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1.
Genome Med ; 11(1): 48, 2019 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-31349857

RESUMO

BACKGROUND: Although mosaic variation has been known to cause disease for decades, high-throughput sequencing technologies with the analytical sensitivity to consistently detect variants at reduced allelic fractions have only recently emerged as routine clinical diagnostic tests. To date, few systematic analyses of mosaic variants detected by diagnostic exome sequencing for diverse clinical indications have been performed. METHODS: To investigate the frequency, type, allelic fraction, and phenotypic consequences of clinically relevant somatic mosaic single nucleotide variants (SNVs) and characteristics of the corresponding genes, we retrospectively queried reported mosaic variants from a cohort of ~ 12,000 samples submitted for clinical exome sequencing (ES) at Baylor Genetics. RESULTS: We found 120 mosaic variants involving 107 genes, including 80 mosaic SNVs in proband samples and 40 in parental/grandparental samples. Average mosaic alternate allele fraction (AAF) detected in autosomes and in X-linked disease genes in females was 18.2% compared with 34.8% in X-linked disease genes in males. Of these mosaic variants, 74 variants (61.7%) were classified as pathogenic or likely pathogenic and 46 (38.3%) as variants of uncertain significance. Mosaic variants occurred in disease genes associated with autosomal dominant (AD) or AD/autosomal recessive (AR) (67/120, 55.8%), X-linked (33/120, 27.5%), AD/somatic (10/120, 8.3%), and AR (8/120, 6.7%) inheritance. Of note, 1.7% (2/120) of variants were found in genes in which only somatic events have been described. Nine genes had recurrent mosaic events in unrelated individuals which accounted for 18.3% (22/120) of all detected mosaic variants in this study. The proband group was enriched for mosaicism affecting Ras signaling pathway genes. CONCLUSIONS: In sum, an estimated 1.5% of all molecular diagnoses made in this cohort could be attributed to a mosaic variant detected in the proband, while parental mosaicism was identified in 0.3% of families analyzed. As ES design favors breadth over depth of coverage, this estimate of the prevalence of mosaic variants likely represents an underestimate of the total number of clinically relevant mosaic variants in our cohort.

2.
J Genet Couns ; 28(2): 213-228, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30964584

RESUMO

There are approximately 7,000 rare diseases affecting 25-30 million Americans, with 80% estimated to have a genetic basis. This presents a challenge for genetics practitioners to determine appropriate testing, make accurate diagnoses, and conduct up-to-date patient management. Exome sequencing (ES) is a comprehensive diagnostic approach, but only 25%-41% of the patients receive a molecular diagnosis. The remaining three-fifths to three-quarters of patients undergoing ES remain undiagnosed. The Stanford Center for Undiagnosed Diseases (CUD), a clinical site of the Undiagnosed Diseases Network, evaluates patients with undiagnosed and rare diseases using a combination of methods including ES. Frequently these patients have non-diagnostic ES results, but strategic follow-up techniques identify diagnoses in a subset. We present techniques used at the CUD that can be adopted by genetics providers in clinical follow-up of cases where ES is non-diagnostic. Solved case examples illustrate different types of non-diagnostic results and the additional techniques that led to a diagnosis. Frequent approaches include segregation analysis, data reanalysis, genome sequencing, additional variant identification, careful phenotype-disease correlation, confirmatory testing, and case matching. We also discuss prioritization of cases for additional analyses.

3.
Genome Med ; 10(1): 74, 2018 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-30266093

RESUMO

BACKGROUND: Exome sequencing is now being incorporated into clinical care for pediatric and adult populations, but its integration into prenatal diagnosis has been more limited. One reason for this is the paucity of information about the clinical utility of exome sequencing in the prenatal setting. METHODS: We retrospectively reviewed indications, results, time to results (turnaround time, TAT), and impact of exome results for 146 consecutive "fetal exomes" performed in a clinical diagnostic laboratory between March 2012 and November 2017. We define a fetal exome as one performed on a sample obtained from a fetus or a product of conception with at least one structural anomaly detected by prenatal imaging or autopsy. Statistical comparisons were performed using Fisher's exact test. RESULTS: Prenatal exome yielded an overall molecular diagnostic rate of 32% (n = 46/146). Of the 46 molecular diagnoses, 50% were autosomal dominant disorders (n = 23/46), 41% were autosomal recessive disorders (n = 19/46), and 9% were X-linked disorders (n = 4/46). The molecular diagnostic rate was highest for fetuses with anomalies affecting multiple organ systems and for fetuses with craniofacial anomalies. Out of 146 cases, a prenatal trio exome option designed for ongoing pregnancies was performed on 62 fetal specimens, resulting in a diagnostic yield of 35% with an average TAT of 14 days for initial reporting (excluding tissue culture time). The molecular diagnoses led to refined recurrence risk estimates, altered medical management, and informed reproductive planning for families. CONCLUSION: Exome sequencing is a useful diagnostic tool when fetal structural anomalies suggest a genetic etiology, but other standard prenatal genetic tests did not provide a diagnosis.

