RESUMEN
Recurrent miscarriage (RM) affects millions of couples globally, and half of them have no demonstrated etiology. Genome sequencing (GS) is an enhanced and novel cytogenetic tool to define the contribution of chromosomal abnormalities in human diseases. In this study we evaluated its utility in RM-affected couples. We performed low-pass GS retrospectively for 1,090 RM-affected couples, all of whom had routine chromosome analysis. A customized sequencing and interpretation pipeline was developed to identify chromosomal rearrangements and deletions/duplications with confirmation by fluorescence in situ hybridization, chromosomal microarray analysis, and PCR studies. Low-pass GS yielded results in 1,077 of 1,090 couples (98.8%) and detected 127 chromosomal abnormalities in 11.7% (126/1,077) of couples; both members of one couple were identified with inversions. Of the 126 couples, 39.7% (50/126) had received former diagnostic results by karyotyping characteristic of normal human male or female karyotypes. Low-pass GS revealed additional chromosomal abnormalities in 50 (4.0%) couples, including eight with balanced translocations and 42 inversions. Follow-up studies of these couples showed a higher miscarriage/fetal-anomaly rate of 5/10 (50%) compared to 21/93 (22.6%) in couples with normal GS, resulting in a relative risk of 2.2 (95% confidence interval, 1.1 to 4.6). In these couples, this protocol significantly increased the diagnostic yield of chromosomal abnormalities per couple (11.7%) in comparison to chromosome analysis (8.0%, chi-square test p = 0.000751). In summary, low-pass GS identified underlying chromosomal aberrations in 1 in 9 RM-affected couples, enabling identification of a subgroup of couples with increased risk of subsequent miscarriage who would benefit from a personalized intervention.
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Aborto Habitual/diagnóstico , Aborto Habitual/genética , Aberraciones Cromosómicas , Secuenciación Completa del Genoma/métodos , Adulto , Femenino , Estudios de Seguimiento , Humanos , Cariotipificación , Masculino , Embarazo , Pronóstico , Estudios RetrospectivosRESUMEN
BACKGROUND: Exon-targeted microarrays can detect small (<1000 bp) intragenic copy number variants (CNVs), including those that affect only a single exon. This genome-wide high-sensitivity approach increases the molecular diagnosis for conditions with known disease-associated genes, enables better genotype-phenotype correlations, and facilitates variant allele detection allowing novel disease gene discovery. METHODS: We retrospectively analyzed data from 63,127 patients referred for clinical chromosomal microarray analysis (CMA) at Baylor Genetics laboratories, including 46,755 individuals tested using exon-targeted arrays, from 2007 to 2017. Small CNVs harboring a single gene or two to five non-disease-associated genes were identified; the genes involved were evaluated for a potential disease association. RESULTS: In this clinical population, among rare CNVs involving any single gene reported in 7200 patients (11%), we identified 145 de novo autosomal CNVs (117 losses and 28 intragenic gains), 257 X-linked deletion CNVs in males, and 1049 inherited autosomal CNVs (878 losses and 171 intragenic gains); 111 known disease genes were potentially disrupted by de novo autosomal or X-linked (in males) single-gene CNVs. Ninety-one genes, either recently proposed as candidate disease genes or not yet associated with diseases, were disrupted by 147 single-gene CNVs, including 37 de novo deletions and ten de novo intragenic duplications on autosomes and 100 X-linked CNVs in males. Clinical features in individuals with de novo or X-linked CNVs encompassing at most five genes (224 bp to 1.6 Mb in size) were compared to those in individuals with larger-sized deletions (up to 5 Mb in size) in the internal CMA database or loss-of-function single nucleotide variants (SNVs) detected by clinical or research whole-exome sequencing (WES). This enabled the identification of recently published genes (BPTF, NONO, PSMD12, TANGO2, and TRIP12), novel candidate disease genes (ARGLU1 and STK3), and further confirmation of disease association for two recently proposed disease genes (MEIS2 and PTCHD1). Notably, exon-targeted CMA detected several pathogenic single-exon CNVs missed by clinical WES analyses. CONCLUSIONS: Together, these data document the efficacy of exon-targeted CMA for detection of genic and exonic CNVs, complementing and extending WES in clinical diagnostics, and the potential for discovery of novel disease genes by genome-wide assay.
