Progress Update: Base editing of DUX4 somatic polyadenylation signal

Report by Darina Šikrová

The focus of this grant is to test a new therapeutic strategy for FSHD by selectively mutating an important DNA regulatory element for DUX4 mRNA production (known as polyadenylation signal) in a targeted and efficient manner. Polyadenylation signals are crucial elements for efficient processing of mRNA transcripts and mutations in polyadenylation signals have been known to result in lower mRNA production. If this would be applicable for DUX4 gene, the benefit of such "hit-and-run" approach is, in theory, a one-time delivery of the therapeutic component leading ultimately to continuous lower DUX4 production in muscles and thus avoiding a life-long repeated treatment interventions as is required for therapeutic strategies such as small molecules or antisense oligonucleotides. To achieve this we use a modified CRISPR/Cas9 technology which allows for more precise and controlled mutagenesis outcome at the desired locus.

We have tested this strategy in four different myogenic cell lines derived from four donors with a diagnosis of FSHD (2x FSHD1 and 2X FSHD2) and observed in three of them the desired effect. We are currently investigating the cause of a lack of effect in the fourth sample. Common feature between these three cell lines which differentiate them from the fourth cell line is the D4Z4 specific variant.

To test this approach in an FSHD mouse model, we are currently exploring and optimizing different delivery methods. Once available, we will first determine the optimal conditions in tissue culture and next proceed to animal models.

See grant Base editing of DUX4 somatic polyadenylation signal