Whole exome sequencing identifies mutations in 10% of patients with familial non-syndromic cleft lip and/or palate in genes mutated in well-known syndromes.

BACKGROUND: Oral clefts, that is, clefts of the lip and/or cleft palate (CL/P), are the most common craniofacial birth defects with an approximate incidence of ~1/700. To date, physicians stratify patients with oral clefts into either syndromic CL/P (syCL/P) or non-syndromic CL/P (nsCL/P) depending on whether the CL/P is associated with another anomaly or not. In general, patients with syCL/P follow Mendelian inheritance, while those with nsCL/P have a complex aetiology and, as such, do not adhere to Mendelian inheritance. Genome-wide association studies have identified approximately 30 risk loci for nsCL/P, which could explain a small fraction of heritability. METHODS: To identify variants causing nsCL/P, we conducted whole exome sequencing on 84 individuals with nsCL/P, drawn from multiplex families (n=46). RESULTS: We identified rare damaging variants in four genes known to be mutated in syCL/P: TP63 (one family), TBX1 (one family), LRP6 (one family) and GRHL3 (two families), and clinical reassessment confirmed the isolated nature of their CL/P. CONCLUSION: These data demonstrate that patients with CL/P without cardinal signs of a syndrome may still carry a mutation in a gene linked to syCL/P. Rare coding and non-coding variants in syCL/P genes could in part explain the controversial question of 'missing heritability' for nsCL/P. Therefore, gene panels designed for diagnostic testing of syCL/P should be used for patients with nsCL/P, especially when there is at least third-degree family history. This would allow a more precise management, follow-up and genetic counselling. Moreover, stratified cohorts would allow hunting for genetic modifiers.

Unmasking familial CPX by WES and identification of novel clinical signs.

Mutations in the T-Box transcription factor gene TBX22 are found in X-linked Cleft Palate with or without Ankyloglossia syndrome (CPX syndrome). In addition to X-linked inheritance, ankyloglossia, present in the majority of CPX patients, is an important diagnostic marker, but it is frequently missed or unreported, as it is a "minor" feature. Other described anomalies include cleft lip, micro and/or hypodontia, and features of CHARGE syndrome. We conducted whole exome sequencing (WES) on 22 individuals from 17 "a priori" non-syndromic cleft lip and/or cleft palate (CL/P) families. We filtered the data for heterozygous pathogenic variants within a set of predefined candidate genes. Two canonical splice-site mutations were found in TBX22. Detailed re-phenotyping of the two probands and their families unravelled orofacial features previously not associated with the CPX phenotypic spectrum: choanal atresia, Pierre-Robin sequence, and overgrowths on the posterior edge of the hard palate, on each side of the palatal midline. This study emphasizes the importance of WES analysis in familial CLP cases, combined with deep (reverse) phenotyping in "a priori" non-syndromic clefts.

Efficacy of treatment with belladonna in children with severe pallid breath-holding spells.

IntroductionPallid breath-holding spells are common and dramatic forms of recurrent syncope in infancy. They are very stressful despite their harmless nature and sometimes require treatment. OBJECTIVE: The objective of this study was to evaluate the efficacy of belladonna in severe breath-holding spells. METHODS: This is a multicentric, retrospective series involving 84 children with severe pallid breath-holding spells. Inclusion criteria were >1 pallid breath-holding spell with loss of consciousness, paediatric cardiology evaluation, and follow-up >6 months. In total, 45 patients received belladonna and 39 patients did not receive treatment, according to physician preference. RESULTS: Mean age was 11 months, ranging from 4 to 18 months, with 54% of males. Mean spell duration was 30 seconds (interquartile range 15, 60), and the frequency was four episodes per month (interquartile range 0.5, 6.5). Comparison of baseline characteristics between groups showed similar demographics, with the single difference in the severity of the spells, being more severe in the treated group. When comparing the treated and non-treated groups at 3 months, only two (5%) patients had a complete remission in the first group, whereas 20 (44%) had remission in the belladonna group (p<0.01). When considering the characteristics of the spells before and after the initiation of treatment with belladonna, 75% of the patients presented a positive response, with 44% of the patients presenting with complete resolution of the spells (p<0.01). No major adverse reaction was reported, with only 5% minor adverse events. CONCLUSIONS: Belladonna is highly effective to alleviate severe breath-holding spells in young children, without any major adverse effects.