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Variaciones en el Número de Copia de ADN , Exones , Enfermedades Genéticas Congénitas , Estudios de Cohortes , Genoma Humano , Proteínas de Homeodominio/genética , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas de la Membrana/genética , Trastornos del Neurodesarrollo/genética , Proteínas Serina-Treonina Quinasas/genética , Estudios Retrospectivos , Serina-Treonina Quinasa 3 , Factores de Transcripción/genética , Secuenciación Completa del GenomaRESUMEN
Objective To assess the additive value of prenatal chromosomal microarray analysis (CMA) for all indications and the likelihood of detecting pathologic copy number variations (CNVs) based on specific indications. Methods A retrospective analysis was performed on amniocentesis and chorionic villi sampling results obtained between 2010 and 2014 in a single institution. A total of 3,314 consecutive patients undergoing invasive genetic testing for different indications were offered CMA in addition to standard karyotype. The prevalence of pathologic CNVs was compared between patients with low-risk indications and those with high-risk indications. Likewise, the prevalence of pathologic CNVs among patients with different sonographic abnormalities was calculated and compared with the low-risk group. Chi-square and Fisher exact tests were used for statistical analysis. Results The prevalence of pathologic CNVs was significantly higher in patients with high-risk indications and specifically those with sonographic abnormalities, compared with the low-risk group (2.8 and 5.9% vs. 0.4%, respectively; all p < 0.05). Conclusion Prenatal CMA detected clinically relevant CNVs in fetuses with a normal karyotype. Major structural malformations and nuchal translucency (NT) ≥ 3.0 mm are associated with the highest risk for a CMA abnormality. Nevertheless, the prevalence of pathologic CNVs in the low-risk population was high enough (1:250) to consider genetic counseling in this group.
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Aberraciones Cromosómicas , Anomalías Congénitas/diagnóstico por imagen , Anomalías Congénitas/genética , Variaciones en el Número de Copia de ADN , Pruebas Genéticas/métodos , Análisis por Micromatrices , Amniocentesis , Muestra de la Vellosidad Coriónica , Femenino , Asesoramiento Genético , Humanos , Cariotipo , Masculino , Embarazo , Estudios Retrospectivos , Factores de Riesgo , Ultrasonografía PrenatalRESUMEN
OBJECTIVES: The phenotype for 10q22q23 duplication is diverse, ranging from intellectual disability and dysmorphism to normal development. Interpreting the clinical significance of the duplication identified in this region is difficult, especially in the prenatal setting. This study aimed to characterize the prenatal findings associated with this submicroscopic imbalance and discuss the dilemmas in predicting the phenotype of 10q22q23 duplications. METHODS: This is a retrospective study of three cases of 10q22q23 duplications diagnosed prenatally by chromosomal microarray analysis. Detailed pregnancy outcome and pediatric follow-up were documented. RESULTS: The genotypic and phenotypic features of the reported cases were discussed. 10q22q23 duplications are associated with an unpredictable and variable phenotypic outcome. Despite there was no phenotype found to be shared by 50% of the duplication cases, congenital heart defects, hypotelorism, and developmental delays including speech and motor delay seem to be more common. CONCLUSIONS: The phenotype of 10q22q23 duplication is highly variable prenatally and postnatally. Identification of additional affected individuals with similar duplications is needed to provide further insights into the pathogenesis of this microduplication. © 2016 John Wiley & Sons, Ltd.
