Heterozygosity for loss-of-function variants in LZTR1 is associated with isolated multiple café-au-lait macules.

PURPOSE: Pathogenic LZTR1 variants cause schwannomatosis and dominant/recessive Noonan syndrome (NS). We aim to establish an association between heterozygous loss-of-function (LoF) LZTR1 alleles and isolated multiple café-au-lait macules (CaLMs). METHODS: 849 unrelated participants with multiple CaLMs, lacking pathogenic/likely pathogenic NF1 and SPRED1 variants, underwent RASopathy gene panel sequencing. Data on 125 individuals with heterozygous LZTR1 variants were collected for characterizing their clinical features and the associated molecular spectrum. In vitro functional assessment was performed on a representative panel of missense variants and small in-frame deletions. RESULTS: Analysis revealed heterozygous LZTR1 variants in 6.0% (51/849) of participants, exceeding the general population prevalence. LZTR1-related CaLMs varied in number, displayed sharp or irregular borders, and were generally isolated, but occasionally associated with features recurring in RASopathies. In two families, CaLMs and schwannomas co-occurred. The molecular spectrum mainly consisted of truncating variants, indicating LoF. These variants substantially overlapped with those occurring in schwannomatosis and recessive NS. Functional characterization showed accelerated protein degradation or mislocalization, and failure to downregulate MAPK signaling. CONCLUSION: Our findings expand the phenotypic variability associated with LZTR1 variants, which, in addition to conferring susceptibility to schwannomatosis and causing dominant and recessive NS, occur in individuals with isolated multiple CaLMs.

Heterozygous Pathogenic Nonsense Variant in the ATM Gene in a Family with Unusually High Gastric Cancer Susceptibility.

Germline pathogenic variants (PVs) in the Ataxia Telangiectasia mutated (ATM) gene (MIM* 607585) increase the risk for breast, pancreatic, gastric, and prostatic cancer and, to a reduced extent, ovarian and colon cancer and melanoma, with moderate penetrance and variable expressivity. We describe a family presenting early-onset gastric cancer and harboring a heterozygous pathogenic ATM variant. The proband had gastric cancer (age 45) and reported a sister deceased due to diffuse gastric cancer (age 30) and another sister who developed diffuse gastric cancer (age 52) and ovarian serous cancer. Next generation sequencing for cancer susceptibility genes (APC, ATM, BRD1, BRIP1, CDH1, CDK4, CDKN2A, CHEK2, EPCAM, MLH1, MRE11, MSH2, MSH6, MUTYH, NBN, PALB2, PMS2, PTEN, RAD50, RAD51C, RAD51D, RECQL1, SMAD4, STK11, and TP53) was performed. Molecular analysis identified the truncating c.5944C>T, p.(Gln1982*) variant in the ATM (NM_000051.3; NP_000042.3) in the proband. The variant had segregated in the living affected sister and in the unaffected daughter of the deceased affected sister. Familial early-onset gastric cancer is an unusual presentation for ATM-related malignancies. Individual variants may result in different specific risks. Genotype-phenotype correlations are challenging given the low penetrance and variable expressivity. Careful family history assessments are pivotal for prevention planning and are strengthened by the availability of molecular diagnoses.

Clinical variability in DYNC2H1-related skeletal ciliopathies includes Ellis-van Creveld syndrome.

Deleterious variants of DYNC2H1 gene are associated with a wide spectrum of skeletal ciliopathies (SC). We used targeted parallel sequencing to analyze 25 molecularly unsolved families with different SCs. Deleterious DYNC2H1 variants were found in six sporadic patients and two monozygotic (MZ) twins. Clinical diagnoses included short rib-polydactyly type 3 in two cases, and asphyxiating thoracic dystrophy (ATD) in one case. Remarkably, clinical diagnosis fitted with EvC, mixed ATD/EvC and short rib-polydactyly/EvC phenotypes in three sporadic patients and the MZ twins. EvC/EvC-like features always occurred in compound heterozygotes sharing a previously unreported splice site change (c.6140-5A>G) or compound heterozygotes for two missense variants. These results expand the DYNC2H1 mutational repertoire and its clinical spectrum, suggesting that EvC may be occasionally caused by DYNC2H1 variants presumably acting as hypomorphic alleles.

Mosaic genome-wide paternal uniparental disomy after discordant results from primary fetal samples and cultured cells.

Mosaic genome-wide paternal uniparental disomy (GWpUPD) is a rare condition in which two euploid cell lines coexist in the same individual, one with biparental content and one with genome-wide paternal isodisomy. We report a complex prenatal diagnosis with discordant results from cultured and uncultured samples. A pregnant woman was referred for placental mesenchymal dysplasia and fetal omphalocele. Karyotype, array-CGH and Beckwith-Wiedemann Syndrome (BWS) testing (methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) of 11p15) performed on amniocytes were negative. After intrauterine fetal demise, the clinical suspicion persisted and BWS MS-MLPA was repeated on cultured cells from umbilical cord and amniotic fluid, revealing a mosaicism for KvH19 hypermethylation/KCNQ1OT1:TSS:DMR hypomethylation. These results, along with microsatellite analysis of the BWS region, were consistent with mosaic paternal 11p15 isodisomy. A concurrent maternal contamination exclusion test, analyzing polymorphic microsatellite markers on multiple chromosomes, showed an imbalance in favor of paternal alleles at all examined loci on cultured amniocytes and umbilical cord samples. This led to suspicion of mosaic GWpUPD, later confirmed by SNP-array, identifying a mosaic genome-wide paternal isodisomy affecting 60% of fetal cells. The assessment of mosaic GWpUPD requires multiple approaches beyond the current established diagnostic processes, also entertaining possible low-rate mosaicism. Clinical acumen and an integrated testing approach are the key to a successful diagnosis.

