Key points

Introduction

The pace of the study of inherited skin diseases is rapidly accelerating. Massively parallel (next-generation) sequencing technologies have created paradigm shifts in how these conditions are investigated. The time it takes to generate near-complete genetic data on an individual is now measured in days to weeks and the cost is decreasing rapidly. Advancement is ongoing and evolving third-generation sequencing technologies have generated speculation of generating a full human genome sequence overnight. The list of new techniques, fueled by the maturation of massively parallel DNA sequencing and bioinformatics technologies to handle the resultant data deluge, is continually growing, including linkage analysis, homozygosity mapping, case-control association, whole-genome genotyping, targeted resequencing, and whole-exome sequencing. New technologies bring new challenges. The genetic causes of many inherited skin conditions are being rapidly elucidated, leading to a deluge of genes and new data. Even for investigators in the field, keeping up with all of the new mutations and pathways at play is impractical. The subsequent deluge of “new genetic cause for disease X” articles being submitted to journals has led the Nature family of journals to publish new, more stringent guidelines for accepting articles reporting the genetic cause of a disease. However, it is critical to realize that the identification of causative genes is just the beginning of the journey to understanding the pathomechanism of an inherited skin disease. This article highlights a few of the many new discoveries in inherited skin diseases from the past year.

Introduction

The pace of the study of inherited skin diseases is rapidly accelerating. Massively parallel (next-generation) sequencing technologies have created paradigm shifts in how these conditions are investigated. The time it takes to generate near-complete genetic data on an individual is now measured in days to weeks and the cost is decreasing rapidly. Advancement is ongoing and evolving third-generation sequencing technologies have generated speculation of generating a full human genome sequence overnight. The list of new techniques, fueled by the maturation of massively parallel DNA sequencing and bioinformatics technologies to handle the resultant data deluge, is continually growing, including linkage analysis, homozygosity mapping, case-control association, whole-genome genotyping, targeted resequencing, and whole-exome sequencing. New technologies bring new challenges. The genetic causes of many inherited skin conditions are being rapidly elucidated, leading to a deluge of genes and new data. Even for investigators in the field, keeping up with all of the new mutations and pathways at play is impractical. The subsequent deluge of “new genetic cause for disease X” articles being submitted to journals has led the Nature family of journals to publish new, more stringent guidelines for accepting articles reporting the genetic cause of a disease. However, it is critical to realize that the identification of causative genes is just the beginning of the journey to understanding the pathomechanism of an inherited skin disease. This article highlights a few of the many new discoveries in inherited skin diseases from the past year.

Epidermolysis bullosa

A group of skin disorders characterized by skin fragility, blistering, and erosion, epidermolysis bullosa (EB) occurs because of inherited defects in structural proteins critical to maintaining cell structure and adhesion in the epidermis and dermoepidermal junction. Many of the EB subtypes exhibit great morbidity and some exhibit great mortality.

There are several exciting new developments in EB. Several recent reports have added to the understanding of EB subtypes, including the great range in severity that occurs in different forms of EB. A second patient was recently reported with a mild form of EB simplex caused by mutations in dystonin (DST; OMIM 113810 ), the gene encoding bullous pemphigoid antigen type 1 (BPAG1). The first patient with a DST mutation, long hypothesized as a potential cause of EB, was reported in 2010; however, the patient also had mutations in NOTCH3 (OMIM 600276 ) leading to neurologic features that were likely not related to the cutaneous phenotype. However, given the concomitant diagnoses it was difficult for investigators to determine what phenotypic findings were typical for BPAG1 mutations. In the newly reported patient (with a different BPAG1 mutation), neurologic features were absent and AR mutations in the DST gene led to a very mild EB simplex phenotype with blistering only with warmth or trauma.

