Progress Update: DUX4 mRNA silencing using CRISPR-Cas13

Posted by Friends of FSH Research on Apr 20, 2021

Lay progress update submitted by Afrooz Rashnonejad
See original grant DUX4 mRNA silencing using CRISPR-Cas13

In this report, we focused on investigating the efficacy of CRISPR-Cas13b knockdown of DUX4 expression in an adult FSHD mouse model. To do that, we intramuscularly co-injected AAV-DUX4, AAV-Cas13b, and AAV-gRNA1 into one leg of C57BL/6 mice. The other leg was injected with only AAV-DUX4 as control. CRISPR-Cas13b reduced DUX4-induced muscle damage to 25% of the total damage level seen in the untreated legs. DUX4 and DUX4-associated biomarker expression levels decreased significantly in treated muscles. Cas13b and gRNA1 expression were detected up to 6 months in injected mouse muscles, demonstrating long-term expression and stability of CRISPR components in animal models. Finally, using specific staining methods, we showed that Cas13b and gRNA have been expressed in many muscle fibers due to the ability of AAV vectors to deliver therapeutic agents into muscle.

In conclusion, our in vitro and in vivo results using FHSD derived myotubes and a co-injected adult FSHD mouse model suggest promise for the AAV-CRISPR-Cas13 system to target and reduce DUX4 and to prevent DUX4-mediated cell death. However, the system still requires more optimization, such as increasing the system efficacy in vivo and investigating Cas13 off-target effects. We would like to thank the Friends of FSH Research for supporting the initial phase of this study.