RESUMEN

Tetrasomy X or 48,XXXX is a rare sex chromosome aneuploidy. The parental origin of tetrasomy X in a female patient with developmental delay was analyzed; all four X chromosomes were derived from the mother, and there were no paternally derived sex chromosomes. This finding indicates a rare incidental co-occurrence of maternal and paternal nondisjunction or polysomy rescue. The mechanism of 48,XXYY, which is related to developmental delay in males, was analyzed for comparison.

RESUMEN

Droplet digital PCR (ddPCR) is an emerging method for the absolute quantification of PCR products, and it can detect DNA copy numbers accurately. It analyzes the end-point absolute fluorescence signals of the PCR-positive droplets and calculates the target concentration. EvaGreen is a nonspecific double-stranded DNA-binding fluorescent dye, and the ddPCR system also supports assays using this cost-effective hydrolysis probe. Here, we describe a simple method of quantification for DNA copy numbers using the EvaGreen single-color fluorescent design.

Asunto(s)

Variaciones en el Número de Copia de ADN , Genómica , Colorantes Fluorescentes , Reacción en Cadena de la Polimerasa , ADN/genética

RESUMEN

Many disease-causing genes have been identified by determining the breakpoints of balanced chromosomal translocations. Recent progress in genomic analysis has accelerated the analysis of chromosomal translocation-breakpoints at the nucleotide level. Using a long-read whole-genome sequence, we analyzed the breakpoints of the cytogenetically balanced chromosomal translocation t(5;15)(q21;26.3), which was confirmed to be of de novo origin, in a patient with a neurodevelopmental disorder. The results showed complex rearrangements with seven fragments consisting of five breakpoint-junctions (BJs). Four of the five BJs showed microhomologies of 1-3-bp, and only one BJ displayed a signature of blunt-end ligation, indicating chromothripsis as the underlying mechanism. Although the BJs did not disrupt any disease-causing gene, the clinical features of the patient were compatible with MEF2C haploinsufficiency syndrome. Complex rearrangements were located approximately 2.5-Mb downstream of MEF2C. Therefore, position effects were considered the mechanism of the occurrence of MEF2C haploinsufficiency syndrome.

Asunto(s)

Trastornos del Neurodesarrollo , Translocación Genética , Humanos , Masculino , Lactante , Encéfalo/patología , Trastornos del Neurodesarrollo/genética

RESUMEN

MCT8 deficiency is an X-linked recessive disorder. We report the case of a 2-year-old Japanese boy with MCT8 deficiency caused by a novel frameshift variant, NM_006517.5(SLC16A2_v001):c.966dup [p.(Ile323Hisfs*57)]. He presented no head control and spoke no meaningful words, indicating severe developmental delay. Although missense or in-frame mutations of SLC16A2 are usually related to milder phenotypes and later-onset pyramidal signs, loss-of-function mutations are expected to cause severe clinical symptoms.

RESUMEN

Chromosomal triplications can be classified into recurrent and nonrecurrent triplications. Most of the nonrecurrent triplications are embedded in duplicated segments, and duplication-inverted triplication-duplication (DUP-TRP/INV-DUP) has been established as one of the mechanisms of triplication. This study aimed to reveal the underlying mechanism of the TRP-DUP-TRP pattern of chromosomal aberrations, in which the appearance of moving averages obtained through array-based comparative genomic hybridization analysis is similar to the shadows of the caldera volcano-like pattern, which were first identified in two patients with neurodevelopmental disabilities. For this purpose, whole-genome sequencing using long-read Nanopore sequencing was carried out to confirm breakpoint junctions. Custom array analysis and Sanger sequencing were also used to detect all breakpoint junctions. As a result, the TRP-DUP-TRP pattern consisted of only two patterns of breakpoint junctions in both patients. In patient 1, microhomologies were identified in breakpoint junctions. In patient 2, more complex architectures with insertional segments were identified. Thus, replication-based mechanisms were considered as a mechanism of the TRP-DUP-TRP pattern.