A case of lethal congenital EB resulted from mutations in the junction plakoglobin (JUP; OMIM 173325 ) gene encoding plakoglobin (PLG). PLG plays a critical role in desmosomal function via interaction with desmoplakin, which serves to tether intermediate filaments to the desmosomal plaque. PLG is also present in adherens junctions. Previously, JUP mutations had been detected in Naxos syndrome (OMIM 601214 ), which exhibits wooly hair, palmoplantar keratoderma, and arrhythmogenic heart disease. The infant reported in this study exhibited severe skin fragility from birth with diffuse epidermal separation and massive transcutaneous water loss. The JUP mutation in this patient was unique and, instead of leading to a truncated plakoglobin protein as occurs in Naxos syndrome, led to complete PLG loss in the skin. Few desmosomes could be detected and those present were malformed. Because PLG plays a role in both desmosomal and adherens junction adhesion, its absence led to a complete loss of adhesion structures between keratinocytes.

A new subtype of EB has been described in three patients with homozygous mutations in the integrin (ITG) α3 gene (OMIM 605025 ). ITGα6 (OMIM 147556 ) and ITGβ4 (OMIM 147557 ) mutations are known to cause subtypes of junctional EB (JEB) often associated with pyloric atresia (OMIM 226730 ) and, although phenotypic variability exists, many of these patients exhibit severe skin disease and die during infancy. Patients with integrin α3 mutations had significant multiorgan disease, including a congenital nephrotic syndrome with significant morbidity, and interstitial lung disease that is progressive and eventually lethal. Skin blistering was not a prominent feature in the immediate neonatal period and began after several months of age. Skin blistering worsened with time, with increasing fragility with trauma. Blistered areas healed slowly, with residual erythema but not scarring. The mucosa was free from lesions. Hairs were fine and sparse and, with time, onychodystrophy of the great toenails was noted as well as distal onycholysis of the fingernails. Respiratory distress or other pulmonary issues were typically the presenting complaint, with nephrotic syndrome and its resultant sequelae noted during laboratory evaluation for the respiratory problems. Ultrastructural studies did not correspond with any previously reported EB subtypes but showed disruption within the basement membrane zone. Thus, an infantile onset progressive EB phenotype should prompt investigation for internal organ involvement.

A major source of mortality in patients with recessive dystrophic EB (RDEB) is metastatic squamous cell carcinoma. A newly published study investigates why cutaneous squamous cell carcinomas (cSCC) in patients with RDEB have a much higher rate of metastasis than standard cSCC or that seen in association with other inherited disorders such as xeroderma pigmentosum. The investigators hypothesized that type VII collagen deficiency created a permissive matrix environment for the growth and spread of cSCC. Increased type XII collagen, thrombospondin-1, and Wnt-5A were noted with deficiency of type VII collagen. Re-expression of normal type VII collagen decreased levels of these proteins, and reduced tumor cell invasion in culture and tumor growth in vivo. Although the details of these interactions still must be clarified, this suggests that, if type VII collagen can be replaced in patients with RDEB, skin fragility and blistering can be affected and, possibly, the risk of cSCC and the permissive growth environment can be improved.

Revertant mosaicism, in which a secondary mutation occurs in a patient with an inherited disease that corrects or overcomes the primary inherited mutation, was considered a very rare event. Studies of non-Herlitz JEB (OMIM 226650 ) patients have shed greater light on the frequency of revertant mosaicism in these patients. Revertant mosaicism seems to be common in patients with generalized non-Herlitz JEB—possibly occurring in most patients. In patients with preexisting localized JEB, however, revertant lesions do not seem to be frequent. There does not seem to be a preference for which genetic repair mechanism is used. Such patients offer great promise as potential donors of their own reverted fibroblasts or keratinocytes for application to blistering areas as a form of natural gene therapy.

Translation of the understanding of the underlying genetic causes of inherited skin disease into therapies has always been a major goal of research. Chronic ulcers often become a problem in EB patients and treating them can be challenging. Jonkman and colleagues describe success using a punch grafting procedure using autologous donor skin for chronic ulcers in patients with laminin-332 deficient non-Herlitz JEB (OMIM 226650 ). Previous studies have described increased type VII collagen expression and skin adhesion in RDEB patients after injection with allogeneic fibroblasts; however, the mechanism of this increase was unclear. A recent report notes increased cytokine expression after allogeneic fibroblast injection, including heparin binding–EGF-like growth factor (HB-EGF), which upregulates the patient’s own COL7A1 expression. The effect of recombinant HB-EGF on chronic EB ulcers has not been reported but is an obvious area for future study.

