CBS mutations are good predictors for B6-responsiveness : a study based on the analysis of 35 brazilian classical homocystinuria patients

Background Classical homocystinuria (HCU) is a monogenic disease caused by the deficient activity of cystathionine β‐synthase (CβS). The objective of this study was to identify the CBS mutations in Brazilian patients with HCU. Methods gDNA samples were obtained for 35 patients (30 families) with biochemically confirmed diagnosis of HCU. All exons and exon‐intron boundaries of CBS gene were sequenced. Gene expression analysis by qRT‐PCR was performed in six patients. Novel missense point mutations were expressed in E. coli by site‐directed mutagenesis. Results Parental consanguinity was reported in 16 families, and pyridoxine responsiveness in five (15%) patients. Among individuals from the same family, all presented the same phenotype. Both pathogenic mutations were identified in 29/30 patients. Twenty‐one different mutations were detected in nine exons and three introns; being six common mutations. Most prevalent were p.Ile278Thr (18.2%), p.Trp323Ter (11.3%), p.Thr191Met (11.3%), and c.828+1G>A (11.3%). Eight novel mutations were found [c.2T>C, c.209+1delG, c.284T>C, c.329A>T, c.444delG, c.864_868delGAG c.989_991delAGG, and c.1223+5G>T]. Enzyme activity in E. coli‐expressed mutations was 1.5% for c.329A>T and 17.5% for c.284T>C. qRT‐PCR analysis revealed reduced gene expression in all evaluated genotypes: [c.209+1delG; c.572C>T]; [c.2T>C; c.828+1G>A]; [c.828+1G>A; c.1126G>A]; [c.833T>C; c.989_991delAGG]; [c.1058C>T; c.146C>T]; and [c.444delG; c.444delG]. The expected phenotype according to the genotype (pyridoxine responsiveness) matched in all cases. Conclusions Most patients studied were pyridoxine nonresponsive and presented early manifestations, suggesting severe phenotypes. Many private mutations were observed, but the four most prevalent mutations together accounted for over 50% of mutated alleles. A good genotype–phenotype relationship was observed within families and for the four most common mutations

Molecular analysis of the CTSK gene in a cohort of 33 brazilian families with Pycnodysostosis from a cluster in a brazilian northeast region

Pycnodysostosis is an autosomal recessive skeletal dysplasia, the prevalence of which is estimated to be low (1 per million). Nevertheless, in recent years we have found 27 affected individuals from 22 families in Ceara State, a region of the Brazilian Northeast, giving a local prevalence of 3 per million. This local prevalence associated with a high parental consanguinity, suggesting a possible founder effect, prompted us to perform a molecular investigation of these families to test this hypothesis. The CTSK gene was sequenced by the Sanger method in the patients and their parents. In addition to 18 families from Ceara, this study also included 15 families from other Brazilian regions. We also investigated the origin of each family from the birthplace of the parents and/or grandparents. We have studied 39 patients, including 33 probands and 6 sibs, from 33 families with pycnodysostosis and identified six mutations, five previously described (c.436G>C, c.580G>A, c.721C>T, c.830C>T and c.953G>A) and one novel frameshift (c.83dupT). This frameshift variant seems to have a single origin in Ceara State, since the haplotype study using the polymorphic markers D1S2344, D1S442, D1S498 and D1S2715 suggested a common origin. Most of the mutations were found in homozygosity in the patients from Ceara (83.3 %) while in other states the mutations were found in homozygosity in half of patients. We have also shown that most of the families currently living outside of Ceara have northeastern ancestors, suggesting a dispersion of these mutations from the Brazilian Northeast. The high frequency of pycnodysostosis in Ceara State is the consequence of the high inbreeding in that region. Several mutations, probably introduced a long time ago in Ceara, must have spread due to consanguineous marriages and internal population migration. However, the novel mutation seems to have a single origin in Ceara, suggestive of a founder effect21CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQCOORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP402008/2010-3; 590148/2011-73300017023p62015/22145-

Molecular analysis of the CTSK gene in a cohort of 33 Brazilian families with pycnodysostosis from a cluster in a Brazilian Northeast region

