃疗法预测平台的方案。

Protocol for using MYOD1-transduced human urine-derived cells as a predictive platform for exon skipping therapy in Duchenne muscular dystrophy.

作者信息

Kunitake Katsuhiko, Ishizuka Takami, Takeshita Eri, Komaki Hirofumi, Aoki Yoshitsugu

机构信息

Department of Molecular Therapy, National Institute of Neuroscience, National Center of Neurology and Psychiatry (NCNP), Kodaira, Tokyo 187-8502, Japan.

Translational Medical Center, National Center of Neurology and Psychiatry (NCNP), Kodaira, Tokyo 187-8502, Japan.

出版信息

STAR Protoc. 2025 Jun 20;6(2):103856. doi: 10.1016/j.xpro.2025.103856. Epub 2025 Jun 6.

DOI:10.1016/j.xpro.2025.103856

PMID:40483693

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12173603/

Abstract

Antisense oligonucleotide (ASO)-based exon skipping is a splice-modulating therapy effective for Duchenne muscular dystrophy (DMD) caused by dystrophin deficiency. Here, we present a protocol for evaluating exon skipping efficacy in MYOD1-transduced human urine-derived cells (MYOD1-UDCs) from patients. We describe steps for isolating UDCs, selecting CD90-positive cells, inducing myogenic differentiation, and assessing the restoration of DMD mRNA and proteins after exon skipping. This platform enhances the predictability of ASO screening, promoting early-stage drug discovery and translational research in DMD. For complete details on the use and execution of this protocol, please refer to Komaki et al..

摘要

基于反义寡核苷酸（ASO）的外显子跳跃是一种对由肌营养不良蛋白缺乏引起的杜氏肌营养不良症（DMD）有效的剪接调节疗法。在此，我们展示了一种评估来自患者的MYOD1转导的人尿源细胞（MYOD1-UDCs）中外显子跳跃效率的方案。我们描述了分离UDCs、选择CD90阳性细胞、诱导肌源性分化以及评估外显子跳跃后DMD mRNA和蛋白质恢复情况的步骤。该平台提高了ASO筛选的可预测性，促进了DMD的早期药物发现和转化研究。有关本方案使用和执行的完整详细信息，请参考Komaki等人的研究。