4.
Am J Obstet Gynecol ; 217(6): 691.e1-691.e6, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-29032050

RESUMO

BACKGROUND: Since its debut in 2011, cell-free fetal DNA screening has undergone rapid expansion with respect to both utilization and coverage. However, conclusive data regarding the clinical validity and utility of this screening tool, both for the originally included common autosomal and sex-chromosomal aneuploidies as well as the more recently added chromosomal microdeletion syndromes, have lagged behind. Thus, there is a continued need to educate clinicians and patients about the current benefits and limitations of this screening tool to inform pre- and posttest counseling, pre/perinatal decision making, and medical risk assessment/management. OBJECTIVE: The objective of this study was to determine the positive predictive value and false-positive rates for different chromosomal abnormalities identified by cell-free fetal DNA screening using a large data set of diagnostic testing results on invasive samples submitted to the laboratory for confirmatory studies. STUDY DESIGN: We tested 712 patient samples sent to our laboratory to confirm a cell-free fetal DNA screening result, indicating high risk for a chromosome abnormality. We compiled data from all cases in which the indication for confirmatory testing was a positive cell-free fetal DNA screen, including the common trisomies, sex chromosomal aneuploidies, microdeletion syndromes, and other large genome-wide copy number abnormalities. Testing modalities included fluorescence in situ hybridization, G-banded karyotype, and/or chromosomal microarray analysis performed on chorionic villus samples, amniotic fluid, or postnatally obtained blood samples. Positive predictive values and false-positive rates were calculated from tabulated data. RESULTS: The positive predictive values for trisomy 13, 18, and 21 were consistent with previous reports at 45%, 76%, and 84%, respectively. For the microdeletion syndrome regions, positive predictive values ranged from 0% for detection of Cri-du-Chat syndrome and Prader-Willi/Angelman syndrome to 14% for 1p36 deletion syndrome and 21% for 22q11.2 deletion syndrome. Detection of sex chromosomal aneuploidies had positive predictive values of 26% for monosomy X, 50% for 47,XXX, and 86% for 47,XXY. CONCLUSION: The positive predictive values for detection of common autosomal and sex chromosomal aneuploidies by cell-free fetal DNA screening were comparable with other studies. Identification of microdeletions was associated with lower positive predictive values and higher false-positive rates, likely because of the low prevalence of the individual targeted microdeletion syndromes in the general population. Although the obtained positive predictive values compare favorably with those seen in traditional screening approaches for common aneuploidies, they highlight the importance of educating clinicians and patients on the limitations of cell-free fetal DNA screening tests. Improvement of the cell-free fetal DNA screening technology and continued monitoring of its performance after introduction into clinical practice will be important to fully establish its clinical utility. Nonetheless, our data provide valuable information that may aid result interpretation, patient counseling, and clinical decision making/management.


Assuntos
Ácidos Nucleicos Livres/sangue , Transtornos Cromossômicos/sangue , Amniocentese , Síndrome de Angelman/sangue , Síndrome de Angelman/diagnóstico , Síndrome de Angelman/genética , Amostra da Vilosidade Coriônica , Transtornos Cromossômicos/diagnóstico , Transtornos Cromossômicos/genética , Cromossomos Humanos X/genética , Síndrome do Miado do Gato/sangue , Síndrome do Miado do Gato/diagnóstico , Síndrome do Miado do Gato/genética , Síndrome de Down/sangue , Síndrome de Down/diagnóstico , Síndrome de Down/genética , Feminino , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Síndrome de Klinefelter/sangue , Síndrome de Klinefelter/diagnóstico , Síndrome de Klinefelter/genética , Análise em Microsséries , Síndrome de Prader-Willi/sangue , Síndrome de Prader-Willi/diagnóstico , Síndrome de Prader-Willi/genética , Valor Preditivo dos Testes , Gravidez , Diagnóstico Pré-Natal , Aberrações dos Cromossomos Sexuais , Transtornos do Cromossomo Sexual no Desenvolvimento Sexual/sangue , Transtornos do Cromossomo Sexual no Desenvolvimento Sexual/diagnóstico , Transtornos do Cromossomo Sexual no Desenvolvimento Sexual/genética , Trissomia/diagnóstico , Trissomia/genética , Síndrome da Trissomia do Cromossomo 13/sangue , Síndrome da Trissomia do Cromossomo 13/diagnóstico , Síndrome da Trissomia do Cromossomo 13/genética , Síndrome da Trissomía do Cromossomo 18/sangue , Síndrome da Trissomía do Cromossomo 18/diagnóstico , Síndrome da Trissomía do Cromossomo 18/genética , Síndrome de Turner/sangue , Síndrome de Turner/diagnóstico , Síndrome de Turner/genética
6.
Hum Genet ; 136(4): 377-386, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28251352

