Identification of the DNA methylation signature of Mowat-Wilson syndrome.

Mowat-Wilson syndrome (MOWS) is a rare congenital disease caused by haploinsufficiency of ZEB2, encoding a transcription factor required for neurodevelopment. MOWS is characterized by intellectual disability, epilepsy, typical facial phenotype and other anomalies, such as short stature, Hirschsprung disease, brain and heart defects. Despite some recognizable features, MOWS rarity and phenotypic variability may complicate its diagnosis, particularly in the neonatal period. In order to define a novel diagnostic biomarker for MOWS, we determined the genome-wide DNA methylation profile of DNA samples from 29 individuals with confirmed clinical and molecular diagnosis. Through multidimensional scaling and hierarchical clustering analysis, we identified and validated a DNA methylation signature involving 296 differentially methylated probes as part of the broader MOWS DNA methylation profile. The prevalence of hypomethylated CpG sites agrees with the main role of ZEB2 as a transcriptional repressor, while differential methylation within the ZEB2 locus supports the previously proposed autoregulation ability. Correlation studies compared the MOWS cohort with 56 previously described DNA methylation profiles of other neurodevelopmental disorders, further validating the specificity of this biomarker. In conclusion, MOWS DNA methylation signature is highly sensitive and reproducible, providing a useful tool to facilitate diagnosis.

Prenatal Array-CGH Detection of 3q26.32q26.33 Interstitial Deletion Encompassing the SOX2 Gene: Ultrasound, Pathological, and Cytogenetic Findings.

Background: SOX2 disorders are associated with anophthalmia-esophageal-genital syndrome or microphthalmia, syndromic 3 (MCOPS3- # 206900). Case Report: We describe a third fetal case with a de novo 3q26.32q26.33 deletion extending for 4.31 Mb, detected in a 15-week fetus. After legal interruption of pregnancy, at autopsy, the fetus presented bilateral microphthalmia, right cleft lip and palate, bilateral cerebral ventriculomegaly and dilated third ventricle, microcystic left lung, and intestinal malrotation. Histologically, the left lung showed congenital pulmonary airway malformation (CPAM) type 2. Retinal dysplasia was found in both eyes. Discussion/Conclusion: The human SOX2 gene (OMIM #184429) is located on chromosome 3 at position q26.3-27 and encodes a transcription factor involved in the development of the central and peripheral nervous systems, retina, and lung. In our case, the combination of cerebral, retinal, and pulmonary anomalies, not previously described, are consistent with SOX2 haploinsufficiency due to chromosomal deletion.

Case Report: Sequential postzygotic HRAS mutation and gains of the paternal chromosome 11 carrying the mutated allele in a patient with epidermal nevus and rhabdomyosarcoma: evidence of a multiple-hit mechanism involving HRAS in oncogenic transformation.

We report a 7-year-old boy born with epidermal nevi (EN) arranged according to Blaschko's lines involving the face and head, right upper limb, chest, and left lower limb, who developed a left paratesticular embryonal rhabdomyosarcoma at 18 months of age. Parallel sequencing identified a gain-of-function variant (c.37G>C, p.Gly13Arg) of HRAS in both epidermal nevus and tumor but not in leukocytes or buccal mucosal epithelial cells, indicating its postzygotic origin. The variant accounted for 33% and 92% of the total reads in the nevus and tumor DNA specimens, respectively, supporting additional somatic hits in the latter. DNA methylation (DNAm) profiling of the tumor documented a signature consistent with embryonal rhabdomyosarcoma and CNV array analysis inferred from the DNAm arrays and subsequent MLPA analysis demonstrated copy number gains of the entire paternal chromosome 11 carrying the mutated HRAS allele, likely as the result of paternal unidisomy followed by subsequent gain(s) of the paternal chromosome in the tumor. Other structural rearrangements were observed in the tumours, while no additional pathogenic variants affecting genes with role in the RAS-MAPK and PI3K-AKT-MTOR pathways were identified. Our findings provide further evidence of the contribution of "gene dosage" to the multistep process driving cell transformation associated with hyperactive HRAS function.