In the case of dystrophic EB, there have been several exciting developments focused on investigating and developing therapeutic approaches for the disease. Bone marrow–derived adult stem cells were known to play a role in maintenance of skin homeostasis; however, more recent work confirmed that these cells could generate and differentiate into skin cell types necessary for skin repair after wounding. This observation was taken to its translational conclusion with the report of a clinical trial of allogeneic bone marrow transplantation (BMT) in seven children with RDEB in 2010. Although the results of the successful BMTs were encouraging, with demonstration of detectable type VII collagen staining and clinical improvement in the patients, which lasted for more than 1 year after BMT, two of the seven children died due to complications from the BMT. Newer trials are underway that use conditioning regimens of significantly reduced intensity. Induced pluripotent stem cells (iPSCs) can be generated from skin fibroblasts and can serve as a renewable source of autologous stem cells. They may also be differentiated into keratinocytes. Initial methods of iPSC generation involved integration of viral transgenes that raised concerns that in vivo use could lead to tumor development. Currently, work is underway to develop methods of generating integration-free iPSCs. Another hurdle to the use of iPSCs is developing efficient and safe ways to correct the genetic defects present in patient-derived iPSCs. Efforts are underway to use customized zinc finger nucleases that are able to increase the incidence of homologous recombination at targeted mutation sites up to 10,000-fold. Alternatively, as noted previously, patients with revertant mosaic skin lesions offer a potential source of autologous-corrected iPSCs. Direct replacement of type VII collagen using purified recombinant type VII collagen, topically and injected intravenously, is also under study. Previously, IV infusion of recombinant type VII collagen was reported to home to wounded skin and lead to functional anchoring fibril formation in a type VII collagen knockout mouse. Recently, similar results were described in human RDEB skin grafted onto athymic nude mice. An important caveat to these therapies is the concern for the potential development of antibodies against type VII collagen. Recent work suggests that patients with RDEB may have preexisting anti–type VII collagen antibodies, even when type VII collagen is not detectable in the skin. Although the source of these antibodies is not clear, one possibility is that many RDEB patients have subclinical patches of revertant fibroblasts or keratinocytes that make small amounts of type VII collagen that serve as a source for the antibody development (similar to the growing understanding of revertant mosaicism in non-Herlitz JEB). Currently, the clinical relevance of these antibodies is not known.

Ichthyoses

The elucidation of the genetic causes of a variety of Mendelian disorders of cornification (MeDOC) has bolstered support for the bricks and mortar model of the epidermis. Although the role of each individual component detected to date is not completely understood, as other players are discovered the interconnections between the elements become clearer. Again, this is allowing the development of pathomechanism-based therapies for these patients.

The genetic abnormalities underlying a form of lamellar ichthyosis found in golden retriever dogs led to the discovery of a new genetic cause of ichthyosis. The ichthyosis in the retrievers is similar to autosomal recessive congenital ichthyosis (ARCI) and occurs because of homozygous mutations in patatin-like phospholipase domain-containing protein 1 (PNPLA1; OMIM 612121 ). Although the specific function of PNPLA1 is unknown, it seems important for epidermal lipid barrier formation. On discovery of this mutation, the investigators screened ARCI patients without known mutations and found six patients from two families with PNPLA1 mutations. Affected patients could present as collodion babies and evolve into a mild-to-moderate nonbullous congenital ichthyosiform erythroderma. Erythema and keratosis involved the skin diffusely, including the flexures. A mild palmoplantar keratoderma was present and some patients had pseudosyndactyly of the second to third toes. Although acitretin therapy improved scaling, it worsened the erythema.