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Duplicación Cromosómica/genética , Cromosomas Humanos Par 10/genética , Fenotipo , Diagnóstico Prenatal/métodos , Anomalías Congénitas/genética , Discapacidades del Desarrollo/genética , Enfermedades en Gemelos/genética , Femenino , Retardo del Crecimiento Fetal/genética , Genotipo , Humanos , Hibridación Fluorescente in Situ , Análisis por Micromatrices , Medida de Translucencia Nucal , Embarazo , Embarazo Gemelar/genética , Estudios Retrospectivos , Ultrasonografía PrenatalRESUMEN
PURPOSE: We sought to investigate the utility of chromosomal microarray analysis (CMA) for prenatal diagnosis of oral clefts, as compared with traditional chromosome analysis, for improved prenatal genetic counseling and discovery of a potential correlation between genotype and oral cleft. METHODS: This retrospective analysis encompassed 270 prenatal oral cleft cases with documented detailed ultrasound findings and CMA results from four referral centers. Detection rates for pathogenic copy-number variants (CNVs) were calculated and compared with cases for which chromosome analysis was also performed. RESULTS: The overall detection rate was 14.8% (40/270) for pathogenic CNVs by CMA, 7.2% (9/125) for the nonsyndromic cases, and 21.4% (31/145) for the syndromic cases. Of the nonsyndromic cases with ultrasound soft markers, 20% (5/25) were identified with pathogenic CNVs. CMA showed an improved detection rate of 15.3% (29/190) compared with 10.5% (20/190) for chromosome analysis. CONCLUSION: This study not only highlights the improved detection of chromosomal defects by CMA in prenatal oral clefts but also deepens our understanding of oral clefts. The results suggest that CMA is highly recommended in prenatal invasive genetic testing not only for syndromic oral cleft cases but also for nonsyndromic cases with soft markers. Candidate genes including CRKL, AKAP8, SYDE1, BRD4 are worthy of further investigation regarding their role in human palatogenesis.Genet Med 18 10, 1052-1055.
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Fisura del Paladar/diagnóstico , Variaciones en el Número de Copia de ADN/genética , Asesoramiento Genético , Diagnóstico Prenatal , Aberraciones Cromosómicas , Fisura del Paladar/genética , Fisura del Paladar/fisiopatología , Estudios de Cohortes , Femenino , Genómica , Humanos , Cariotipificación , Análisis por Micromatrices , EmbarazoRESUMEN
New human mutations are thought to originate in germ cells, thus making a recurrence of the same mutation in a sibling exceedingly rare. However, increasing sensitivity of genomic technologies has anecdotally revealed mosaicism for mutations in somatic tissues of apparently healthy parents. Such somatically mosaic parents might also have germline mosaicism that can potentially cause unexpected intergenerational recurrences. Here, we show that somatic mosaicism for transmitted mutations among parents of children with simplex genetic disease is more common than currently appreciated. Using the sensitivity of individual-specific breakpoint PCR, we prospectively screened 100 families with children affected by genomic disorders due to rare deletion copy-number variants (CNVs) determined to be de novo by clinical analysis of parental DNA. Surprisingly, we identified four cases of low-level somatic mosaicism for the transmitted CNV in DNA isolated from parental blood. Integrated probabilistic modeling of gametogenesis developed in response to our observations predicts that mutations in parental blood increase recurrence risk substantially more than parental mutations confined to the germline. Moreover, despite the fact that maternally transmitted mutations are the minority of alleles, our model suggests that sexual dimorphisms in gametogenesis result in a greater proportion of somatically mosaic transmitting mothers who are thus at increased risk of recurrence. Therefore, somatic mosaicism together with sexual differences in gametogenesis might explain a considerable fraction of unexpected recurrences of X-linked recessive disease. Overall, our results underscore an important role for somatic mosaicism and mitotic replicative mutational mechanisms in transmission genetics.
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Variaciones en el Número de Copia de ADN/genética , Gametogénesis/genética , Enfermedades Genéticas Congénitas/genética , Células Germinativas/citología , Mutación de Línea Germinal/genética , Mosaicismo , División Celular , Femenino , Genómica , Humanos , Masculino , Modelos Genéticos , Mutación , Linaje , Estudios Prospectivos , Recurrencia , Riesgo , Caracteres Sexuales , Síndrome de Smith-Magenis/genéticaRESUMEN
A genetic etiology for autism spectrum disorders (ASDs) was first suggested from twin studies reported in the 1970s. The identification of gene mutations in syndromic ASDs provided evidence to support a genetic cause of ASDs. More recently, genome-wide copy number variant and sequence analyses have uncovered a list of rare and highly penetrant copy number variants (CNVs) or single nucleotide variants (SNVs) associated with ASDs, which has strengthened the claim of a genetic etiology for ASDs. Findings from research studies in the genetics of ASD now support an important role for molecular diagnostics in the clinical genetics evaluation of ASDs. Various molecular diagnostic assays including single gene tests, targeted multiple gene panels and copy number analysis should all be considered in the clinical genetics evaluation of ASDs. Whole exome sequencing could also be considered in selected clinical cases. However, the challenge that remains is to determine the causal role of genetic variants identified through molecular testing. Variable expressivity, pleiotropic effects and incomplete penetrance associated with CNVs and SNVs also present significant challenges for genetic counseling and prenatal diagnosis.