RESUMO

Impairment of ubiquitin-proteasome system activity involving ubiquitin ligase genes UBE3A, UBE3B, and HUWE1 and deubiquitinating enzyme genes USP7 and USP9X has been reported in patients with neurodevelopmental delays. To date, only a handful of single-nucleotide variants (SNVs) and copy-number variants (CNVs) involving TRIP12, encoding a member of the HECT domain E3 ubiquitin ligases family on chromosome 2q36.3 have been reported. Using chromosomal microarray analysis and whole-exome sequencing (WES), we have identified, respectively, five deletion CNVs and four inactivating SNVs (two frameshifts, one missense, and one splicing) in TRIP12. Seven of these variants were found to be de novo; parental studies could not be completed in two families. Quantitative PCR analyses of the splicing mutation showed a dramatically decreased level of TRIP12 mRNA in the proband compared to the family controls, indicating a loss-of-function mechanism. The shared clinical features include intellectual disability with or without autistic spectrum disorders, speech delay, and facial dysmorphism. Our findings demonstrate that E3 ubiquitin ligase TRIP12 plays an important role in nervous system development and function. The nine presented pathogenic variants further document that TRIP12 haploinsufficiency causes a childhood-onset neurodevelopmental disorder. Finally, our data enable expansion of the phenotypic spectrum of ubiquitin-proteasome dependent disorders.


Assuntos
Transtorno do Espectro Autista/genética , Proteínas de Transporte/genética , Facies , Haploinsuficiência , Deficiência Intelectual/genética , Transtornos do Desenvolvimento da Linguagem/genética , Ubiquitina-Proteína Ligases/genética , Adolescente , Transtorno do Espectro Autista/complicações , Criança , Pré-Escolar , Estudos de Coortes , Variações do Número de Cópias de DNA , Feminino , Humanos , Lactente , Deficiência Intelectual/complicações , Transtornos do Desenvolvimento da Linguagem/complicações , Masculino
7.
Hum Mutat ; 35(12): 1407-17, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25212744

RESUMO

Angelman syndrome is a neurodevelopmental disorder caused by a deficiency of the imprinted and maternally expressed UBE3A gene. Although de novo genetic and epigenetic imprinting defects of UBE3A genomic locus account for majority of Angelman diagnoses, approximately 10% of individuals affected with Angelman syndrome are a result of UBE3A loss-of-function mutations occurring on the expressed maternal chromosome. The variants described in this manuscript represent the analysis of 2,515 patients referred for UBE3A gene sequencing at our institution, along with a comprehensive review of the UBE3A mutation literature. Of these, 267 (10.62%) patients had a report issued for detection of a UBE3A gene nucleotide variant, which in many cases involved family studies resulting in reclassification of variants of unknown clinical significance (VUS). Overall, 111 (4.41%) probands had a nucleotide change classified as pathogenic or strongly favored to be pathogenic, 29 (1.15%) had a VUS, and 126 (5.0%) had a nucleotide change classified as benign or strongly favored to be benign. All variants and their clinical interpretations are submitted to NCBI ClinVar, a freely accessible human variation and phenotype database.


Assuntos
Síndrome de Angelman/genética , Mutação , Ubiquitina-Proteína Ligases/genética , Feminino , Humanos , Masculino , Linhagem
8.
N Engl J Med ; 369(16): 1502-11, 2013 Oct 17.
Artigo em Inglês | MEDLINE | ID: mdl-24088041

RESUMO

BACKGROUND: Whole-exome sequencing is a diagnostic approach for the identification of molecular defects in patients with suspected genetic disorders. METHODS: We developed technical, bioinformatic, interpretive, and validation pipelines for whole-exome sequencing in a certified clinical laboratory to identify sequence variants underlying disease phenotypes in patients. RESULTS: We present data on the first 250 probands for whom referring physicians ordered whole-exome sequencing. Patients presented with a range of phenotypes suggesting potential genetic causes. Approximately 80% were children with neurologic phenotypes. Insurance coverage was similar to that for established genetic tests. We identified 86 mutated alleles that were highly likely to be causative in 62 of the 250 patients, achieving a 25% molecular diagnostic rate (95% confidence interval, 20 to 31). Among the 62 patients, 33 had autosomal dominant disease, 16 had autosomal recessive disease, and 9 had X-linked disease. A total of 4 probands received two nonoverlapping molecular diagnoses, which potentially challenged the clinical diagnosis that had been made on the basis of history and physical examination. A total of 83% of the autosomal dominant mutant alleles and 40% of the X-linked mutant alleles occurred de novo. Recurrent clinical phenotypes occurred in patients with mutations that were highly likely to be causative in the same genes and in different genes responsible for genetically heterogeneous disorders. CONCLUSIONS: Whole-exome sequencing identified the underlying genetic defect in 25% of consecutive patients referred for evaluation of a possible genetic condition. (Funded by the National Human Genome Research Institute.).


