Leila Youssefian

Klippel–Trenaunay syndrome belongs to the PIK3CA-related overgrowth spectrum (PROS)

Klippel–Trenaunay syndrome (KTS), originally described as a triad of cutaneous capillary malformation, bone and soft‐tissue hypertrophy, as well as venous and lymphatic malformations, has been considered by dermatologists as a distinct diagnostic entity. However, cases with KTS have also been reported to have neurological disorders, developmental delay and digital abnormalities, indicating multisystem involvement. Recently, a number of overgrowth syndromes, with overlapping phenotypic features with KTS, have been identified; these include MCAP and CLOVES syndromes as well as fibroadipose hyperplasia. These conditions harbour mutations in the PIK3CA gene, and they have been included in the PIK3CA‐related overgrowth spectrum (PROS). Based on recent demonstrations of PIK3CA mutations also in KTS, it appears that, rather than being a distinct diagnostic entity, KTS belongs to PROS. These observations have potential diagnostic and therapeutic implications for KTS.

Molecular pathology of the basement membrane zone in heritable blistering diseases:: The paradigm of epidermolysis bullosa.

Epidermolysis bullosa (EB), a phenotypically heterogeneous group of skin fragility disorders, is characterized by blistering and erosions with considerable morbidity and mortality. Mutations in as many as 18 distinct genes expressed at the cutaneous basement membrane zone have been shown to be associated with the blistering phenotype, attesting to the role of the corresponding proteins in providing stable association of the epidermis to the dermis through adhesion at the dermo-epidermal basement membrane zone. Thus, different forms of EB have been highly instructive in providing information on the physiological functions of these proteins as integral components of the supramolecular adhesion complexes. In addition, precise information of the underlying genes and distinct mutations in families with EB has been helpful in subclassification of the disease with prognostic implications, as well as for prenatal testing and preimplantation genetic diagnosis. Furthermore, knowledge of the types of mutations is a prerequisite for application of allele-specific treatment approaches that have been recently developed, including read-through of premature termination codon mutations and chaperone-facilitated intracellular transport of conformationally altered proteins to proper physiologic subcellular location. Collectively, EB serves as a paradigm of heritable skin diseases in which significant progress has been made in identifying the underlying genetic bases and associated aberrant pathways leading from mutations to the phenotype, thus allowing application of precision medicine for this, currently intractable group of diseases.

Dystrophic Epidermolysis Bullosa: COL7A1 Mutation Landscape in a Multi-Ethnic Cohort of 152 Extended Families with High Degree of Customary Consanguineous Marriages

Dystrophic epidermolysis bullosa is a heritable skin disease manifesting with sub-lamina densa blistering, erosions, and chronic ulcers. COL7A1, encoding type VII collagen, has been identified as the candidate gene for dystrophic epidermolysis bullosa. In this study, we have identified COL7A1 mutations in a large multi-ethnic cohort of 152 extended Iranian families with high degree of consanguinity. The patients were diagnosed by clinical manifestations, histopathology, and immunoepitope mapping. Mutation detection consisted of a combination of single nucleotide polymorphism-based whole-genome homozygosity mapping, Sanger sequencing, and gene-targeted next-generation sequencing. A total of 104 distinct mutations in COL7A1 were identified in 149 of 152 families (98%), 56 (53%) of them being previously unreported. Ninety percent of these mutations were homozygous recessive, reflecting consanguinity in these families. Three recurrent mutations were identified in five or more families, and haplotype analysis suggested a founder effect in two of them. In conclusion, COL7A1 harbored mutations in the overwhelming majority of patients with dystrophic epidermolysis bullosa, and most of them in this Iranian cohort were consistent with autosomal recessive inheritance. The mutation profile attests to the impact of consanguinity in these families.

Gene-Targeted Next Generation Sequencing Identifies PNPLA1 Mutations in Patients with a Phenotypic Spectrum of Autosomal Recessive Congenital Ichthyosis: The Impact of Consanguinity

Autosomal recessive congenital ichthyosis is a heterogeneous group of disorders associated with mutations in at least nine distinct genes. To ascertain the molecular basis of ichthyosis patients in Iran, a country of approximately 80 million people with a high prevalence of customary consanguineous marriages, we have developed a gene-targeted next generation sequencing array consisting of 38 genes reported in association with ichthyosis phenotypes. In a subset of nine extended consanguineous families, we found homozygous missense mutations in the PNPLA1 gene, six of them being distinct and, to our knowledge, previously unpublished. This gene encodes an enzyme with lipid hydrolase activity, important for development and maintenance of the barrier function of the epidermis. These six mutations, as well as four previously published mutations, reside exclusively within the patatin-like subdomain of PNPLA1 containing the catalytic site. The mutations clustered around the active center of the enzyme or resided at the surface of the protein possibly involved in the protein-protein interactions. Clinical features of the patients showed considerable intra- and interfamilial heterogeneity. Knowledge of the specific mutations allows identification of heterozygous carriers, assisting in genetic counseling, prenatal testing, and preimplantation genetic diagnosis in extended families at risk of recurrence of this disorder, the incidence of which is significantly increased in consanguineous marriages.

