Phenotypic Variability in Novel Doublecortin Gene Variants Associated with Subcortical Band Heterotopia.

Doublecortin, encoded by the DCX gene, plays a crucial role in the neuronal migration process during brain development. Pathogenic variants of the DCX gene are the major causes of the "lissencephaly (LIS) spectrum", which comprehends a milder phenotype like Subcortical Band Heterotopia (SBH) in heterozygous female subjects. We performed targeted sequencing in three unrelated female cases with SBH. We identified three DCX-related variants: a novel missense (c.601A>G: p.Lys201Glu), a novel nonsense (c.210C>G: p.Tyr70*), and a previously identified nonsense (c.907C>T: p.Arg303*) variant. The novel c.601A>G: p.Lys201Glu variant shows a mother-daughter transmission pattern across four generations. The proband exhibits focal epilepsy and achieved seizure freedom with a combination of oxcarbazepine and levetiracetam. All other affected members have no history of epileptic seizures. Brain MRIs of the affected members shows predominant fronto-central SBH with mixed pachygyria on the overlying cortex. The two nonsense variants were identified in two unrelated probands with SBH, severe drug-resistant epilepsy and intellectual disability. These novel DCX variants further expand the genotypic-phenotypic correlations of lissencephaly spectrum disorders. Our documented phenotypic descriptions of three unrelated families provide valuable insights and stimulate further discussions on DCX-SBH cases.

Generation of an induced pluripotent stem cell line (SDQLCHi067-A) from a patient with subcortical band heterotopia harboring a heterozygous mutation in DCX gene.

Subcortical band heterotopia (SHB) is a rare severe brain developmental malformation caused by deficient neuronal migration during the development of cerebral cortex. Here, a human induced pluripotent stem cell (iPSCs) line was established from a 4-year-1-month-old girl with SHB carrying a heterozygous mutation (c.568A > G, p.K190E) in DCX. The generated iPSC line showed the ability to differentiate into three lineages in vitro and was confirmed by pluripotency markers and the original gene mutation.

Leveraging multiple approaches for the detection of pathogenic deep intronic variants in developmental and epileptic encephalopathies: A case report.

About 50% of individuals with developmental and epileptic encephalopathies (DEEs) are unsolved following genetic testing. Deep intronic variants, defined as >100 bp from exon-intron junctions, contribute to disease by affecting the splicing of mRNAs in clinically relevant genes. Identifying deep intronic pathogenic variants is challenging and resource intensive, and interpretation is difficult due to limited functional annotations. We aimed to identify deep intronic variants in individuals suspected to have unsolved single gene DEEs. In a research cohort of unsolved cases of DEEs, we searched for children with a DEE syndrome predominantly caused by variants in specific genes in >80% of described cases. We identified two children with Dravet syndrome and one individual with classic lissencephaly. Multiple sequencing and bioinformatics strategies were employed to interrogate intronic regions in SCN1A and PAFAH1B1. A novel de novo deep intronic 12 kb deletion in PAFAH1B1 was identified in the individual with lissencephaly. We showed experimentally that the deletion disrupts mRNA splicing, which results in partial intron retention after exon 2 and disruption of the highly conserved LisH motif. We demonstrate that targeted interrogation of deep intronic regions using multiple genomics technologies, coupled with functional analysis, can reveal hidden causes of unsolved monogenic DEE syndromes. PLAIN LANGUAGE SUMMARY: Deep intronic variants can cause disease by affecting the splicing of mRNAs in clinically relevant genes. A deep intronic deletion that caused abnormal splicing of the PAFAH1B1 gene was identified in a patient with classic lissencephaly. Our findings reinforce that targeted interrogation of deep intronic regions and functional analysis can reveal hidden causes of unsolved epilepsy syndromes.

Further expansion and confirmation of phenotype in rare loss of YWHAE gene distinct from Miller-Dieker syndrome.