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Trastornos Generalizados del Desarrollo Infantil , Hibridación Genómica Comparativa , Pruebas Genéticas , Genómica , Análisis de Secuencia por Matrices de Oligonucleótidos , Niño , Trastornos Generalizados del Desarrollo Infantil/diagnóstico , Trastornos Generalizados del Desarrollo Infantil/genética , Variaciones en el Número de Copia de ADN , Humanos , Polimorfismo de Nucleótido Simple , Análisis de Secuencia de ADNRESUMEN
Somatic chromosomal mosaicism arising from post-zygotic errors is known to cause several well-defined genetic syndromes as well as contribute to phenotypic variation in diseases. However, somatic mosaicism is often under-diagnosed due to challenges in detection. We evaluated 10,362 patients with a custom-designed, exon-targeted whole-genome oligonucleotide array and detected somatic mosaicism in a total of 57 cases (0.55%). The mosaicism was characterized and confirmed by fluorescence in situ hybridization (FISH) and/or chromosome analysis. Different categories of abnormal cell lines were detected: (1) aneuploidy, including sex chromosome abnormalities and isochromosomes (22 cases), (2) ring or marker chromosomes (12 cases), (3) single deletion/duplication copy number variations (CNVs) (11 cases), (4) multiple deletion/duplication CNVs (5 cases), (5) exonic CNVs (4 cases), and (6) unbalanced translocations (3 cases). Levels of mosaicism calculated based on the array data were in good concordance with those observed by FISH (10-93%). Of the 14 cases evaluated concurrently by chromosome analysis, mosaicism was detected solely by the array in 4 cases (29%). In summary, our exon-targeted array further expands the diagnostic capability of high-resolution array comparative genomic hybridization in detecting mosaicism for cytogenetic abnormalities as well as small CNVs in disease-causing genes.
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Hibridación Genómica Comparativa/métodos , Exones , Mosaicismo , Adolescente , Adulto , Aneuploidia , Línea Celular , Niño , Preescolar , Aberraciones Cromosómicas , Trastornos de los Cromosomas/diagnóstico , Trastornos de los Cromosomas/genética , Femenino , Humanos , Hibridación Fluorescente in Situ , Masculino , Mutación , Adulto JovenRESUMEN
In clinical diagnostics, both array comparative genomic hybridization (array CGH) and single nucleotide polymorphism (SNP) genotyping have proven to be powerful genomic technologies utilized for the evaluation of developmental delay, multiple congenital anomalies, and neuropsychiatric disorders. Differences in the ability to resolve genomic changes between these arrays may constitute an implementation challenge for clinicians: which platform (SNP vs array CGH) might best detect the underlying genetic cause for the disease in the patient? While only SNP arrays enable the detection of copy number neutral regions of absence of heterozygosity (AOH), they have limited ability to detect single-exon copy number variants (CNVs) due to the distribution of SNPs across the genome. To provide comprehensive clinical testing for both CNVs and copy-neutral AOH, we enhanced our custom-designed high-resolution oligonucleotide array that has exon-targeted coverage of 1860 genes with 60,000 SNP probes, referred to as Chromosomal Microarray Analysis - Comprehensive (CMA-COMP). Of the 3240 cases evaluated by this array, clinically significant CNVs were detected in 445 cases including 21 cases with exonic events. In addition, 162 cases (5.0%) showed at least one AOH region >10 Mb. We demonstrate that even though this array has a lower density of SNP probes than other commercially available SNP arrays, it reliably detected AOH events >10 Mb as well as exonic CNVs beyond the detection limitations of SNP genotyping. Thus, combining SNP probes and exon-targeted array CGH into one platform provides clinically useful genetic screening in an efficient manner.