Genomic Breakpoints' Characterization of a Large CHEK2 Duplication in an Italian Family with Hereditary Breast Cancer.

CHEK2 (checkpoint kinase 2; MIM# 604373) is a tumor suppressor gene that encodes a serine threonine kinase involved in pathways such as DNA repair, cell cycle arrest, mitosis, and apoptosis. Pathogenic variants in CHEK2 contribute to a moderately increased risk of breast and other cancers. Several variant classes have been reported, either point mutations or large intragenic rearrangements. However, a significant portion of reported variants has an uncertain clinical significance. We report an intragenic CHEK2 duplication, ranging from intron 5 to intron 13, identified in an Italian family with hereditary breast cancer. Using long range PCR, with duplication-specific primers, we were able to ascertain the genomic breakpoint. We also performed a real-time PCR to assess a possible loss-of-function effect. The genomic characterization of large intragenic rearrangements in cancer susceptibility genes is important for the clinical management of the carriers and for a better classification of rare variants. The molecular definition of breakpoints allows for the prediction of the impact of the variant on transcripts and proteins, aiding in its characterization and clinical classification.

A Pain in the Neck: Lessons Learnt from Genetic Testing in Fetuses Detected with Nuchal Fluid Collections, Increased Nuchal Translucency versus Cystic Hygroma-Systematic Review of the Literature, Meta-Analysis and Case Series.

Fetal Nuchal fluid collections can manifest with two distinct presentations attributable to the same phenotypic spectrum: increased nuchal translucency (iNT) and cystic hygroma. The prenatal detection of these findings should prompt an accurate assessment through genetic counseling and testing, including karyotype, chromosomal microarray analysis (CMA) and multigene RASopathy panel. We performed a systematic review of the literature and meta-analysis, to calculate diagnostic yields of genetic testing in fetuses with iNT and cystic hygroma. We compared the results with a cohort of 96 fetuses with these isolated findings. Fetuses with isolated NT ≥ 2.5 mm showed karyotype anomalies in 22.76% of cases and CMA presented an incremental detection rate of 2.35%. Fetuses with isolated NT ≥ 3 mm presented aneuploidies in 14.36% of cases and CMA had an incremental detection rate of 3.89%. When the isolated NT measured at least 3.5 mm the diagnostic yield of karyotyping was 34.35%, the incremental CMA detection rate was 4.1%, the incremental diagnostic rate of the RASopathy panel was 1.44% and it was 2.44% for exome sequencing. Interestingly, CMA presents a considerable diagnostic yield in the group of fetuses with NT ≥ 3.5 mm. Similarly, exome sequencing appears to show promising results and could be considered after a negative CMA result.

When to test fetuses for RASopathies? Proposition from a systematic analysis of 352 multicenter cases and a postnatal cohort.

PURPOSE: Recent studies have identified suggestive prenatal features of RASopathies (e.g., increased nuchal translucency [NT], cystic hygroma [CH], hydrops, effusions, congenital heart diseases [CHD], polyhydramnios, renal anomalies). Our objective is to clarify indications for RASopathy prenatal testing. We compare genotype distributions between pre- and postnatal populations and propose genotype-phenotype correlations. METHODS: Three hundred fifty-two chromosomal microarray-negative cases sent for prenatal RASopathy testing between 2012 and 2019 were collected. For most, 11 RASopathy genes were tested. Postnatal cohorts (25 patients with available prenatal information and 108 institutional database genotypes) and the NSeuroNet database were used for genotypic comparisons. RESULTS: The overall diagnostic yield was 14% (50/352), with rates >20% for effusions, hydrops, and CHD. Diagnostic yield was significantly improved in presence of hypertrophic cardiomyopathy (HCM), persistent or associated CH, any suggestive finding combined with renal anomaly or polyhydramnios, or ≥2 ultrasound findings. Largest prenatal contributors of pathogenic variants were PTPN11 (30%), RIT1 (16%), RAF1 (14%), and HRAS (12%), which considerably differ from their prevalence in postnatal populations. HRAS, LZTR1, and RAF1 variants correlated with hydrops/effusions, and RIT1 with prenatal onset HCM. CONCLUSION: After normal chromosomal microarray, RASopathies should be considered when any ultrasound finding of lymphatic dysplasia or suggestive CHD is found alone or in association.

Recurrent prenatal PIEZO1-related lymphatic dysplasia: Expanding molecular and ultrasound findings.

Generalized lymphatic dysplasia (GLD), characterized by lymphedema, lymphangiectasias, chylothorax, effusions, represents a recognized cause of fetal hydrops. We describe for the first time recurrent pregnancies showing different ultrasound presentations of lymphatic dysplasia. The first fetus displayed diffuse subcutaneous cysts and septations while the second one presented fetal hydrops. Exome sequencing results at 18 gestational weeks in the second pregnancy showed compound heterozygosity for two novel PIEZO1 variants, afterwards detected also in the first fetus and in the heterozygous parents. Both ultrasound and genetic findings expand the current knowledge of PIEZO1-related GLD. We suggest exome sequencing in hydropic fetuses with normal cytogenetics and in pregnancies with recurrent hydrops/lymphatic dysplasia.