Different presentations of late-detected phenylketonuria in two brothers with the same R408W/R111X genotype in the PAH gene.

Although the clinical heterogeneity of phenylketonuria (PKU) is well established, some questions about this condition remain. Subjects from the same family who share the same mutations in the phenylalanine hydroxylase (PAH) gene are expected to display similar disease courses, and therefore, when blood phenylalanine (Phe) levels, genotype and dietary treatment are all similar, differences in patient outcomes require additional explanations. The present authors describe two entirely different courses of late-detected PKU in two brothers with the same R408W/R111X genotype in the PAH gene. The older sibling was diagnosed with PKU at the age of 4 years and given treatment. His IQ was 97 at 26 years of age and moderate involvement of periventricular white matter was detected. The younger brother was diagnosed with PKU at the age of 11 months and given treatment. His IQ was < 25 at 22 years of age and severe dysmyelination changes were found by magnetic resonance imaging. The differences in the courses of the disease between these two brothers appear to be related to variations in their blood-brain barriers.

Should newborn mutation scanning for hyperphenylalaninaemia and galactosaemia be implemented? A Polish experience.

OBJECTIVE: To elucidate whether screening for mutations causing hyperphenylalaninaemia (HPA) and classic galactosaemia could provide important, additional information on a clinical phenotype. METHOD: Genotypes that cause disease at the phenylalanine hydroxylase (PAH) gene and galactose-1-phosphate uridyltransferase (GALT) gene in a group of 101 hyperphenylalaninaemic and 77 patients with classic galactosaemia were established. The PAH and GALT mutations were identified in genomic DNA extracted from whole blood leucocytes using single stranded conformational analysis and direct fluorescent sequencing of polymerase chain reaction (PCR) products. RESULTS: Mild HPA and mild phenylketonurea (PKU) were caused by divergent genotypes. In the studied group a total of 26 different mild and intermediate PAH mutations were identified, most of them being rare ones. Classic galactosaemia was caused by two frequent mutations, accounting for 82% of all mutated alleles. CONCLUSIONS: Identification of mild or intermediate mutations causing HPA could provide fast and reliable information about future clinical outcome of a newborn infant. Molecular diagnosis of HPA should be preceded by biochemical analysis and implemented to differentiate mild forms of HPA and cases of ambiguous classification. Because of multiple rare mutations scattered on all exons, scanning of the entire PAH coding sequence could be useful and cost beneficial. Routine genotyping is not proposed in classic phenylketonuria and classic galactosaemia, as it provides limited additional, prospective information on the clinical phenotype.

[Standards for diagnosis and treatment of phenylketonuria].

The aim of this study is to unify the diagnostic and therapeutic standards in phenylketonuria. The course of the disease, diagnostic methods as well as treatment procedures is presented. Standards of treatment are based on the experiences and research carried out by the Screening Tests Unit of the Department of Public Health and the Clinical Department of Paediatrics of the National Research Institute of Mother and Child in Warsaw.

[Non-ketonic hyperglycinemia]. / Niekwasicza hiperglicynemia (NKH)

The author presents the possible pathomechanism of brain damage in NKH and the clinical course of severe - newborn and infants - atypical type of the disease. Characteristic for the newborn type EEG patterns has been discussed. Among the detailed presentation of diagnostic methods it was stressed that besides high concentration of glycine in blood and urine, the necessary parameters are high glycine ratio of cerebro-spinal fluid to blood serum, and the detailed differentiation with many transient and persistent hyperglycinemic states, the most important of them being organic acidurias which need the estimation of urinary organic acids. Possibilities of prenatal diagnostic and molecular identification are presented. All the treatment methods used up to now have been presented with special attention to natrium benzoate and dextrametorphan being used together.

[Serum tyrosine in children with phenylketonuria and mild hyperphenylalaninemia]. / Tyrozyna w surowicy krwi u dzieci z fenyloketonuria i lagodna hiperfenyloalaninemia.

