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

TL;DR

This paper introduces a protocol using patient-derived urine cells to test exon skipping therapies for Duchenne muscular dystrophy, improving drug screening predictability.

Contribution

A novel platform using MYOD1-transduced urine-derived cells for evaluating exon skipping therapy in Duchenne muscular dystrophy.

Findings

MYOD1-transduced urine-derived cells can be used to assess exon skipping efficacy in DMD.

CD90-positive cells show high myogenic potency and are suitable for drug screening.

The platform improves predictability for antisense oligonucleotide screening in DMD.

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.1

•Protocol to isolate UDCs from DMD patient-derived urine•Instructions for FACS of CD90-positive UDCs, which possess high myogenic potency•Steps for evaluating ASO efficacy in MYOD1-converted UDCs

Protocol to isolate UDCs from DMD patient-derived urine

Instructions for FACS of CD90-positive UDCs, which possess high myogenic potency

Steps for evaluating ASO efficacy in MYOD1-converted UDCs

Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

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.

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