The cause for a type of neuroichthyosis syndrome was identified in several Saudi families whose children who presented with a diffuse ichthyosis very similar to Sjögren-Larsson syndrome (SLS; OMIM 270200 ). However, the associated neurologic phenotype in these patients was more severe than SLS, with seizures, mental retardation, and significant spasticity. These patients had been diagnosed as having ichthyosis, spastic quadriplegia, and mental retardation (ISQMR; OMIM 614457 ), or pseudo-SLS. Patients can present with a collodion membrane at birth. All of these patients were profoundly developmentally delayed from infancy. Seizures began in the first months of life and were unresponsive to anti-epileptics. Bilateral inguinal hernias were noted. The ichthyotic skin changes were similar to SLS and often acrally accentuated. In these patients, AR mutations in the elongation of very long chain fatty acid-like 4 gene (ELOVL4; OMIM 605512 ), which encodes a very long chain fatty acid synthase, were detected. Heterozygous mutations in ELOVL4 cause macular degeneration in humans (autosomal dominant [AD] macular dystrophy or Stargardt disease 3; OMIM 600110 ). However, ocular findings in these patients were difficult to assess but seemed absent to mild. Very long chain fatty acids play important roles in cell membrane structure and in cell signaling.

Historically, the peeling skin syndromes (PSSs) have been a poorly understood group of rare AR syndromes characterized by episodic to continual superficial skin peeling. Acral and generalized variants are recognized with subdivision of the generalized forms into inflammatory and noninflammatory subtypes. New genetic approaches have recently allowed characterization and clarification of several of these syndromes. Some years ago, transglutaminase 5 (OMIM 603805 ) mutations were discovered to be the cause of the acral PSS variant (OMIM 609796 ) that can mimic mild forms of EB simplex. More recently, mutations in corneodesmosin (CDSN; OMIM 602593 ) were found to underlie the inflammatory variant of PSS (OMIM 270300 ) that is associated with allergies and significant atopic features. Most recently, a large family with noninflammatory AR PSS (PSS type A) was studied using whole genome homozygosity mapping followed by whole-exome sequencing of a single proband. Using this approach, a homozygous missense mutation in the carbohydrate sulfotransferase 8 (CHST8; OMIM 610190 ) gene was detected. CHST8 encodes an N-acetylgalactosamine-4-O-sulfotransferase (GalNAc4-ST1) localized to the Golgi transmembrane and expressed in normal human epidermis. Cells expressing the mutant enzyme had decreased levels of total sulfated glycosoaminoglycans and the investigators postulate that increased turnover of the mutant enzyme was responsible. Additionally, expression levels of GalNAc4-ST1 increase in the upper epidermal layers, suggesting a role in epidermal differentiation.

An AR subtype of ichthyosis termed exfoliative ichthyosis (OMIM 607936 ) shares some features with the PSS. Affected individuals develop dry, scaling skin diffusely over the body with peeling on the palms and soles. Using a combination of genetic approaches, including whole-genome homozygosity mapping, candidate-gene analysis, and deep sequencing, loss of function (LOF) mutations in cystatin A (OMIM 184600 ), a protease inhibitor, were determined to be the genetic cause of exfoliative ichthyosis. Two homozygous mutations, one a splice-site and the other a nonsense mutation were detected. Microscopic analysis revealed skin disruption at the basal or suprabasal epidermal layers. Further investigation suggests that a cell-cell adhesion defect exists when cystatin A is missing. This parallels the understanding of protease inhibitor function in the normal desquamation of the epidermis as revealed by Netherton syndrome (NS; OMIM 256500 ). In NS, deficiency of the serine protease inhibitor (OMIM 605010 ) allows overactive and ongoing epidermal protease activity to continually degrade the epidermal barrier, resulting in the cutaneous stigmata of NS, the desquamative elements of which may have clinical similarity with exfoliative ichthyosis and other variants of PSS. A better understanding of the causes of these syndromes will enhance their clinical recognition and prevent misdiagnosis of affected patients. Also, given the role of the individual affected proteins in very specific sites of epidermal barrier function, detailed clinical and molecular study of patients with these individual disorders and with filaggrin deficiency may shed light on the exact mechanisms by which some epidermal barrier defects contribute to an inflammatory phenotype (atopic dermatitis; CDSN deficiency) while others do not.