Serum tyrosine concentration, Phe/Tyr scores and psychomotor/mental development scores were analysed in 32 children with phenylketonuria (PKU) and 39 with mild hyperphenylalaninaemia. Observation period included the first 6 years of life. Tendency to tyrosine deficiency was observed; stronger in dietary treated PKU patients than in those with mild hyperphenylalaninaemia. Statistically significant differences between patient groups were found only in 3 and 6 years old children (lower tyrosine values in PKU patients). It was observed that evaluation of Phe/Tyr score value might be usefull in differentiation between PKU and mild hyperphenylalaninaemia. Moreover, the above score may help in the evaluation of hypo- and hyperalimentation state in the course of dietary treatment. The level of tyrosine deficiency in the analysed patient groups did not influence their normal intellectual development.

Mutations in exon 3 of the PAH gene causing mild hyperphenylalaninemia.

Phenylketonuria (PKU), an autosomal recessive disorder caused by a deficiency of hepatic phenylalanine hydroxylase (PAH), is clinically very heterogeneous. On molecular level more than 350 mutations in the PAH gene are known to date, which in different genotype combinations could account for biochemical and clinical variability. Mutations located in exon 3 coding for a part of the regulatory domain of the PAH enzyme cause classical PKU, mild PKU, and mild hyperphenylalaninemia (MHP). We describe the phenotypic effects of seven mutations in exon 3 of the PAH gene (R68G, R68S, R71H, S87R, P89S, I95F, and A104D). We propose that mutations located between amino acid positions 71 through 94 cause MHP.

[Mutations causing hereditary hyperphenylalaninemia]. / Mutacje powodujace dziedziczna hiperfenyloalaninemie.

Mutations in the genes encoding different parts of phenylalanine hydroxylation system cause persistent hyperphenylalaninaemia. The most frequent form of hyperphenylalaninaemia is caused by mutations in the PAH gene. The most common variant result from defect of tetrahydrobiopterin synthase. Mutations in the PAH and PTS genes in the Polish population are presented. Genotype--phenotype correlations are discussed.

[Evaluation of the usefulness for neonatal mass screening in light of 35 years personal experience]. / Ocena przydatnosci badan przesiewowych u noworodków w swietle 35 lat doswiadczen wlasnych.

The results and the significance of neonatal mass-screening programmes for inborn errors of metabolism, conducted by the National Research Institute of Mother and Child (NRIMC), are discussed. As the first in Poland, in 1964, mass-screening for phenylketonuria (PKU) was introduced. The BIA-Guthrie test was used. Other Guthrie tests (GBIA) were applied in homocystinuria, tyrosinemia, histidinemia and leucinosis (Maple Syrup Urine Disease-MSUD). In the middle of the 60. the Beutler and Baluda test was introduced for galactosaemia, as well as the Efron urine test in infant screening for different inborn errors of metabolism. In the middle of the 70., neonatal mass-screening for cystic fibrosis (CF, mucoviscidosis) was started. Meconium tests and the sweat test with ion selective chloride electrode were used. Apart from inborn errors of metabolism, we also introduced a screening programme for neuroblastoma in which vaniline mandelic acid (VMA) in urine was estimated and for congenital hypothyroidism were TSH level was assessed. The results of screening are shown in the tables and in the figures. In our opinion the best clinical results are obtained with screening for congenital hypothyroidism and for PKU, since very early detection and treatment in these diseases prevents severe mental retardation. We therefore consider that both these screening programmes should be treated as obligatory examinations in all neonates. Taking into consideration the fact that there are different types of hyperhenylalaninemias, the principles of differential diagnosis are discussed. Molecular genetic investigations, carried out in the NRIMC Department of Genetics proved to be a very important procedure in the verification of diagnosis of different mutations. The authors also discuss the problem of dietary treatment duration in PKU. In our opinion the hypophenyloalanine diet regimen in girls, should not be discontinued during adolescence, since there is the problem of maternal PKU and the possibility of foetal damage. The results of our own investigations of maternal PKU are discussed. The significance of mass-screening for galactosemia is still under discussion. In our opinion, mass-screening for galactosemia is not useful and we have discontinued it. Selective screening has been started combined with molecular genetic studies in high risk families. In the future, we plan to prepare guidelines on the principles of diagnosis and treatment of galactosemia in children and women in the reproductive age. Mass-screening for cystic fibrosis is also still under discussion. The results of the early screening programmes were not satisfactory and the tests were discontinued. In 1998, after reorganisation of the whole system, CF screening, using tripsin-radioimmune assays, was started again. The new screening programme is combined with molecular genetic investigation of different mutations. It is still too early to assess the importance and success of this CF mass-screening programme. We decided to discontinue the screening for homocystinuria, histidinemia, tyrosinemia, leucinosis and for neuroblastoma, since these programmes did not comply with criteria of mass-screening. In 1997, major reorganisation of screening programmes for inborn errors of metabolism, at NRIMC, was undertaken. The Guthrie test for PKU was changed to a quantitative colorimetric method. The immuno-luminometric method is used for TSH estimation. The whole system is based on complete computer control of all the steps of screening, from blood sampling on filter paper until the final diagnosis. The advantages of this modern system of organisation of the screening programme are discussed.