Assuntos
Exoma , Doenças Genéticas Inatas/diagnóstico , Testes Genéticos/métodos , Análise de Sequência de DNA/métodos , Adolescente , Criança , Pré-Escolar , Genes Dominantes , Genes Recessivos , Genes Ligados ao Cromossomo X , Doenças Genéticas Inatas/genética , Humanos , Mutação , Fenótipo , Adulto Jovem
9.
J Hum Genet ; 56(7): 516-23, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21593745

RESUMO

Mucopolysaccharidosis type II (MPS II) is caused by mutations in the IDS gene, which encodes the lysosomal enzyme iduronate-2-sulfatase. In ∼20% of MPS II patients the disorder is caused by gross IDS structural rearrangements. We identified two male cases harboring complex rearrangements involving the IDS gene and the nearby pseudogene, IDSP1, which has been annotated as a low-copy repeat (LCR). In both cases the rearrangement included a partial deletion of IDS and an inverted insertion of the neighboring region. In silico analyses revealed the presence of repetitive elements as well as LCRs at the junctions of rearrangements. Our models illustrate two alternative consequences of rearrangements initiated by non-allelic homologous recombination of LCRs: resolution by a second recombination event (that is, Alu-mediated recombination), or resolution by non-homologous end joining repair. These complex rearrangements have the potential to be recurrent and may be present among those MSP II cases with previously uncharacterized aberrations involving IDS.


Assuntos
Elementos Alu , Rearranjo Gênico , Glicoproteínas/genética , Recombinação Genética , Criança , Pré-Escolar , Feminino , Loci Gênicos , Humanos , Masculino , Pessoa de Meia-Idade , Mucopolissacaridose II/genética , Mutação Puntual , Duplicações Segmentares Genômicas
10.
Genet Test Mol Biomarkers ; 14(5): 709-13, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20854095

RESUMO

Focal dermal hypoplasia (FDH) is an X-linked dominant disorder caused by mutations in the gene PORCN, which encodes a protein required for the secretion and signaling of Wnt proteins. While deletions are responsible for a small percentage of FDH-causing mutations, the vast majority of mutations are single-nucleotide substitutions or small deletions or insertions that can be identified by sequence analysis. In 2007, we implemented a PORCN gene sequencing test for individuals with a clinical diagnosis of FDH. To date, we have detected 12 novel PORCN mutations and 6 previously reported mutations in 53 such unrelated patients. The pathogenic PORCN mutations included nine nonsense mutations, three missense mutations, one small deletion, two small duplications, and three splice-site mutations. Of these mutations, two were found in affected men and were mosaic; one of these was found in three other affected women. The remaining 16 mutations were found only in women. All the mutations detected in women were presumed heterozygous. In addition to the disease-causing mutations, eight nucleotide variants of unknown significance were identified. Further characterization of these variants suggests that four of them are pathogenic mutations. These findings add to the heterogeneity of mutations in the PORCN gene that cause FDH.


Assuntos
Hipoplasia Dérmica Focal/genética , Proteínas de Membrana/genética , Mutação , Aciltransferases , Cromossomos Humanos X/genética , Códon sem Sentido , Análise Mutacional de DNA , Feminino , Hipoplasia Dérmica Focal/diagnóstico , Duplicação Gênica , Heterogeneidade Genética , Variação Genética , Genótipo , Humanos , Masculino , Mutação de Sentido Incorreto , Linhagem , Reação em Cadeia da Polimerase , Sítios de Splice de RNA/genética , Análise de Sequência de DNA , Deleção de Sequência
11.
Prenat Diagn ; 29(1): 29-39, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19012303