FAF1, a gene that is disrupted in cleft palate and has conserved function in zebrafish.

Cranial neural crest (CNC) is a multipotent migratory cell population that gives rise to most of the craniofacial bones. An intricate network mediates CNC formation, epithelial-mesenchymal transition, migration along distinct paths, and differentiation. Errors in these processes lead to craniofacial abnormalities, including cleft lip and palate. Clefts are the most common congenital craniofacial defects. Patients have complications with feeding, speech, hearing, and dental and psychological development. Affected by both genetic predisposition and environmental factors, the complex etiology of clefts remains largely unknown. Here we show that Fas-associated factor-1 (FAF1) is disrupted and that its expression is decreased in a Pierre Robin family with an inherited translocation. Furthermore, the locus is strongly associated with cleft palate and shows an increased relative risk. Expression studies show that faf1 is highly expressed in zebrafish cartilages during embryogenesis. Knockdown of zebrafish faf1 leads to pharyngeal cartilage defects and jaw abnormality as a result of a failure of CNC to differentiate into and express cartilage-specific markers, such as sox9a and col2a1. Administration of faf1 mRNA rescues this phenotype. Our findings therefore identify FAF1 as a regulator of CNC differentiation and show that it predisposes humans to cleft palate and is necessary for lower jaw development in zebrafish.

An unusual cause of fetal hypomobility:congenital central hypoventilation syndrome associated with hirschsprung disease.

Co-occurrence of congenital central hypoventilation syndrome and Hirschsprung disease is known as Haddad syndrome. Affected patients develop with variable expressivity a dysfunction of the autonomic nervous system. We report the natural history of a full-term newborn infant presenting multiple features of autonomic system dysfunction that were already noted antenatally. The presence of a nonpolyalanine repeat expansion mutation in the PHOX2B gene confirmed postnatally the diagnosis of Haddad syndrome. This case suggests that patients presenting with autonomic system dysfunction may already present signs of the disease during the fetal period. Furthermore, antenatal presentations may correlate with a more severe presentation of the disease. In conclusion, antenatal signs of dysautonomy should stimulate multidisciplinary prenatal approach to orientate proper postnatal intervention and facilitate treatment strategies.

Pregnancy and neonatal outcome following an antenatal diagnosis of cleft lip and palate.

AIM: To identify the significance of associated antenatal ultrasound findings on long-term prognosis following the antenatal diagnosis of cleft lip/palate [CL(P)]. PATIENTS AND METHODS: Retrospective case note analysis of patients seen at a single tertiary referral centre with a diagnosis of CL(P). The patients were classified as those with unilateral or bilateral clefts and then further subdivided according to the presence of associated anomalies, and these were related to pregnancy and neonatal outcome. RESULTS: A total of 125 singleton pregnancies were seen at the antenatal diagnostic unit, 14 of which were subsequently lost to follow-up. Eighty-two (65.6%) had a diagnosis of unilateral CL(P) and 43 (34.4%) a bilateral CL(P). Seventy-five foetuses (67.5%) had no other anomalies detected on antenatal ultrasound. Seventeen patients (15%) underwent a termination of pregnancy. A normal postnatal outcome was seen in 79% of liveborn infants overall. Only 50% of foetuses diagnosed with a single minor anomaly and 4% of the foetuses in whom more than two minor anomalies or one major anomaly had been detected on ultrasound had a normal postnatal outcome. Infants with bilateral CL(P) had a significantly reduced incidence of a normal postnatal course (60% vs. 87.5%, P<0.01). CONCLUSION: In cases of CL(P), there is a high incidence of associated anomalies detected on antenatal ultrasound and these significantly increase the risk of poor neonatal outcome.