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Hibridación Genómica Comparativa/métodos , Variaciones en el Número de Copia de ADN/genética , Técnicas de Genotipaje/métodos , Polimorfismo de Nucleótido Simple/genética , Genoma Humano , Genómica , Heterocigoto , Humanos , Análisis de Secuencia por Matrices de OligonucleótidosRESUMEN
PURPOSE: Chromosomal microarray analysis enables the detection of microdeletions/duplications and has become the standard in clinical diagnostic testing for individuals with congenital anomalies and developmental disabilities. In the era of genomic arrays, the value of traditional chromosome analysis needs to be reassessed. METHODS: We studied 3,710 unrelated patients by chromosomal microarray analysis and chromosome analysis simultaneously and compared the results. RESULTS: We found that chromosomal microarray analysis detected the chromosomal imbalances that were identified by chromosome analysis with the exception of six cases (0.16%) that had mosaic abnormalities. Of note, one case showed mosaicism for two abnormal cell lines, resulting in a balanced net effect and a normal chromosomal microarray analysis. Further structural abnormalities such as unbalanced translocations, rings, and complex rearrangements were subsequently clarified by chromosome analysis in 18% of the cases with abnormal chromosomal microarray analysis results. Apparently balanced rearrangements were detected by chromosome analysis in 30 cases (0.8%). CONCLUSION: Our data demonstrate that although chromosomal microarray analysis should be the first-tier test for clinical diagnosis of chromosome abnormalities, chromosome analysis remains valuable in the detection of mosaicism and delineation of chromosomal structural rearrangements.
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Aberraciones Cromosómicas , Análisis Citogenético , Genómica , Análisis de Secuencia por Matrices de Oligonucleótidos , Bandeo Cromosómico , Análisis Citogenético/métodos , Variaciones en el Número de Copia de ADN , Genómica/métodos , Humanos , Hibridación Fluorescente in Situ , Cariotipificación , Mosaicismo , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Translocación GenéticaRESUMEN
OBJECTIVE: To evaluate the results of prenatal chromosomal microarray analysis (CMA) on >1000 fetal samples referred for testing at our institution and to compare these data to published reports. METHODS: High resolution CMA was offered to women undergoing amniocentesis or chorionic villus sampling. Parental samples were obtained concurrently to exclude maternal cell contamination and assist interpretation of copy number variations. RESULTS: Clinically significant copy number variations were observed in 85/1115 cases (7.6%) overall, and in 45/1075 cases (4.2 %) if 40 abnormal cases with known chromosome abnormalities or familial genomic imbalances were excluded. Eighteen of the 1115 cases had variants of unclear clinical significance (1.6%). Indications yielding the most clinically significant findings were abnormal karyotype/fluorescence in situ hybridization (26/61, 42.6%), family history of chromosomal abnormality (13/137, 9.5%), abnormal ultrasound (38/410, 9.3%), abnormal serum screening (2/37, 5.4%) and advanced maternal age (5/394, 1.3%). Of 1075 cases having no previously known cytogenetic abnormality or family history, 18 (1.7%) had clinically significant genomic changes undetectable by conventional prenatal chromosome analysis. CONCLUSION: Current experience confirms that the detection rate of CMA for prenatal chromosomal abnormalities surpasses that of conventional karyotype analysis and continues to improve with higher resolution arrays, while maintaining a low frequency of results of unclear clinical significance.
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Amniocentesis/métodos , Muestra de la Vellosidad Coriónica/métodos , Trastornos de los Cromosomas/diagnóstico , Análisis de Secuencia por Matrices de Oligonucleótidos , Ultrasonografía Prenatal , Cariotipo Anormal , Adulto , Líquido Amniótico/química , Líquido Amniótico/citología , Células Cultivadas , Aberraciones Cromosómicas , Trastornos de los Cromosomas/epidemiología , Trastornos de los Cromosomas/genética , ADN/análisis , Femenino , Dosificación de Gen , Humanos , Hibridación Fluorescente in Situ , Valor Predictivo de las Pruebas , Embarazo , Texas/epidemiologíaRESUMEN
Oligonucleotide array-based comparative genomic hybridization (aCGH) targeted to coding exons of genes of interest has been proven to be a valuable diagnostic tool to complement with Sanger sequencing for the detection of large deletions/duplications. We have developed a custom designed oligonucleotide aCGH platform for this purpose. This array platform provides tiled coverage of the entire mitochondrial genome and high-density coverage of a set of nuclear genes involving mitochondrial and metabolic disorders and can be used to evaluate large deletions in targeted genes. A total of 1280 DNA samples from patients suspected of having mitochondrial or metabolic disorders were evaluated using this targeted aCGH. We detected 40 (3%) pathogenic large deletions in unrelated individuals, including 6 in genes responsible for mitochondrial DNA (mtDNA) depletion syndromes, 23 in urea cycle genes, 11 in metabolic and related genes. Deletion breakpoints have been confirmed in 31 cases by PCR and sequencing. The possible deletion mechanism has been discussed. These results illustrate the successful utilization of targeted aCGH to detect large deletions in nuclear and mitochondrial genomes. This technology is particularly useful as a complementary diagnostic test in the context of a recessive disease when only one mutant allele is found by sequencing. For female carriers of X-linked disorders, if sequencing analysis does not detect point mutations, targeted aCGH should be considered for the detection of large heterozygous deletions.
