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Xeroderma pigmentosum (XP) is characterized by defective repair of ultraviolet radiation(UVR)-induced DNA damage. Patients have UVR hypersensitivity and increased skin cancer risk. Effective photoprotection has reduced childhood cancer-related deaths, but revealed adolescence-onset neurodegeneration, arising through unknown mechanisms. Here, we investigate XP neurodegeneration using pluripotent stem cells derived from XP patients and healthy relatives, performing functional multi-omics on samples during neuronal differentiation. We find endoplasmic reticulum stress is upregulated, preceded by oxidative stress, causing substantial 5’,8-cyclopurine and 8-oxopurine DNA damage. Critically, XP neurons exhibit inappropriate downregulation of the protein clearance ubiquitin-proteasome system (UPS). Chemical enhancement of UPS activity improves phenotypes, albeit inadequately, implying that early detection/prevention strategies are necessary to produce clinically impactful outcomes. Thus, we develop an early detection assay predicting neurodegeneration in at-risk patients.

Background: Cockayne syndrome (CS) is a DNA repair disorder primarily associated with pathogenic variants in ERCC6 and ERCC8. As in other Mendelian disorders, there are a number of genetically unsolved CS cases. Methods: We ascertained five individuals with monoallelic pathogenic variants in MORC2, previously associated with three dominantly inherited phenotypes: an axonal form of Charcot-Marie-Tooth disease type 2Z; a syndrome of developmental delay, impaired growth, dysmorphic facies, and axonal neuropathy; and a rare form of spinal muscular atrophy. Results: One of these individuals bore a strong phenotypic resemblance to CS. We then identified monoallelic pathogenic MORC2 variants in three of five genetically unsolved individuals with a clinical diagnosis of CS. In total, we identified eight individuals with MORC2-related disorder, four of whom had clinical features strongly suggestive of CS. Conclusions: Our findings indicate that some forms of MORC2-related disorder have phenotypic similarities to CS, including features of accelerated aging. Unlike classic DNA repair disorders, MORC2-related disorder does not appear to be associated with a defect in transcription-coupled nucleotide excision repair and follows a dominant pattern of inheritance with variants typically arising de novo. Such de novo pathogenic variants present particular challenges with regard to both initial gene discovery and diagnostic evaluations. MORC2 should be included in diagnostic genetic test panels targeting the evaluation of microcephaly and/or suspected DNA repair disorders. Future studies of MORC2 and its protein product, coupled with further phenotypic characterization, will help to optimize the diagnosis, understanding, and therapy of the associated disorders.

Hypotrichosis 12 (HYPT12) is an autosomal dominant, nonsyndromic hypotrichosis, caused by a pathogenic variant in the RPL21 gene encoding ribosomal protein L21, although only two pedigrees harbouring the amino acid substitution, p.Arg32Gln, have been reported previously. We present the case of a 44-year-old White British man with progressive hair loss since the age of 10 months, affecting his scalp, eyebrow, eyelashes and most of his body. Similar hair loss also affected several members of his family, with likely autosomal dominant inheritance. Using whole-exome sequencing, we identified a rare heterozygous missense variant (NM_000982.3:c.127A > G, NP_000973.2:p.Lys43Glu) in RPL21, and subsequent Sanger sequencing confirmed segregation of this variant in affected family members.

Solar urticaria is a rare, immunologically-mediated photodermatosis in which activation of cutaneous mast cells is triggered by specific wavelengths of solar electromagnetic radiation. This manifests clinically as the rapid development of cutaneous itch, erythema and wheal formation after several minutes of sun exposure. Disease mechanisms in solar urticaria remain incompletely elucidated and there have been few recent investigations of its pathobiology. Historic passive transfer experiments performed during the twentieth century provide support for a "photoallergy" model of disease pathogenesis, wherein molecular alteration of a putative chromophore by solar electromagnetic radiation produces mast cell activation via an IgE-dependent mechanism. However, this model does not account for several observations made during passive transfer experiments, nor does it explain a range of subsequent clinical and photobiological observations made in solar urticaria patients. Furthermore, increased understanding of the molecular dynamics underpinning cutaneous mast cell responses highlights the need to reformulate our understanding of solar urticaria pathogenesis in the context of this contemporary scientific landscape. In this review, we discuss the current understanding of solar urticaria pathogenesis and, by incorporating recent scientific and clinical observations, develop new hypotheses to drive future investigation into this intriguing disorder.