Identification of Mutations Causing 6-Pyruvoyl- Tetrahydrobiopterin Synthase Deficiency in Polish Patients With Variant Hyperphenylalaninemia.

Background: 6-Pyruvoyl-tetrahydrobiopterin synthase (PTPS) is required for biosynthesis of tetrahydrobiopterin, the cofactor of various enzymes including the hepatic phenylalanine hydroxylase. Mutations in the PTS gene result in a variant type of hyperphenylalaninemia, requiring cofactor replacement therapy for treatment. Methods and Results: Four Polish patients with PTPS deficiency were screened for mutations in the PTS gene. Three novel mutations E35G, N36K, and F100V were identified. In one patient, a known mutation D136V was identified in both PTS alleles. Conclusions: Mutation D136V present in both alleles was proposed to be connected with a mild form of PTPS deficiency. The other three mutations were found in heterozygous patients with a central type of PTPS deficiency. D136V mutation is a common mutation in the Polish population.

[Maternal PKU syndrome as an obstetric problem: literature review and own clinical experience]. / Zespól fenyloketonurii matczynej jako problem polozniczy--przedstawienie wspólczesnych pogladów i wlasnych doswiadczen klinicznych.

Maternal phenylketonuria (M-PKU) is a syndrome of embryo- and fetopathy observed in the offsprings of mothers with increased blood level of phenylalanine. These women fall into two groups: phenylketonuria (PKU) and mild hyperphenylalaninemia (MHP). The Phe--level safe for the fetus is 4-6 mg%. Typical for M-PKU syndrome is: microcephalia, mental retardation, intrauterine growth retardation, congenital heart diseases and other anomalies like esophageal atresia, meningocoele, Pierre-Robin syndrome, cataract. The only way to prevent this syndrome is Phe--restricted diet that should be initiated before conception. We reviewed updated literature on the pathogenesis of this syndrome, clinic, possibility of prophylaxis and treatment. We present also 5 pregnancies of 3 patients with PKU, treated in National Research Institute of Mother and Child in Warsaw. On that ground we propose the scheme of prevention of maternal PKU syndrome.

Molecular basis of mild hyperphenylalaninaemia in Poland.

The major cause of the different forms of hyperphenylalaninaemia (HPA) is mutations in the gene encoding phenylalanine hydroxylase (PAH). The aim of this study was to determine the mutations responsible for mild forms of HPA and to relate different clinical phenotypes of HPA patients to their PAH genotypes. Four "mild" mutations, including the most frequent A403V and R297H mutations, occurred exclusively in mild hyperphenylalaninaemia (MHP). Mutations A104D, R243Q, R241H, and Y414C were detected in patients with mild phenylketonuria (mild PKU) only. These results may be useful in establishing a molecular differential diagnosis for PAH deficiency in Poland.

