Monoallelic and biallelic variants in LEF1 are associated with a new syndrome combining ectodermal dysplasia and limb malformations caused by altered WNT signaling.

PURPOSE: LEF1 encodes a transcription factor acting downstream of the WNT-ß-catenin signaling pathway. It was recently suspected as a candidate for ectodermal dysplasia in 2 individuals carrying 4q35 microdeletions. We report on 12 individuals harboring LEF1 variants. METHODS: High-throughput sequencing was employed to delineate the genetic underpinnings of the disease. Cellular consequences were characterized by immunofluorescence, immunoblotting, pulldown assays, and/or RNA sequencing. RESULTS: Monoallelic variants in LEF1 were detected in 11 affected individuals from 4 unrelated families, and a biallelic variant was detected in an affected individual from a consanguineous family. The phenotypic spectrum includes various limb malformations, such as radial ray defects, polydactyly or split hand/foot, and ectodermal dysplasia. Depending on the type and location of LEF1 variants, the inheritance of this novel Mendelian condition can be either autosomal dominant or recessive. Our functional data indicate that 2 molecular mechanisms are at play: haploinsufficiency or loss of DNA binding are responsible for a mild to moderate phenotype, whereas loss of ß-catenin binding caused by biallelic variants is associated with a severe phenotype. Transcriptomic studies reveal an alteration of WNT signaling. CONCLUSION: Our findings establish mono- and biallelic variants in LEF1 as a cause for a novel syndrome comprising limb malformations and ectodermal dysplasia.

Identification of limb-specific Lmx1b auto-regulatory modules with Nail-patella syndrome pathogenicity.

LMX1B haploinsufficiency causes Nail-patella syndrome (NPS; MIM 161200), characterized by nail dysplasia, absent/hypoplastic patellae, chronic kidney disease, and glaucoma. Accordingly in mice, Lmx1b has been shown to play crucial roles in the development of the limb, kidney and eye. Although one functional allele of Lmx1b appears adequate for development, Lmx1b null mice display ventral-ventral distal limbs with abnormal kidney, eye and cerebellar development, more disruptive, but fully concordant with NPS. In Lmx1b functional knockouts (KOs), Lmx1b transcription in the limb is decreased nearly 6-fold, indicating autoregulation. Herein, we report on two conserved Lmx1b-associated cis-regulatory modules (LARM1 and LARM2) that are bound by Lmx1b, amplify Lmx1b expression with unique spatial modularity in the limb, and are necessary for Lmx1b-mediated limb dorsalization. These enhancers, being conserved across vertebrates (including coelacanth, but not other fish species), and required for normal locomotion, provide a unique opportunity to study the role of dorsalization in the fin to limb transition. We also report on two NPS patient families with normal LMX1B coding sequence, but with loss-of-function variations in the LARM1/2 region, stressing the role of regulatory modules in disease pathogenesis.

TAR syndrome: Clinical and molecular characterization of a cohort of 26 patients and description of novel noncoding variants of RBM8A.

Thrombocytopenia-absent radius (TAR) syndrome is characterized by radial defect and neonatal thrombocytopenia. It is caused by biallelic variants of RBM8A gene (1q21.1) with the association of a null allele and a hypomorphic noncoding variant. RBM8A encodes Y14, a core protein of the exon junction complex involved in messenger RNA maturation. To date, only two hypomorphic variants have been identified. We report on a cohort of 26 patients affected with TAR syndrome and carrying biallelic variants in RBM8A. Half patients carried a 1q21.1 deletion and one of the two known hypomorphic variants. Four novel noncoding variants of RBM8A were identified in the remaining patients. We developed experimental models enabling their functional characterization in vitro. Two variants, located respectively in the 5'-untranslated region (5'-UTR) and 3'-UTR regions, are responsible for a diminished expression whereas two intronic variants alter splicing. Our results bring new insights into the molecular knowledge of TAR syndrome and enabled us to propose genetic counseling for patients' families.

WNT10B variants in split hand/foot malformation: Report of three novel families and review of the literature.