Deep phenotyping of the neuroimaging and skeletal features in KBG syndrome: a study of 53 patients and review of the literature.

BACKGROUND: KBG syndrome is caused by haploinsufficiency of ANKRD11 and is characterised by macrodontia of upper central incisors, distinctive facial features, short stature, skeletal anomalies, developmental delay, brain malformations and seizures. The central nervous system (CNS) and skeletal features remain poorly defined. METHODS: CNS and/or skeletal imaging were collected from molecularly confirmed individuals with KBG syndrome through an international network. We evaluated the original imaging and compared our results with data in the literature. RESULTS: We identified 53 individuals, 44 with CNS and 40 with skeletal imaging. Common CNS findings included incomplete hippocampal inversion and posterior fossa malformations; these were significantly more common than previously reported (63.4% and 65.9% vs 1.1% and 24.7%, respectively). Additional features included patulous internal auditory canal, never described before in KBG syndrome, and the recurrence of ventriculomegaly, encephalic cysts, empty sella and low-lying conus medullaris. We found no correlation between these structural anomalies and epilepsy or intellectual disability. Prevalent skeletal findings comprised abnormalities of the spine including scoliosis, coccygeal anomalies and cervical ribs. Hand X-rays revealed frequent abnormalities of carpal bone morphology and maturation, including a greater delay in ossification compared with metacarpal/phalanx bones. CONCLUSION: This cohort enabled us to describe the prevalence of very heterogeneous neuroradiological and skeletal anomalies in KBG syndrome. Knowledge of the spectrum of such anomalies will aid diagnostic accuracy, improve patient care and provide a reference for future research on the effects of ANKRD11 variants in skeletal and brain development.

Case report: Expanding the phenotype of FOXP1-related intellectual disability syndrome and hyperkinetic movement disorder in differential diagnosis with epileptic seizures.

Objective: We aimed to report on previously unappreciated clinical features associated with FOXP1-related intellectual disability (ID) syndrome, a rare neurodevelopmental disorder characterized by global developmental delay, intellectual disability, and language delay, with or without autistic features. Methods: We performed whole-exome sequencing (WES) to molecularly characterize an individual presenting with ID, epilepsy, autism spectrum disorder, behavioral problems, and facial dysmorphisms as major features. Results: WES allowed us to identify a previously unreported de novo splice site variant, c.1429-1G>T (NM_032682.6), in the FOXP1 gene (OMIM*605515) as the causative event underlying the phenotype. Clinical reassessment of the patient and revision of the literature allowed us to refine the phenotype associated with FOXP1 haploinsufficiency, including hyperkinetic movement disorder and flat angiomas as associated features. Interestingly, the patient also has an asymmetric face and choanal atresia and a novel de novo variant of the CHD7 gene. Conclusion: We suggest that FOXP1-related ID syndrome may also predispose to the development of hyperkinetic movement disorders and flat angiomas. These features could therefore require specific management of this condition.

Developmental and epileptic encephalopathy in a young Italian woman with a de novo missense variant in the CLCN4 gene: A case report.

INTRODUCTION: Raynaud-Claes syndrome is a very rare X-linked condition, characterized by intellectual disability, impaired language development, brain abnormalities, facial dysmorphisms and drug-resistant epilepsy. It is caused by loss-of-function variants in the CLCN4 gene, which encodes the 2Cl-/H + exchanger ClC-4, prominently expressed in the hippocampus and cerebellum. Different genotypic variants have been described, each exhibiting specific phenotypic characteristics. The loss-of-function variant p.Gly544Arg in the CLCN4 gene has been described in only two male probands, but there are no reports on phenotypic characterization in females. CASE PRESENTATION: We present a 30-year-old Italian woman with early-onset drug-resistant epilepsy, developmental and epileptic encephalopathy, developmental delay, absence of verbal language development, behavioral impairment with autistic features, and clusters of seizures during catamenial periods. The interictal EEG showed slight inconstant slowing of the background rhythm, with abnormal frontal predominant mu like rhythm and generalized spike and polyspike wave discharges, which increased in frequency during drowsiness. A brain MRI showed slight cranio-encephalic asymmetry and a smaller size of the left hippocampus. The whole exome sequencing (WES) revealed a de novo heterozygous c.1630G > A variant in the CLCN4 gene, resulting in the amino acid substitution p.Gly544Arg (rs587777161), consistent with Raynaud-Claes syndrome. DISCUSSION AND CONCLUSION: Our patient is the first case of a de novo p.Gly544Arg variant of the CLCN4 gene in a female proband, confirming that female patients with Raynaud-Claes syndrome can be as severely affected as the male counterparts. Our case expands the phenotypic characterization of different genotypic CLCN4 variants, which can become crucial in the future for early diagnosis if targeted therapy becomes available.