Frequencies of the most common mutations responsible for phenylketonuria in Poland.

We screened 91 Polish phenylketonuric (PKU) children for the presence of 18 common mutations in the phenylalanine hydroxylase (PAH) gene, and 75.7% of PAH alleles were identified. The R408W mutation accounted for 54.9% of PAH mutant alleles. In the other 20.8%, eight mutations were detected: R158Q (6.6%), IVS10 (4.9%), IVS12 (2.7%), R261Q (2.2%), G272ter (1.65%), Y414C (1.1%), R252W (1.1%) and P281L (0.54%). Correlations between genotype and clinical phenotype were described.

[Evaluation of amino acids in plasma and amniotic fluid of women from genetic risk groups]. / Ocena aminogramu osocza i plynu owodniowego u kobiet z grupy ryzyka genetycznego.

Plasma and amniotic fluid amino acids were assayed in 20 women of genetic risk groups in the second trimester of pregnancy. The age of patients ranged from 19 to 38 years. Indication for amnio-puncture were: chromosomal aberration or neural tube defect in previous pregnancy or age of pregnant women over 35 years. Blood and amniotic fluid were obtained with transabdominal amnio-puncture performed routinely in prenatal diagnosis. Amino acids were assayed with ion exchange column chromatography, using automatic amino acids analyzer LKB 4400. The obtained results were compared with plasma and amniotic fluid amino acids patterns in healthy women at the same trimester of pregnancy. The comparison of plasma aminograms revealed increased concentrations (above 100%) of cystine and alanine in the examined group. In women with neural tube defect in previous pregnancy and in those over 35 years of age, increased proline, leucine and valine (above 50%) concentrations were found. Decreased concentrations of glutamic acid (about 60%) was detected in women with chromosomal aberration in previous pregnancy and in those older than 35 years. Amniotic fluid amino acid pattern showed in all three patients groups decreased values arginine and ornitine (amino acids of the urea cycle), and also of cystine and taurine. In women with neural tube defect in previous pregnancy and in those older than 35, decreased concentration of lysine, serine and leucine was found. Also in the above 2 groups increasing tendency in comparison to the values in healthy women was observed for alanine (30-40%) and asparagine (150-215%). No correlation was observed between particular amino acids plasma concentration and amniotic fluid in our patients. Ratios of plasma to the amniotic fluid (P/FA) amino acid concentrations were calculated for the examined group and compared with those in healthy pregnant women. While in the last population most of the analyzed scores is below 1 which points higher amino acid concentration in amniotic fluid than that in plasma, performed analysis revealed; an increase in P/AF ratio by 214% in the group of "neural tube defect" women, alanine, glutamine and ornitine P/AF ratios were 35-50% than the normal value. P/AF ratio for cystine by 176%, for tyrosine--100% and for ornitine and arginine--55-65% respectively higher than normal in patients of "chromosomal aberration". P/AF ratio for glutamine acid was decreased by about 50%. In patients examined because of age above 35 years, P/AF ratio for cystine was increased by 200%, for alanine by 125%, for proline, histidine, ornitine, phenylalanine, I-leucine and metionine were increased by 30%-50%.(ABSTRACT TRUNCATED AT 400 WORDS)

Red blood cell glutathione peroxidase activity as a function of selenium supplementation in dietary treated children with phenylketonuria.

Red blood cell glutathione peroxidase (GSH-Px) activity of six patients with phenylketonuria treated with aminoacid mixture and protein hydrolysate diets was significantly lower (11.2 IU/g Hb) than that of 21 age-matched healthy children (14.9 IU/g Hb). When the diets were supplemented with yeast rich in selenium the red blood cell GSH-Px activity increased already after one month of treatment to 16.1 IU/g Hb (P less than 0.001) and remained at that level during subsequent two months of selenium supplementation. A high significantly positive correlation has been found between calculated red blood cell selenium level and GSH-Px activity within three months of selenium supplementation.