RESUMO

OBJECTIVE: To evaluate the use of array comparative genomic hybridization (aCGH) for prenatal diagnosis, including assessment of variants of uncertain significance, and the ability to detect abnormalities not detected by karyotype, and vice versa. METHODS: Women undergoing amniocentesis or chorionic villus sampling (CVS) for karyotype were offered aCGH analysis using a targeted microarray. Parental samples were obtained concurrently to exclude maternal cell contamination and determine if copy number variants (CNVs) were de novo, or inherited prior to issuing a report. RESULTS: We analyzed 300 samples, most were amniotic fluid (82%) and CVS (17%). The most common indications were advanced maternal age (N=123) and abnormal ultrasound findings (N=84). We detected 58 CNVs (19.3%). Of these, 40 (13.3%) were interpreted as likely benign, 15 (5.0%) were of defined pathological significance, while 3 (1.0%) were of uncertain clinical significance. For seven (approximately 2.3% or 1/43), aCGH contributed important new information. For two of these (1% or approximately 1/150), the abnormality would not have been detected without aCGH analysis. CONCLUSION: Although aCGH-detected benign inherited variants in 13.3% of cases, these did not present major counseling difficulties, and the procedure is an improved diagnostic tool for prenatal detection of chromosomal abnormalities.


Assuntos
Hibridização Genômica Comparativa , Dosagem de Genes/genética , Diagnóstico Pré-Natal/métodos , Adulto , Amniocentese , Amostra da Vilosidade Coriônica , Feminino , Aconselhamento Genético , Humanos , Cariotipagem , Análise em Microsséries , Gravidez
12.
Psychooncology ; 17(8): 783-9, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18688785

RESUMO

OBJECTIVE: Li-Fraumeni syndrome (LFS) confers an increased risk of multiple types of cancer in both children and adults. Clinical genetic testing for deleterious germline p53 gene mutations can identify most LFS-affected families. We evaluated factors associated with cancer-specific distress and perceived self-efficacy in coping with a positive genetic test result among persons at risk of having deleterious p53 mutations. METHODS: One hundred thirty-five persons from 15 LFS-affected families were invited to take part in a study that offered p53 genetic counseling and testing and to complete psychosocial measures. RESULTS: Participants (n=92) were more likely to be younger and female than nonparticipants (n=43). In multivariate analyses, greater cancer-specific distress was associated with having a lower quality of life, a higher perceived risk of having a p53 mutation, no personal history of cancer and a greater number of first degree relatives (FDRs) affected with cancer. Lower perceived self-efficacy in coping with a positive test result was associated with greater cancer worry, higher decisional conflict about p53 testing and having no personal history of cancer. CONCLUSIONS: Individual perceptions about cancer risk and p53 genetic testing, as well as personal experience with FDRs' cancer diagnoses and deaths, should be addressed during the counseling and testing process for LFS-affected families.


Assuntos
Aconselhamento Genético/métodos , Síndrome de Li-Fraumeni/genética , Neoplasias/diagnóstico , Neoplasias/genética , Mutação Puntual/genética , Adaptação Psicológica , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Conflito (Psicologia) , Tomada de Decisões , Feminino , Seguimentos , Genes p53 , Humanos , Pessoa de Meia-Idade , Qualidade de Vida/psicologia , Fatores de Risco , Autoeficácia , Inquéritos e Questionários , Adulto Jovem
13.
Prenat Diagn ; 27(12): 1112-7, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17849500

RESUMO

OBJECTIVES: Use high-resolution genome analysis to clarify the genomic integrity in a fetus with a cytogenetically balanced translocation t(2;9)(q11.2;q34.3). METHODS: High resolution molecular cytogenetic analyses including G-banded chromosome analysis, fluorescence in situ hybridization (FISH), and array-comparative genomic hybridization (CGH) were performed on cultured cells, and DNA extracted from chorionic villus sample (CVS), amniotic fluid cells and fetal tissue. In addition, a custom fosmid-based tiling path 9q34.3 microarray with a resolution of 35-40 kb was used for array-CGH. RESULTS: GTG-banding analysis showed an apparently balanced de novo translocation between the long arms of chromosomes 2 and 9; t(2;9)(q11.2;q34.3). Array-CGH using a targeted chromosomal microarray analysis (CMA) uncovered a submicroscopic deletion of the subtelomeric region of 9q34.3 revealing the unbalanced nature of the rearrangement. These results were confirmed independently by FISH. The deletion was delimited to 2.7 Mb in size using the 9q34.3 fosmid-based tiling path array-CGH. CONCLUSION: Array-CGH is a powerful tool for rapid detection of genomic imbalances associated with microdeletion/duplication syndromes and for the evaluation of de novo apparently balanced translocation to enable high-resolution genomic analysis at the breakpoints. Prenatal diagnosis of chromosomal rearrangements involving dosage-sensitive genomic regions is an important adjuvant to prenatal care and provides more accurate information for counseling and informed decision making.


Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 9 , Feto/ultraestrutura , Hibridização de Ácido Nucleico/métodos , Diagnóstico Pré-Natal/métodos , Translocação Genética , Adulto , Feminino , Humanos , Gravidez
14.
Vaccine ; 25(35): 6458-73, 2007 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-17651872

RESUMO

Gene expression in human peripheral blood mononuclear cells was systematically evaluated following smallpox and yellow fever vaccination, and naturally occurring upper respiratory infection (URI). All three infections were characterized by the induction of many interferon stimulated genes, as well as enhanced expression of genes involved in proteolysis and antigen presentation. Vaccinia infection was also characterized by a distinct expression signature composed of up-regulation of monocyte response genes, with repression of genes expressed by B and T-cells. In contrast, the yellow fever host response was characterized by a suppression of ribosomal and translation factors, distinguishing this infection from vaccinia and URI. No significant URI-specific signature was observed, perhaps reflecting greater heterogeneity in the study population and etiological agents. Taken together, these data suggest that specific host gene expression signatures may be identified that distinguish one or a small number of virus agents.


Assuntos
Perfilação da Expressão Gênica , Monócitos/metabolismo , Monócitos/virologia , Infecções Respiratórias/genética , Vacinação , Vaccinia/genética , Vacinas Virais/imunologia , Febre Amarela/genética , Adolescente , Adulto , DNA Complementar/biossíntese , DNA Complementar/genética , Interpretação Estatística de Dados , Feminino , Humanos , Masculino , Análise de Sequência com Séries de Oligonucleotídeos , RNA Viral/biossíntese , RNA Viral/genética , Análise de Regressão , Infecções Respiratórias/virologia , Vacina Antivariólica/imunologia , Vaccinia/virologia , Vírus Vaccinia/imunologia , Febre Amarela/virologia , Vacina contra Febre Amarela/imunologia , Vírus da Febre Amarela/imunologia
15.
PLoS One ; 2(3): e327, 2007 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-17389918

RESUMO

BACKGROUND: Array Comparative Genomic Hybridization (a-CGH) is a powerful molecular cytogenetic tool to detect genomic imbalances and study disease mechanism and pathogenesis. We report our experience with the clinical implementation of this high resolution human genome analysis, referred to as Chromosomal Microarray Analysis (CMA). METHODS AND FINDINGS: CMA was performed clinically on 2513 postnatal samples from patients referred with a variety of clinical phenotypes. The initial 775 samples were studied using CMA array version 4 and the remaining 1738 samples were analyzed with CMA version 5 containing expanded genomic coverage. Overall, CMA identified clinically relevant genomic imbalances in 8.5% of patients: 7.6% using V4 and 8.9% using V5. Among 117 cases referred for additional investigation of a known cytogenetically detectable rearrangement, CMA identified the majority (92.5%) of the genomic imbalances. Importantly, abnormal CMA findings were observed in 5.2% of patients (98/1872) with normal karyotypes/FISH results, and V5, with expanded genomic coverage, enabled a higher detection rate in this category than V4. For cases without cytogenetic results available, 8.0% (42/524) abnormal CMA results were detected; again, V5 demonstrated an increased ability to detect abnormality. Improved diagnostic potential of CMA is illustrated by 90 cases identified with 51 cryptic microdeletions and 39 predicted apparent reciprocal microduplications in 13 specific chromosomal regions associated with 11 known genomic disorders. In addition, CMA identified copy number variations (CNVs) of uncertain significance in 262 probands; however, parental studies usually facilitated clinical interpretation. Of these, 217 were interpreted as familial variants and 11 were determined to be de novo; the remaining 34 await parental studies to resolve the clinical significance. CONCLUSIONS: This large set of clinical results demonstrates the significantly improved sensitivity of CMA for the detection of clinically relevant genomic imbalances and highlights the need for comprehensive genetic counseling to facilitate accurate clinical correlation and interpretation.


Assuntos
Cromossomos Humanos/genética , Hibridização Genômica Comparativa/métodos , Análise de Sequência com Séries de Oligonucleotídeos , Desequilíbrio Alélico/genética , Variação Genética , Genoma Humano , Humanos , Recém-Nascido , Cariotipagem , Análise em Microsséries/métodos , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Fenótipo , Valores de Referência
16.
Genet Med ; 8(12): 784-92, 2006 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17172942