Split-hand/foot malformation (SHFM) is a genetically heterogeneous congenital limb malformation typically limited to a defect of the central rays of the autopod, presenting as a median cleft of hands and feet. It can be associated with long bone deficiency or included in more complex syndromes. Among the numerous genetic causes, WNT10B homozygous variants have been recently identified in consanguineous families, but remain still rarely described (SHFM6; MIM225300). We report on three novel SHFM families harboring WNT10B variants and review the literature, allowing us to highlight some clinical findings. The feet are more severely affected than the hands and there is a frequent asymmetry without obvious side-bias. Syndactyly of third-fourth fingers was a frequent finding (62%). Polydactyly, which was classically described in SHFM6, was only present in 27% of patients. No genotype-phenotype correlation is delineated but heterozygous individuals might have mild features of SHFM, suggesting a dose-effect of the WNT10B loss-of-function.

Duplication of 10q24 locus: broadening the clinical and radiological spectrum.

Split-hand-split-foot malformation (SHFM) is a rare condition that occurs in 1 in 8500-25,000 newborns and accounts for 15% of all limb reduction defects. SHFM is heterogeneous and can be isolated, associated with other malformations, or syndromic. The mode of inheritance is mostly autosomal dominant with incomplete penetrance, but can be X-linked or autosomal recessive. Seven loci are currently known: SHFM1 at 7q21.2q22.1 (DLX5 gene), SHFM2 at Xq26, SHFM3 at 10q24q25, SHFM4 at 3q27 (TP63 gene), SHFM5 at 2q31 and SHFM6 as a result of variants in WNT10B (chromosome 12q13). Duplications at 17p13.3 are seen in SHFM when isolated or associated with long bone deficiency. Tandem genomic duplications at chromosome 10q24 involving at least the DACTYLIN gene are associated with SHFM3. No point variant in any of the genes residing within the region has been identified so far, but duplication of exon 1 of the BTRC gene may explain the phenotype, with likely complex alterations of gene regulation mechanisms that would impair limb morphogenesis. We report on 32 new index cases identified by array-CGH and/or by qPCR, including some prenatal ones, leading to termination for the most severe. Twenty-two cases were presenting with SHFM and 7 with monodactyly only. Three had an overlapping phenotype. Additional findings were identified in 5 (renal dysplasia, cutis aplasia, hypogonadism and agenesis of corpus callosum with hydrocephalus). We present their clinical and radiological findings and review the literature on this rearrangement that seems to be one of the most frequent cause of SHFM.

Blepharocheilodontic syndrome is a CDH1 pathway-related disorder due to mutations in CDH1 and CTNND1.

PURPOSE: Blepharocheilodontic (BCD) syndrome is a rare autosomal dominant condition characterized by eyelid malformations, cleft lip/palate, and ectodermal dysplasia. The molecular basis of BCD syndrome remains unknown. METHODS: We recruited 11 patients from 8 families and performed exome sequencing for 5 families with de novo BCD syndrome cases and targeted Sanger sequencing in the 3 remaining families. RESULTS: We identified five CDH1 heterozygous missense mutations and three CTNND1 heterozygous truncating mutations leading to loss-of-function or haploinsufficiency. Establishment of detailed genotype-phenotype correlations was not possible because of the size of the cohort; however, the phenotype seems to appear more severe in case of CDH1 mutations. Functional analysis of CDH1 mutations confirmed their deleterious impact and suggested accelerated E-cadherin degradation. CONCLUSION: Mutations in CDH1 encoding the E-cadherin were previously reported in hereditary diffuse gastric cancer as well as in nonsyndromic cleft lip/palate. Mutations in CTNND1 have never been reported before. The encoded protein, p120ctn, prevents E-cadherin endocytosis and stabilizes its localization at the cell surface. Conditional deletion of Cdh1 and Ctnnd1 in various animal models induces features reminiscent of BCD syndrome and underlines critical role of the E-cadherin-p120ctn interaction in eyelid, craniofacial, and tooth development. Our data assert BCD syndrome as a CDH1 pathway-related disorder due to mutations in CDH1 and CTNND1 and widen the phenotypic spectrum of E-cadherin anomalies.Genet Med advance online publication 09 March 2017.

Nail-Patella Syndrome: clinical and molecular data in 55 families raising the hypothesis of a genetic heterogeneity.