Characterization of BRCA Deficiency in Ovarian Cancer.

BRCA testing is recommended in all Ovarian Cancer (OC) patients, but the optimal approach is debated. The landscape of BRCA alterations was explored in 30 consecutive OC patients: 6 (20.0%) carried germline pathogenic variants, 1 (3.3%) a somatic mutation of BRCA2, 2 (6.7%) unclassified germline variants in BRCA1, and 5 (16.7%) hypermethylation of the BRCA1 promoter. Overall, 12 patients (40.0%) showed BRCA deficit (BD), due to inactivation of both alleles of either BRCA1 or BRCA2, while 18 (60.0%) had undetected/unclear BRCA deficit (BU). Regarding sequence changes, analysis performed on Formalin-Fixed-Paraffin-Embedded tissue through a validated diagnostic protocol showed 100% accuracy, compared with 96.3% for Snap-Frozen tissue and 77.8% for the pre-diagnostic Formalin-Fixed-Paraffin-Embedded protocol. BD tumors, compared to BU, showed a significantly higher rate of small genomic rearrangements. After a median follow-up of 60.3 months, the mean PFS was 54.9 ± 27.2 months in BD patients and 34.6 ± 26.7 months in BU patients (p = 0.055). The analysis of other cancer genes in BU patients identified a carrier of a pathogenic germline variant in RAD51C. Thus, BRCA sequencing alone may miss tumors potentially responsive to specific treatments (due to BRCA1 promoter methylation or mutations in other genes) while unvalidated FFPE approaches may yield false-positive results.

Prenatal Clinical Findings in RASA1-Related Capillary Malformation-Arteriovenous Malformation Syndrome.

Pathogenic variants in RASA1 are typically associated with a clinical condition called "capillary malformation-arteriovenous malformation" (CM-AVM) syndrome, an autosomal dominant genetic disease characterized by a broad phenotypic variability, even within families. In CM-AVM syndrome, multifocal capillary and arteriovenous malformations are mainly localized in the central nervous system, spine and skin. Although CM-AVM syndrome has been widely described in the literature, only 21 cases with prenatal onset of clinical features have been reported thus far. Here, we report four pediatric cases of molecularly confirmed CM-AVM syndrome which manifested during the prenatal period. Polyhydramnios, non-immune hydrops fetalis and chylothorax are only a few possible aspects of this condition, but a correct interpretation of these prenatal signs is essential due to the possible fatal consequences of unrecognized encephalic and thoracoabdominal deep vascular malformations in newborns and in family members carrying the same RASA1 variant.

Bi-allelic variants in the ESAM tight-junction gene cause a neurodevelopmental disorder associated with fetal intracranial hemorrhage.