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Hibridación Genómica Comparativa , Enfermedades Metabólicas/diagnóstico , Enfermedades Metabólicas/genética , Enfermedades Mitocondriales/diagnóstico , Enfermedades Mitocondriales/genética , Adolescente , Adulto , Secuencia de Bases , Niño , Preescolar , Rotura Cromosómica , Variaciones en el Número de Copia de ADN , ADN Mitocondrial , Femenino , Eliminación de Gen , Genoma Humano , Genotipo , Humanos , Lactante , Masculino , Datos de Secuencia Molecular , Ornitina Carbamoiltransferasa/genética , Trastornos Innatos del Ciclo de la Urea/genética , Adulto JovenRESUMEN
We report 24 unrelated individuals with deletions and 17 additional cases with duplications at 10q11.21q21.1 identified by chromosomal microarray analysis. The rearrangements range in size from 0.3 to 12 Mb. Nineteen of the deletions and eight duplications are flanked by large, directly oriented segmental duplications of >98% sequence identity, suggesting that nonallelic homologous recombination (NAHR) caused these genomic rearrangements. Nine individuals with deletions and five with duplications have additional copy number changes. Detailed clinical evaluation of 20 patients with deletions revealed variable clinical features, with developmental delay (DD) and/or intellectual disability (ID) as the only features common to a majority of individuals. We suggest that some of the other features present in more than one patient with deletion, including hypotonia, sleep apnea, chronic constipation, gastroesophageal and vesicoureteral refluxes, epilepsy, ataxia, dysphagia, nystagmus, and ptosis may result from deletion of the CHAT gene, encoding choline acetyltransferase, and the SLC18A3 gene, mapping in the first intron of CHAT and encoding vesicular acetylcholine transporter. The phenotypic diversity and presence of the deletion in apparently normal carrier parents suggest that subjects carrying 10q11.21q11.23 deletions may exhibit variable phenotypic expressivity and incomplete penetrance influenced by additional genetic and nongenetic modifiers.
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Anomalías Múltiples/genética , Aberraciones Cromosómicas , Factores de Crecimiento Nervioso/genética , Duplicaciones Segmentarias en el Genoma/genética , Eliminación de Secuencia , Proteínas de Transporte Vesicular de Acetilcolina/genética , Niño , Preescolar , Mapeo Cromosómico , Cromosomas Humanos Par 10 , Variaciones en el Número de Copia de ADN , Discapacidades del Desarrollo/complicaciones , Discapacidades del Desarrollo/genética , Femenino , Variación Genética , Recombinación Homóloga , Humanos , Hibridación Fluorescente in Situ , Lactante , Discapacidad Intelectual/complicaciones , Discapacidad Intelectual/genética , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , PenetranciaRESUMEN
Metabolic disorders are inborn errors that often present in the neonatal period with a devastating clinical course. If not treated promptly, these diseases can result in severe, irreversible disease or death. Determining the molecular defects in metabolic diseases is important in providing a definitive diagnosis for patient management. Therefore, prenatal diagnosis for families with known mutations causing metabolic disorders is crucial for timely intervention. Here we present three families in which standard Sanger sequencing failed to provide a definitive diagnosis, but the detection of genomic deletions by array comparative genomic hybridization (CGH) specifically targeted to mitochondrial and metabolic disease genes, MitoMet®, was fundamental in providing accurate prenatal diagnosis. In addition, to our knowledge, two deletions are the smallest detected by oligonucleotide array CGH reported for their respective genes, OTC and ARG1. These data highlight the importance of targeted array CGH in patients with suspected metabolic disorders and incomplete or negative sequencing results, as well as its emerging role in prenatal diagnosis.