RESUMO

PURPOSE: Mutations in the MECP2 gene are associated with Rett syndrome, an X-linked mental retardation disorder in females. Mutations also cause variable neurodevelopmental phenotypes in rare affected males. Recent clinical testing for MECP2 gene rearrangements revealed that entire MECP2 gene duplication occurs in some males manifesting a progressive neurodevelopmental syndrome. METHODS: Clinical testing through quantitative DNA methods and chromosomal microarray analysis in our laboratories identified seven male patients with increased MECP2 gene copy number. RESULTS: Duplication of the entire MECP2 gene was found in six patients, and MECP2 triplication was found in one patient with the most severe phenotype. The Xq28 duplications observed in these males are unique and vary in size from approximately 200 kb to 2.2 Mb. Three of the mothers who were tested were asymptomatic duplication carriers with skewed X-inactivation. In silico analysis of the Xq28 flanking region showed numerous low-copy repeats with potential roles in recombination. CONCLUSIONS: These collective data suggest that increased MECP2 gene copy number is mainly responsible for the neurodevelopmental phenotypes in these males. These findings underscore the allelic and phenotypic heterogeneity associated with the MECP2 gene and highlight the value of molecular analysis for patient diagnosis, family members at risk, and genetic counseling.


Assuntos
Deficiências do Desenvolvimento/genética , Dosagem de Genes , Duplicação Gênica , Proteína 2 de Ligação a Metil-CpG/genética , Adolescente , Criança , Pré-Escolar , Aberrações Cromossômicas , Cromossomos Humanos X , Testes Genéticos , Humanos , Lactente , Masculino
17.
Am J Med Genet A ; 140(22): 2401-15, 2006 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-17041943

RESUMO

Mutations in GJB2 are associated with hereditary hearing loss. DNA sequencing of GJB2 in a cohort of hearing impaired patients and a multi-ethnic control group is reported. Among 610 hearing impaired cases, 43 DNA sequence variations were identified in the coding region of GJB2 including 24 mutations, 8 polymorphisms, 3 unclassified variants (G4D, R127C, M163V), 1 controversial variant (V37I), and 7 novel variants (G12C, N14D, V63A, T86M, L132V, D159, 592_600delinsCAGTGTTCATGACATTC). Sixteen non-coding sequence variations were also identified among cases including the IVS1+1A>G mutation, 2 polymorphisms, and 13 novel variants. A diagnosis of GJB2-associated hearing loss was confirmed for 63 cases (10.3%). Heterozygous mutations were found in 39 cases (6.4%). Eleven cases carrying novel or unclassified variants (1.8 %) and 18 cases carrying the controversial V37I variant were identified (3%). In addition, 294 control subjects from 4 ethnic groups were sequenced for GJB2. Thirteen sequence variations in the coding region of GJB2 were identified among controls including 2 mutations, 6 polymorphisms, 2 unclassified variants (G4D, T123N), 1 controversial variant (V37I), and 2 novel variants (R127L, V207L). Nine sequence variations were identified among controls in the non-coding regions in and around GJB2 exon 2. Of particular interest among controls were the variability in carrier rates and ethnic stratification of alleles, and the complex genotypes among Asians, 47% of whom carried two to four sequence variations in the coding region of GJB2. These data provide new information about carrier rates for GJB2-based hearing loss in various ethnic groups and contribute to evaluation of the pathogenicity of the controversial V37I variant.


Assuntos
Conexinas/genética , DNA/genética , Perda Auditiva Neurossensorial/genética , Mutação , Alelos , Sequência de Aminoácidos , Sequência de Bases , Estudos de Casos e Controles , Estudos de Coortes , Conexina 26 , Análise Mutacional de DNA , Grupos Étnicos/genética , Feminino , Frequência do Gene , Variação Genética , Genótipo , Heterozigoto , Humanos , Masculino , Mutação de Sentido Incorreto , Análise de Sequência de DNA
18.
Genet Med ; 8(4): 226-33, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16617243

RESUMO

PURPOSE: Li-Fraumeni syndrome (LFS) is associated with p53 germline mutations, and carriers are at increased risk for multiple primary cancers. We evaluated outcomes following the administration of a video-based decision aid (DA) prior to clinical p53 genetic counseling and testing among persons who had previously participated in cancer genetics research. METHODS: Fifty-seven individuals at risk for a known p53 mutation completed baseline and post-DA measures of psychological outcomes, plus knowledge and attitudes regarding p53 genetic testing. Counseling and testing uptake also was recorded. RESULTS: At baseline, multivariate analysis showed that greater testing intention was associated with lower decisional conflict (P < 0.01). Compared with baseline data, multivariate analyses of post-DA outcomes showed that knowledge about LFS and genetic testing increased and decisional conflict related to testing decreased (P < 0.001). Mean cancer worries scores decreased among all participants (P < 0.001), and mean depression scores decreased for males (P < 0.05). Thirty-nine (68%) completed pre-test genetic counseling and 23 (40%) subsequently gave a blood sample for clinical genetic testing. CONCLUSION: This intervention was useful as an initial outreach and educational method for families considering p53 genetic testing, and may improve knowledge about LFS as well as psychological outcomes.