Nail-Patella Syndrome (NPS) is a rare autosomal dominant condition comprising nail and skeletal anomalies. Skeletal features include dysplastic patellae and iliac horns, as well as scapula and elbow dysplasia. Nephropathy and glaucoma or intra-ocular hypertension can sometimes be present. NPS is due to variants affecting function in LMX1B, which encodes a LIM-homeodomain protein critical for limb, kidney and eye development. We describe the phenotype and the molecular data of 55 index patients and their 39 relatives presenting with typical NPS. We identified 38 different LMX1B anomalies, 19 of which were not reported before. In our series, 9% of families are not carriers of a LMX1B genomic alteration after extensive study of the coding and non-coding regions of the gene. One of the families showed no linkage to the LMX1B locus, raising the hypothesis of a genetic heterogeneity.

9q33.3q34.11 microdeletion: new contiguous gene syndrome encompassing STXBP1, LMX1B and ENG genes assessed using reverse phenotyping.

The increasing use of array-CGH in malformation syndromes with intellectual disability could lead to the description of new contiguous gene syndrome by the analysis of the gene content of the microdeletion and reverse phenotyping. Thanks to a national and international call for collaboration by Achropuce and Decipher, we recruited four patients carrying de novo overlapping deletions of chromosome 9q33.3q34.11, including the STXBP1, the LMX1B and the ENG genes. We restrained the selection to these three genes because the effects of their haploinsufficency are well described in the literature and easily recognizable clinically. All deletions were detected by array-CGH and confirmed by FISH. The patients display common clinical features, including intellectual disability with epilepsy, owing to the presence of STXBP1 within the deletion, nail dysplasia and bone malformations, in particular patellar abnormalities attributed to LMX1B deletion, epistaxis and cutaneous-mucous telangiectasias explained by ENG haploinsufficiency and common facial dysmorphism. This systematic analysis of the genes comprised in the deletion allowed us to identify genes whose haploinsufficiency is expected to lead to disease manifestations and complications that require personalized follow-up, in particular for renal, eye, ear, vascular and neurological manifestations.

ELN gene triplication responsible for familial supravalvular aortic aneurysm.

Supravalvular aortic aneurysms are less frequent than abdominal ones. Among Supravalvular aortic aneurysm aetiologies, we focused on dystrophic lesions as they can be secondary to genetic causes such as elastin anomaly. We report on a familial 7q11.23 triplication - including the ELN gene - segregating with a supravalvular aortic aneurysm. During her first pregnancy, our index patient was diagnosed with tuberous sclerosis and with a Supravalvular aortic aneurysm. The foetus was affected equally. For the second pregnancy, parents applied for preimplantation diagnosis, and a subsequent prenatal diagnosis was offered to the couple, comprising TSC1 molecular analysis, karyotype, and multiplex ligation probe amplification. TSC1 mutation was not found on foetal deoxyribo nucleic acid. Foetal karyotype was normal, but multiplex ligation probe amplification detected a 7q11.23 duplication. Quantitative-polymerase chain reaction and array-comparative genomic hybridisation carried out to further assess this chromosome imbalance subsequently identified a 7q11.23 triplication involving ELN and LIMK1. Foetal heart ultrasound identified a Supravalvular aortic aneurysm. A familial screening was offered for the 7q11.23 triplication and, when found, heart ultrasound was performed. The triplication was diagnosed in our index case as well as in her first child. Of the 17 individuals from this family, 11 have the triplication. Of the 11 individuals with the triplication, 10 were identified to have a supravalvular aortic aneurysm. Of them, two individuals received a medical treatment and one individual needed surgery. We provide evidence of supravalvular aortic aneurysm segregating with 7q11.23 triplication in this family. We would therefore recommend cardiac surveillance for individuals with 7q11.23 triplication. It would also be interesting to offer a quantitative-polymerase chain reaction or an array-comparative genomic hybridisation to a larger cohort of patients presenting with isolated supravalvular aortic aneurysm, as it may provide further information.

[Hereditary predisposition to cancers of the digestive tract, breast, gynecological and gonadal: focus on the Peutz-Jeghers]. / Prédisposition héréditaire aux cancers digestifs, mammaires, gynécologiques et gonadiques : état des lieux du syndrome de Peutz-Jeghers.