Fibroadipose Hyperplasia versus Proteus Syndrome: Segmental Overgrowth with a Mosaic Mutation in the PIK3CA Gene

Hassan Vahidnezhad, Mohammadreza Barzegar, Qiaoli Li, Soheila Sotoudeh, Ameneh Yazdanfar, Amir Hooshang Ehsani, Abdol-Mohammad Kajbafzadeh, Nikoo Mozafari, Nasser Ebrahimi Daryani, Farzaneh Agha-hosseini, Sirous Zeinali and Jouni Uitto Tehran University of Medical Sciences, Tehran, Iran; Thomas Jefferson University, Philadelphia, PA, USA; Pasteur Institute of Iran, Tehran, Iran; Shahid Beheshti University of Medical Sciences, Tehran, Iran and Hamedan University of Medical Sciences, Hamedan, Iran E-mail: Jouni.Uitto@jefferson.edu

Recessive mutation in tetraspanin CD151 causes Kindler syndrome-like epidermolysis bullosa with multi-systemic manifestations including nephropathy

Epidermolysis bullosa (EB) is caused by mutations in as many as 19 distinct genes. We have developed a next-generation sequencing (NGS) panel targeting genes known to be mutated in skin fragility disorders, including tetraspanin CD151 expressed in keratinocytes at the dermal-epidermal junction. The NGS panel was applied to a cohort of 92 consanguineous families of unknown subtype of EB. In one family, a homozygous donor splice site mutation in CD151 (NM_139029; c.351+2T>C) at the exon 5/intron 5 border was identified, and RT-PCR and whole transcriptome analysis by RNA-seq confirmed deletion of the entire exon 5 encoding 25 amino acids. Immunofluorescence of probands skin and Western blot of skin proteins with a monoclonal antibody revealed complete absence of CD151. Transmission electron microscopy showed intracellular disruption and cell-cell dysadhesion of keratinocytes in the lower epidermis. Clinical examination of the 33-year old proband, initially diagnosed as Kindler syndrome, revealed widespread blistering, particularly on pretibial areas, poikiloderma, nail dystrophy, loss of teeth, early onset alopecia, and esophageal webbing and strictures. The patient also had history of nephropathy with proteinuria. Collectively, the results suggest that biallelic loss-of-function mutations in CD151 underlie an autosomal recessive mechano-bullous disease with systemic features. Thus, CD151 should be considered as the 20th causative, EB-associated gene.

Multigene Next-Generation Sequencing Panel Identifies Pathogenic Variants in Patients with Unknown Subtype of Epidermolysis Bullosa: Subclassification with Prognostic Implications

Please cite this article as: Vahidnezhad H, Youssefian L, Saeidian AH, Touati A, Sotoudeh S, Abiri M, Barzgar M, Aghazadeh N, Mahmoudi H, Norouz-zadeh S, Hamid M, Zahabiyon M, Bagherian H, Zeinali S, Fortina P, Uitto J, Multigene Next Generation Sequencing Panel Identifies Pathogenic Variants in Patients with Unknown Subtype of Epidermolysis Bullosa: Subclassification with Prognostic Implications, The Journal of Investigative Dermatology (2017), doi: 10.1016/j.jid.2017.07.830.

Genotypic Heterogeneity and the Mode of Inheritance in Epidermolysis Bullosa

Epidermolysis bullosa (EB) comprises a clinically heterogeneousgroupofdisorderscharacterizedbyfragilityof skin, leading to formation of blisters, erosions, and chronic ulcers. The cutaneousmanifestations, together with extracutaneous complications, cause considerable morbidity and in some cases premature death.1,2 Epidermolysis bullosa is anorphandisease (defined in theUnited States as adiagnosiswith <200000affected individuals), yet thereareup to40000affected individuals in theUnitedStates and asmany as half a million patients globally. The disease is characteristically diagnosed at birth or during the early postnatal period, and there is currently no effective and specific treatment beyond prevention of trauma, appropriate wound care, and prevention of infections. Thus, EB imposes amajor burden for global health care, and the cost of the treatment of a severely affected patient in the United States can approach

Lipoid proteinosis: phenotypic heterogeneity in Iranian families with c.507delT mutation in ECM1