Esophageal Trachea, a Unique Foregut Malformation Requiring Multistage Surgical Reconstruction: Case Report.

Abnormal connections between the esophagus and low respiratory tract can result from embryological defects in foregut development. Beyond well-known malformations, including tracheo-esophageal fistula and laryngo-tracheo-esophageal cleft, rarer anomalies have also been reported, including communicating bronchopulmonary foregut malformations and tracheal atresia. Herein, we describe a case of what we have called "esophageal trachea," which, to our knowledge, has yet to be reported. A full-term neonate was born in our institution presenting with a foregut malformation involving both the middle esophagus and the distal trachea, which were found to be longitudinally merged into a common segment, 3 cm in length, located just above the carina and consisted of esophageal tissue without cartilaginous rings. At birth, the esophagus and trachea were surgically separated via right thoracotomy, the common segment kept on the tracheal side only, creating a residual long-gap esophageal atresia. The resulting severe tracheomalacia was treated via simultaneous posterior splinting of such diseased segment using an autologous pericardium patch, as well as by anterior aortopexy. Terminal esophagostomy and gastrostomy were created at that stage due to the long distance between esophageal segments. Between ages 18 and 24 months, the patient underwent native esophageal reconstruction using a multistage traction-and-growth surgical strategy that combined Kimura extra-thoracic esophageal elongations at the upper esophagus and Foker external traction at the distal esophagus. Ten months after esophageal reconstruction, prolonged, refractory, and severe tracheomalacia was further treated via anterior external stenting using a semitubular ringed Gore-Tex® prosthesis, through simultaneous median sternotomy and tracheoscopy. Currently, 2 years after the last surgery, respiratory stabilization, and full oral feeding were stably achieved. Multidisciplinary management was crucial for assuring lifesaving procedures, correctly assessing anatomy, and planning for multiple sequential surgical approaches that aimed to restore long-term respiratory and digestive functions.

Likely Pathogenic Variants in One Third of Non-Syndromic Discontinuous Cleft Lip and Palate Patients.

Oral clefts are composed of cleft of the lip, cleft of the lip and palate, or cleft of the palate, and they are associated with a wide range of expression and severity. When cleft of the palate is associated with cleft of the lip with preservation of the primary palate, it defines an atypical phenotype called discontinuous cleft. Although this phenotype may represent 5% of clefts of the lip and/or palate (CLP), it is rarely specifically referred to and its pathophysiology is unknown. We conducted whole exome sequencing (WES) and apply a candidate gene approach to non-syndromic discontinuous CLP individuals in order to identify genes and deleterious variants that could underlie this phenotype. We discovered loss-of-function variants in two out of the seven individuals, implicating FGFR1 and DLG1 genes, which represents almost one third of this cohort. Whole exome sequencing of clinically well-defined subgroups of CLP, such as discontinuous cleft, is a relevant approach to study CLP etiopathogenesis. It could facilitate more accurate clinical, epidemiological and fundamental research, ultimately resulting in better diagnosis and care of CLP patients. Non-syndromic discontinuous cleft lip and palate seems to have a strong genetic basis.

Postcricoid hemangioma: an overlooked cause of dysphagia in infants?-a case report.

Feeding and swallowing disorders in children remain a major challenge owing to a wide differential diagnosis. Hemangioma of the upper aerodigestive tract represents one of the numerous non-neoplastic causes of dysphagia. We report two cases of postcricoid hemangioma causing inhalation and recurrent respiratory infections, treated successfully with systemic corticotherapy alone. To our knowledge, these are the second and third cases described in the literature. After a short review of the literature, the diagnostic procedures are discussed and a management strategy is proposed for this clinical entity, by far underestimated.