Genetic profiles of phenylalanine hydroxylase gene mutations in neonates with phenylketonuria in Xinjiang / 中国新生儿科杂志

Objective:To study the genetic profiles of phenylalanine hydroxylase (PAH) gene mutations in neonates with phenylketonuria (PKU) in Xinjiang.Methods:From January 2015 to December 2021,neonates born and genetically diagnosed with PKU in our region were retrospectively included. The genetic profiles of different ethnic groups were analyzed and compared with PKU patients from central, northwest and northern regions of China.Results:A total of 131 neonates with PKU were enrolled, including 82 Han, 25 Hui and 20 Uyghur patients, 4 cases of other ethnic groups. 46, 20 and 14 types of pathogenic variants were detected in each ethnic group with detection rates of 95.1% (156/164), 66.0% (33/50), and 60.0% (24/40), respectively. The variants were mainly missense mutations and located in exons 2, 3, 6,7 and 11. The most common loci in Hui patients were c.158G>A (18.2%), c.728G>A (18.2%) and c.898G>T (9.1%). The most common loci in Uyghur patients were c.158G>A (33.3%), c.355C>T (12.5%) and c.1068C>A (8.3%). c. 898G>T might be most unique in Hui patients and c.355C>T most unique in Uyghur patients in Xinjiang. A novel variant of PAH gene, c.828G>C (p.M276I) in exon 7 was identified. Compared with northern, central and northwestern regions of China, PKU patients in Xinjiang had significantly higher incidence of c.158G>A mutation and lower incidence of c.728G>A mutation ( P<0.05). Conclusions:Missense mutations of PAH gene are common in some regions of Xinjiang. The compositions of PAH gene variations are similar to northwest and northern China with significant differences in hotspots of mutations.

Adecuación del aporte de proteína natural en el tratamiento de la fenilcetonuria: una revisión bibliográfica / Adequacy of natural protein intake in patients with phenylketonuria: bibliographic review

Introducción: la fenilcetonuria (PKU) es el error congénito del metabolismo de las proteínas más frecuente. El tratamiento dietético consiste en un plan de alimentación con una ingesta de proteínas naturales restringida, un sustituto proteico libre o de bajo contenido en fenilalanina (Phe) y el aporte de alimentos muy bajos en proteínas. El objetivo principal de este trabajo fue investigar si es posible aumentar la ingesta de proteína natural (PN) que se indica a los pacientes con PKU manteniendo los dosajes de Phe en sangre en rangos de seguridad. Materiales y método: se buscaron en 6 bases de datos electrónicas artículos publicados. Se identificaron un total de 154 artículos de Pub Med por intervalo de años desde 1999 a 2020. Se eligieron 15 artículos que se adaptaron a los criterios de inclusión y exclusión y respondían al objeto de estudio de esta revisión bibliográfica. Resultados: hay varios factores que pueden influenciar la estimación de la tolerancia de Phe como la severidad del fenotipo del paciente, la edad, el rango de seguridad de Phe en sangre, la prescripción de Phe y la adherencia al sustituto proteico. Si los niveles de Phe en sangre se mantienen en forma constante dentro del rango adecuado y por un período determinado, se debería considerar un incremento de la ingesta de Phe. El aumento de la ingesta de PN deberá ser realizado de manera controlada, individual y evaluando en forma constante el impacto en los dosajes de Phe en sangre. Conclusión: optimizar la ingesta de PN ofrece una mejora en la calidad de vida de pacientes con PKU, facilita la capacidad del paciente para socializar y contribuye a una mejor adherencia a la dieta(AU).