BACKGROUND: The U.K. 100,000 Genomes Project is in the process of investigating the role of genome sequencing in patients with undiagnosed rare diseases after usual care and the alignment of this research with health care implementation in the U.K. National Health Service. Other parts of this project focus on patients with cancer and infection.METHODS: We conducted a pilot study involving 4660 participants from 2183 families, among whom 161 disorders covering a broad spectrum of rare diseases were present. We collected data on clinical features with the use of Human Phenotype Ontology terms, undertook genome sequencing, applied automated variant prioritization on the basis of applied virtual gene panels and phenotypes, and identified novel pathogenic variants through research analysis.RESULTS: Diagnostic yields varied among family structures and were highest in family trios (both parents and a proband) and families with larger pedigrees. Diagnostic yields were much higher for disorders likely to have a monogenic cause (35%) than for disorders likely to have a complex cause (11%). Diagnostic yields for intellectual disability, hearing disorders, and vision disorders ranged from 40 to 55%. We made genetic diagnoses in 25% of the probands. A total of 14% of the diagnoses were made by means of the combination of research and automated approaches, which was critical for cases in which we found etiologic noncoding, structural, and mitochondrial genome variants and coding variants poorly covered by exome sequencing. Cohortwide burden testing across 57,000 genomes enabled the discovery of three new disease genes and 19 new associations. Of the genetic diagnoses that we made, 25% had immediate ramifications for clinical decision making for the patients or their relatives.CONCLUSIONS: Our pilot study of genome sequencing in a national health care system showed an increase in diagnostic yield across a range of rare diseases. (Funded by the National Institute for Health Research and others.).

Xeroderma pigmentosum (XP) is a genetic disorder caused by mutations in genes of the Nucleotide Excision Repair (NER) pathway (groups A-G) or in Translesion Synthesis (TLS) DNA polymerase η (V). XP is associated with an increased skin cancer risk, reaching, for some groups, several thousand-fold compared to the general population. Here, we analyzed 38 skin cancer genomes from five XP groups. We found that the activity of NER determines heterogeneity of the mutation rates across skin cancer genomes and that transcription-coupled NER extends beyond the gene boundaries reducing the intergenic mutation rate. Mutational profile in XP-V tumors and experiments withPOLH-KO cell line revealed the role of polymerase η in the error-free bypass of (i) rare TpG and TpA DNA lesions, (ii) 3’ nucleotides in pyrimidine dimers, and (iii) TpT photodimers. Our study unravels the genetic basis of skin cancer risk in XP and provides insights into the mechanisms reducing UV-induced mutagenesis in the general population.

Pediatric autoimmune bullous disease is a rare group of blistering skin disorders in children that result from autoimmunity against intercellular and basement membrane antigens in the skin and mucous membranes. Most pediatric cases are treated with oral corticosteroids or longer-term immunosuppressants such as azathioprine or mycophenolate mofetil. Immunomodulating drugs such as rituximab are increasingly being considered as options for refractory disease.

Images of solar urticaria in which localized skin areas remained refractory to sun exposure following omalizumab injections.

Patients with classic hydroa vacciniforme–like lymphoproliferative disorder (HVLPD) typically have high levels of Epstein-Barr virus (EBV) DNA in T cells and/or natural killer (NK) cells in blood and skin lesions induced by sun exposure that are infiltrated with EBV-infected lymphocytes. HVLPD is very rare in the United States and Europe but more common in Asia and South America. The disease can progress to a systemic form that may result in fatal lymphoma. We report our 11-year experience with 16 HVLPD patients from the United States and England and found that whites were less likely to develop systemic EBV disease (1/10) than nonwhites (5/6). All (10/10) of the white patients were generally in good health at last follow-up, while two-thirds (4/6) of the nonwhite patients required hematopoietic stem cell transplantation. Nonwhite patients had later age of onset of HVLPD than white patients (median age, 8 vs 5 years) and higher levels of EBV DNA (median, 1 515 000 vs 250 000 copies/ml) and more often had low numbers of NK cells (83% vs 50% of patients) and T-cell clones in the blood (83% vs 30% of patients). RNA-sequencing analysis of an HVLPD skin lesion in a white patient compared with his normal skin showed increased expression of interferon-γ and chemokines that attract T cells and NK cells. Thus, white patients with HVLPD were less likely to have systemic disease with EBV and had a much better prognosis than nonwhite patients. This trial was registered at www.clinicaltrials.gov as #NCT00369421 and #NCT00032513.

