P28 In vivo targeting of basal keratinocytes using lipid nanoparticles in a dominant dystrophic epidermolysis bullosa mouse model
Prashant Kumar, Ina Guri-Lamce, Carina Graham, Isabelle Benson, Nadira Ali, Panicos Shanagris, Stephen L Hart, Matthew Caley, John A McGrath, Emanuel Rognoni, Joanna Jacków-MalinowskaAbstract
Introduction and aims
Gene editing holds strong therapeutic promise for inherited skin disorders, but translation to in vivo application requires clinically relevant animal models. Dystrophic epidermolysis bullosa (DEB), caused by pathogenic COL7A1 variants, is a severe condition for which existing mouse models are poorly suited to testing gene-editing therapies. This study aimed to establish and characterize a dominant DEB (DDEB) mouse model for in vivo gene-editing evaluation and to assess the feasibility of lipid nanoparticle (LNP)-mediated delivery to skin cells.
Methods
A DDEB mouse model heterozygous for the Col7a1 exon 73 c.6085G>C variant was utilized. Phenotypic assessment focused on blistering severity and wound-healing dynamics. Cells isolated from mouse skin biopsies were analysed by Western blot to assess intracellular accumulation and extracellular secretion of type VII collagen. Topical delivery of self-formulated LNPs encapsulating mCherry mRNA was tested on wounded mouse skin to evaluate cellular targeting and delivery efficiency. In parallel, two gene-editing strategies were established in vitro: a deaminase-free, glycosylase-based cytosine base editor (gCBE) to correct the C>G mutation, and CRISPR-Cas9-mediated knockout of the mutant allele to mitigate the dominant-negative effect.
Results
The DDEB mouse model exhibited a mild baseline blistering phenotype, enabling longer-term studies, but showed delayed wound healing following injury, providing a suitable context for therapeutic testing. Western blot analysis revealed intracellular accumulation of C7 with reduced extracellular secretion. Topical application of LNPs resulted in mCherry expression in basal keratinocytes within the migrating epithelial tongue of healing wounds, indicating effective delivery to a therapeutically relevant target cell population. Gene-editing approaches are being established in vitro, with ongoing optimization to identify the most efficient strategy.
Conclusions
This DDEB mouse model provides a clinically relevant platform for in vivo gene-editing studies. Our results demonstrate topical LNP-mediated delivery to basal keratinocytes, supporting further development of gene-editing therapies for DDEB and other inherited skin disorders.