Introduction: phenylketonuria (PKU) is the most frequent inborn error of protein metabolism. The dietary treatment consists of a diet with a restricted natural protein intake, a free or low phenylalanine (Phe) protein substitute, and the intake of low protein food. The main objective of this work is to analyze if it is possible to increase the natural protein (NP) intake prescribed to PKU patients while maintaining blood Phe dosages within safe range. Materials and method: studies published were searched in 6 electronic data- basis. A total of 154 Pub Med articles were identified by range of years from 1999 to 2020. Fifteen articles which met the inclusion and exclusion criteria and responded to the objective of this bibliographic review were chosen. Results: several factors may influence Phe tolerance, such as severity of the patient´s phenotype, age, blood Phe safe range, Phe prescription and adherence to protein substitute. If Phe blood levels remain constantly within safe range and for a certain period, an increase of Phe intake should be considered. Increase of NP intake must be carried out in a controlled manner, individually and constantly evaluating blood Phe levels. Conclusion: optimizing NP intake offers the PKU patient an improvement in quality of life, facilitates the patient´s ability to socialize and contributes to a better adherence to the diet(AU).

Physical, intellectual and genetic characteristics of 53 neonates with phenylketonuria / 预防医学

Objective@#To investigate the physical and intellectual development and mutation characteristics of the phenylalanine hydroxylase (PAH) gene among 53 newborns with phenylketonuria (PKU), so as to provide insights into the management and genetic counseling of PKU@*Methods@#The medical records of 54 children with definitive diagnosis of PKU and standardized therapy until 2 years at the Center for Neonatal Disease Screening of Shanxi Children' s Hospital from 2018 to 2021 were collected. Newborns' body weight and height developments were evaluated using the World Health Organization growth chart (2006 version), and the intellectual development was assessed using the national criteria of Development Behavior Assessment Scale among Children at Ages of 0 to 6 Years (WS/T 580-2017). The gene mutations were detected among neonates and their children, and the physical, intellectual developments and genetic characteristics of neonates with PKU were descriptively analyzed.@*Results@#The 53 PKU cases included 29 male children and 24 female children, 36 cases with classic PKU and 17 cases with mild PKU, and 30 cases from rural areas and 23 cases from urban areas. The study subjects had a median age of 30 (10) d at initial therapy, and a mean blood phenylalanine concentration of (1 507±685) μmol/L at definitive diagnosis. There were 52 cases with normal height developments (98.11%), and all cases had normal weight and intellectual developments. The mean developmental functional quotient (DFQ) was significantly greater among urban children with PKU than among rural children [(94.92±8.57) vs. (87.65±6.57); t=-3.498, P=0.001], and the mean DFQ was significantly higher among children with mild PKU than among those with classic PKU [(95.55±8.76) vs. (88.57±7.11); t=-3.095, P=0.003]. There were 37 mutations detected in the PAH gene, which were mainly distributed in exons 3, 6, 7, 11, 12 and intron 4. Three high-frequency mutation sites were detected, including c.728G>A, c.611A>G and c.1197A>T, including three novel mutations (c.674C>G, c.1316-2A>C and c.1069T>C).@*Conclusions@#Following standardized treatment, the children with PKU have comparable physical and intellectual developments as compared to normal children. c.728G>A, c.611A>G and c.1197A>T were predominant mutations in the PAH gene among these 53 children with PKU, and three novel mutations were identified, including c.674C>G, c.1316-2A>C and c.1069T>C.

Preliminary screening of therapeutic drugs for phenylalanine hydroxylase deficiency / 中国基层医药

Objective:To screen new drugs for treatment of phenylalanine hydroxylase deficiency.Methods:From October 2019 to October 2020, virtual drug screening was performed in Center of Genetic Medicine, Nanjing Maternity and Child Health Care Hospital, Women's Hospital of Nanjing Medical University computer according to the characteristics of the binding ability of phenylalanine hydroxylase to drug spatial structure. Ten candidate drugs were screened from the FDA drug library (including 2 697 kinds of active pharmaceutical ingredients). A eukaryotic expression system was used to determine the effects of drugs on the activity of phenylalanine hydroxylase at the molecular level. Drug-sensitive mutants were screened.Results:Among the 10 candidate drugs, neoplasm hydrochloride, fluocinonide acetate and risperidone increased 23% [ t = 18.21, P < 0.001, vs. non-drug-treated phenylalanine hydroxylase group (i.e., only solvent and no drug added to the reaction system)], 21% ( t = 3.44, P < 0.05, vs. non-drug-treated phenylalanine hydroxylase group), 31% ( t = 19.57, P < 0.001, vs. non-drug-treated phenylalanine hydroxylase group) of the activity of phenylalanine hydroxylase. The remaining drugs exhibited weak even inhibitory effects on the activity of phenylalanine hydroxylase. 25% of p.D101N mutant could be activated by risperidone ( t = 15.86, P < 0.001, vs. non-drug-treated p.D101N mutant group). Conclusion:Neoplasm hydrochloride, fluocinonide acetate and risperidone can be used as potential therapeutic drugs for phenylalanine hydroxylase deficiency, and p.D101N mutant can be used as the drug-sensitive mutation site.