The blood-brain barrier (BBB) is an essential gatekeeper for the central nervous system and incidence of neurodevelopmental disorders (NDDs) is higher in infants with a history of intracerebral hemorrhage (ICH). We discovered a rare disease trait in thirteen individuals, including four fetuses, from eight unrelated families associated with homozygous loss-of-function variant alleles of ESAM which encodes an endothelial cell adhesion molecule. The c.115del (p.Arg39Glyfs∗33) variant, identified in six individuals from four independent families of Southeastern Anatolia, severely impaired the in vitro tubulogenic process of endothelial colony-forming cells, recapitulating previous evidence in null mice, and caused lack of ESAM expression in the capillary endothelial cells of damaged brain. Affected individuals with bi-allelic ESAM variants showed profound global developmental delay/unspecified intellectual disability, epilepsy, absent or severely delayed speech, varying degrees of spasticity, ventriculomegaly, and ICH/cerebral calcifications, the latter being also observed in the fetuses. Phenotypic traits observed in individuals with bi-allelic ESAM variants overlap very closely with other known conditions characterized by endothelial dysfunction due to mutation of genes encoding tight junction molecules. Our findings emphasize the role of brain endothelial dysfunction in NDDs and contribute to the expansion of an emerging group of diseases that we propose to rename as "tightjunctionopathies."

ANKLE2-related microcephaly: A variable microcephaly syndrome resembling Zika infection.

OBJECTIVE: This study delineates the clinical and molecular spectrum of ANKLE2-related microcephaly (MIC), as well as highlights shared pathological mechanisms between ANKLE2 and the Zika virus. METHODS: We identified 12 individuals with MIC and variants in ANKLE2 with a broad range of features. Probands underwent thorough phenotypic evaluations, developmental assessments, and anthropometric measurements. Brain imaging studies were systematically reviewed for developmental abnormalities. We functionally interrogated a subset of identified ANKLE2 variants in Drosophila melanogaster. RESULTS: All individuals had MIC (z-score ≤ -3), including nine with congenital MIC. We identified a broad range of brain abnormalities including simplified cortical gyral pattern, full or partial callosal agenesis, increased extra-axial spaces, hypomyelination, cerebellar vermis hypoplasia, and enlarged cisterna magna. All probands had developmental delays in at least one domain, with speech and language delays being the most common. Six probands had skin findings characteristic of ANKLE2 including hyper- and hypopigmented macules. Only one individual had scalp rugae. Functional characterization in Drosophila recapitulated the human MIC phenotype. Of the four variants tested, p.Val229Gly, p.Arg236*, and p.Arg536Cys acted as partial-loss-of-function variants, whereas the c.1421-1G>C splicing variant demonstrated a strong loss-of-function effect. INTERPRETATION: Deleterious variants in the ANKLE2 gene cause a unique MIC syndrome characterized by congenital or postnatal MIC, a broad range of structural brain abnormalities, and skin pigmentary changes. Thorough functional characterization has identified shared pathogenic mechanisms between ANKLE2-related MIC and congenital Zika virus infection. This study further highlights the importance of a thorough diagnostic evaluation including molecular diagnostic testing in individuals with MIC.

MCPH1: A Novel Case Report and a Review of the Literature.

Microcephaly primary hereditary (MCPH) is a congenital disease characterized by nonsyndromic reduction in brain size due to impaired neurogenesis, often associated with a variable degree of intellectual disability (ID). The genetic etiology of MCPH is heterogeneous and comprises more than 20 loci, nearly all following a recessive inheritance pattern. The first causative gene identified, MCPH1 or Microcephalin, encodes a centrosomal protein that modulates chromosome condensation and cell cycle progression. It is also involved in DNA damage response and telomere maintenance in the nucleus. Despite numerous studies on MCPH1 function, MCPH1-affected individuals are rare and the available clinical reports are not sufficient to define the natural history of the disease. Here, we present a novel patient with congenital microcephaly, ID, language delay, short stature, and other minor features such as strabismus. magnetic resonance imaging revealed ventriculomegaly, simplified gyral pattern in the frontal lobes, and a neuronal migration defect. Genetic testing detected a homozygous deletion of exons 1-8 of MCPH1. We compare the patients' characteristics with a list of features from MCPH1 cases described in the literature, in an effort to provide additional clues for a comprehensive definition of disease presentation and evolution.

Gingival Overgrowths Revealing PTEN Hamartoma Tumor Syndrome: Report of Novel PTEN Pathogenic Variants.