In another promising example of bench to bedside translational research, the skin lesions of congenital hemidysplasia, ichthyosis, and limb defects (CHILD) syndrome (OMIM 308050 ), an X-linked dominant (XLD) disorder improved with topical therapy. CHILD syndrome is caused by defects in a key component of the cholesterol biosynthesis pathway, the NAD(P)H steroid dehydrogenase-like protein (OMIM 300275 ). Patients with CHILD syndrome exhibit chronic recalcitrant inflammatory segmental skin lesions, often hemilateral with associated ipsilateral limb defects. These lesions cause significant morbidity for patients and have not responded to standard systemic or topical therapies apart from sporadic reports of improvement with destructive or surgical interventions. Because the mammalian NAD(P)H steroid dehydrogenase-like protein defect leads to impaired cholesterol production, investigators first attempted topical application of cholesterol to the skin lesions to replace the deficiency, but there was no benefit. However, the addition of topical lovastatin to block overproduction of cholesterol precursors, in addition to topical cholesterol, led to significant improvement in one treated case with minimal side effects.

Inflammatory and/or immunologic skin diseases

Several recent studies have demonstrated the power of new genetic analysis strategies to identify genes of importance in cutaneous and systemic inflammatory pathways. One report describes two siblings of Lebanese descent from a consanguineous marriage with neonatal inflammatory skin and bowel disease (OMIM 614328 ) that developed skin lesions by the second day of life. These cutaneous lesions began as perioral and perianal erythema with fissuring and a generalized pustular rash that transitioned into psoriasiform erythroderma with periodic flares of erythema, scaling, and widespread pustules. Diarrhea began in the first week of life. In the neonatal period the diarrhea led to failure to thrive. It was bloody, clinically consistent with malabsorption, and worsened with flares in the cutaneous disease and with gastrointestinal infections. Compensatory mechanisms for the defect seem to exist in humans as there is some normalization of the gut phenotype with time. The children were prone to Staphylococcus aureus infections with recurrent blepharitis and otitis externa. Their hairs were short or broken and eyelashes and eyebrows were wiry and rough. Nail abnormalities were noted, including thickening with frequent paronychia due to recurrent candida and pseudomonal infections. The sister died at 12 years from parvovirus B19–associated myocarditis and the brother had asymptomatic left ventricular dilatation. Using single nucleotide polymorphism (SNP)-homozygosity mapping, the investigators detected three large regions of homozygosity. Probes from all exons in these regions of the genome were included on a capture array and exons from these regions in the affected brother were sequenced. After known SNPs were removed, a total of 22 unique SNPs were detected in coding regions. Parallel assessment of the data for insertion-deletion variations detected a new deletion (4bp) in a disintegrin and metalloproteinase 17 (ADAM17; tumor necrosis factor α [TNF-α] converting enzyme [TACE]; OMIM 603639 ), that segregated with the disease in an AR manner. This mutation predicted a severely truncated protein.

Keratinocytes from the surviving sibling expressed ADAM10 (OMIM 602192 ), which cleaves some of the same substrates as ADAM17. Desmoglein 2 (DSG2; OMIM 125671 ) is a known direct target of both. Increased DSG2 expression was present in the patient’s skin suggesting a role for ADAM17 in regulating DSG2 availability at cell junctions. DSG2 is the predominant DSG in cardiac myocytes and mutations in DSG2 are associated with arrhythmogenic and dilated cardiomyopathies. Abnormal DSG2 expression may be related to the cardiac findings noted in these siblings. Because ADAM17 encodes a TACE, abnormal TNF-α signaling would be predicted to result from ADAM17 mutations and PBMCs from the surviving sibling showed impaired TNF-α production after stimulation. The low level of TNF-α detected may be from low-level production via ADAM10. The investigators also note that lack of TNF-α may have contributed to the fatal outcome in the sister because it is known to be cardioprotective in acute myocarditis.