Peutz-Jeghers syndrome (PJS) is a rare autosomal dominant disease due to mutations in the tumor suppressor gene STK11. PJS is characterized by periorificial hyperpigmented macules (lentiginosis) and hamartomatous polyposis. Polyps can be located anywhere in the gastrointestinal tract, but are preferably observed in the small bowel (70-90%), the colon (50%) and the stomach (25%). They tend to be cancerous in a particular sequence hamartoma-dysplasia-cancer. The diagnosis is often made in the first or second decade following the appearance of lentigines or upon the occurrence of complications due to polyps (obstruction, intussusception, occult bleeding responsible for anemia). Furthermore PJS is associated with a significant increase in cancer risk (relative risk of 89% over the life according to the most recent series). Digestive cancers are the more frequent with cumulative incidences of 55% for gastro-intestinal cancer (39% for colorectal cancer, 13% for small bowel cancer and between 11 and 36% for pancreatic cancer, respectively). There is also an increased risk of non digestive cancers. In particular the risk of breast cancer is similar to that of patients carrying deleterious BRCA1 or BRCA2 mutations (cumulative incidence of 45%). Gynecological and gonadal tumors are frequent as well and can be more (adenoma malignum) or less aggressive (ovarian sex cord tumors with annular tubules and testicular tumors with calcified Sertoli cells). Finally the frequency of lung cancer is moderately increased. Recommendations for screening and management based on retrospective series in the literature have led to various strategies. The aim of this paper is to summarize the clinical and molecular diagnostic criteria of PJS as well as recommendations on screening strategies, management and monitoring.

Phenotypic similarities and differences in patients with a p.Met112Ile mutation in SOX10.

Waardenburg syndrome (WS) is characterized by an association of pigmentation abnormalities and sensorineural hearing loss. Four types, defined on clinical grounds, have been delineated, but this phenotypic classification correlates imperfectly with known molecular anomalies. SOX10 mutations have been found in patients with type II and type IV WS (i.e., with Hirschsprung disease), more complex syndromes, and partial forms of the disease. The phenotype induced by SOX10 mutations is highly variable and, except for the neurological forms of the disease, no genotype-phenotype correlation has been characterized to date. There is no mutation hotspot in SOX10 and most cases are sporadic, making it particularly difficult to correlate the phenotypic and genetic variability. This study reports on three independent families with SOX10 mutations predicted to result in the same missense mutation at the protein level (p.Met112Ile), offering a rare opportunity to improve our understanding of the mechanisms underlying phenotypic variability. The pigmentation defects of these patients are very similar, and the neurological symptoms showed a somewhat similar evolution over time, indicating a potential partial genotype-phenotype correlation. However, variability in gastrointestinal symptoms suggests that other genetic factors contribute to the expression of these phenotypes. No correlation between the rs2435357 polymorphism of RET and the expression of Hirschsprung disease was found. In addition, one of the patients has esophageal achalasia, which has rarely been described in WS.

A novel mutation in CDMP1 causes brachydactyly type C with "angel-shaped phalanx". A genotype-phenotype correlation in the mutational spectrum.

Brachydactyly type C (BDC), a well-recognized autosomal dominant hand malformation, displays brachymesophalangy of the second, third, and fifth fingers, a short first metacarpal, hyperphalangy, and ulnar deviation of the index finger. An "angel-shaped phalanx" is a distinctive radiological sign that can be found in BDC and other skeletal dysplasias, such as angel-shaped phalango-epiphyseal dysplasia (ASPED), an autosomal dominant skeletal abnormality characterized by a typical angel-shaped phalanx, brachydactyly, specific radiological findings, abnormal dentition, hip dysplasia, and delayed bone age. BDC and ASPED result from mutations in the CDMP1 gene. We report here a Mexican patient with BDC and clinical features of ASPED who carries a novel mutation in CDMP1, confirming that BDC and ASPED are part of the CDMP1 mutational spectrum. Based on the large number of clinical features in common, we suggest that both anomalies are part of the same clinical spectrum. Supported by an extensive review of the literature, a possible genotype-phenotype correlation in the mutational spectrum of this gene is proposed.

Thrombocytopenia-absent radius (TAR) syndrome: a clinical genetic series of 14 further cases. impact of the associated 1q21.1 deletion on the genetic counselling.