Phenylalanine Hydroxylase (PAH) Genotyping in PKU Argentine Patients

Abstract Phenylketonuria (PKU, OMIM 261600) is predominantly caused by mutations in the PAH gene. One hundred and three Argentine PKU patients were studied by Sanger sequencing; 101 were completely characterized (90.3% were compound heterozygotes). Fifty-four different pathogenic variants were identified. Mutations were distributed all along the PAH gene but concentrated in exon 7 (26%), 12 (12%), 11 (10%), and 6 (10%). 77% were missense, and 77% affected the enzyme catalytic domain, nine mutations accounted for 57% of 179 studied alleles: p.Arg261Gln (Allele frequency(AF):10.6%), c.1066-11G>A (AF:9,5%), p.Arg408Trp (AF:8,3%), p.Tyr414Cys (AF:5,5%), p.Ala403Val, p.Val388Met, and p.Arg158Gln (AF: 5% each), p.Leu48Ser, and p.Ile65Thr (AF:4% each). The predicted phenotype was assigned by Guldberg´s arbitrary value (AV) and compared with the clinical phenotype based in tolerance to Phe intake. 29.1% (n:30) were hyperphenylalaninemias, 18.5% (n:19) mild-PKU, 27.2% (n:28) moderate-PKU and 25.2 % (n:26) classical-PKU. Genotype/phenotype correlation was statistically significant (p<0.001) Overall concordance was 62,5%: 93.3% in hyperphenylalaninemia, 64.7% in mild-PKU and 65.4% in classical patients. The moderate-PKU showed a weak concordance (17%) with milder AV prediction than clinical assessment. 74% of discordant moderate patients harbored p.Arg261Gln, and p.Val388Met. Our cohort is highly heterogeneous, with predominant Mediterranean influence (mainly Spanish), but with differences with other Latin-American countries.

Characteristics of gene variants among patients with hyperphenylalaninemia from Quanzhou region of Fujian province / 中华医学遗传学杂志

Objective@#To explore the spectrum of genetic variants among patients with hyperphenylalaninemia (HPA) from Quanzhou area of Fujian province.@*Methods@#For 63 children affected with HPA, next generation sequencing was used to identify potential variants in PAH, PTS, PCBD1, QDPR, SPR and GCH1 genes.@*Results@#Fifty two variants underlying phenylalanine hydroxylase deficiency (PAHD) and 13 variants underlying 6-pyruvoyl tetrahydropterin synthase deficiency (PTPSD) were identified. Two patients carried variants of both PAH and PTS genes. The most common variants of the PAH gene were R53H (21.69%), R241C (18.07%), R243Q (12.05%) and EX6-96A>G (7.23%), which were mainly located in exons 7 (32.53%), 2 (21.69%), 6 (9.64%) and 12 (9.64%). The L227M variant of the PAH gene was unreported previously. N52S (35.00%), P87S (25.00%), IVS1-291A>G (10.00%) and T67M (10.00%) variants were the most common variants for the PTS gene and were mainly located in exons 2 (35.00%) and 5 (35.00%).@*Conclusion@#The variant spectrum underlying HPA in Quanzhou area showed a geographical specificity. A novel variant of the PAH gene (L227M) has been detected.