Deletion of 17p13.3 has varying degrees of severity on brain development based on precise location and size of the deletion. The most severe phenotype is Miller-Dieker syndrome (MDS) which is characterized by lissencephaly, dysmorphic facial features, growth failure, developmental disability, and often early death. Haploinsufficiency of PAFAH1B1 is responsible for the characteristic lissencephaly in MDS. The precise role of YWHAE haploinsufficiency in MDS is unclear. Case reports are beginning to elucidate the phenotypes of individuals with 17p13.3 deletions that have deletion of YWHAE but do not include deletion of PAFAH1B1. Through our clinical genetics practice, we identified four individuals with 17p13.3 deletion that include YWHAE but not PAFAH1B1. These patients have a similar phenotype of dysmorphic facial features, developmental delay, and leukoencephalopathy. In a review of the literature, we identified 19 patients with 17p13.3 microdeletion sparing PAFAH1B1 but deleting YWHAE. Haploinsufficiency of YWHAE is associated with brain abnormalities including cystic changes. These individuals have high frequency of epilepsy, intellectual disability, and dysmorphic facial features including prominent forehead, epicanthal folds, and broad nasal root. We conclude that deletion of 17p13.3 excluding PAFAH1B1 but including YWHAE is associated with a consistent phenotype and should be considered a distinct condition from MDS.

Clinical findings and genetic analysis of patients with copy number variants involving 17p13.3 using a single nucleotide polymorphism array: a single-center experience.

BACKGROUND: 17p13.3 microdeletions or microduplications (collectively known as copy number variants or CNVs) have been described in individuals with neurodevelopmental disorders. However, 17p13.3 CNVs were rarely reported in fetuses. This study aims to investigate the clinical significance of 17p13.3 CNVs with varied sizes and gene content in prenatal and postnatal samples. METHODS: Eight cases with 17p13.3 CNVs out of 8806 samples that had been subjected to single nucleotide polymorphism array analysis were retrospectively analyzed, along with karyotyping, clinical features, and follow-up. RESULTS: Eight cases with 17p13.3 CNVs consisted of five fetuses, one aborted embryo and two probands manifested severe congenital defects. The indications of prenatal testing varied considerably for the five fetuses, including ultrasound abnormalities (n = 3), segmental deletions indicated by non-invasive prenatal testing (n = 1), and intellectual disability in the mother of one fetus (n = 1). Of them, two and six harbored copy number gains and losses involving 17p13.3, respectively. The size of the detected 17p13.3 CNVs ranged from 576 kb to 5.7 Mb. Case 1 was diagnosed with 17p13.3 duplication syndrome, and cases 4, 6, and 7 with Miller-Dieker syndrome (MDS). Microdeletions of the 17p13.3 region in two cases (cases 5 and 8) involving YWHAE and CRK, sparing PAFAH1B1, were classified as pathogenic. Case 2 harbored a 576 kb microduplication, encompassing YWHAE and CRK but not PAFAH1B1, which was of maternal origin and considered a variant of uncertain significance. Case 3 carried one 74.2 Mb mosaic duplication of approximately 3.5 on chromosome 17p13.2q25.3, and two deletions at 17p13.3p13.2 and 17q25.3. The karyotype of case 3 was 46,XY,r(17)(p13q25). For five fetuses, only case 2 continued gestation and showed normal development at the age of 15 months; the others were subjected to termination of pregnancy. CONCLUSION: The clinical findings of 17p13.3 microdeletions or microduplications varied among subjects, and 17p13.3 CNVs often differ in size and gene content. Microdeletions or microduplications containing the typical MDS region, as well as the microdeletions involving YWHAE and CRK, could be classified as pathogenic. The clinical significance of small duplications including YWHAE and CRK but not PAFAH1B1 remains uncertain, for which parental testing and clinical heterogeneity should be considered in genetic counseling.

Bi-allelic CAMSAP1 variants cause a clinically recognizable neuronal migration disorder.