Background: Pycnodysostosis is an autosomal recessive skeletal dysplasia, the prevalence of which is estimated to be low (1 per million). Nevertheless, in recent years we have found 27 affected individuals from 22 families in Ceara State, a region of the Brazilian Northeast, giving a local prevalence of 3 per million. This local prevalence associated with a high parental consanguinity, suggesting a possible founder effect, prompted us to perform a molecular investigation of these families to test this hypothesis. Methods: The CTSK gene was sequenced by the Sanger method in the patients and their parents. In addition to 18 families from Ceara, this study also included 15 families from other Brazilian regions. We also investigated the origin of each family from the birthplace of the parents and/or grandparents. Results: We have studied 39 patients, including 33 probands and 6 sibs, from 33 families with pycnodysostosis and identified six mutations, five previously described (c.436G>C, c.580G>A, c.721C>T, c.830C>T and c.953G>A) and one novel frameshift (c.83dupT). This frameshift variant seems to have a single origin in Ceara State, since the haplotype study using the polymorphic markers D1S2344, D1S442, D1S498 and D1S2715 suggested a common origin. Most of the mutations were found in homozygosity in the patients from Ceara (83.3 %) while in other states the mutations were found in homozygosity in half of patients. We have also shown that most of the families currently living outside of Ceara have northeastern ancestors, suggesting a dispersion of these mutations from the Brazilian Northeast. Conclusions: The high frequency of pycnodysostosis in Ceara State is the consequence of the high inbreeding in that region. Several mutations, probably introduced a long time ago in Ceara, must have spread due to consanguineous marriages and internal population migration. However, the novel mutation seems to have a single origin in Ceara, suggestive of a founder effect.CNPq [402008/2010-3, 590148/2011-7]CAPES [3300017023p6]Fapesp [2015/22145-6]Univ Estadual Campinas, Fac Med Sci, Dept Med Genet, Skeletal Dysplasia Grp, Campinas, SP, BrazilChildrens Hosp Albert Sabin, Fortaleza, Ceara, BrazilMed Sci Fac Juazeiro do Norte FMJ, Juazeiro Do Norte, CE, BrazilUniv Estadual Campinas, Dept Med Genet, Fac Med Sci, Perinatal Genet Program, Campinas, SP, BrazilUniv Fed Campina Grande, Campina Grande, PB, BrazilChildrens Clin, City Hall Guarulhos, Guarulhos, SP, BrazilFed Univ Sao Carlos UFSCAR, Dept Med, Sao Carlos, SP, BrazilUniv Sao Paulo FCMUSP, Fac Med Sci, Childrens Inst, Med Genet Unit, Sao Paulo, SP, BrazilChildrens Hosp Juvencio Mattos, Sao Luis, MA, BrazilClin Hosp Porto Alegre, Med Genet Serv, Porto Alegre, RS, BrazilUniv Fed Sao Paulo, Ctr Genet Med, Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Ctr Genet Med, Sao Paulo, SP, BrazilCNPq:402008/2010-3590148/2011-7CAPES: 3300017023p6]FAPESP:2015/22145-6Web of Scienc

Diagnosis and management of classica homocystinuria in Brazil: a summary of 72 late-diagnosed patients

This study described a broad clinical characterization of classical homocystinuria (HCU) in Brazil. This was a cross-sectional, observational study including clinical and biochemical data from 72 patients (60 families) from Brazil (South, n = 13; Southeast, n = 37; Northeast, n = 8; North, n = 1; and Midwest, n = 1). Parental consanguinity was reported in 42% of families. Ocular manifestations were the earliest detected symptom (53% of cases), the main reason for diagnostic suspicion (63% of cases), and the most prevalent manifestation at diagnosis (67% of cases). Pyridoxine responsiveness was observed in 14% of patients. Only 22% of nonresponsive patients on treatment had total homocysteine levels <100 mmol/L. Most commonly used treatment strategies were pyridoxine (93% of patients), folic acid (90%), betaine (74%), vitamin B12 (27%), and low-methionine diet + metabolic formula (17%). Most patients diagnosed with HCU in Brazil are late diagnosed, express a severe phenotype, and poor metabolic control. Milder forms of HCU are likely underrepresented due to underdiagnosis.SILVA, L. C. S. da. Universidade Federal do Par

Alternative genomic diagnoses for individuals with a clinical diagnosis of Dubowitz syndrome.

Dubowitz syndrome (DubS) is considered a recognizable syndrome characterized by a distinctive facial appearance and deficits in growth and development. There have been over 200 individuals reported with Dubowitz or a "Dubowitz-like" condition, although no single gene has been implicated as responsible for its cause. We have performed exome (ES) or genome sequencing (GS) for 31 individuals clinically diagnosed with DubS. After genome-wide sequencing, rare variant filtering and computational and Mendelian genomic analyses, a presumptive molecular diagnosis was made in 13/27 (48%) families. The molecular diagnoses included biallelic variants in SKIV2L, SLC35C1, BRCA1, NSUN2; de novo variants in ARID1B, ARID1A, CREBBP, POGZ, TAF1, HDAC8, and copy-number variation at1p36.11(ARID1A), 8q22.2(VPS13B), Xp22, and Xq13(HDAC8). Variants of unknown significance in known disease genes, and also in genes of uncertain significance, were observed in 7/27 (26%) additional families. Only one gene, HDAC8, could explain the phenotype in more than one family (N = 2). All but two of the genomic diagnoses were for genes discovered, or for conditions recognized, since the introduction of next-generation sequencing. Overall, the DubS-like clinical phenotype is associated with extensive locus heterogeneity and the molecular diagnoses made are for emerging clinical conditions sharing characteristic features that overlap the DubS phenotype

Alternative genomic diagnoses for individuals with a clinical diagnosis of Dubowitz syndrome

Abstract Dubowitz syndrome (DubS) is considered a recognizable syndrome characterized by a distinctive facial appearance and deficits in growth and development. There have been over 200 individuals reported with Dubowitz or a “Dubowitz-like” condition, although no single gene has been implicated as responsible for its cause. We have performed exome (ES) or genome sequencing (GS) for 31 individuals clinically diagnosed with DubS. After genome-wide sequencing, rare variant filtering and computational and Mendelian genomic analyses, a presumptive molecular diagnosis was made in 13/27 (48%) families. The molecular diagnoses included biallelic variants in SKIV2L, SLC35C1, BRCA1, NSUN2; de novo variants in ARID1B, ARID1A, CREBBP, POGZ, TAF1, HDAC8, and copy-number variation at1p36.11(ARID1A), 8q22.2(VPS13B), Xp22, and Xq13(HDAC8). Variants of unknown significance in known disease genes, and also in genes of uncertain significance, were observed in 7/27 (26%) additional families. Only one gene, HDAC8, could explain the phenotype in more than one family (N = 2). All but two of the genomic diagnoses were for genes discovered, or for conditions recognized, since the introduction of next-generation sequencing. Overall, the DubS-like clinical phenotype is associated with extensive locus heterogeneity and the molecular diagnoses made are for emerging clinical conditions sharing characteristic features that overlap the DubS phenotype