Abstract Details

Heterozygous mutations in the VCP gene result in a combination of hereditary Inclusion Body Myopathy, Paget’s Disease of Bone, Frontotemporal Dementia and Amyotrophic Lateral Sclerosis named as VCP disease, or MSP. There is no cure for the disease, but one of the highly promising treatment strategies is CRISPR/Cas9 gene-editing technology.  

Valosin-containing protein (VCP) multisystem proteinopathy (MSP) is a neurodegenerative and ultimately fatal disease. Given the genetic background of the disease (dominant negative pathology) we invented a novel method called REMEDY for allele-specific targeting of heterozygous mutations.

In patient-derived fibroblast, three of the four gRNAs designed showed specificity to the mutant allele, and they were tested in VCPR155H/+ mouse myoblasts. All three gRNAs resulted in complete deletion of the mutant allele (100%). Interestingly, we observed a slight correction of the mutant allele as well, and without the need for an exogenous DNA template. Therefore, the frequency of the wild-type allele in the cells targeted by one of the gRNAs increased from 50% (normalized levels) to 58% measured by Sanger sequencing and next generation sequencing (NGS) and analyzed by ICE (interference of CRISPR Edits) and CRISPResso. 

We demonstrated precise targeting of the mutant allele in VCP patient-derived fibroblast and in mouse-derived myoblasts. Our approach has the potential to significantly advance the therapeutics and correct VCP disease in patients-derived cells.