Study of the phenylalanine hydroxylase gene variants in patients with phenylketonuria from Jiangxi province / 中华医学遗传学杂志

Objective@#To delineate the variants spectrum of phenytalanine hydroxylase (PAH) gene among 78 unrelated patients with phenylketonuria (PKU) from Jiangxi province.@*Methods@#The 13 exons and flanking intronic regions of the PAH gene were subjected to PCR amplification and sequencing.@*Results@#A total of 143 variants were detected among the 156 alleles, which included 54 types of variants, which yielded a detection rate of 91.7%. Common variants have included R243Q (26/143, 18.2%), R408Q (10/143, 7.0%), EX6-96A>G (8/143, 5.6%), IVS4-1G>A (7/143, 4.9%), R241C (7/143, 4.9%) and V399V (7/143, 4.9%). In addition, 6 novel variants were detected, which included IVS4-3T>G, Q172H, C284Y, V291L, V329del, and L430R. The variants consisted of missense, splicing, nonsense and deletion variants, which have mainly located in exons 7 (45, 31.5%), 12 (17, 11.9%), 11 (16, 11.2%) and 6 (14, 9.8%).@*Conclusion@#Variants of the PAH gene identified in Jiangxi province mainly involve exons 7, 12, 11 and 6, with the most common variants being R243Q and R408Q. Six novel variants were identified.

Mutation spectrum and new mutation of phenylalanine hydroxylase gene in Guangxi region / 中华实用儿科临床杂志

Objective To investigate the characteristics of the phenylalanine hydroxylase( PAH)gene muta﹣tions in patients With phenylketonuria(PKU)in Guangxi region,in order to provide clinical data for genetic counseling and prenatal gene diagnosis. Methods Thirty－seven children diagnosed as PKU in the Maternal and Children＇s Hos﹣pital of Guangxi Zhuang Autonomous Region Were enrolled in the study betWeen January 2009 and December 2017. Ve﹣nous blood Was collected and the PAH gene sequence Was determined by Sanger sequencing after amplification With the polymerase chain reaction technique. The neW gene mutations Were defined based on the national and international literature revieW and databases. MeanWhile,100 healthy individuals Were selected as the control group for gene sequen﹣cing to confirm Whether the mutation Was a neW one. Results Thirty－seven cases of PKU Were detected for 68 muta﹣tions,With the detection rate being 91. 89%(68／74). Six mutations Were identified in exon 7,Which accounted for 31. 08% of all,exon 12(18. 92%),exon 8(10. 81%)and exon 6(10. 81%)folloWed. A total of 25 different muta﹣tions Were identified Which including 14 missense mutations(56. 00%),7 nonsense mutations(28. 00%),3 splicing junction mutations(12. 00%),and 1 deletion mutation(4. 00%). The most common mutations included c. 1223G＞A (p. R408Q),c. 728G＞A(p. R243Q)and c. 721C＞T( p. R241C),accounting for 14. 86%,13. 51%,and 10. 81%, respectively. After querying international databases,including PAH mutation database and Human Gene Mutation Data﹣base and forecasting softWare,three kinds of mutations c. 314C＞ T(p. T105I),c. 583A＞ G(p. K195E),c . 851G＞A(p. C284Y)Were verified as novel PAH gene mutations. Conclusions The mutation spectrum of the PAH gene in Guangxi has been identified. And 3 kinds of mutations have been identified. This may accumulate valuable information for gene diagnosis and prenatal diagnosis of PKU in Guangxi region.

Deficiencia de fenilalanina hidroxilasa: espectro clínico y estado actual del diagnóstico en colombia / Phenylalanine hydroxylase deficiency: clinical spectrum and current diagnosis in colombia

RESUMEN Las mutaciones del gen PAH generan deficiencia de la enzima fenilalanina hidroxilasa. Su actividad final varía desde una actividad casi nula o indetectable en la fenilcetonuria clásica hasta una actividad residual del 10 al 35% de la normal. Esta alteración corresponde al error innato del metabolismo de los aminoácidos más frecuente, afectando a 1 de cada 10.000 personas. Las diferentes cantidades de fenilalanina en sangre se traducen en un espectro amplio de manifestaciones clínicas que incluyen retraso global del desarrollo, discapacidad intelectual, convulsiones, rasgos autistas y comportamiento agresivo en los casos más graves. El diagnóstico temprano a través de los programas de tamizaje neonatal se considera prioritario pues las intervenciones oportunas evitan el daño del sistema nervioso central. Conclusiones: El diagnóstico en Colombia es tardío, las intervenciones realizadas a partir de ese momento son fútiles pues el deterioro cognitivo es irreparable, por lo tanto es imperativa la realización de pruebas diagnósticas tempranas cuando aún las intervenciones médicas pueden impactar la mejoría clínica del paciente con disminución importante de la morbilidad propia de esta patología, convirtiéndose en una necesidad la ampliación del programa de tamizaje neonatal, el cual estaría amparado bajo la ley colombiana de enfermedades huérfanas.