Thrombocytopenia-absent radius Syndrome (TAR) is a rare congenital malformation syndrome of complicated transmission. 1q21.1 deletion is necessary but not sufficient for its expression. We report the result of a French multicentric clinical study, and we emphasized on the role of the associated 1q21.1 deletion in the diagnosis and the genetic counselling of our patients. We gathered information on 14 patients presenting with TAR syndrome and referred for genetic counselling in six different university hospitals (8 foetuses, 1 child and 5 adults). Clinical or pathology details, as well as skeletal X-rays were analyzed. Genetic studies were performed by Array-CGH, and Quantitative Multiplex PCR. We demonstrated the very variable phenotypes of TAR syndrome. Female:male ratio was ∼2:1. All patients presented with bilateral radial aplasia/hypoplasia with preserved thumbs. Phocomelia and lower limb anomalies were present in 28% of the cases. We reported the first case of cystic hygroma on affected foetus. 1q21.1 deletions ranging from 330 to 1100 kb were identified in all affected patients. Most of them were inherited from one healthy parent (80%). The identification of a 1q21.1 deletion allowed confirmation of TAR syndrome diagnosis, particularly in foetuses and in atypical phenotypes. Additionally, it allowed accurate genetic counselling, especially when it occurred de novo. These findings allowed discussing the diagnostic criteria and management towards TAR syndrome.

Duplication at chromosome 2q31.1-q31.2 in a family presenting syndactyly and nystagmus.

HOXD genes encode transcription factors involved in the antero-posterior patterning of the limb bud and in the specification of fingers. During the embryo development, HOXD genes are expressed, following a spatio-temporal colinearity that involves at least three regions, centrometric and telomeric to this cluster. Here, we describe a father and a daughter presenting a 3-4 hand bilateral syndactyly associated with a nystagmus. Array-comparative genomic hybridisation showed a 3.8 Mb duplication at 2q31.1-q31.2, comprising 27 genes including the entire HOXD cluster. We performed expression studies in lymphoblasts by reverse transcription-PCR and observed an HOXD13 and HOXD10 overexpression, whereas the HOXD12 expression was decreased. HOXD13 and HOXD10 overexpression, associated with a misregulation of at least HOXD12, may therefore induce the syndactyly. Deletions of the HOXD cluster and its regulatory sequences induce hand malformations and, particularly, finger anomalies. Recently, smaller duplications of the same region have been reported in association with a mesomelic dysplasia, type Kantaputra. We discuss the variable phenotypes associated with such 2q duplications.

Further delineation of the 17p13.3 microdeletion involving YWHAE but distal to PAFAH1B1: four additional patients.

BACKGROUND: The 17p13.3 deletion syndrome (or Miller-Dieker syndrome, MDS, MIM 247200) is characterized by lissencephaly, mental retardation and facial dysmorphism. The phenotype is attributed to haploinsufficiency of two genes present in the minimal critical region of MDS: PAFAH1B1 (formerly referred to as LIS1) and YWHAE. Whereas isolated PAFAH1B1 deletion causes lissencephaly, YWHAE is a candidate for the dysmorphic phenotype associated with MDS. OBJECTIVE: We describe clinical, neuroradiological and molecular data in four patients with a 17p13.3 deletion distal to PAFAH1B1 involving YWHAE. RESULTS: All patients presented with mild or moderate developmental delay and pre and/or post-natal growth retardation. Patients A, B and C had neuro-imaging anomalies: leucoencephalopathy with macrocephaly (patients A and C), Chiari type 1 malformation (patient A) and paraventricular cysts (patient C). Patient B had patent ductus arteriosus and pulmonary arterial hypertension. Patient C had unilateral club foot. Patient D had enlarged Virchow Robin spaces, microcornea, and chorioretinal and lens coloboma. Array-CGH revealed de novo terminal 17p13.3 deletions for patient A and B, and showed interstitial 17p13.3 deletions of 1.4 Mb for patient C and of 0.5 Mb for patient D. In all patients, PAFAH1B1 was not deleted. CONCLUSION: Our patients confirm that 17p deletion distal to PAFAH1B1 have a distinctive phenotype : mild mental retardation, moderate to severe growth restriction, white matter abnormalities and developmental defects including Chiari type 1 malformation and coloboma. Our patients contribute to the delineation and clinical characterization of 17p13.3 deletion distal to PAFAH1B1 and highlight the role of the region containing YWHAE in brain and eye development and in somatic growth.