A recent report elucidates the genetic cause of an AD cold urticarial syndrome (familial cold autoinflammatory syndrome 3; OMIM 614468 ). Affected patients develop cold urticaria with generalized cold exposure rather than by touching cold objects. Similarly, patients with this subtype of cold urticaria had negative skin testing with ice-cube and cold-water immersion but positive skin testing using evaporative cooling and generalized cold air exposure. Many patients developed additional features, including a common variable immunodeficiency (75%); a susceptibility to infections (56%), especially sinopulmonary; and autoimmune disorders (56%), such as thyroiditis. Some patients also developed granulomatous skin disease. The cold urticaria began early in life and was chronic. Abnormalities in patients included low IgM and IgA, low natural killer (NK) cells, decreased circulating CD19+ B cells, and IgA+ and IgG+ class-switched memory B cells. IgE was elevated in most patients. Antinuclear antibodies were present in 62%. Linkage analysis identified a single chromosomal region; however, whole-genome sequencing of one patient did not reveal any abnormalities. Analysis of a second family narrowed the linkage interval, allowing a candidate gene approach, which identified family-specific deletions in the PLCG2 gene (OMIM 600220 ), which encodes phospholipase Cγ ₂ (PLCγ ₂ ), a signaling molecule expressed in B cells, NK cells, and mast cells. The disease-causing heterozygous deletions in PLCG2 occur in an autoinhibitory domain, leading to constitutive phospholipase activity and resulting in a gain of function (GOF) phenotype. Spontaneous degranulation below 20°C was noted in mast cells transfected with mutant PLCG2, a finding not present in controls. Also, these defects were temperature sensitive in B cells with enhanced signaling and cellular activation. The investigators propose the moniker PLCγ ₂ -associated antibody deficiency and immune dysregulation (PLAID) to identify this syndrome.

The cause of various forms of chronic mucocutaneous candidiasis (CMC) syndromes and their causal interrelationships has been clarified greatly in several publications in the past year. Patients with CMC typically exhibit localized candida infections instead of systemic infection, due to defective cellular immunity. Defects in Th17-mediated immune function tend to result in fungal infections, reflecting the importance of Th17 cell function in the skin and/or mucosa. The pathway from fungal recognition to Th17 antifungal response involves several steps. Defects in these individual steps lead to various CMC syndromes. DECTIN1 (OMIM 606264 ) is a fungal pattern recognition receptor and fungal binding to dectin1 leads to signaling mediated by CARD9 (OMIM 607212 ). Defects in either can lead to CMC, semidominant (see later discussion) or AR in the case of dectin1 defects (OMIM 613108 ) and AR with CARD9 defects (OMIM 212050 ). In the case of DECTIN1, heterozygous carriers also showed increased Candida colonization and infections, though not at the level seen with CMC, suggesting a semidominant effect. Signal transducer and activator of transcription 3 (STAT3; OMIM 102582 ) plays a key role in Th17 cell differentiation and LOF in STAT3 impairs Th17 cell differentiation leading to AD hyperimmunoglobulin E syndrome (AD-HIES; OMIM 147060 ). Patients with AR-HIES (OMIM 243700 ) due to DOCK8 (OMIM 611432 ) mutations also exhibit impaired Th17 cell activity—albeit at a different point than that seen with STAT3 mutations. STAT1-dependent (OMIM 600555 ) pathways inhibit Th17 cell production and heterozygous mutations that result in GOF in STAT1 decrease Th17 cell number and activity, leading to AD CMC (OMIM 614162 ). Interleukin 17 (IL-17) production is critical and mutations in the genes encoding this cytokine IL-17F (OMIM 606496 ) and its receptor IL-17RA (OMIM 605461 ) inhibit Th17 cell effector function and lead to CMC, AD (OMIM 613956 ), and AR (OMIM 613953 ), respectively. The final step of antifungal immune response involves effective neutrophil function and impaired neutrophil production or activity can result in CMC, such as that seen with severe congenital neutropenia.

Large genome wide association studies (GWAS) have been performed for a variety of multigenic inflammatory skin disorders. Psoriasis was at the forefront of this research and loci identified in these studies are now being better characterized. The identity of one of these loci, PSORS2 (OMIM 602723 ), has long been sought. Recent reports describe GOF mutations in caspase recruitment domain family member 14 (CARD14; OMIM 607211 ) as the PSORS2 gene. The GOF mutations in CARD14 lead to enhanced NF-kB activation. Identified through analysis of rare families with strong penetrance of psoriasis through multiple generations, examination of large numbers of psoriasis patients from multiple cohorts has identified a variety of CARD14 variants and is beginning to shed light on their relation to NF-kB activation and possibly other mechanisms involved in the generation of psoriatic lesions.