Asunto(s)

Rotura Cromosómica , Reordenamiento Génico/genética , Genoma Humano , Preescolar , Aberraciones Cromosómicas , Hibridación Genómica Comparativa , Femenino , Dosificación de Gen , Humanos , Lactante , Recién Nacido , Masculino , Secuenciación de Nanoporos

RESUMEN

Coffin-Siris syndrome (CSS) is a congenital anomaly syndrome characterized by developmental delay, coarse facial features, and hypoplasia of the fifth digit's nail or phalanges. Herein, we report a case of the 8-year-old female patient who showed developmental delay associated with dysplasia in the macular and large toe area. Comprehensive genomic analysis showed no possible candidate variants, but the subsequent genomic copy number analysis revealed a novel exonic deletion in the coding region of AT-rich interactive domain-containing protein 1B (ARID1B), a gene responsible for CSS. Genomic copy number analysis can aid in diagnosing CSS by confirming undiagnosed exonic deletions in ARID1B. Furthermore, this is the first report of CSS associated with bilateral macular dysplasia.

Asunto(s)

Anomalías Múltiples/diagnóstico , Anomalías Múltiples/genética , Proteínas de Unión al ADN/genética , Exones , Cara/anomalías , Deformidades Congénitas de la Mano/diagnóstico , Deformidades Congénitas de la Mano/genética , Discapacidad Intelectual/diagnóstico , Discapacidad Intelectual/genética , Mácula Lútea/anomalías , Micrognatismo/diagnóstico , Micrognatismo/genética , Cuello/anomalías , Fenotipo , Eliminación de Secuencia , Factores de Transcripción/genética , Niño , Femenino , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos

RESUMEN

Comprehensive genomic analysis was performed in a patient with mild psychomotor developmental delay, elevated creatine kinase, and white matter abnormalities. The results revealed biallelic pathogenic variants in the gene related to merosin-deficient congenital muscular dystrophy, NM_000426.3(LAMA2):c.1338_1339del [p.Gly447Phefs*7] and c.2749 + 2dup, which consist of compound heterozygous involvement with predicted loss-of-function and splicing abnormalities.

RESUMEN

The chromosomal region critical in Down syndrome has long been analyzed through genotype-phenotype correlation studies using data from many patients with partial trisomy 21. Owing to that, a relatively small region of human chromosome 21 (35.9 ~ 38.0 Mb) has been considered as Down syndrome critical region (DSCR). In this study, microarray-based comparative genomic hybridization analysis identified complex rearrangements of chromosome 21 in a patient manifesting clinical features partially overlapped with that of Down syndrome. Although the patient did not show up-slanting palpebral fissures and single transverse palmar creases, other symptoms were consistent with Down syndrome. Rearrangements were analyzed by whole-genome sequencing using Nanopore long-read sequencing. The analysis revealed that chromosome 21 was fragmented into seven segments and reassembled by six connected points. Among 12 breakpoints, 5 are located within the short region and overlapped with repeated segments. The rearrangement resulted in a maximum gain of five copies, but no region showed loss of genomic copy numbers. Breakpoint-junctions showed no homologous region. Based on these findings, chromoanasynthesis was considered as the mechanism. Although the distal 21q22.13 region was not included in the aberrant regions, some of the genes located on the duplicated regions, SOD1, SON, ITSN1, RCAN1, and RUNX1, were considered as possible candidate genes for clinical features of the patient. We discussed the critical region for Down syndrome, with the literature review.

Asunto(s)

Cromosomas Humanos Par 21/genética , Cromosomas Humanos/genética , Síndrome de Down/genética , Proteínas Adaptadoras del Transporte Vesicular/genética , Aberraciones Cromosómicas , Hibridación Genómica Comparativa/métodos , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Proteínas de Unión al ADN/genética , Síndrome de Down/fisiopatología , Femenino , Dosificación de Gen/genética , Estudios de Asociación Genética , Humanos , Hibridación Fluorescente in Situ/métodos , Lactante , Análisis por Micromatrices/métodos , Antígenos de Histocompatibilidad Menor/genética , Proteínas Musculares/genética , Superóxido Dismutasa-1/genética , Secuenciación Completa del Genoma

RESUMEN

The widespread use of genomic copy number analysis has revealed many previously unknown genomic structural variations, including some which are more complex. In this study, three consecutive microdeletions were identified in the same chromosome by microarray-based comparative genomic hybridization (aCGH) analysis for a patient with a neurodevelopmental disorder. Subsequent fluorescence in situ hybridization (FISH) analyses unexpectedly suggested complicated translocations and inversions. For better understanding of the mechanism, breakpoint junctions were analyzed by nanopore sequencing, as a new long-read whole-genome sequencing (WGS) tool. The results revealed a new chromosomal disruption, giving rise to four junctions in chromosome 7. According the sequencing results of breakpoint junctions, all junctions were considered as the consequence of multiple double-strand breaks and the reassembly of DNA fragments by nonhomologous end-joining, indicating chromothripsis. KMT2E, located within the deletion region, was considered as the gene responsible for the clinical features of the patient. Combinatory usage of aCGH and FISH analyses would be recommended for interpretation of structural variations analyzed through WGS.