300000peryear (email communication;November24,2015; Brett Kopelan, executive director of the dystrophic epidermolysis bullosa research association [DEBRA] of America). The heritable forms of EB have been divided into 4 broad categories based on the level of blister formation within the dermoepidermal basement membrane zone (BMZ) as determined by immunoepitope mapping or diagnostic transmission electronmicroscopy (Table). In the classic simplex (EBS) junctional (JEB) anddystrophic (DEB) formsofEB, tissue separation is within the basal cell layer of epidermis, within the lamina lucida of the BMZ, and sublamina densa, respectively, whereas the Kindler syndrome (KS), which has been recently classified as a form of EB, can demonstrate multiple levels of blistering.Asmanyas 18different geneshavenowbeenshown to harbormutations resulting in EB phenotypes (Table).3 The tremendous clinical variability noted in patients with EB can be explained by the topographic location of expression of the mutant geneswithin the BMZ, the types and combinations of mutations in these genes, and their interactionswith environmental factors, particularly trauma, at the genome/environment interface. The mode of inheritance of EB can be variably either autosomal dominant (AD) or autosomal recessive (AR).1 Thehistorical suggestions of X-linked forms of EB have been discounted by molecular diagnostics. Specifically, EBS is AD in most cases, although approximately 25%of familiesmayhave anARmodeof inheritance.Thedystrophic formscanbeeither AD or AR, the dominant forms being usually less severe, whereas the most devastating type, recessive dystrophic EB, thegeneralized severe, is inherited in anARpattern. The junctional forms aswell asKShavebeenuniformlydescribedwith AR inheritance.4,5 In this issue, Turcan and coworkers6 report a novel heterozygousmissensemutation in the ITGB4gene,whichcosegregates with the JEB phenotype in an extended family in an apparent AD pattern. This gene encodes an integrin polypeptide (β4), which physiologically dimerizes with another integrin polypeptide (α6) to form a cell surface receptor, α6β4 integrin (Figure).Thus,bothpolypeptides,α6andβ4,are required for formation of this protein that is critical for cell-matrix interaction through their association with laminin 332.7 These proteins form hemidesmosomes, multiprotein attachment complexes stabilizing the adherence of the epidermis to the underlying basement membrane. This mutation was found only in the affected patients and was not present in unaffectedmembers of the family. The authors6 conclude that this study highlights, for the first time, the possibility of a dominant mode of inheritance in the JEB, thus expanding the genotypic heterogeneity and extending the genotype-phenotypecorrelations forEB.They6alsopostulate,butdonotprove, that the effect of thismutation is dominant negative. In other words, the mutated gene product interferes with the functionof thenormal product, thusdestabilizing the assembly of hemidesmosomes and rendering them nonfuncational (Figure). A large number ofmutations in the candidate genes have beenpreviously identified indifferent formsofEB,andthemutations in the same gene can result either in an AD or AR inheritance pattern.3 For example, most mutations in the keratin 14 gene (KRT14) result inAD inheritance, but 14 cases have been shown toharbormutations in this gene in amanner that results inARmodeof inheritance. Similarly, Turcanet al6 now demonstrate that a heterozygous missense mutation in the ITGB4 gene isAD,while all previously publishedmutations in the samegeneareAR.What then is theexplanation for themutations in the same gene giving rise to 2 different forms of inheritance? In all previously published cases of ITGB4, the mutations cause loss of function, as a result of premature termination codon of translation resulting in synthesis of truncated and nonfunctional polypeptides, for example.4 In heterozygous carriers of such mutations, half of the protein is truncated and nonfunctional whereas the normal allele results in the synthesis of functional full-length polypeptide (Figure, C). In this case, half of the physiological level of the protein is sufficient to maintain normal integrity of the BMZ, and only in complete absence of the functional protein, as a result of either homozygous or compound heterozygous Related article page 558 Opinion

Phenotypic spectrum of autosomal recessive congenital ichthyosis due to PNPLA1 mutation

Lipoid proteinosis (LP) is a rare autosomal recessive genodermatosis caused by loss‐of‐function mutations in the ECM1 gene, and previous studies have noted phenotypic variability. In this study, we examined 12 patients representing three Iranian families for clinical manifestations and genotyped them for mutations in ECM1. LP was diagnosed with characteristic mucocutaneous and neurologic manifestations. Five patients were also subjected to magnetic resonance imaging (MRI)/computed tomography (CT) scan of the central nervous system. DNA was isolated from peripheral blood from patients and their clinically unaffected relatives, and mutations in ECM1 were sought by PCR‐based amplification of all exons and flanking intronic sequences, followed by bidirectional Sanger sequencing. Significant phenotypic variability in this multisystem disorder, including presence of convulsions and epilepsy in about half of the patients was noted. In most cases, this was associated with calcifications in the brain detected by MRI/CT scans. Genotyping of the affected individuals in three families from the central region of Iran revealed presence of homozygous c.507delT mutation in ECM1, reflecting the observed consanguinity in these families. This large cohort revealed extensive phenotypic variability in individuals with the same mutation in ECM1. This observation suggests a role for genetic and epigenetic as well as environmental modulation of the phenotype. Identification of mutations allows screening of unaffected individuals for presence or absence of this mutation in extended LP families, with implications for genetic counseling.