Xeroderma pigmentosum (XP) encompasses a group of rare diseases characterized in most cases by malfunction of nucleotide excision repair (NER), which results in an increased sensitivity to UV radiation in affected individuals. Approximately 25-30% of XP patients present with neurological symptoms, such as sensorineural deafness, mental deterioration and ataxia. Although it is known that dysfunctional DNA repair is the primary pathogenesis in XP, growing evidence suggests that mitochondrial pathophysiology may also occur. This appears to be secondary to dysfunctional NER but may contribute to the neurodegenerative process in these patients. The available pharmacological treatments in XP mostly target the dermal manifestations of the disease. In the present review, we outline how current understanding of the pathophysiology of XP could be used to develop novel therapies to counteract the neurological symptoms. Moreover, the coexistence of cancer and neurodegeneration present in XP led us to focus on possible new avenues targeting mitochondrial pathophysiology. LINKED ARTICLES: This article is part of a themed section on Mitochondrial Pharmacology: Featured Mechanisms and Approaches for Therapy Translation. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.22/issuetoc.

A teenage girl had the rare combined phenotype of xeroderma pigmentosum and trichothiodystrophy, resulting from mutations in the XPD (ERCC2) gene involved in nucleotide excision repair (NER). After treatment with antibiotics, including metronidazole for recurrent infections, she showed signs of acute and severe hepatotoxicity, which gradually resolved after withdrawal of the treatment. Cultured skin fibroblasts from the patient revealed cellular sensitivity to killing by metronidazole compared with cells from a range of other donors. This reveals that the metronidazole sensitivity was an intrinsic property of her cells. It is well recognized that patients with Cockayne syndrome, another NER disorder, are at high risk of metronidazole-induced hepatotoxicity, but this had not been reported in individuals with other NER disorders. We would urge extreme caution in the use of metronidazole in the management of individuals with the xeroderma pigmentosum and trichothiodystrophy overlap or trichothiodystrophy phenotypes.

Linked Comment: P. Wolf. Br J Dermatol 2022; 187:285–286.

Supplemental Digital Content is available in the text., Background: Approximately 25% of patients with pulmonary arterial hypertension (PAH) have been found to harbor rare mutations in disease-causing genes. To identify missing heritability in PAH, we integrated deep phenotyping with whole-genome sequencing data using Bayesian statistics. Methods: We analyzed 13 037 participants enrolled in the NBR study (NIHR BioResource—Rare Diseases), of which 1148 were recruited to the PAH domain. To test for genetic associations between genes and selected phenotypes of pulmonary hypertension, we used the Bayesian rare variant association method BeviMed. Results: Heterozygous, high impact, likely loss-of-function variants in the kinase insert domain receptor (KDR) gene were strongly associated with significantly reduced transfer coefficient for carbon monoxide (posterior probability=0.989) and older age at diagnosis (posterior probability=0.912). We also provide evidence for familial segregation of a rare nonsense KDR variant with these phenotypes. On computed tomographic imaging of the lungs, a range of parenchymal abnormalities were observed in the 5 patients harboring these predicted deleterious variants in KDR. Four additional PAH cases with rare likely loss-of-function variants in KDR were independently identified in the US PAH Biobank cohort with similar phenotypic characteristics. Conclusions: The Bayesian inference approach allowed us to independently validate KDR, which encodes for the VEGFR2 (vascular endothelial growth factor receptor 2), as a novel PAH candidate gene. Furthermore, this approach specifically associated high impact likely loss-of-function variants in the genetically constrained gene with distinct phenotypes. These findings provide evidence for KDR being a clinically actionable PAH gene and further support the central role of the vascular endothelium in the pathobiology of PAH.

Xeroderma pigmentosum (XP) is a well-studied disorder of (in most cases) nucleotide excision repair. The establishment in 2010 of a multidisciplinary XP clinic in the UK has enabled us to make a detailed analysis of genotype-phenotype relationships in XP patients and in several instances to make confident prognostic predictions. Splicing mutations in XPA and XPD and a specific amino acid change in XPD are associated with mild phenotypes, and individuals assigned to the XP-F group appear to have reduced pigmentation changes and a lower susceptibility to skin cancer than XPs in other groups. In an XP-C patient with advanced metastatic cancer arising from an angiosarcoma, molecular analysis of the tumour DNA suggested that immunotherapy, not normally recommended for angiosarcomas, might in this case be successful, and indeed the patient showed a dramatic recovery following immunotherapy treatment. These studies show that molecular analyses can improve the management, prognoses and therapy for individuals with XP.