Non-centrosomal microtubules are essential cytoskeletal filaments that are important for neurite formation, axonal transport, and neuronal migration. They require stabilization by microtubule minus-end-targeting proteins including the CAMSAP family of molecules. Using exome sequencing on samples from five unrelated families, we show that bi-allelic CAMSAP1 loss-of-function variants cause a clinically recognizable, syndromic neuronal migration disorder. The cardinal clinical features of the syndrome include a characteristic craniofacial appearance, primary microcephaly, severe neurodevelopmental delay, cortical visual impairment, and seizures. The neuroradiological phenotype comprises a highly recognizable combination of classic lissencephaly with a posterior more severe than anterior gradient similar to PAFAH1B1(LIS1)-related lissencephaly and severe hypoplasia or absence of the corpus callosum; dysplasia of the basal ganglia, hippocampus, and midbrain; and cerebellar hypodysplasia, similar to the tubulinopathies, a group of monogenic tubulin-associated disorders of cortical dysgenesis. Neural cell rosette lineages derived from affected individuals displayed findings consistent with these phenotypes, including abnormal morphology, decreased cell proliferation, and neuronal differentiation. Camsap1-null mice displayed increased perinatal mortality, and RNAScope studies identified high expression levels in the brain throughout neurogenesis and in facial structures, consistent with the mouse and human neurodevelopmental and craniofacial phenotypes. Together our findings confirm a fundamental role of CAMSAP1 in neuronal migration and brain development and define bi-allelic variants as a cause of a clinically distinct neurodevelopmental disorder in humans and mice.

Prenatal diagnosis of Miller-Dieker syndrome/PAFAH1B1-related lissencephaly: Ultrasonography and genetically investigative results.

OBJECTIVE: To present the experience on prenatal diagnosis of Miller-Dieker syndrome (MDS)/PAFAH1B1-related lissencephaly to further determine fetal phenotypes of this syndrome. STUDY DESIGN: This was a retrospective study of ten pregnancies with fetal MDS/PAFAH1B1-related lissencephaly identified by chromosomal microarray (CMA)/exome sequencing (ES). Clinical and laboratory data were collected and reviewed for these cases, including maternal demographics, prenatal sonographic findings, CMA or ES results and pregnancy outcomes. RESULTS: Two cases were diagnosed in the first trimester because of an increased nuchal translucency. The remaining eight cases were identified at late gestation, including four in the second trimester because of fetal cardiac anomalies or ventriculomegaly, and four in the third trimester because of ventriculomegaly. CMA revealed 17p13.3 deletions in nine cases, and ES detected a de novo PAFAH1B1 missense mutation in one case. CONCLUSION: The prenatal presentation of MDS/PAFAH1B1-related lissencephaly depended on the gestational age when the diagnosis was made. Mild ventriculomegaly was the most common prenatal sonographic sign identified in cases of MDS/PAFAH1B1-related lissencephaly. It is important that fetal MRI and invasive testing with CMA should be considered in fetuses with apparently 'isolated' mild ventriculomegaly.

Lissencephaly with Congenital Hypothyroidism: A Case Report.

Lissencephaly is a malformation of cortical development associated with deficient neuronal migration and abnormal formation of cerebral convolutions or gyri. The lissencephaly spectrum consists of agyria, pachygyria, and subcortical band heterotopia. At least 19 genes have been identified in the causation of lissencephaly and related syndrome. Lissencephaly is associated with many other congenital disorders but the association of lissencephaly with congenital hypothyroidism is rarely reported. We report a case of a 10-year-old girl having lissencephaly with congenital hypothyroidism. Early diagnosis of lissencephaly and genetic counselling can be made in suspected cases and further possible interventions can be taken. Also, regular follow-up, monitoring, and better conservative management lead to a better prognosis. Keywords: congenital abnormalities; hypothyroidism; lissencephaly; neuronal migration disorders.