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Hibridación Genómica Comparativa , Enfermedades Metabólicas/diagnóstico , Enfermedades Metabólicas/genética , Diagnóstico Prenatal , Arginasa/genética , Secuencia de Bases , Niño , Cromosomas Humanos X/genética , Femenino , Eliminación de Gen , Heterocigoto , Humanos , Hiperargininemia/diagnóstico , Recién Nacido , Masculino , Enfermedades Metabólicas/enzimología , Ornitina Carbamoiltransferasa/genética , Linaje , EmbarazoRESUMEN
CPSI deficiency usually results in severe hyperammonemia presenting in the first days of life warranting prompt diagnosis. Most CPS1 defects are non-recurrent, private mutations, including point mutation, small insertions and deletions. In this study, we report the detection of large deletions varying from 1.4 kb to >130 kb in the CPS1 gene of 4 unrelated patients by targeted array CGH. These results underscore the importance of analysis of large deletions when only one mutation or no mutations are identified in cases where CPSI deficiency is strongly indicated.
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Carbamoil-Fosfato Sintasa (Amoniaco)/genética , Enfermedad por Deficiencia de Carbamoil-Fosfato Sintasa I/diagnóstico , Eliminación de Gen , Secuencia de Bases , Enfermedad por Deficiencia de Carbamoil-Fosfato Sintasa I/genética , Preescolar , Resultado Fatal , Femenino , Heterocigoto , Humanos , Recién Nacido , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , Polimorfismo de Nucleótido SimpleRESUMEN
We demonstrate the utility of an exon coverage microarray platform in detecting intragenic deletions: one in exons 24-27 of the EP300 gene and another in exons 27 and 28 of the CREBBP gene in two patients with Rubinstein-Taybi syndrome (RSTS). RSTS is a heterogeneous disorder in which approximately 45-55% of cases result from deletion or mutations in the CREBBP gene and an unknown portion of cases result from gene changes in EP300. The first case is a 3-year-old female with an exonic deletion of the EP300 gene who has classic facial features of RSTS without the thumb and great toe anomalies, consistent with the milder skeletal phenotype that has been described in other RSTS cases with EP300 mutations. In addition, the mother of this patient also had preeclampsia during pregnancy, which has been infrequently reported. The second case is a newborn male who has the classical features of RSTS. Our results illustrate that exon-targeted array comparative genomic hybridization (aCGH) is a powerful tool for detecting clinically significant intragenic rearrangements that would be otherwise missed by aCGH platforms lacking sufficient exonic coverage or sequencing of the gene of interest.
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Proteína de Unión a CREB/genética , Proteína p300 Asociada a E1A/genética , Exones/genética , Eliminación de Gen , Síndrome de Rubinstein-Taybi/genética , Preescolar , Hibridación Genómica Comparativa/métodos , Femenino , Humanos , Recién Nacido , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , EmbarazoRESUMEN
Array comparative genomic hybridization (aCGH) is a powerful tool for the molecular elucidation and diagnosis of disorders resulting from genomic copy-number variation (CNV). However, intragenic deletions or duplications--those including genomic intervals of a size smaller than a gene--have remained beyond the detection limit of most clinical aCGH analyses. Increasing array probe number improves genomic resolution, although higher cost may limit implementation, and enhanced detection of benign CNV can confound clinical interpretation. We designed an array with exonic coverage of selected disease and candidate genes and used it clinically to identify losses or gains throughout the genome involving at least one exon and as small as several hundred base pairs in size. In some patients, the detected copy-number change occurs within a gene known to be causative of the observed clinical phenotype, demonstrating the ability of this array to detect clinically relevant CNVs with subkilobase resolution. In summary, we demonstrate the utility of a custom-designed, exon-targeted oligonucleotide array to detect intragenic copy-number changes in patients with various clinical phenotypes.
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Hibridación Genómica Comparativa/métodos , Variaciones en el Número de Copia de ADN/genética , Exones/genética , Adolescente , Secuencia de Bases , Niño , Preescolar , Puntos de Rotura del Cromosoma , Femenino , Estudios de Asociación Genética , Humanos , Lactante , Recién Nacido , Masculino , Datos de Secuencia Molecular , Análisis de Secuencia de ADN , Eliminación de Secuencia/genética , Adulto JovenRESUMEN
Due to the lack of robust diagnostic methods and limited resolution of conventional microscopy, submicroscopic genomic duplication copy number variants (CNVs) have been long underascertained. The development of array CGH has enabled detection of microduplications with nearly the same sensitivity as microdeletions and thus allowing them to be routinely identified throughout the human genome. However, in contrast to microdeletions, clinical interpretation of microduplications more often presents a diagnostic dilemma, as the functional impact of these genomic alterations is not well understood. Microduplications are especially difficult to interpret when they encompass several genes or a portion of a gene. Determining their significance involves investigative teamwork between both the diagnostic laboratory and the clinician. We present the steps for interpreting the clinical significance of microduplications and representative examples of these challenging cases.