PTEN hamartoma tumor syndrome (PHTS), is a spectrum of disorders caused by mutations of PTEN, in which non-cancerous growths, called hamartomas, develop in different areas of the body, often including the oral mucosa. PHTS also implies a recognized increased risk of malignancies, as PTEN is a tumor suppressor gene capable of inhibiting progression of several cancers. One of the main and most common clinical manifestation of PHTS are gingival overgrowths presenting as warty lumps. The current study describes patients with gingival or mucosal enlargements leading to the diagnosis of PHTS associated to novel PTEN pathogenic variants. Patients referred to us for gingival lumps suggestive of PHTS associated overgrowths were submitted to genetic analysis in the PTEN gene. Two related and two unrelated patients were investigated. PTEN novel pathogenic variant was found in all of them. Two patients also fulfilled diagnostic criteria of Cowden syndrome (CS). Mucocutaneous lesions, and particularly diffuse gingival overgrowths, are both early and major clinical signs revealing a potential diagnosis of PHTS. Further genetic and clinical assessments are needed in order to confirm and clarify the diagnosis within the PHTS spectrum, including, among others, the CS. A correct interpretation of oral clinical features potentially associated to PHTS is mandatory for diagnosis and a surgical approach can be useful just in case of impairment of periodontal health or for aesthetic needs. The increased risk of malignancies associated to PHTS makes a correct diagnosis pivotal to set up an appropriate lifelong surveillance, aiming at secondary cancer prevention.

Posterior Lissencephaly Associated with Subcortical Band Heterotopia Due to a Variation in the CEP85L Gene: A Case Report and Refining of the Phenotypic Spectrum.

Lissencephaly describes a group of conditions characterized by the absence of normal cerebral convolutions and abnormalities of cortical development. To date, at least 20 genes have been identified as involved in the pathogenesis of this condition. Variants in CEP85L, encoding a protein involved in the regulation of neuronal migration, have been recently described as causative of lissencephaly with a posterior-prevalent involvement of the cerebral cortex and an autosomal dominant pattern of inheritance. Here, we describe a 3-year-old boy with slightly delayed psychomotor development and mild dysmorphic features, including bitemporal narrowing, protruding ears with up-lifted lobes and posterior plagiocephaly. Brain MRI at birth identified type 1 lissencephaly, prevalently in the temporo-occipito-parietal regions of both hemispheres with "double-cortex" (Dobyns' 1-2 degree) periventricular band alterations. Whole-exome sequencing revealed a previously unreported de novo pathogenic variant in the CEP85L gene (NM_001042475.3:c.232+1del). Only 20 patients have been reported as carriers of pathogenic CEP85L variants to date. They show lissencephaly with prevalent posterior involvement, variable cognitive deficits and epilepsy. The present case report indicates the clinical variability associated with CEP85L variants that are not invariantly associated with severe phenotypes and poor outcome, and underscores the importance of including this gene in diagnostic panels for lissencephaly.

PTEN Hamartoma Tumor Syndrome: Skin Manifestations and Insights Into Their Molecular Pathogenesis.

Germline PTEN pathogenic variants cause a spectrum of disorders collectively labeled PTEN Hamartoma Tumor Syndrome (PHTS) and featured by hamartomas, developmental anomalies and increased cancer risk. Studies on experimental models provided evidence that PTEN is a "haploinsufficient" tumor-suppressor gene, however, mechanisms involved in the pathogenesis of clinical manifestations in PHTS patients remain elusive. Beyond analyzing clinical and molecular features of a series of 20 Italian PHTS patients, we performed molecular investigations to explore the mechanisms involved in the pathogenesis of PTEN-associated manifestations, with special focus on mucocutaneous manifestations. Typical mucocutaneous features were present in all patients assessed, confirming that these are the most important clue to the diagnosis. The most frequent were papules located in the trunk or extremities (73.7%), oral mucosa papules (68.4%), acral/palmoplantar keratosis and facial papules (both 57.9%), according with literature data. Molecular analyses on one trichilemmoma suggested that the wild-type PTEN allele was retained and expressed, reinforcing the evidence that PTEN does not require a second somatic hit to initiate pathogenic processes. Unexpectedly, one patient also displayed a cutaneous phenotype consistent with atypical mole/melanoma syndrome; no variants were detected in known melanoma genes, but Whole Exome Sequencing showed the rare truncating variant c.495G>A in the CDH13 gene that might have cooperated with PTEN-haploinsufficiency to generate such phenotype. Our findings confirm the reproducibility of known PHTS manifestations in real-world practice, highlighting the role of mucocutaneous manifestations in facilitating prompt diagnosis of the syndrome, and provide some insights into the pathogenic process induced by PTEN alterations, which may contribute to its understanding.