Lissencephaly: Update on diagnostics and clinical management.

Lissencephaly represents a spectrum of rare malformations of cortical development including agyria, pachygyria and subcortical band heterotopia. The progress in molecular genetics has led to identification of 31 lissencephaly-associated genes with the overall diagnostic yield over 80%. In this review, we focus on clinical and molecular diagnosis of lissencephaly and summarize the current knowledge on histopathological changes and their correlation with the MRI imaging. Additionally we provide the overview of clinical follow-up recommendations and available data on epilepsy management in patients with lissencephaly.

Genetic causes underlying grey matter heterotopia.

Grey matter heterotopia (GMH) can cause of seizures and are associated with a wide range of neurodevelopmental disorders and syndromes. They are caused by a failure of neuronal migration during fetal development, leading to clusters of neurons that have not reached their final destination in the cerebral cortex. We have performed an extensive literature search in Pubmed, OMIM, and Google scholar and provide an overview of known genetic associations with periventricular nodular heterotopia (PNVH), subcortical band heterotopia (SBH) and other subcortical heterotopia (SUBH). We classified the heterotopias as PVNH, SBH, SUBH or other and collected the genetic information, frequency, imaging features and salient features in tables for every subtype of heterotopia. This resulted in 105 PVNH, 16 SBH and 25 SUBH gene/locus associations, making a total of 146 genes and chromosomal loci. Our study emphasizes the extreme genetic heterogeneity underlying GMH. It will aid the clinician in establishing an differential diagnosis and eventually a molecular diagnosis in GMH patients. A diagnosis enables proper counseling of prognosis and recurrence risks, and enables individualized patient management.

[Prenatal diagnosis and genetic analysis of a fetus with Miller-Dieker syndrome].

OBJECTIVE: To explore the genetic basis for a fetus with lissencephaly. METHODS: Genomic DNA was extracted from amniotic fluid sample and subjected to copy number variation (CNV) analysis. RESULTS: The fetus was found to harbor a heterozygous 5.2 Mb deletion at 17p13.3p13.2, which encompassed the whole critical region of Miller-Dieker syndrome (MDS) (chr17: 1-2 588 909). CONCLUSION: The fetus was diagnosed with MDS. Deletion of the PAFAH1B1 gene may account for the lissencephaly found in the fetus.

Crk Haploinsufficiency Is Associated with Intrauterine Growth Retardation and Severe Postnatal Growth Failure.

BACKGROUND: Children with 17p13.3 microdeletions including the YWHAE gene show intrauterine growth restriction, craniofacial dysmorphisms, postnatal growth failure, and cognitive impairment. This region is characterized by genomic instability and has been associated with isolated lissencephaly sequence and Miller-Dieker syndrome characterized by facial dysmorphisms, microcephaly, short stature, seizures, cardiac malformations, and agyria. Whilst brain abnormalities are secondary to YWHAE deficiency, the cause of pre- and postnatal growth failure has not been identified yet. CASE PRESENTATION: We describe 2 patients (patient 1 15 years and patient 2 11 years and 10 months) referred to our Center of Pediatric Endocrinology for intrauterine growth retardation with de novo 17p13.3 deletion. In vitro assays showed a defect in CRK expression and GH/IGF1 signaling. rhGH therapy was effective in partially reducing the deficit in height in patient 1 and induced catch-up growth in patient 2. CONCLUSION: Our results suggest that 17p13.3 microdeletion involving CRK affects both GH and IGF1 signaling ultimately leading to pre- and postnatal growth retardation, secondary to partial insensitivity to GH/IGF1. rhGH therapy may be considered to reduce the height deficit in these patients, though data on adult height are lacking.

Prenatal diagnosis of Miller-Dieker syndrome by chromosomal microarray.