Asunto(s)

Aberraciones Cromosómicas , Hibridación Genómica Comparativa/métodos , Proteínas de Unión al ADN/genética , Genómica/métodos , Hibridación Fluorescente in Situ/métodos , Trastornos del Neurodesarrollo/genética , Rotura Cromosómica , Inversión Cromosómica , Roturas del ADN de Doble Cadena , Variaciones en el Número de Copia de ADN , Reparación del ADN por Unión de Extremidades , Femenino , Estudios de Asociación Genética , Genoma Humano , Humanos , Secuenciación de Nanoporos , Análisis de Secuencia por Matrices de Oligonucleótidos , Inversión de Secuencia , Translocación Genética , Secuenciación Completa del Genoma , Adulto Joven

RESUMEN

Primrose syndrome is a congenital malformation syndrome characterized by intellectual disability, developmental delay, progressive muscle wasting, and ear lobe calcification. Mutations in the ZBTB20 gene have been established as being accountable for this syndrome. In this study, a novel de novo ZBTB20 mutation, NM_001164342.2:c.1945C>T (p.Leu649Phe), has been identified through whole exome sequencing (WES) in a female patient presenting a typical Primrose phenotype. Because the present patient exhibited recurrent otitis media, detailed immunological examinations were performed in this study and subnormal immunoglobulin levels were firstly identified in a Primrose patient. Anatomical anomaly of the inner ear has never been reported in this patient and WES data did not include any relevant variants causally linked with the immunologic defect. Thus, there is a possibility of a relation between an unclassified immunodeficiency with selective IgG2 deficiency and Primrose syndrome and this may be the reason of recurrent otitis media frequently observed in Primrose patients. Because subnormal levels of IgG2 in this patient might be caused by an unrelated and still uncharacterized genetic cause, further studies are required to prove the causal link between aberrant ZBTB20 function and immunodeficiency.

Asunto(s)

Anomalías Múltiples/genética , Calcinosis/genética , Discapacidades del Desarrollo/genética , Enfermedades del Oído/genética , Predisposición Genética a la Enfermedad , Discapacidad Intelectual/genética , Atrofia Muscular/genética , Proteínas del Tejido Nervioso/genética , Factores de Transcripción/genética , Anomalías Múltiples/diagnóstico , Anomalías Múltiples/patología , Calcinosis/diagnóstico , Calcinosis/patología , Niño , Discapacidades del Desarrollo/diagnóstico , Discapacidades del Desarrollo/patología , Enfermedades del Oído/diagnóstico , Enfermedades del Oído/patología , Femenino , Humanos , Inmunoglobulina G/genética , Discapacidad Intelectual/diagnóstico , Discapacidad Intelectual/patología , Masculino , Atrofia Muscular/diagnóstico , Atrofia Muscular/patología , Mutación Missense/genética , Fenotipo , Secuenciación del Exoma

RESUMEN

Genomic copy number variations (CNVs) can be detected by chromosomal microarray testing. However, upon final diagnosis, other methods may be recommended for a validation method to confirm CNVs. Trio analyses or carrier detection in family members are also frequently required. Previously, fluorescence in situ hybridization and/or quantitative PCR have been used; however, these methods present limitations. The purpose of this study was to establish a simple and rapid method to detect genomic copy numbers. We utilized droplet digital PCR (dPCR) with an intercalation method. Thirteen patients, who were diagnosed with MECP2 duplications via chromosomal microarray testing, were enrolled in this study. Four of their female relatives, who were verified as carriers of MECP2 duplications, were also included. Genomic copy numbers of MECP2 and IRAK1 were analyzed in comparison with reference genes: XIST and AR on the X-chromosome, and RPP30 and RPPH1 on the autosomal chromosomes. As a result, genomic copy numbers of MECP2 were rapidly and precisely detected by the dPCR system established in this study. This method can be widely applied as a diagnostic method to confirm CNVs on other chromosomal regions.