ABSTRACT Mutations in the PAH gene generate phenylalanine hydroxylase enzyme deficiency. Its final activity varies from almost null or undetectable in classical phenylketonuria to a residual activity of 10 to 35% of normal activity. This alteration corresponds to the innate more frequent error of the metabolism of the amino acids, affecting 1 of every 10,000 people. Different amounts of phenylalanine in blood translate into a broad spectrum of clinical manifestations including global developmental delay, intellectual disability, seizures, autistic traits, and aggressive behavior in the most severe cases. Early diagnosis through neonatal screening programs is considered a priority because timely interventions avoid damage to the central nervous system. Conclusions: The diagnosis in Colombia is belated, the interventions made from that moment are futile because the cognitive deterioration is irreparable. Therefore, it is imperative to carry out early diagnostic tests when medical interventions can still impact the clinical improvement of the patient with an important decrease of the morbidity characteristic of this pathology, making it necessary to expand the neonatal screening program which would be protected under the Colombian law of orphan diseases.

Prenatal diagnosis and PAH gene mutations in 55 pedigrees with phenylketonuria / 中华围产医学杂志

Objective To investigate phenylalanine hydroxylase ( PA H ) gene mutations and to perform prenatal diagnosis in 55 pedigrees with classical phenylketonuria (PKU). Methods Subjects of this study were 55 probands diagnosed with PKU in the Gansu Provincial Maternal and Child Health Care Hospital from 2013 to 2017 and their pedigrees. Sanger sequencing/Multiplex ligation-dependent probe amplification (MLPA) was used to investigate PA H gene mutations in these probands and their parents. Sanger sequencing/MLPA, linkage analysis of three common short tandem repeats (STR) including PAH-26, PAH-STR and PAH-32 in the PA H gene and paternity testing were used in combination for prenatal diagnosis of 60 fetuses in the 55 pedigrees. Results Among the 110 alleles in the 55 probands, 108 mutant alleles (98.2%) were found by Sanger sequencing. The 108 mutant alleles located in 38 regions resulting in 22 missense mutations, nine splice site mutations, five nonsense mutations and two microdeletion. The most common mutations were c.728G>A (22.2%, 24/108), c.442-1G>A (5.6%, 6/108), c.611A>G (5.6%, 6/108), c.764T>C (5.6%, 6/108), c.1068C>A (5.6%, 6/108) and c.331C>T (4.6%, 5/108). Loss of heterozygosity in 4-5 and 4-7 exons were detected by MLPA in two probands, in which only one mutation was unidentified. Prenatal diagnosis for the 60 fetuses were successfully performed. Among them, 17 fetuses (28.3%) were affected, 29 fetuses (48.3%) were heterozygous carriers and fetuses 14(23.4%) were unaffected ones. Conclusions Combination of Sanger sequencing/MLPA, linkage analysis and paternity testing could provide accurate prenatal diagnosis in pedigrees with PKU.

Phenylalanine hydroxylase gene mutations analysis on phenylketonuria patients of Han Chinese people in Huaihai region / 中华实用儿科临床杂志

