## 4 Discussion
In this regard, many scientists are determining the efficacy of certain drugs, ointments, and gene therapy in treating Epidermolysis bullosa, especially in _COL7A1_ mutation-affected patients. [PERSON] et al., 2017 [20] used a (CRISPR/Cas9) method with microinjection into NOD/SCID IL2 rncull (NSG) embryos to swiftly construct an immunodeficient Col7a1/animal model of RDEB for experimental purposes. They also recognized the ability of a newly discovered skin-resident immunomodulatory dermal mesenchymal stem cell, ABCB5, to minimize disease and significantly increase the life expectancy of these (RDEB NSG) mice by reducing skin inflammatory myeloid-derived penetration. Double-strand breaks are induced via Nuclease-based genome editing at certain genomic sites ([PERSON], 2019). Several approaches for modifying the genomes of fibroblasts and keratinocytes for therapeutic purposes have been successful [22; 23]. Changes in mutant allele reading frames for knockout [24; 25; 26] or coding sequence restoration [27; 28; 29] have been shown to be very effective. Dual RNP-mediated genome editing is efficient and homogenous, eliminating the need for plasmid-based delivery and selecting properly altered single-cell clones [30]. Moreover, recent advancements have uncovered more comprehensive, expectable consequences ensuing single targeting through CRISPR/Cas9 at various loci [31; 32]. [PERSON] used a CRISPR/Cas9 nuclease as a ribonucleoprotein to induce a consistent, predicted single adenine sense-strand insertion at the target position in primary wild-type and recessive DEB keratinocytes [33]. This shows that _COL7A1_ editing based on precise end-joining-mediated DNA repair is an efficient strategy for reversing the characteristics of DEB associated with the disease, regardless of the inheritance pattern of the disease. Similarly, [PERSON] et al., 2022 showed for the first time the potential of endogenous 50 trans-splicing as a useful tool for beneficially modulating the RDEB-phenotype by correcting pathogenic mutations within the COL7A1 gene, thereby addressing an essential need of this patient population [34]. In this study, Family A was diagnosed as being affected by epidermolysis bullosa recessive Dystrophic EB, generalized severe type, which is a rare sub-type of DEB, associated with mutations of the _COL7A1_ gene. Clinically, it is characterized by sublamina densa blistering that leads to erosions and poor wound healing, persistent ulcers, and significant scarring, notably on the hands and feet, which are exposed to frequent trauma [35]. The proband, a 4-year-old boy (V-2) in Family A, had a history of abnormal blister formation on the skin, erosions and atypical scarring with erythemat hypersurfacequels, and keloids affecting his trunk (Figure 2(Aa-c)), arms, hands, feet, elbows, knees, and genitalia. He also had nail dystrophy, mitten deformities, pseudo syndactyly and dysphagia, as described in previous studies [17; 36]. WES revealed a novel missense mutation variant c.7034G\(>\)A, p. Gly2345 Asp in exon 91 of _COL7A1_ (Table 1). This variant c. 7034G\(>\)A was predicted to be disease producing by Mutation Taster, damaging by Polyphen-2, SIFT and Fathmm, and pathogenic by VarSome and MudPred, and was shown to have decreased stability by MUPRO (Table 2). In Family B, homozygous novel frameshift sequence variant c385 del (p. His129 MetfsTer18) in the 3 rd exon of the _COL7A1_ gene was identified by WES, which results in generalized severe RDEB in the affected members of the family (Table 1). The proband IV-4 demonstrated abnormal generalized blistering of the skin and extensive scarring of the whole body. Involvement of the mucosal membranes has been documented in some patients, causing difficulty with oral feeding [36], as observed in our patient. Opening of the mouth was restricted, along with ankyloglossia and feeding difficulties. The limbs show onychysis, poly syndactly, and restriction of movement due to flexion contracture of joints.