Asunto(s)

Duplicación de Gen/genética , Quinasas Asociadas a Receptores de Interleucina-1/genética , Proteína 2 de Unión a Metil-CpG/genética , Reacción en Cadena de la Polimerasa/métodos , Autoantígenos/genética , Aberraciones Cromosómicas , Variaciones en el Número de Copia de ADN/genética , Femenino , Genoma/genética , Humanos , Hibridación Fluorescente in Situ , Masculino , Proteína 2 de Unión a Metil-CpG/aislamiento & purificación , Análisis por Micromatrices , Linaje , ARN Largo no Codificante/genética , Receptores Androgénicos/genética , Ribonucleasa P/genética

RESUMEN

Pelizaeus-Merzbacher disease (PMD) is an X-linked, recessively inherited disorder associated with hypomyelination in the brain white matter. Mutations involving the proteolipid protein 1 gene (PLP1) located on Xq22.2 are responsible for PMD. PLP1 duplication is the major genetic abnormality in PMD patients. In this study, we utilized droplet-digital polymerase chain reaction (ddPCR) as a potential method to detect PLP1 duplications. Samples from four PMD patients and one of their mothers were used as positive controls. They had been previously diagnosed as having an additional PLP1 copy by chromosomal microarray testing. Genomic copy number of PLP1 was analyzed in triplicate experiments and compared with reference genes XIST and AR on the X-chromosome, and RPP30 and RPPH1 on the autosomes. As a result, precise results were obtained for each triplicate procedure. Thus, we concluded that triplicate experiments are no longer necessary. Compared to other methods, including fluorescence in-situ hybridization, multiplex ligation-dependent probe amplification, chromosomal microarray testing, and quantitative PCR, we were able to establish ddPCR results rapidly with very small amounts of DNA. In conclusion, we showed that ddPCR can be a potential diagnostic tool to confirm genomic copy number as a routine clinical application, including in prenatal diagnostic settings.

RESUMEN

BACKGROUND: Recently, many genes related to neurodevelopmental disorders have been identified by high-throughput genomic analysis; however, a comprehensive understanding of the mechanism underlying neurodevelopmental disorders remains to be established. To further understand these underlying mechanisms, we performed a comprehensive genomic analysis of patients with undiagnosed neurodevelopmental disorders. METHODS: Genomic analysis using next-generation sequencing with a targeted panel was performed for a total of 133 Japanese patients (male/female, 81/52) with previously undiagnosed neurodevelopmental disorders, including developmental delay (DD), intellectual disability (ID), autism spectrum disorder (ASD), and epilepsy. Genomic copy numbers were also analyzed using the eXome Hidden Markov Model (XHMM). RESULTS: Thirty-nine patients (29.3%) exhibited pathogenic or likely pathogenic findings with single-gene variants or chromosomal aberrations. Among them, 20 patients were presented here. Pathogenic or likely pathogenic variants were identified in 18 genes, including ACTG1, CACNA1A, CHD2, CDKL5, DNMT3A, EHMT1, GABRB3, GABRG2, GRIN2B, KCNQ3, KDM5C, MED13L, SCN2A, SHANK3, SMARCA2, STXBP1, SYNGAP1, and TBL1XR1. CONCLUSION: A diagnostic yield of 29.3% in this study was nearly the same as that previously reported from other countries. Thus, we suggest that there is no difference in genomic backgrounds in Japanese patients with undiagnosed neurodevelopmental disabilities. Although most of the patients possessed de novo variants, one of the patients showed an X-linked inheritance pattern. As X-linked recessive disorders exhibit the possibility of recurrent occurrence in the family, comprehensive molecular diagnosis is important for genetic counseling.

Asunto(s)

Trastornos del Neurodesarrollo/genética , Pueblo Asiatico/genética , Preescolar , Variaciones en el Número de Copia de ADN , Femenino , Predisposición Genética a la Enfermedad , Humanos , Japón , Masculino , Polimorfismo de Nucleótido Simple

RESUMEN

Pelizaeus-Merzbacher disease (PMD) is an X-linked recessive disorder caused by abnormalities in the gene PLP1. Most females harboring heterozygous PLP1 abnormalities are basically asymptomatic. However, as a result of abnormal patterns of X-chromosome inactivation, it is possible for some female carriers to be symptomatic. Whole-exome sequencing of a female patient with unknown spastic paraplegia was performed to obtain a molecular diagnosis. As a result, a de novo heterozygous single-nucleotide deletion in PLP1 [NM_000533.5(PLP1_v001):c.783del; p.Thr262Leufs*20] was identified. RNA sequencing was performed in a patient-derived lymphoblastoid cell line, confirming mono-allelic expression of the mutated allele and abnormal inactivation of the wild-type allele. The patient-derived lymphoblastoid cell line was then treated with VX680 or 5azadC, which resulted in restored expression of the wild-type allele. These two agents thus have the potential to reverse inappropriately-skewed inactivation of the X-chromosome.