Normal human skin is photosensitive in that it reddens following acute sunlight exposure and tans and thickens following chronic sunlight exposure. Skin cancer, particularly non-melanoma skin cancer, is also a consequence of high cumulative sun exposure in genetically predisposed normal individuals (predominantly those with fair skin). Outside the range of normal photosensitivity, there are several conditions in which patients exhibit diverse abnormal cutaneous reactions to sunlight. These are broadly described together as the photosensitivity disorders, but in fact they comprise a very heterogeneous group of skin conditions. Abnormal cutaneous photosensitive responses range from easy sunburn (as in drug phototoxicity and the DNA repair photodermatoses) and pain (erythropoietic protoporphyria), through to complex inflammatory responses such as urticaria, eczema, or epidermal necrosis induced by specific wavelengths of sunlight, the so-called idiopathic photodermatoses.

Importance Phototherapy is one of the mainstays of treatment for early mycosis fungoides (MF). The most common modalities are psoralen–UV-A (PUVA) and narrowband UV-B (NBUVB). Objective To compare the efficacy and adverse effects of PUVA vs NBUVB in early-stage MF. Data Sources A systematic review was performed by searching Cochrane Database of Systematic Reviews, Cochrane Central Register of Controlled Trials, Ovid Medline, PubMed, Cochrane Library, American College of Physicians ACP Journal Club, and Database of Abstracts of Review of Effectiveness from inception to March 30, 2018.UV A,PUVA,mycosis fungoides,Sezary syndrome,cutaneous T-cell lymphoma,UV B, andUVBwere used as either key words or MeSH terms. Study Selection Studies of cohorts with histologically confirmed early-stage MF, defined as stages IA, IB, and IIA, that compared PUVA vs NBUVB, had at least 10 patients in each comparator group, and reported outcomes of response to therapy. Exclusion criteria were studies with patients with stage IIB or higher MF, pediatric patients, fewer than 10 in each comparator group, noncomparative studies, case reports, and abstract studies. Data Extraction and Synthesis The Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) reporting guideline was followed. Data were pooled using a random-effects model with odds ratio (OR) as effect size. Main Outcomes and Measures Main outcomes were complete response rate, partial response rate, disease recurrence, and adverse effects, including erythema, nausea, pruritus, phototoxic effects, dyspepsia, and pain. Results Seven studies were included with a total of 778 patients (405 of 724 [55.9%] men; mean age, 52 years); 527 were treated with PUVA and 251 with NBUVB. Most of the included studies were of poor to moderate quality. Any response was found in 479 of the 527 (90.9%) patients treated with PUVA vs 220 of 251 (87.6%) treated with NBUVB (OR, 1.40; 95% CI, 0.84-2.34;P = .20). Complete response was found in 389 of 527 (73.8%) patients who received PUVA vs 156 of 251 (62.2%) who received NBUVB, which was statistically significant (OR, 1.68; 95% CI, 1.02-2.76;P = .04). Partial response was similar (90 of 501 [18.0%] vs 64 of 233 [27.5%]; OR, 0.58; 95% CI, 0.33-1.04;P = .07). No significant difference was found between PUVA and NBUVB in terms of adverse effects of erythema (38 of 527 [7.2%] vs 17 of 251 [6.7%];P = .54), nausea (10 of 527 [1.9%] vs 3 of 251 [1.2%];P = .72), pruritus (2 of 527 [0.4%] vs 4 of 251 [1.7%];P = .26), phototoxic effects (7 of 527 [1.4%] vs 2 of 251 [0.9%];P = .72), dyspepsia (6 of 527 [1.2%] vs 0 of 251 [0%];P = .59), or pain (0 of 527 [0%] vs 2 of 251 [0.9%];P = .50). Conclusions and Relevance The findings suggest that PUVA is a potential alternative to NBUVB in the management of early-stage MF. These findings have implications for clinicians involved in the management of early-stage MF.

The xeroderma pigmentosum (XP) population in the UK is around 100 patients. Since 2010, their medical care has been provided by a single national multidisciplinary clinical service, which cares for patients of all ages.

Xeroderma pigmentosum is characterized by cutaneous, ophthalmological, and neurological features. Although it is typical of childhood, late presentations can mimic different neurodegenerative conditions. We report two families presenting as Huntington's disease‐like syndromes. The first case (group G) presented with neuropsychiatric features, cognitive decline and chorea. Typical lentigines were only noticed after the neurological disease started. The second case (group B) presented adult‐onset chorea and neuropsychiatric symptoms after an aggressive ocular melanoma. Xeroderma pigmentosum can manifest as a Huntington's Disease‐like syndrome. Classic dermatological and oncological features have to be investigated in choreic patients with negative genetic tests for Huntington's disease‐like phenotypes.

