The term epidermolysis bullosa (EB) encompasses an inherited clinically heterogenous group of skin conditions with a frequency of 1 in 17,000 live births (1). At least 20 different genes are associated with the diagnosis of EB, which displays profound genetic heterogeneity, further emphasized by the phenomenon of one gene leading to both autosomal-dominant and autosomal-recessive pattern inheritance with different phenotypes (2,3). The involved genes encode structural proteins, essential for intraepidermal adhesion, dermo-epidermal anchorage and skin integrity (3,4). Thus, the main clinical symptom is mechanical skin fragility presenting as mucocutaneous blistering, erosions and ulceration caused by minimal trauma or friction (5). Extracutaneous manifestation (oral, oesophageal, tracheal, genitourinary and ocular mucous membranes) has also been documented. According to the most recent consensus reclassification from 2020, the 4 main types of EB based on the lesions' location are EB simplex, junctional EB, dystrophic EB (DEB) and Kindler's EB (4). Currently, the most effective diagnostic approach for EB includes transmission electron microscopy and immunofluorescence antigen mapping of affected skin, along with DNA mutational analysis. The genetic test is a particularly important component of the diagnostic process for diagnostic confirmation (with a subtype), as it also provides the basis for proper genetic counseling and allows the application of prenatal diagnosis. At present, next-generation sequencing (NGS) is considered the preferable strategy, as it offers the ability to screen for multiple sequence variations and is less expensive (5).

The current study presents a case report of a female patient with a clinical diagnosis of dystrophic epidermolysis bullosa, identifying a novel variant in the type VII collagen α1 chain (COL7A1) gene.

DEB is the second most common type of EB, accounting for 30% of all cases (8). It can be inherited in an autosomal-dominant and autosomal-recessive manner. Both modes of inheritance are attributed to mutations in the COL7A1 gene, which encodes type VII collagen. The latter is an essential component of the anchoring fibrils and is responsible for epidermal basement membrane attachment to the dermal extracellular matrix proteins (9). Thus, the anchoring fibrils in patients with DEB are morphologically altered and lead to sub-lamina densa plane of tissue separation (10). The clinical manifestation exhibits great variability-from isolated nail dystrophy through to mild localized skin blistering to generalized blistering with extracutaneous involvement (11). The dominant subtype of DEB (DDEB) tends to exhibit less severe symptoms. Typical manifestations include blistering limited to hands, feet, knees, elbows associated with milia, atrophic scarring and nail dystrophy. Еxtracutaneous involvement is rare (12). Recessive DEB (RDEB) is less prevalent (1). The most common RDEB subtypes are the severe generalized type (formerly known as Hallopeau-Siemens) and intermediate RDEB (formerly known as non-Hallopeau-Siemens) (13). Severe RDEB manifests as congenital widespread skin blistering resulting in extensive scarring with pseudosyndactyly, alopecia, extracutaneous involvement such as corneal abrasions, mucous membrane blistering, oesophageal strictures, kidney problems and cardiomyopathy (13,14). Intermediate RDEB exhibits less severe blistering with oral, dental, hair and nail changes (14,15). Extracutaneous complications and pseudosyndactyly are less common (16). Both subtypes are associated with an increased risk of squamous cell carcinoma (15). The COL7A1 gene displays great allelic heterogeneity, as >800 genetic variants have been documented (17). It is difficult to define clear genotype-phenotype correlations, given the massive number of listed pathogenic variants and the clinical overlapping among DEB subtypes.

However, certain patterns stand out. Multiple mutations occur in exons 73-75, where 75% of DDEB pathogenic variants reside. DDEB is predominantly a result of glycine substitutions within the triple helix of the COL7A1 gene, which affects the stability of fibrils involved in dermo-epidermal anchorage (10,18). Severe RDEB is generally associated with premature termination codon (PTC) in both COL7A1 alleles due to nonsense, frameshift and splice-site mutations, leading to an insufficiency or a total lack of production of type VII collagen. Intermediate RDEB results from compound heterozygosity of the PTC variant on one allele and missense variant on the other or simultaneous presence of two missense variants on both alleles, i.e. at least one missense variant in the COL7A1 gene is always present. The expected outcome is the production of somewhat functional but abnormal type VII collagen (10,14). Clinical manifestation of DDEB and intermediate RDEB tends to be indistinguishable, posing diagnostic difficulties, particularly if there is no family history of EB (19).

In the presented case, the patient was assigned a diagnosis of DEB. There was no family history of similar skin conditions. The presented manifestation was relatively mild and classifying it preliminarily as dominant or recessive was difficult. Therefore, a segregation approach was considered most suitable for diagnostic precision-the patient and the patient's parents were simultaneously tested via NGS. The sequence data analysis revealed a compound heterozygous state in the patient's COL7A1 gene associated with the recessive form of DEB. Segregation analysis of the patient's parents further revealed their heterozygous carrier status and thus established the parental origin of both COL7A1 variants. The paternal variant was found to be c.6022C>T p.(Arg2008Cys). It is a missense located in exon 73 that leads to substitution of the amino acid arginine (basic and polar) by cysteine (neutral and slightly polar) at codon 2,008 of the COL7A1 protein. The Arg2008 amino acid is considered to be clinically significant. Reported variants that disrupt it have been classified as pathogenic. In silico analysis shows that the variant has a deleterious effect on the protein encoded by the COL7A1 gene. It has been reported in individuals affected by DEB (https://www.ncbi.nlm.nih.gov/clinvar/variation/522791/). According to the Genome Aggregation Database (GnomAD; https://gnomad.broadinstitute.org/), the frequency of the c.6022C>T p.(Arg2008Cys) variant regarding the non-Finnish European population is 0.000009. Based on the American College of Medical Genetics and Genomics (ACMG) guidelines (7), it was classified as pathogenic. The variant of maternal origin was found to be c.3474del p.(Val1160Ter). It is a novel one that had not been identified before the analysis. Its frequency is unknown as it is absent from the GnomAD database. The variant is a deletion of a single nucleotide that leads to the formation of a PTC in exon 26, out of 118 exons. It is expected to result in an absent or truncated protein product. According to the ACMG guidelines, it was predicted to be likely pathogenic.

In agreement with the literature data, the case presents as an RDEB with a milder manifestation associated with a missense and a PTC variant in a compound heterozygous state. The missense mutation is situated in exon 73, even though the type of DEB is not dominant. The inconclusive phenotype and lack of family history necessitated DNA testing to establish the genetic origin of DEB in the patient along with the right pattern of inheritance. Not only did it provide an exact genetic diagnosis, but it also allowed for informed reproductive choices to be undertaken within the family. Establishing the carrier status of patients' parents for variants in the COL7A1 gene determines a 25% risk for each of their subsequent pregnancies to end up with an affected offspring. Concerning the optimization of patient reproduction, it is recommended that the patient's future partner is tested for their COL7A1 gene carrier status. The latter is necessary to estimate the risk for an affected offspring. If no pathogenic variants in the COL7A1 gene of the partner are detected, the risk is virtually zero. Detection of carrier status determines a 50% risk for an affected offspring.

In conclusion, the presented case with detected variants in the COL7A1 gene may contribute to the establishment of more certain genotype-phenotype correlations. The overlapping of symptoms in different subtypes of DEB requires genetic testing via NGS to determine the exact inheritance pattern. The latter is essential information for accurate genetic counseling regarding the subsequent reproduction of the patients' parents and the patients' own reproduction.