Whole Exome Sequencing Is the Minimal Technological Approach in Probands Born to Consanguineous Couples.

We report on two siblings suffering from different pathogenic conditions, born to consanguineous parents. A multigene panel for brain malformations and microcephaly identified the homozygous splicing variant NM_005886.3:c.1416+1del in the KATNB1 gene in the older sister. On the other hand, exome sequencing revealed the homozygous frameshift variant NM_005245.4:c.9729del in the FAT1 gene in the younger sister, who had a more complex phenotype: in addition to bilateral anophthalmia and heart defects, she showed a right split foot with 4 toes, 5 metacarpals, second toe duplication and preaxial polydactyly on the right hand. These features have been never reported before in patients with pathogenic FAT1 variants and support the role of this gene in the development of limb buds. Notably, each parent was heterozygous for both of these variants, which were ultra-rare and rare, respectively. This study raises awareness about the value of using whole exome/genome sequencing rather than targeted gene panels when testing affected offspring born to consanguineous couples. In this way, exomic data from the parents are also made available for carrier screening, to identify heterozygous pathogenetic and likely pathogenetic variants in genes responsible for other recessive conditions, which may pose a risk for subsequent pregnancies.

Detecting Variants in the NBN Gene While Testing for Hereditary Breast Cancer: What to Do Next?

The NBN gene has been included in breast cancer (BC) multigene panels based on early studies suggesting an increased BC risk for carriers, though not confirmed by recent research. To evaluate the impact of NBN analysis, we assessed the results of NBN sequencing in 116 BRCA-negative BC patients and reviewed the literature. Three patients (2.6%) carried potentially relevant variants: two, apparently unrelated, carried the frameshift variant c.156_157delTT and another one the c.628G>T variant. The latter was subsequently found in 4/1390 (0.3%) BC cases and 8/1580 (0.5%) controls in an independent sample, which, together with in silico predictions, provided evidence against its pathogenicity. Conversely, the rare c.156_157delTT variant was absent in the case-control set; moreover, a 50% reduction of NBN expression was demonstrated in one carrier. However, in one family it failed to co-segregate with BC, while the other carrier was found to harbor also a probably pathogenic TP53 variant that may explain her phenotype. Therefore, the c.156_157delTT, although functionally deleterious, was not supported as a cancer-predisposing defect. Pathogenic/likely pathogenic NBN variants were detected by multigene panels in 31/12314 (0.25%) patients included in 15 studies. The risk of misinterpretation of such findings is substantial and supports the exclusion of NBN from multigene panels.

Colorectal polyposis as a clue to the diagnosis of Cowden syndrome: Report of two cases and literature review.

Cowden Syndrome (CS) is an autosomal dominant disorder characterized by hamartomatous growth in several organs and by an increased risk of malignancies, which makes its recognition essential to undertake risk reduction measures. Although the involvement of gastrointestinal tract is extremely common, awareness of this entity among gastroenterologists appears limited. We report on two unrelated patients: a 46-year-old male and a 38-year-old woman, who were referred to the Genetic Clinic because of the endoscopic finding of multiple colorectal polyps. Despite both displayed striking clinical (and, in the first case, familial) manifestations of Cowden Syndrome (PTEN Hamartoma Tumor Syndrome-PHTS), they had not been recognized before. Diagnosis of PHTS was confirmed by the detection of causative PTEN variants. Pathological examination of the polyps showed multiple histology types: hyperplastic, juvenile, serrated and lymphoid. Hyperplastic polyps analyzed from both patients failed to show BRAF V600E and KRAS codon 12/13 mutations, which provides evidence against their potential to evolve to colorectal cancer through the serrated pathway. We then reviewed the literature on gastrointestinal polyps detected in patients with Cowden Syndrome, in order to provide a comprehensive scenario of presentations: among a total of 568 patients reported in the literature, 91.7 % presented with colon polyps, with 63.0 % having two or more different histological types of polyps; besides, 58.5 % had extra-colonic polyps (located either in stomach and/or in small intestine). Finding multiple polyps with mixed and/or unusual histology should alert gastroenterologists and pathologists about the possible diagnosis of Cowden Syndrome and prompt the search for other manifestations of this condition in the patient.