© 2014 The Authors. Somatic mutations in cancer are more frequent in heterochromatic and late-replicating regions of the genome. We report that regional disparities in mutation density are virtually abolished within transcriptionally silent genomic regions of cutaneous squamous cell carcinomas (cSCCs) arising in an XPC-/-background. XPC-/-cells lack global genome nucleotide excision repair (GG-NER), thus establishing differential access of DNA repair machinery within chromatin-rich regions of the genome as the primarycause for the regional disparity. Strikingly, we find that increasing levels of transcription reduce mutation prevalence on both strands of gene bodies embedded within H3K9me3-dense regions, and only to those levels observed in H3K9me3-sparse regions, also in an XPC-dependent manner. Therefore, transcription appears to reduce mutation prevalence specifically by relieving the constraints imposed by chromatin structure on DNA repair. We model this relationship among transcription, chromatin state, and DNA repair, revealing a new, personalized determinant of cancer risk.

Chronic actinic dermatitis (CAD) is a debilitating photodermatosis. First line therapy consists of strict photoprotection and topical corticosteroids. Second line therapy uses systemic immunosuppression. However an alternative is needed for patients with severe CAD who cannot use systemic immunosuppressants.1,2 Case reports and small case series suggest that PUVA may be effective.3-6 This article is protected by copyright. All rights reserved.+

“Mutational signatures” are patterns of mutations that report DNA damage and subsequent repair processes that have occurred. Whole-genome sequencing (WGS) can provide additional information to standard diagnostic techniques and can identify therapeutic targets. A 32-yr-old male with xeroderma pigmentosum developed metastatic angiosarcoma that was unresponsive to three lines of conventional sarcoma therapies. WGS was performed on his primary cancer revealing a hypermutated tumor, including clonal ultraviolet radiation-induced mutational patterns (Signature 7) and subclonal signatures of mutated DNA polymerase epsilon (POLE) (Signature 10). These signatures are associated with response to immune checkpoint blockade. Immunohistochemistry confirmed high PD-L1 expression in metastatic deposits. The anti-PD-1 monoclonal antibody pembrolizumab was commenced off-label given the POLE mutation and high mutational load. After four cycles, there was a significant reduction in his disease with almost complete resolution of the metastatic deposits. This case highlights the importance of WGS in the analysis, interpretation, and treatment of cancers. We anticipate that as WGS becomes integral to the cancer diagnostic pathway, treatments will be stratified to the individual based on their unique genomic and/or transcriptomic profile, enhancing classical approaches of histologically driven treatment decisions.

Background Xeroderma pigmentosum (XP) is a rare autosomal recessive disorder of DNA repair. It is divided into eight complementation groups: XP-A to XP-G (classical XP) and XP variant (XP-V). Severe and prolonged sunburn reactions on minimal sun exposure have been considered a cardinal feature of classical XP. However, it has recently become clear that not all patients have abnormal sunburn reactions. Objectives To examine sunburn reactions in a cohort of patients with XP and correlate this to the complementation group. Methods Sixty patients with XP attending the U.K. National XP Service from 2010 to 2012 were studied. Their history of burning after minimal sun exposure was assessed using a newly developed sunburn severity score. The age at which the first skin cancer was histologically diagnosed in each patient, and the presence of any neurological abnormality, was also recorded. Results Sunburn severity scores were abnormally high in patients with XP-A, XP-D, XP-F and XP-G compared with non-XP controls. There was no significant difference in sunburn score of patients with XP-C, XP-E and XP-V compared with controls (P > 0·05). Patients with XP-C, XP-E and XP-V were more likely to have skin cancer diagnosed at an earlier age than those with severe sunburn on minimal sun exposure. In addition, patients with XP with severe sunburn had an increased frequency of neurological abnormalities. Conclusions Not all patients with XP have a history of severe and prolonged sunburn on minimal sun exposure. The normal sunburn response of patients with XP-C, XP-E and XP-V may relate to the preservation of transcription-coupled DNA repair in these groups. Those with a history of severe sunburn on minimal sun exposure developed their first skin cancer at an older age compared with patients with XP-C, XP-E and XP-V, but they had an increased frequency of neurological abnormalities. Physicians need to be aware that about half of all patients with XP will present without a history of abnormal sunburn. What's already known about this topic? Xeroderma pigmentosum (XP) is a rare autosomal recessive disorder of DNA repair. It is characterized by pigmentary skin changes, significantly increased risk of skin cancer, and progressive neurological disease in about 25% of cases. It is subdivided into eight complementation groups: XP-A to XP-G (classical XP) and XP variant (XP-V). Severe and prolonged sunburn reactions on minimal sun exposure have previously been considered a cardinal feature of classical XP. Recent data from a cohort of patients with XP at the National Institutes of Health in the U.S.A. have shown that about a third of their patients with XP have never sunburned. What does this study add? Not all patients with classical XP have severe and prolonged sunburn reactions. There is a correlation between response to sun exposure and XP complementation group. Patients with XP-C, XP-E and XP-V have sunburn reactions normal for skin-type compared with unaffected controls. Physicians need to be aware that about half of all patients with XP will present without a history of abnormal sunburn. This has important diagnostic and prognostic implications. See also the Commentary by Kraemer et al

