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Gene & Protein in Disease Gene therapy progress for DEB

transcription template together with targeting guide Spontaneous DEB animal models have been identified
RNA. Compared to base editors, PE systems have a more in various species, including sheep, 90,91 cats, dogs,
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flexible targeting scope and can enable gene modification mice, cattle, horses, goats, and rats. These models
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in a precise and targeted manner. However, up to now, arise from naturally occurring genetic mutations that lead
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interest in applying PE to gene therapy for RDEB has been to DEB-like blistering phenotypes. Such models provide
limited. Hong et al. explored the potential of a PE system valuable insights into the pathology and mechanisms
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in correcting two mutations (c.2005C>T and c.3631C>T) underlying DEB. Among DEB animal models, the
in RDEBF. Although gene correction efficiency was only transgenic mouse model (Table 1) stands out for its utility
10.5% in the bulk population of cells, the formation of AFs in dissecting molecular mechanisms and testing gene-,
was observed at the dermo-epidermal junction in mice protein-, and cell-based therapies. The Col7a1 gene-
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grafted with skin equivalents consisting of RDEB patient- knockout mouse has been instrumental in pilot studies.
derived keratinocytes and PE-corrected RDEBF. Steinbeck However, its short lifespan poses challenges for long-
et al. reported a twin PE (twinPE) strategy utilizing the term follow-up and limits the interpretation of results.
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PEmax and PE6 primer editors to replace the target exon To mitigate this issue, hypomorphic mice with reduced
with recombinase attachment sequences, subsequently C7 expression are used in preclinical studies. These
reintroducing a wild-type copy of the target exon to restore mice exhibit milder phenotypes, allowing the majority
the C7 production in RDEB skin cells. to survive up to 20 weeks with improved husbandry
practices. 100,101 In addition, an inducible Col7a1-knockout
3. DEB models for gene therapy validation mouse model has been developed to further explore the
Understanding pathophysiology and developing effective role of C7 in wound re-epithelialization and granulation
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treatments for DEB requires robust model systems that tissue maturation. Recent advancements in genomic
accurately mimic human genomic and clinical features. engineering have significantly enhanced the generation
Various animal models, including in vitro, ex vivo, and of specific animal models for DEB research. Transgenic
mouse models have been established with common human
in vivo systems, have been established to facilitate DEB mutations, providing a more accurate representation of
research.
the relationship between phenotype and genotype, as well
In vitro models serve as the initial proof of concept for as the secondary effects of the environment. 103,104 These
studying pathophysiology and validating curative therapies. models are crucial for understanding the complex interplay
DEB patient-derived skin cells, specifically keratinocytes, between genetic mutations and clinical manifestations
and fibroblasts, are cultured in 2D environments to study of DEB. However, to meet the demand for validating the
cellular behaviors and test potential treatments. 49,83,84 rapid development of gene therapies, a more suitable DEB
While these models provide essential insights, they lack animal model is still needed.
the complexity of living tissue. To bridge the gap between
simple cell cultures and complex in vivo systems, ex vivo 4. Administration of gene therapy for DEB
models are employed. Epidermal skin equivalents and full- One of the primary challenges in gene therapy is the
thickness human skin equivalents are commonly used in effective delivery of treatments to the target site. For skin
DEB research. 78,85-87 These models simulate the architecture conditions, there are various ways to apply the treatment
and inter-cellular interactions of human skin. By studying directly to the skin or wound area. Genetically modified
these skin equivalents, researchers can better understand cells can be transplanted or grafted onto DEB patients
DEB’s impact on skin integrity and evaluate therapeutic (Figure 1). However, this presents several challenges
strategies. and risks. For instance, grafting techniques entail the
For a comprehensive analysis, in vivo models are risk of engraftment failure, infection, and general
indispensable. Engineered epidermal sheets transplanted anesthesia requirements while also being labor-intensive.
onto immunodeficient mice serve as an effective in vivo Alternatively, treatment can be applied by intradermal
model for DEB, particularly valuable for evaluating the injection, but this may cause intolerable pain in chronic
efficacy of cell therapies. 28,88,89 Furthermore, this model wounds that often present with scarring and fibrosis as
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enables in vivo editing of the human COL7A1 gene in complications of DEB.
mice, facilitating the study of gene therapies within a living Topical treatments are an attractive approach, as they
organism. However, the time-consuming process, low can be applied directly to the wound during routine
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graft success rate, and the necessity of obtaining primary bandage changing. However, topical skin treatment therapy
skin cells from DEB patients have limited the accessibility development is hindered by a lack of an ideal delivery system
of these models for researchers working on DEB. for genetic material, particularly for delivery in chronic

Volume 3 Issue 3 (2024) 6 doi: 10.36922/gpd.4047