HDAC8 Loss of Function and SHOX Haploinsufficiency: Two Independent Genetic Defects Responsible for a Complex Phenotype.

We report a patient with developmental delay, brachydactyly type E, short stature, and tetralogy of Fallot. Brachydactyly-mental retardation syndrome (BDMR) was suspected based on the phenotype; however, array CGH excluded a 2q37 deletion, but identified a deletion encompassing the SHOX gene. BDMR is characterized by cognitive impairment, skeletal abnormalities involving hands and feet, short stature, and overweight. Most affected individuals carry relatively large 2q37 deletions encompassing HDAC4. This gene encodes a histone deacetylase involved in epigenetic regulation of cell growth and differentiation, specifically during endochondral bone formation in chondrocyte hypertrophy. Since SHOX haploinsufficiency can cause skeletal defects and short stature but would not fully explain the clinical picture of this patient, exome sequencing was performed, and a heterozygous HDAC8 frameshift mutation was identified. HDAC8 is a distinct histone deacetylase involved in cohesin recycling and is responsible for an X-linked dominant Cornelia de Lange-like phenotype. A new blended clinical phenotype may be explained by the result of a dual molecular diagnosis, which represents a combination of 2 independent genetic defects, with relevant implications for genetic counseling, clinical management, and prognosis.

Dealing With BRCA1/2 Unclassified Variants in a Cancer Genetics Clinic: Does Cosegregation Analysis Help?

Background: Detection of variants of uncertain significance (VUSs) in BRCA1 and BRCA2 genes poses relevant challenges for counseling and managing patients. VUS carriers should be managed similarly to probands with no BRCA1/2 variants detected, and predictive genetic testing in relatives is discouraged. However, miscomprehension of VUSs is common and can lead to inaccurate risk perception and biased decisions about prophylactic surgery. Therefore, efforts are needed to improve VUS evaluation and communication at an individual level. Aims: We aimed at investigating whether cosegregation analysis, integrated with a careful review of available functional data and in silico predictions, may improve VUSs interpretation and counseling in individual families. Methods: Patients with Breast Cancer (BC) and/or Ovarian Cancer (OC) fulfilling established criteria were offered genetic counseling and BRCA1/2 testing; VUSs identified in index cases were checked in other relatives affected by BC/OC whenever possible. As an alternative, if BC/OC clustered only in one branch of the family, the parental origin of the VUS was investigated. Public prediction tools and databases were used to collect additional information on the variants analyzed. Results: Out of 1045 patients undergoing BRCA1/2 testing in the period October 2011-April 2018, 66 (6.3%) carried class 3 VUSs. Cosegregation analysis was performed for 13 VUSs in 11 kindreds. Seven VUSs (53.8%) did not cosegregate with breast/ovarian cancer in the family, which provided evidence against their role in cancer clustering in those families. Among the 6 cosegregating VUSs, for two (BRCA1 c.5152+2T>G and BRCA2 c.7975A>G) additional evidence exists from databases and in silico tools supporting their pathogenicity, which reinforces the hypothesis they may have had a predisposing effect in respective families. For the remaining four VUSs (31%), cosegregation analysis failed to provide relevant information. Conclusion: Our findings suggest that cosegregation analysis in a clinical context may be helpful to improve test result interpretation in the specific family and, therefore, should be offered whenever possible. Besides, obtaining and sharing cosegregation data helps gathering evidence that may eventually contribute to VUS classification.