Xeroderma pigmentosum (XP) is a rare DNA repair disorder characterized by increased susceptibility to UV radiation (UVR)-induced skin pigmentation, skin cancers, ocular surface disease, and, in some patients, sunburn and neurological degeneration. Genetically, it is assigned to eight complementation groups (XP-A to -G and variant). For the last 5 y, the UK national multidisciplinary XP service has provided follow-up for 89 XP patients, representing most of the XP patients in the United Kingdom. Causative mutations, DNA repair levels, and more than 60 clinical variables relating to dermatology, ophthalmology, and neurology have been measured, using scoring systems to categorize disease severity. This deep phenotyping has revealed unanticipated heterogeneity of clinical features, between and within complementation groups. Skin cancer is most common in XP-C, XP-E, and XP-V patients, previously considered to be the milder groups based on cellular analyses. These patients have normal sunburn reactions and are therefore diagnosed later and are less likely to adhere to UVR protection. XP-C patients are specifically hypersensitive to ocular damage, and XP-F and XP-G patients appear to be much less susceptible to skin cancer than other XP groups. Within XP groups, different mutations confer susceptibility or resistance to neurological damage. Our findings on this large cohort of XP patients under long-term follow-up reveal that XP is more heterogeneous than has previously been appreciated. Our data now enable provision of personalized prognostic information and management advice for each XP patient, as well as providing new insights into the functions of the XP proteins.

Summary Background Herlitz junctional epidermolysis bullosa (HJEB) is a severe, life-threatening, autosomal recessive blistering skin disease for which no cure is currently available. Prenatal diagnosis for couples at risk is feasible through fetal skin biopsy or analysis of DNA extracted from chorionic villi, but these methods can be applied only after pregnancy has been established. An alternative approach, which involves the analysis of single cells from embryos prior to establishment of pregnancy, is preimplantation genetic diagnosis (PGD). Until now, its clinical uptake has been hindered by lengthy delays in establishing mutation-specific protocols, and by the small amount of template DNA that can be obtained from a single cell. A new method that addresses these problems, preimplantation genetic haplotyping (PGH), relies on whole genome amplification followed by haplotyping of multiple polymorphic markers using standard DNA-based polymerase chain reaction (PCR) assays. Objectives To design and validate a generic PGH assay for HJEB and to transfer this into clinical practice. Materials and methods We established a multiplex PCR-based PGH assay involving 16 markers within and flanking the LAMB3 gene (the most frequently mutated gene in HJEB). The assay was then validated in 10 families with at least one previously affected offspring. After licensing by the Human Fertilisation and Embryology Authority (HFEA), the new test was used for PGD in a couple at risk of HJEB. Results The chromosome 1 LAMB3 markers within the assay were shown to be of sufficient heterogeneity to have widespread application for preimplantation testing of HJEB. In one couple that were heterozygous carriers of nonsense mutations in LAMB3, we used the new assay to identify unaffected embryos in a series of PGD cycles. Pregnancy was established in the third PGD cycle and a healthy, unaffected child was born. DNA analysis of cord blood confirmed the predicted single-cell mutation status of wild-type LAMB3 alleles. Conclusions PGH represents a major step forward in widening the scope and availability of preimplantation testing for serious mapped single-gene disorders. We have established a generic test that is suitable for the majority of couples at risk of HJEB.

Chronic graft-versus-host disease (GVHD) is a frequent complication after allogeneic hematopoietic stem cell transplantation (HSCT). Approximately 10% of patients with GVHD develop sclerodermatous changes, which can cause significant morbidity and are often refractory to standard systemic immunosuppression. We present two cases of sclerodermatous GVHD. The first is a 39-year-old man, who had a matched sibling, undergoing allogeneic HSCT for severe aplastic anemia. The second patient is a 7-year-old boy, who had an allogeneic HSCT from his HLA-identical mother for acute myeloid leukemia (AML). Both patients presented with widespread sclerotic changes, resulting in joint contractures and significant functional difficulties. Studies have shown UVA1 phototherapy to be a promising and well tolerated treatment modality in patients with sclerotic skin diseases. Both of our patients were treated with UVA1, which resulted in a significant skin softening, improvement in joint mobility and quality of life. UVA1 appears to be an effective treatment for refractory sclerodermatous GVHD; however, long-term clinical studies in larger groups are needed to accurately evaluate its efficacy and safety.

