[Mutation analysis for a methylmalonic acidemia pedigree without proband by high-throughput sequencing].

OBJECTIVE: To apply high-throughput sequencing for the detection of potential mutation in a methylmalonic academia pedigree for which no proband was available. METHODS: For a couple who had previously given birth to an affected child, 14 genes were re-sequenced by high-throughput sequencing. Suspected mutations were validated by Sanger sequencing. Specific mutations were tested for amniotic fluid sample from the fetus. RESULTS: High-throughput sequencing suggested that the husband has carried a heterozygous mutation of the MUT gene (Exon 3: c.729_730insTT; p.Asp244Leufs*39), while the wife also carried a heterozygous mutation of the MUT gene (Exon 5: c.914T>C; p.Leu305Ser). Both mutations were confirmed by Sanger sequencing. Testing of amniotic sample suggested that the fetus has carried neither mutation. Follow-up has found no sign of methylmalonic academia in the neonate. CONCLUSION: High-throughput sequencing is a sensitive method to screen a bunch of genes in a single test. For autosomal recessive diseases, when no proband is available, carrier testing for both parents with high-throughput sequencing can provide an alternative approach, though great caution should be taken in the setting of prenatal diagnosis.

Heterozygosis in aromatic amino acid decarboxylase deficiency: Evidence for a positive interallelic complementation between R347Q and R358H mutations.

Aromatic amino acid or Dopa decarboxylase (AADC or DDC) is a homodimeric pyridoxal 5'-phosphate (PLP) enzyme responsible for the generation of the neurotransmitters dopamine and serotonin. AADC deficiency is a rare inborn disease caused by mutations of the AADC gene leading to a defect of AADC enzyme and resulting in impaired dopamine and serotonin synthesis. Until now, only the molecular effects of homozygous mutations were analyzed. However, although heterozygous carriers of AADC deficiency were identified, the molecular aspects of their enzymatic phenotypes are not yet investigated. Here, we focus our attention on the R347Q/R358H and R347Q/R160W heterozygous mutations, and report for the first time the isolation and characterization, in the purified recombinant form, of the R347Q/R358H heterodimer and of the R358H homodimer. The results, integrated with those already known of the R347Q homodimeric variant, provide evidence that (i) the R358H mutation strongly reduces the PLP-binding affinity and the catalytic activity, and (ii) a positive interallelic complementation exists between the R347Q and the R358H mutations. Bioinformatics analyses provide the structural basis for these data. Unfortunately, the R347Q/R160W heterodimer was not obtained in a sufficient amount to allow its purification and characterization. Nevertheless, the biochemical features of the R160W homodimer give a contribution to the enzymatic phenotype of the heterozygous R347Q/R160W and suggest the possible relevance of Arg160 in the proper folding of human DDC. © 2018 IUBMB Life, 70(3):215-223, 2018.

Prenatal brain disruption in isolated sulfite oxidase deficiency.

BACKGROUND: Isolated sulfite oxidase deficiency (ISOD) is a very rare autosomal recessive inherited neurometabolic disease. The most striking postnatal neuroimaging finding is multicystic encephalomalacia, which occurs rapidly within days to weeks after birth and mimics severe hypoxic-ischemic encephalopathy. The aim of this study was to describe the prenatal neuroimaging features in a neonate and a fetus diagnosed with ISOD. RESULTS: We report an 11-day-old female neonate who presented with feeding difficulties, decreased activity, neonatal seizures, and movement disorders within a few days after birth. Brain MRI at 9 days of age showed cystic lesions over the left frontal and temporal areas, diffuse and evident T2 high signal intensity of bilateral cerebral cortex, and increased T2 signal intensity of the globus pallidi. A pronounced low level of plasma cysteine and normal level of plasma uric acid were noted. Mutation analysis of SUOX revealed homozygous c.1200C > G mutations, resulting in an amino acid substitution of tyrosine to a stop codon (Y400X). The diagnosis of ISOD was made. The brain MRI of a prenatally diagnosed ISOD fetus of the second pregnancy of the mother of the index case showed poor gyration and differentiation of cortical layers without formation of cystic lesions at gestational age 21 weeks. CONCLUSION: Cystic brain destruction might occur prenatally and neurodevelopment of gyration and differentiation of the cortical layers in the developing brain could be affected by sulfite accumulation early during the second trimester in ISOD patients. This is the first description of the prenatal neurodevelopment of brain disruption in ISOD.

[Mutation screening and prenatal diagnosis of methylmalonic academia in a Chinese pedigree by Ion Torrent semiconductor sequencing].

OBJECTIVE: To identify pathogenic mutations in a Chinese pedigree affected with methylmalonic academia for genetic counseling and prenatal diagnosis. METHODS: Molecular analysis of the MUT, MMACHC, MMAA and MMAB genes was performed for the proband with methylmalonic academia by Ion Torrent semiconductor sequencing. Candidate mutations were validated by Sanger sequencing. The couple was offered prenatal diagnosis via analyzing of the fetal DNA through amniocentesis. RESULTS: The proband was found to be compound heterozygous for c.609G>A (p.Trp203X) and c.658-660del AAG (p.Lys220del) mutations, which were inherited respectively from each of his parents. Prenatal diagnosis showed that the fetus has inherited two wild-type parental alleles. CONCLUSION: The targeted Ion Torrent PGM sequencing has detected pathogenic mutations in the Chinese pedigree affected with methylmalonic academia, which has provided molecular evidence for clinical diagnosis, genetic counseling and prenatal diagnosis for the family.

Methylmalonic acidemia (MMA) in pregnancy: a case series and literature review.