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Aberraciones Cromosómicas , Hibridación Genómica Comparativa/métodos , Duplicación de Gen , Exones/genética , Humanos , Secuencias Reguladoras de Ácidos Nucleicos/genéticaRESUMEN
Insertional translocations (ITs) are rare events that require at least three breaks in the chromosomes involved and thus qualify as complex chromosomal rearrangements (CCR). In the current study, we identified 40 ITs from approximately 18,000 clinical cases (1:500) using array-comparative genomic hybridization (aCGH) in conjunction with fluorescence in situ hybridization (FISH) confirmation of the aCGH findings, and parental follow-up studies. Both submicroscopic and microscopically visible IT events were detected. They were divided into three major categories: (1) simple intrachromosomal and interchromosomal IT resulting in pure segmental trisomy, (2) complex IT involving more than one abnormality, (3) deletion inherited from a parent with a balanced IT resulting in pure segmental monosomy. Of the cases in which follow-up parental studies were available, over half showed inheritance from an apparently unaffected parent carrying the same unbalanced rearrangement detected in the propositi, thus decreasing the likelihood that these IT events are clinically relevant. Nevertheless, we identified six cases in which small submicroscopic events were detected involving known disease-associated genes/genomic segments and are likely to be pathogenic. We recommend that copy number gains detected by clinical aCGH analysis should be confirmed using FISH analysis whenever possible in order to determine the physical location of the duplicated segment. We hypothesize that the increased use of aCGH in the clinic will demonstrate that IT occurs more frequently than previously considered but can identify genomic rearrangements with unclear clinical significance.
Asunto(s)
Cromosomas Humanos Par 3/genética , Cromosomas Humanos Par 6/genética , Hibridación Genómica Comparativa/métodos , Hibridación Fluorescente in Situ/métodos , Mutagénesis Insercional/genética , Translocación Genética , Adolescente , Niño , Preescolar , Deleción Cromosómica , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Reproducibilidad de los ResultadosRESUMEN
Thymidine kinase 2 (TK2), encoded by the TK2 gene on chromosome 16q22, is one of the deoxyribonucleoside kinases responsible for the maintenance of mitochondrial deoxyribonucleotide pools. Defects in TK2 mainly cause a myopathic form of the mitochondrial DNA depletion syndrome (MDDS). Currently, only point mutations and small insertions and deletions have been reported in TK2 gene; gross rearrangements of TK2 gene and possible hepatic involvement in patients with TK2 mutations have not been described. We report a non-consanguineous Jordanian family with three deceased siblings due to mtDNA depletion. Sequence analysis of the father detected a heterozygous c.761T>A (p.I254N) mutation in his TK2 gene; however, point mutations in the mother were not detected. Subsequent gene dosage analysis using oligonucleotide array CGH identified an intragenic approximately 5.8-kb deletion encompassing the 5'UTR to intron 2 of her TK2 gene. Sequence analysis confirmed that the deletion spans c.1-495 to c.283-2899 of the TK2 gene (nucleotide 65,136,256-65,142,086 of chromosome 16). Analysis of liver and muscle specimens from one of the deceased infants in this family revealed compound heterozygosity for the paternal point mutation and maternal intragenic deletion. In addition, a significant reduction of the mtDNA content in liver and muscle was detected (10% and 20% of age- and tissue-matched controls, respectively). Prenatal diagnosis was performed in the third pregnancy. The fetus was found to carry both the point mutation and the deletion. This child died 6months after birth due to myopathy. A serum specimen demonstrated elevated liver transaminases in two of the infants from whom results were available. This report expands the mutation spectrum associated with TK2 deficiency. While the myopathic form of MDDS appears to be the main phenotype of TK2 mutations, liver dysfunction may also be a part of the mitochondrial depletion syndrome caused by TK2 gene defects.