Page 102 - ebook

[A. Biochemistry/Molecular Biology] A-51

Development of allele-specific CRISPR gene therapy for

Epidermolysis Bullosa Simplex Dowling-Meara (EBS-DM) with

single-base precision

Yohan An¹, Hyomin Lee², Junho K Hur¹,²*

¹Department of Genetics, College of Medicine, Hanyang University, Seoul 04736, Korea, ²Department of Medicine,

Major in Medical Genetics, Graduate School, Hanyang University, Seoul 04736, Korea

The Dowling-Meara variant of epidermolysis bullosa simplex (EBS-DM) is an inherited skin disorder typified by

blisters that primarily affect the skin after mechanical trauma. While EBS-DM is a severe disease that is associated
with significant morbidity in infancy and early childhood, effective treatment is currently unavailable. The genetic

cause of EBS-DM is dominant-negative mutations within either the KRT5 (12q13.13) or KRT14 (17q12-q21) genes,
encoding keratin 5 and keratin 14, respectively. The keratin proteins are highly abundant in the cytoplasm of basal

keratinocytes in the epidermis, providing cytoskeletal scaffold within the epithelial cells for protection against
mechanical and non-mechanical stresses. If mutations within either KRT5 or KRT5 genes occur, keratin 5 and keratin

14 filament network structures become fragile, which leads to epidermal cytolysis and blistering that are symptoms
of EBS-DM. However, there are currently no therapies approved for the treatment of EBS-DM, besides symptomatic
care. In this regard, gene therapy might be a potential option for corrective therapy for these patients. To this end,

in this study, we used CRISPR Cas9-gRNA to modify mutant KRT5 and KRT14 genes for therapy. We developed

allele-specific CRISPR-Cas9 methods to target specific mutations associated with EBS-DM.