Kindler syndrome (KS) is a rare inherited skin disorder with blistering and poikiloderma as its main clinical features. It is caused by loss-of-function mutations in the C20orf42 (KIND1) gene which encodes kindlin-1, an actin cytoskeleton-focal contact-associated protein which is predominantly expressed in keratinocytes. We investigated the molecular basis of KS in a 16-year-old Indian boy who had additional clinical findings, including scleroatrophic changes of the hands and feet, pseudoainhum and early onset of squamous cell carcinoma on his foot. Immunostaining for kindlin-1 in the patient's skin was completely absent and sequencing of C20orf42 (KIND1) genomic DNA showed a homozygous splice-site mutation at the -6 position, IVS9-6T-->A. Amplification and sequencing of cDNA from the skin revealed aberrant splicing with either deletion of exon 10 or deletion of exons 9, 10 and 11, both of which involve loss of the pleckstrin homology domain of kindlin-1 that is thought to play a role in cytoskeletal attachment and integrin-mediated cell signalling. Pathogenic splice-site mutations at the -6 position are unusual and have rarely been reported for any genetic disorder. Collectively, these findings extend the spectrum of clinical and molecular abnormalities in this rare genodermatosis.

Recent data suggest that individuals with recessive dystrophic epidermolysis bullosa (RDEB) only develop squamous-cell carcinoma (SCC) in the presence of the NC1 domain of type VII collagen. This conclusion was based on experimental work in which cryosections of SCCs from 10 people with RDEB all showed positive type VII collagen immunostaining and observations in a murine model of SCC development in which tumors only occurred using keratinocytes from RDEB subjects that expressed detectable levels of the NC1 domain of the type VII collagen protein. To assess whether the clinical interpretation was valid in another cohort of RDEB patients, we examined expression of type VII collagen in 17 SCC tumors excised from 11 patients. Indirect immunofluorescent staining of SCC cryosections and Western blotting of cultured keratinocyte lysates identified two RDEB individuals who did not express detectable levels of type VII collagen. Mutation analysis revealed that these two patients harbor compound heterozygous nonsense mutations within the region of the COL7A1 gene encoding the NC1 domain. These data suggest that individuals with RDEB can develop SCC regardless of type VII collagen expression and that additional factors have a role in explaining the high incidence of tumors complicating this genodermatosis.

We report an unusual case of an inherited disorder of the desmosomal protein plakophilin 1, resulting in ectodermal dysplasia–skin fragility syndrome. The affected 6-year-old boy had red skin at birth and subsequently developed skin fragility, progressive plantar keratoderma, nail dystrophy, and alopecia. Skin biopsy revealed widening of intercellular spaces in the epidermis and a reduced number of small, poorly formed desmosomes. Mutation analysis of the plakophilin 1 gene PKP1 revealed a homozygous deletion of C at nucleotide 888 within exon 5. This mutation differs from the PKP1 gene pathology reported in 8 previously published individuals with this rare genodermatosis. However, all cases show similar clinical features, highlighting the importance of functional plakophilin 1 in maintaining desmosomal adhesion in skin, as well as the role of this protein in aspects of ectodermal development.

Maintenance of an intact epidermis depends on secure adhesion between adjacent keratinocytes, and between basal keratinocytes and the underlying epidermal basement membrane. The major adhesion units that achieve this are the hemidesmosomes and desmosomes, but when these structures are disrupted, e.g., by gene mutations or autoantibodies, the resilience of the epidermis is lost and blisters develop. Recently, there have been considerable advances in our knowledge of the proteins and glycoproteins that contribute to maintaining keratinocyte adhesion via hemidesmosomes and desmosomes, as well as new insights into the molecular pathogenesis of several inherited and autoimmune blistering skin diseases. These new basic scientific data are clinically relevant, helping to improve patient management and to provide a rationale for developing better and more specific treatments for patients with inherited or acquired blistering skin diseases. In addition, there have also been improvements in our understanding of the organization and assembly of these adhesion structures, and their involvement in signalling pathways, intricately linked to skin development, wound healing and tumour invasion. This review provides an update on the structure and organization of hemidesmosomes and desmosomes, and on the molecular pathology of their various components that result in bullous skin diseases.