OBJECTIVE: To assess the experience on prenatal diagnosis of Miller-Dieker syndrome (MDS) to further delineate the fetal presentation of this syndrome. METHODS: This was a retrospective study. Fetal MDS was diagnosed prenatally by chromosomal microarray (CMA). Clinical data were reviewed for these cases, including maternal characteristics, indications for prenatal diagnosis, sonographic findings, CMA results, and pregnancy outcomes. RESULTS: Four cases were diagnosis as MDS by CMA. The most common sonographic features were ventriculomegaly (3/4) and polyhydramnios (2/4). Deletion sizes ranged from 1.5 to 5.4 Mb. All microdeletions were located at the MDS critical region and showed haploinsufficiency of the YWHAE, CRK, and PAFAH1B1. All patients chose to terminate the pregnancy. Parental chromosome analysis were preformed in three cases and demonstrated that two cases were de novo and one case was caused by inherited derivative chromosomes from parental balanced translocations. CONCLUSION: The most common prenatal ultrasound findings of MDS were ventriculomegaly and polyhydramnios. CMA can improve diagnostic precision for detecting MDS.

Prenatal diagnosis and genetic analysis of a fetus with Miller-Dieker syndrome / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for a fetus with lissencephaly.@*METHODS@#Genomic DNA was extracted from amniotic fluid sample and subjected to copy number variation (CNV) analysis.@*RESULTS@#The fetus was found to harbor a heterozygous 5.2 Mb deletion at 17p13.3p13.2, which encompassed the whole critical region of Miller-Dieker syndrome (MDS) (chr17: 1-2 588 909).@*CONCLUSION@#The fetus was diagnosed with MDS. Deletion of the PAFAH1B1 gene may account for the lissencephaly found in the fetus.

[Prenatal diagnosis of a fetus with Miller-Dieker syndrome].

OBJECTIVE: To carry out genetic diagnosis for a fetus. METHODS: Chromosome G-banding and chromosomal microarray analysis (CMA) were carried out for a fetus with abnormal morphology of lateral cerebral fissure. RESULTS: The karyotype of the fetus was normal, but CMA showed that it has carried a 1.4 Mb deletion at 17p13.3 region, which suggested a diagnosis of Miller-Dieker syndrome (MDS). CONCLUSION: Familiarity with clinical features and proper selection of genetic testing method are crucial for the diagnosis of MDS. Attention should be paid to microdeletions and microduplications which can be missed by conventional chromosomal karyotyping.

17p13.3 microdeletion including YWHAE and CRK genes: towards a clinical characterization.

INTRODUCTION: The short arm of chromosome 17 is characterized by a high density of low copy repeats, creating the opportunity for non-allelic homologous recombination to occur. Microdeletions of the 17p13.3 region are responsible for neuronal migration disorders including isolated lissencephaly sequence and Miller-Dieker syndrome. CASE REPORT: We describe the case of a 4-year and 2-month-old female with peculiar somatic traits and neurodevelopmental delay. At the age of 6 months, she started to present with infantile spasms syndrome; therefore, we administered vigabatrin followed by two cycles of adrenocorticotropic hormone, with good response. The coexistence of epileptic activity, neuropsychological delay, brain imaging abnormalities, and peculiar somatic features oriented us towards the hypothesis of a genetic etiology that could explain her clinical picture. Array CGH identified a 730 Kb deletion in the p13.3 region of the short arm of chromosome 17 including eleven genes, among these are YWHAE and CRK. DISCUSSION: Microdeletions of the 17p13.3 region involving only YWHAE and CRK, sparing PAFAH1B1, result in neurodevelopmental delay, growth retardation, craniofacial dysmorphisms, and mild structural brain abnormalities. Differently from the previously described patients carrying YWHAE and CRK deletions, the main complaint of our patient was represented by seizures. The absence of clear neuronal migration defects and mutations of the PAFAH1B1 gene in our patient underlines the central role of additional genes located in the 17p13.3 chromosomal region in the pathogenesis of epilepsy and helps to expand the phenotype of 17p13.3 microdeletion syndrome.