A 6.9 Mb 1qter deletion/4.4 Mb 18pter duplication in a boy with extreme microcephaly with simplified gyral pattern, vermis hypoplasia and corpus callosum agenesis.

We here report a boy presenting with developmental delay, growth retardation, facial dysmorphisms, vermis hypoplasia, micropolygyria and corpus callosum agenesis. Conventional and high resolution cytogenetic analyses were normal but high resolution oligonucleotide array-CGH, performed at the age of 4 years, allowed the characterisation of a de novo 6.9 Mb 1qter deletion/4.4 Mb 18pter duplication. Numerous 1qter deletions have already been described associated with brain malformations. Among 1q44 deleted genes, AKT3 is the strongest candidate gene for vermis hypoplasia and corpus callosum agenesis.

A new mutation in TP63 is associated with age-related pathology.

Increases in the number of allelic malformation syndromes have led to their classification according to their pathogenesis rather than their clinical specific phenotype. TP63 (also known as TP73L) mutations have been identified in several such syndromes characterized by autosomal dominant transmission and various combinations of ectodermal dysplasia, limb malformations and orofacial clefting. TP63 has not yet been implicated in early aging phenotype in humans, even though p63 activates a program of cellular senescence and p63-compromised mice display features of accelerated aging. We report on a family with four affected adult females presenting with Rapp-Hodgkin syndrome (RHS), an autosomal dominant clinical entity that associates anhidrotic ectodermal dysplasia with cleft lip and palate. Features between RHS and EEC syndrome (ectrodactyly, ectodermal dysplasia and cleft lip/palate) have led to the recent identification of mutations in the TP63 gene, located on 3q27, in this condition. Our patients present typical clinical features of RHS, but also ophthalmic anomalies such as corneal dystrophy and premature menopause (around 30 years). The latter findings have never been reported in this condition, and could be secondary to a new TP63 deletion that has been identified in this family.

Postzygotic mutation and germline mosaicism in the otopalatodigital syndrome spectrum disorders.

The otopalatodigital syndrome (OPD) spectrum disorders are a heterogeneous group of skeletal dysplasias caused by mutations in the X-linked gene, FLNA. All OPD spectrum disorders (otopalatodigital syndromes types 1 and 2, frontometaphyseal dysplasia and Melnick-Needles syndrome) exhibit significant interfamilial variability in their expressivity, especially in female subjects. Factors contributing to this may include allelic heterogeneity, variation in the degree of skewing of X inactivation or, conceivably, mosaicism for the underlying causative mutation. We report here monozygotic twin sisters who are discordant for the severe phenotype, Melnick-Needles syndrome, associated with the heterozygous mutation, 3596C>T. We also describe two brothers with otopalatodigital syndrome type 1 due to the FLNA mutation 620G>A. The mutation is not detectable in the blood leucocytes of their clinically unaffected mother, indicating that she is a germline mosaic for the condition. The description of somatic mutations and germline mosaicism in FLNA has implications for clinical and molecular diagnosis, phenotypic expression and genetic counseling of families with these disorders.

Prenatal phenotypic overlap of Costello syndrome and severe Noonan syndrome by tri-dimensional ultrasonography.

BACKGROUND: Prenatal diagnosis of multiple congenital anomalies is difficult, and usually molecular biology cannot immediately confirm the suspected syndrome. Fetal dysmorphology is useful tool in the diagnosis process, with limitations. METHODS: We report the thorough prenatal investigation by 2D and 3D ultrasonography in a case of suspected Costello syndrome. RESULTS: Prenatal abnormalities were: increased nuchal translucency, polyhydramnios, bilateral pyelectasis and ventriculomegaly. Ultrasonographic morphological fetal face analysis found abnormal thickness of the skin in the prefrontal area, thick dysplastic ears, thick lips and deep-set creases in the hands and feet. As Costello and Noonan syndromes overlap, a PTPN11 analysis was done, with presence of a mutation (T854C). CONCLUSION: Prenatal overlap of feature of severe Noonan syndrome and Costello syndrome is confirmed, with dysmorphological similarities, due to edema of fetal skin in face and extremities.