Asunto(s)

Mutación del Sistema de Lectura , Proteína Proteolipídica de la Mielina/genética , Paraplejía/genética , Enfermedad de Pelizaeus-Merzbacher/genética , Alelos , Línea Celular , Niño , Cromosomas Humanos X/genética , Cromosomas Humanos X/metabolismo , Decitabina/farmacología , Femenino , Humanos , Enfermedad de Pelizaeus-Merzbacher/patología , Enfermedad de Pelizaeus-Merzbacher/terapia , Piperazinas/farmacología , Secuenciación del Exoma

Asunto(s)

Deleción Cromosómica , Discapacidad Intelectual/fisiopatología , Proteínas de Unión al ARN/genética , Espasmos Infantiles/fisiopatología , Cromosomas Humanos Par 1/genética , Humanos , Lactante , Discapacidad Intelectual/genética , Masculino , Espasmos Infantiles/genética

RESUMEN

Because biallelic SZT2 variants have been reported in patients with neurodevelopmental disorders associated with various degrees of developmental delay, intractable seizures, and distinctive features; this condition is recognized as an autosomal recessive disorder. Previously, eleven patients have been reported and most of them have compound heterozygous SZT2 variants, leading to premature termination. In these patients, all reported variants were unique and there were no common pathogenic variants identified. In this study, we identified a paternal uniparental disomy of chromosome 1 in a patient with a neurodevelopmental disorder associated with severe intellectual disability, intractable epilepsy, autistic features, distinctive features, and transient macrocephaly. This resulted in homozygous patterns through chromosome 1. Among the variants in chromosome 1, a rare SZT2 variant, NM_015284.3:c.6553C>T (p.Arg2185Trp), was selected as a powerful candidate variant in this patient. Although the clinical features of this patient are relatively milder than that reported previously, it may be derived from genetic heterogeneity. This is the first report of a homozygous missense SZT2 variant.

RESUMEN

Vici syndrome is a rare, autosomal recessive, multisystem disorder, characterized by agenesis of the corpus callosum, cataracts, psychomotor delay, cardiomyopathy, hypopigmentation, and recurrent infections. Mutations in the ectopic P-granules autophagy protein 5 homolog gene (EPG5), which encodes a key autophagy regulator, are responsible for this syndrome. A 3-year-old Japanese girl manifesting similar symptoms to those found in patients with Vici syndrome showed intractable diarrhea, rather than immunodeficiency. Whole exome sequencing identified only a heterozygous variant in EPG5, NM_020964.2(EPG5):c.3389A > C (p.His1130Pro), which was inherited from her mother. Sequencing analyses of the EPG5 messenger RNA showed only an altered nucleotide "C" at position, c.3389, indicating decreased expression of the wild-type allele. Microarray-based comparative genomic hybridization revealed a de novo microduplication in the exon 1 region. Large exon deletions and duplications of EPG5 have never been reported so far. This was considered the cause of the decreased expression of the wild-type allele. In conclusion, we successfully identified novel compound heterozygous mutations in EPG5 in a patient who was clinically considered to have Vici syndrome.

Asunto(s)

Agenesia del Cuerpo Calloso/diagnóstico , Agenesia del Cuerpo Calloso/genética , Catarata/diagnóstico , Catarata/genética , Exones , Duplicación de Gen , Heterocigoto , Proteínas de Membrana de los Lisosomas/genética , Mutación , Proteínas de Transporte Vesicular/genética , Proteínas Relacionadas con la Autofagia , Encéfalo/anomalías , Encéfalo/diagnóstico por imagen , Preescolar , Femenino , Estudios de Asociación Genética , Pruebas Genéticas , Genómica/métodos , Humanos , Japón , Imagen por Resonancia Magnética , Fenotipo