Development of next-generation AAV vectors for effective CRISPR inhibition of FSHD
• Amount awarded: US$35,000
• FSHD Canada Foundation Contribution: US$17,500
• Date awarded: August 24, 2018 (for six months)
Investigator: Charis L. Himeda, PhD, Research Assistant Professor, University of Nevada, Reno School of Medicine, Reno NV USA
CRISPR technology provides an avenue for specifically targeting and manipulating virtually any DNA sequence in the human genome. This technology is typically used for gene editing, in which DNA is cut and replaced, allowing a disease-causing mutation to be converted to the normal sequence. Alternatively, CRISPR technology can be used to change the expression level of a disease-causing gene (e.g., CRISPR inhibition turns a gene off; CRISPR activation turns a gene on) without cutting the genome. Thus, CRISPR-based technology has the potential to cure the root cause of a disease such as FSHD, rather than merely treating symptoms. In addition, CRISPR-based therapies should have long-term or even permanent beneficial effects. FSHD is caused by mis-expression of the DUX4 gene in skeletal muscle and could potentially be cured by either CRISPR editing or CRISPR inhibition. However, since FSHD is associated with highly repetitive DNA sequences, there are a number of concerns with a CRISPR editing approach that introduces many cuts into the genome. CRISPR inhibition circumvents these problems and is a viable approach for FSHD. In a proof-of-principle study, we demonstrated that CRISPR inhibition components can be successfully targeted to the disease locus to repress DUX4 expression in human FSHD skeletal muscle cells. This new project builds on our continuing work, allowing us to engineer novel and more effective constructs for CRISPR inhibition. We aim to improve skeletal muscle delivery and expression of CRISPR components using adeno-associated virus (AAV) vectors, and to achieve long-term silencing of DUX4 using CRISPR targeting of several different epigenetic regulators.