Molecular genetics and impact of residual in vitro phenylalanine hydroxylase activity on tetrahydrobiopterin responsiveness in Turkish PKU population

Dobrowolski, Steven F.; Heintz, Caroline; Miller, Trent; Ellingson, Clinton; Ellingson, Clifford C.; Özer, Işıl; Gökçay, Gülden; Baykal, T.; Thöny, Beat; Demirkol, Mübeccel; Blau, Nenad

doi:10.1016/j.ymgme.2010.11.158

Cited by 73 publications

(72 citation statements)

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“…Conversely, the response rate among patients with classic PKU (little or no residual PAH activity) is very low. A number of PAH mutations associated with BH4 responsiveness have been identified and genotyping is a useful additional tool for predicting responsiveness [21,22] (see below). Fig.…”

Section: Bh4 Loading Testmentioning

confidence: 99%

Diagnosis, classification, and genetics of phenylketonuria and tetrahydrobiopterin (BH4) deficiencies

Blau

Hennermann

Langenbeck

et al. 2011

Molecular Genetics and Metabolism

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This article summarizes the present knowledge, recent developments, and common pitfalls in the diagnosis, classification, and genetics of hyperphenylalaninemia, including tetrahydrobiopterin (BH4) deficiency. It is a product of the recent workshop organized by the European Phenylketonuria Group in March 2011 in Lisbon, Portugal. Results of the workshop demonstrate that following newborn screening for phenylketonuria (PKU), using tandem mass-spectrometry, every newborn with even slightly elevated blood phenylalanine (Phe) levels needs to be screened for BH4 deficiency. Dried blood spots are the best sample for the simultaneous measurement of amino acids (phenylalanine and tyrosine), pterins (neopterin and biopterin), and dihydropteridine reductase activity from a single specimen. Following diagnosis, the patient's phenotype and individually tailored treatment should be established as soon as possible. Not only blood Phe levels, but also daily tolerance for dietary Phe and potential responsiveness to BH4 are part of the investigations. Efficiency testing with synthetic BH4 (sapropterin dihydrochloride) over several weeks should follow the initial 24-48-hour screening test with 20mg/kg/day BH4. The specific genotype, i.e. the combination of both PAH alleles of the patient, helps or facilitates to determine both the biochemical phenotype (severity of PKU) and the responsiveness to BH4. The rate of Phe metabolic disposal after Phe challenge may be an additional useful tool in the interpretation of phenotype-genotype correlation. This article summarizes the present knowledge, recent developments, and common pitfalls in the diagnosis, classification, and genetics of hyperphenylalaninemia, including tetrahydrobiopterin (BH4) deficiency. It is a product of the recent workshop organized by the European Phenylketonuria Group in March 2011 in Lisbon, Portugal. Results of the workshop demonstrate that following newborn screening for phenylketonuria (PKU), using tandem mass-spectrometry, every newborn with even slightly elevated blood phenylalanine (Phe) levels needs to be screened for BH4 deficiency. Dried blood spots are the best sample for the simultaneous measurement of amino acids (phenylalanine and tyrosine), pterins (neopterin and biopterin), and dihydropteridine reductase activity from a single specimen. Following diagnosis, the patient's phenotype and individually tailored treatment should be established as soon as possible. Not only blood Phe levels, but also daily tolerance for dietary Phe and potential responsiveness to BH4 are part of the investigations. Efficiency testing with synthetic BH4 (sapropterin dihydrochloride) over several weeks should follow the initial 24-48-hour screening test with 20 mg/kg/day BH4. The specific genotype, i.e. the combination of both PAH alleles of the patient, helps or facilitates to determine both the biochemical phenotype (severity of PKU) and the responsiveness to BH4. The rate of Phe metabolic disposal after Phe challenge may be an additional useful tool in the interp...

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Section: Bh4 Loading Testmentioning

confidence: 99%

Diagnosis, classification, and genetics of phenylketonuria and tetrahydrobiopterin (BH4) deficiencies

Blau

Hennermann

Langenbeck

et al. 2011

Molecular Genetics and Metabolism

Self Cite

194

168

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“…Residual in vitro PAH activity was shown to correlate with the patient's phenotype [32]. Thus, knowing patient's residual PAH activity can be relevant for selecting therapeutic options, the likely Phe tolerance, and the expected response to BH 4 [28].…”

Section: Discussionmentioning

confidence: 99%

“…Thus, the main application of this new method will be the quantification of the relative in vitro PAH activity in patients with PKU by expressing mutant proteins in COS-1 cells. A recent study with a large cohort of Turkish PKU patients suggested that both the phenotype and BH 4 -responsiveness are genotype-dependent and that the residual PAH activity is a good predictor for the severity of the disease [28]. Finally, this method can easily be extended to other amino acids since labeled compounds are available as internal standards for quantification.…”