INTRODUCTION: Women with inherited metabolic disorders, including those with previously life-limiting conditions such as MMA, are reaching child-bearing age more often due to advances in early diagnosis and improved pediatric care. Information surrounding maternal and fetal complications associated with the underlying disorders remains largely unexplored. METHODS: Pregnancies affected by maternal MMA were ascertained through study 04-HG-0127 "Clinical and Basic Investigations of Methylmalonic Acidemia and Related Disorders" (clinicaltrials.gov identifier: NCT00078078) and via literature review. Prenatal and delivery records in study participants were reviewed. RESULTS: Seventeen pregnancies were identified in women with isolated MMA, including three abortions, one termination, and 13 completed pregnancies [three cases with cblA (four pregnancies), four cases of mut- (one cobalamin responsive, three non-responsive), five cases with unknown type of MMA]. Seventeen percent (3/17) of the pregnancies resulted in a first trimester abortion, while 38.5% (5/13) of the completed pregnancies resulted in preterm deliveries. A cesarean delivery rate of 53.8% (7/13) was noted among the cohort. Fetal distress or nonreassuring fetal status was the indication for 57% (4/7) cesarean deliveries. One patient was reported to have metabolic crisis as well as episodes of mild hyperammonemia. Malformations or adverse outcomes in the progeny were not observed. CONCLUSION: Although there have been a small number of pregnancies identified in women with MMA, the cumulative results suggest that the majority of pregnancies can be complicated by cesarean delivery and increased risk of prematurity. A pregnancy registry could clarify perinatal complications and define management approaches needed to ensure optimal maternal and fetal outcomes in this growing patient population.

Feasibility of prenatal diagnosis of lysinuric protein intolerance by linkage analysis: a case report.

Lysinuric protein intolerance (LPI) is a rare autosomal recessive defect of cationic amino acid transport (CAA), relatively common in Finland and Italy. After weaning, LPI patients present poor feeding, vomiting and failure to thrive. A severe pulmonary complication and episodes of metabolic imbalance may lead to death. Prenatal diagnosis has not been available due to lack of either biochemical or molecular markers to be used in the fetal period. The LPI locus has recently been assigned to chromosome 14q12, very close to the T-cell receptor alpha-chain (TCRA) locus. We carried out a prenatal diagnosis for LPI by linkage analysis in one LPI Italian family after CVS. For the haplotype analysis 11 DNA markers from the LPI critical region were used (D14S742, D14S50, D14S283, five TCRA intragenic polymorphic sites, D14S990, MYH7 and D14S80). It was concluded that the haplotype analysis indicated that the fetus was healthy as he had inherited the two wild alleles of the LPI locus. After birth, the clearances of CAA were measured and found to be in the normal range, thus confirming the result of the prenatal diagnosis. The prenatal diagnosis of LPI can now be offered to families affected by LPI.

Maternal gonadal mosaicism causing ornithine transcarbamylase deficiency.

Ornithine transcarbamylase (OTC) deficiency (McKusick 311250), an X-linked inherited disorder, often presents in males with severe neonatal onset of hyperammonemia. Maternal gonadal mosaicism in OTC deficiency was postulated previously, but no cases have been reported. We report on a family in which two consecutive males were affected with OTC deficiency, which was proven biochemically with characteristic metabolites and absent enzyme activity in liver. OTC genotyping in both brothers showed a new mutation in exon 6 (Met206Arg: ATG-->AGG), which encodes part of the equatorial H6 alpha-helix. Biochemical investigations confirmed normal results in the mother and grandmother and the absence of OTC activity in the affected males. Genotyping of the mother and grandmother was performed on peripheral blood leukocytes and skin fibroblasts and showed no mutation in the somatic cells. The recurrence of OTC deficiency in offsprings of a woman with normal genotype strongly suggests gonadal mosaicism. Gonadal mosaicism needs to be considered when counseling couples in which the mother has had a previously affected child with OTC deficiency but apparently is not a carrier.

Prenatal molecular evaluation of six fetuses in four unrelated Korean families with ornithine transcarbamylase deficiency.

Ornithine transcarbamylase (OTC) deficiency, an X-linked inborn error of the urea cycle, leads to the accumulation of ammonia, causing neurologic deficits. Clinical management for the patients with OTC deficiency is frustrating and requires a burdensome medical regimen, since they may have impairment and recurrent episodes of hyperammonemia in spite of intensive care. Therefore, prenatal diagnosis of the affected fetus is important in genetic counselling for the family at high risk. In this study, mutations in the OTC gene of three obligate heterozygotes and a proband have been identified in four unrelated families: R141Q, R320X, H214Y, M205T. Each mutation altered restriction recognition sites; TaqI for R141Q, NlaIII for M205T, RsaI for H214Y, BclI for R320X. Based on their molecular defects, prenatal diagnoses of 6 fetuses including one set of fraternal twins were successfully made at the ninth to eleventh week of gestation by polymerase chain reaction (PCR)-restriction digestion using genomic DNA from chorionic villus sampling (CVS). We predicted the outcome of all fetuses prenatally. Among six, four were females and two were males, which were determined by PCR amplification of the sex determining region of the Y chromosome (SRY) gene. Each carried a wild type allele for the corresponding mutant allele. They were also tested postnatally for the mutations to be unaffected.

Ornithine carbamoyl transferase deficiency: a neuropathological study.

A detailed autopsy study of three children with ornithine carbamoyl transferase (OCT) deficiency is presented. Although variable in extent, a basic pattern of neuropathological lesions is discernible. Case 1 shows gross cerebral atrophy, cases 2 and 3 milder lesions in the basal nuclei but also multiple cerebellar heterotopias and delayed myelination. We suggest that the findings may provide evidence that OCT deficiency can have a teratogenic effect in utero and suggest that there is a need to monitor the pregnancies of carriers of this disorder.