Duchenne Muscular Dystrophy Cell Culture Models Created By CRISPR/Cas9 Gene Editing And Their Application To Drug Screening

Abstract

Gene edition methods are an attractive putative therapeutic option for Duchenne muscular dystrophy and they have an immediate application in the generation of research models. To generate two new edited myoblast cultures that could be useful in vitro drug screening, we have optimised a CRISPR/Cas9 gene edition protocol. We have successfully used it in wild type immortalised myoblasts to delete exon 52 of the dystrophin gene: DMD{Delta}52-Model, modelling a common Duchenne muscular dystrophy mutation; and in patients immortalised cultures we have deleted an inhibitory microRNA target region of the utrophin UTR, leading to utrophin upregulation. We have characterised these cultures and, to show their use in the assessment of DMD treatments, we have performed exon skipping in the DMD{Delta}52-Model and have used the unedited cultures/ DMD-UTRN-Model combo to assess utrophin overexpression after drug treatment. While the practical use of DMD{Delta}52-Model is limited to the validation to our gene edition protocol, DMD-UTRN-Model offers a possible therapeutic gene edition target as well as a useful positive control in the screening of utrophin overexpression drugs.