Section: Discussionmentioning

confidence: 99%

“…These mutations have previously been tested in different in vitro systems, but we also analyzed two novel PAH mutations p.E280A and p.Y417C, recently reported in Turkish PKU patients [28] (Figure 3a). While a severe p.E280A mutation exhibits only 15% of the wild-type PAH activity, a mild p.Y417C mutation was almost as active as the wild-type enzyme (76%).…”

Section: Pah Mutations Found In Pku Patientsmentioning

confidence: 99%

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Quantification of phenylalanine hydroxylase activity by isotope-dilution liquid chromatography–electrospray ionization tandem mass spectrometry

Heintz

Troxler

Martı́nez

et al. 2012

Molecular Genetics and Metabolism

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BACKGROUND: Residual phenylalanine hydroxylase (PAH) activity is the key determinant for the phenotype severity in phenylketonuria (PKU) patients and correlates with the patient's genotype. Activity of in vitro expressed mutant PAH may predict the patient's phenotype and response to tetrahydrobiopterin (BH(4)), the cofactor of PAH. METHODS: A robust LC-ESI-MSMS PAH assay for the quantification of phenylalanine and tyrosine was developed. We measured PAH activity a) of the PAH mutations p.Y417C, p.I65T, p.R261Q, p.E280A, p.R158Q, p.R408W, and p.E390G expressed in eukaryotic COS-1 cells; b) in different cell lines (e.g. Huh-7, Hep3B); and c) in liver, brain, and kidney tissue from wild-type and PKU mice. RESULTS: The PAH assay was linear for phenylalanine and tyrosine (r(2) 0.99), with a detection limit of 105 nmol/L for Phe and 398 nmol/L for Tyr. Intra-assay and inter-assay coefficients of variation were <5.3% and <6.2%, respectively, for the p.R158Q variant in lower tyrosine range. Recovery of tyrosine was 100%. Compared to the wild-type enzyme, the highest PAH activity at standard conditions (1 mmol/L L-Phe; 200 mol/L BH(4)) was found for the mutant p.Y417C (76%), followed by p.E390G (54%), p.R261Q (43%), p.I65T (33%), p.E280A (15%), p.R158Q (5%), and p.R408W (2%). A relative high PAH activity was found in kidney (33% of the liver activity), but none in brain. CONCLUSIONS: This novel method is highly sensitive, specific, reproducible, and efficient, allowing the quantification of PAH activity in different cells or tissue extracts using minimum amounts of samples under standardized conditions

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“…The incidence of PKU varies in different populations, with a prevalence of 1 in 10,000 births in Europe and the United States (Dobrowolski et al 2011;Blau et al 2010;Hardelid et al 2007). Its prevalence is higher in northern Europeans and it has an incidence of 1 in 4,500 in the Irish (Blau et al 2010;Magee et al 2002).…”

Section: Introductionmentioning

confidence: 99%

Successful Live Birth following Preimplantation Genetic Diagnosis for Phenylketonuria in Day 3 Embryos by Specific Mutation Analysis and Elective Single Embryo Transfer

et al. 2012

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Phenylketonuria (PKU) is an autosomal recessive inherited metabolic disorder caused by a complete or near-complete deficiency of the liver enzyme phenylalanine hydroxylase (PAH), which converts the amino acid phenylalanine to tyrosine, leading to the increase of blood and tissue concentration of phenylalanine to toxic levels. PKU is not life threatening but is treated through lifelong dietary management. If untreated, it can lead to severe learning disability, brain function abnormalities, behavioural and neurological problems. The non-life threatening nature of PKU has until now caused some debate on whether to licence its detection by preimplantation genetic diagnosis (PGD). We report the first successful live birth in the UK following single cell embryo biopsy and PGD for the detection of two different mutations in the (PAH) gene. This case highlights both an important scientific development as well as the ethical challenge in offering couples who carry PKU this new reproductive option when starting their family.

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Molecular genetics and impact of residual in vitro phenylalanine hydroxylase activity on tetrahydrobiopterin responsiveness in Turkish PKU population

Cited by 73 publications

References 33 publications

Diagnosis, classification, and genetics of phenylketonuria and tetrahydrobiopterin (BH4) deficiencies

Diagnosis, classification, and genetics of phenylketonuria and tetrahydrobiopterin (BH4) deficiencies

Quantification of phenylalanine hydroxylase activity by isotope-dilution liquid chromatography–electrospray ionization tandem mass spectrometry

Successful Live Birth following Preimplantation Genetic Diagnosis for Phenylketonuria in Day 3 Embryos by Specific Mutation Analysis and Elective Single Embryo Transfer

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