Skin fragility-ectodermal dysplasia syndrome is an autosomal recessive disorder caused by loss-of-function mutations in the desmosomal protein, plakophilin 1. Clinically, there may be considerable morbidity from extensive skin erosions and painful fissures on the palms and soles. In the absence of any specific treatment, prenatal diagnosis is an option for couples at reproductive risk of recurrence. In 2000, we developed and applied a single cell nested polymerase chain reaction protocol to test one couple for compound heterozygous plakophilin 1 gene mutations by preimplantation genetic diagnosis (PGD). Although pregnancy was established, an unrelated trisomy 22 led to a spontaneous abortion. However, eight embryos of known genetic status were cryopreserved at that stage, and we planned to undertake subsequent frozen embryo replacement cycles that might lead to the birth of an unaffected child in this family. Embryo cryopreservation was carried out in June 2000 using standard protocols in a three-step freezing procedure. Four embryos were thawed in March 2003, one of which was viable and was used in a frozen embryo replacement cycle, but pregnancy did not occur. The remaining four embryos were thawed in February 2004, two of which were viable (both carriers of the paternal mutation) and these were used in a second frozen embryo replacement cycle, and a singleton pregnancy was established. The child's plakophilin 1 genotype was assessed by direct nucleotide sequencing across the site of both potential mutations. Following two frozen embryo replacement cycles, and almost 4 years after the initial embryo biopsy and mutation analysis, a pregnancy was achieved that progressed to term with the birth of a healthy baby girl. Nucleotide sequencing of cord blood DNA, taken immediately after delivery, showed that the child was a heterozygous carrier of the paternal mutation but not of the maternal mutation. This case demonstrates the value of embryo cryopreservation, which can increase the number of embryo replacement procedures and hence the cumulative pregnancy rate per retrieval cycle. Moreover, this is the first report of successful full-term pregnancy and birth of a healthy baby following exclusion of a severe genodermatosis by PGD. The successful outcome of PGD in this case illustrates what is technically possible for couples at risk of recurrence of a severe inherited skin disease.

Preimplantation genetic haplotyping (PGH) proof-of-principle was demonstrated by multiple displacement amplification (MDA) of single buccal cells from a female donor and genotyping using 12 polymorphic markers within the dystrophin gene; the known paternal genotype enabled identification of the paternal haplotype in the MDA products despite 27% allele dropout. MDA amplified DNA from 49 single human blastomeres with 100% success. The MDA products were genotyped using a total of 57 polymorphic markers for chromosomes 1, 7, 13, 18, 21, X and Y; 72% of alleles amplified providing results at 90% of the loci tested. A PGH cycle was carried out for Duchenne muscular dystrophy. One embryo was biopsied: PGH showed a non-carrier female, which was transferred with no resulting pregnancy. A PGH cycle was carried out for cystic fibrosis. Seven embryos were biopsied and PGH allowed the exclusion of 2 affected embryos; a carrier and a non-carrier embryo were transferred resulting in an on-going twin pregnancy. PGH represents a paradigm shift in embryo diagnosis, as one panel of markers can be used for all carriers of the same monogenic disease, bypassing the need for development of mutation-specific tests, and widening the scope and availability of preimplantation genetic testing.

Herlitz junctional epidermolysis bullosa (JEB) is an autosomal recessive mechanobullous disorder that results from loss-of-function mutations in the genes encoding the basement membrane component, laminin 5. Typically, there are frameshift, splice site or nonsense mutations on both alleles of either the LAMA3, LAMB3 or LAMC2 genes, with affected individuals inheriting one mutated allele from each parent. In this report, we describe a patient with Herlitz JEB in whom DNA analysis revealed homozygosity for the recurrent nonsense mutation R635X in LAMB3, located on chromosome 1q32.2. However, screening of parental DNA showed that although the patient's father was a heterozygous carrier of this mutation, the mother's DNA showed only wild-type sequence. Subsequent genotype analysis using 13 microsatellite markers spanning chromosome 1 revealed that the affected child was homozygous for the entire series of markers tested and that all of the alleles originated from the father. These results indicate that the Herlitz JEB phenotype in this patient is due to complete paternal isodisomy of chromosome 1 and reduction to homozygosity of the mutant LAMB3 gene locus. This is the fourth case of uniparental disomy to be described in Herlitz JEB, but it represents the first example of complete paternal isodisomy for chromosome 1 with a pathogenic mutation in the LAMB3 gene. These findings have important implications for mutation screening in JEB and for genetic counselling.