Assuntos
Tomada de Decisões , Técnicas de Apoio para a Decisão , Família/psicologia , Aconselhamento Genético/psicologia , Testes Genéticos/psicologia , Adulto , Idoso , Feminino , Predisposição Genética para Doença/psicologia , Humanos , Síndrome de Li-Fraumeni/genética , Síndrome de Li-Fraumeni/prevenção & controle , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Mutação , Avaliação de Resultados (Cuidados de Saúde) , Participação do Paciente , Proteína Supressora de Tumor p53/genética
19.
Prenat Diagn ; 25(13): 1188-91, 2005 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-16353282

RESUMO

OBJECTIVES: To report a family with a history of Pelizaeus-Merzbacher disease (PMD) for which prenatal diagnosis of PLP1 gene duplication status was attempted by the use of custom array comparative genomic hybridization (aCGH). METHODS: A 28-year-old woman was referred for genetic counseling for her then current pregnancy because her existing 3-year-old son was diagnosed with a classic form of PMD. At 11 and 3/7 weeks gestation, chorionic villus sampling (CVS) was performed. Custom aCGH and fluorescence in situ hybridization (FISH) analyses were also performed on the DNA from family members. Fetal karyotyping revealed 46,XY. RESULTS: Analysis by aCGH revealed that the male fetus was not duplicated for the PLP1 gene, but confirmed a duplicated PLP1 gene in the 3-year-old son, and that the mother was a duplication carrier. These results were independently confirmed by FISH analysis. aCGH and FISH analyses on DNA and cells derived from cord blood confirmed PLP1 nonduplication in the newborn. CONCLUSION: aCGH is a reliable alternative method for detection of PLP1 copy number for prenatal diagnosis of Pelizaeus-Merzbacher disease.


Assuntos
Duplicação Gênica , Proteínas de Membrana/genética , Proteína Proteolipídica de Mielina/genética , Doença de Pelizaeus-Merzbacher/diagnóstico , Doença de Pelizaeus-Merzbacher/genética , Diagnóstico Pré-Natal , Adulto , Pré-Escolar , Feminino , Feto , Aconselhamento Genético , Humanos , Hibridização in Situ Fluorescente/métodos , Recém-Nascido , Masculino , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Gravidez
20.
Clin Cancer Res ; 11(13): 4689-93, 2005 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-16000562

RESUMO

Heterozygous mutations in one of the DNA mismatch repair genes cause hereditary nonpolyposis colorectal cancer (MIM114500). Turcot syndrome (MIM276300) has been described as the association of central nervous system malignant tumors and familial colorectal cancer and has been reported to be both a dominant and recessive disorder. Homozygous and compound heterozygous mutations in APC, MLH1, MSH2, and PMS2 genes have been reported in five families. Here we describe a nonconsanguineous Pakistani family, including a son with lymphoma and colorectal cancer diagnosed at ages 5 and 8, respectively, and an 8-year-old daughter with glioblastoma multiforme. Both children had features of neurofibromatosis type 1 including atypical café au lait spots and axillary freckling without a family history consistent with neurofibromatosis type 1, familial adenomatous polyposis, or hereditary nonpolyposis colorectal cancer. Mutational analysis was done for MLH1, MSH2, and MSH6 using denaturing high-performance liquid chromatography and sequencing of a blood sample from the daughter. A novel homozygous single base insertion mutation was identified (3634insT) resulting in a premature stop at codon 1,223 in exon 7 of the MSH6 gene. Both parents were found to be heterozygous for the 3634insT mutation. Microsatellite instability testing showed instability in the glioblastoma sample. We report here the first identification of a homozygous mutation in MSH6 in a family with childhood-onset brain tumor, lymphoma, colorectal cancer, and neurofibromatosis type 1 phenotype. Our findings support a role for MSH6 in Turcot syndrome and are consistent with an autosomal recessive mode of inheritance.


Assuntos
Polipose Adenomatosa do Colo/genética , Neoplasias Encefálicas/genética , Proteínas de Ligação a DNA/genética , Mutação , Polipose Adenomatosa do Colo/metabolismo , Polipose Adenomatosa do Colo/patologia , Sequência de Bases , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Criança , Cromatografia Líquida de Alta Pressão/métodos , Análise Mutacional de DNA , Saúde da Família , Feminino , Proteína Glial Fibrilar Ácida/análise , Homozigoto , Humanos , Imuno-Histoquímica , Antígeno Ki-67/análise , Masculino , Repetições de Microssatélites/genética , Mutagênese Insercional , Linhagem , Homologia de Sequência do Ácido Nucleico , Síndrome
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