Objective To determine the spectrum of mutations responsible for Phenylalanine hydroxylase (PAH) deficiency on phenylketonuria (PKU) patients of Han Chinese people in the Huaihai region of central China.Methods One hundred and one patients diagnosed with PKU were referred to Xuzhou Maternity and Child Health Care Hospital for genetic counseling/analysis from January 2003 to December 2013.Thirteen exons of PAH gene mutations,as well as their flanking introns,were identified in 202 of chromosomes using polymerase chain reaction-single strand conformation polymorphism(PCR-SSCP) and sequencing.Results (1) The spectrum was composed of 24 different mutation types,which had been submitted to the National Center for Biotechnology Information(NCBI) dbSNP databases under accession number SS#2137543837_SS#C2137543860.(2)The most commonly affected region was exon 7 and its flanking introns.The most prevalent mutations were c.728G ＞ A (p.R243Q),followed by c.721C ＞ T (p.R241C),c.1155G ＞ C(p.L385L),c.1068C ＞ A(p.Y356X),c.-71A ＞ C(-71A ＞ C) and c.60 + 62C ＞ T (IVS1 +62C ＞T),accounting for 18.317％,8.416％,4.950％,3.960％,3.465％ and 2.970％ of the mutant chromosomes,respectively.(3)Two novel mutations were identified in PAH gene in PKU patients of Han Chinese people:c.60+62C＞T(IVS1 +62C ＞T) and c.782G ＞T(p.R261L).Conclusions The vast majority of PAH mutations identified corresponded to those observed for the PKU populations in the other regions in China,whereas a few are considerably different from others.The mutational spectrum of PAH gene found in patients with PKU in the Huaihai region exhibit regional association.

Analysis of types and metabolic profiles of hyperphenylalaninemia / 中华实用儿科临床杂志

Objective To study the characteristics of hyperphenylalaninemia (HPA) and the differences in blood and urine metabolic index and their correlation.Methods A total of 137 patients with HPA diagnosed by the Pediatric Inherit Metabolism and Endocrine Department,Guangdong Women and Children's Hospital,Guangzhou Medical University from January 2014 to June 2017,were enrolled.Tandem mass spectrometry (MS/MS),gas chromatography/ mass spectrometry (GC-MS) and high performance liquid chromatography (HPLC) were used to analyze the concentration of blood and urine metabolites in children,and the patients were divided into different groups according to the drug load test of tetrahydrobiopterin (BH4) and dihydrobiopterindine reductase (DHPR) deficiency.The HPA metabolite analysis of horizontal concentration by statistical differences and correlation analysis were performed.Results Among the 137 cases of HPA,there were 101 cases (73.7％) of phenylalanine hydroxylase deficiency (PAH),and among them 21 cases (15.3％) were classic phenylketonuria (PKU),37 cases were mild PKU (27.0％),43 cases (31.4％) wcrc mild HPA.Thcrc were 22 cases (16.1％) with BH4 reaction,and 79 cases (57.7％) of non-reactive type.Besides,there were 36 cases (26.3％) of tetrahydrobiopterin deficiency (BH4 D),of which 6-pyruvoyl tetrahydropterin synthase deficiency (PTPS) in 34 cases (24.8％) and dihydrobiopterindine reductase deficiency (DHPR) in 2 cases (1.5％).Urinary phenylacetic acid (r =0.673,P ＜ 0.01),phenyllactic acid (r =0.736,P ＜ 0.01),phenylpyruvic acid (r =0.642,P ＜ 0.01) were significantly correlated with blood phenylalanine (Phe) concentration,and the neopterin (N) (r =0.442,P ＜ 0.01) and biopterin (B) (r =0.398,P ＜ 0.01) had low correlation.Urinary phenylacetic acid,phenyllactic acid and phenylpyruvic acid had no correlation with urinary pterin.There were significant differences among PTPS deficiency group,BH4 response type,and non-reactive type(all P ＜ 0.05),but no significant difference between the BH4 reaction type and the non-reactive group (P ＞ 0.05).Conclusions Through the analysis of the different types of HPA metabolic profiles,it can help to master the incidence and characteristics in the region,within a certain concentration range of blood Phe,the phenylacetic acid,phenyllactic acid,phenylpyruvic acid should not be tested by GC-MS alone.Uterine erythropoietin analysis of BH4D classification and identification of BH4 reaction,non-reactive PKU have a supporting role,so master the metabolic index of various types of concentration and relevance of HPA,it can provide basis for early diagnosis,accurate treatment and follow-up.

Analysis of phenylalanine hydroxylase gene mutations in Han ethnic children with phenylketonuria of Xinjiang region / 临床检验杂志

Objective To investigate the characteristics of phenylalanine hydroxylase (PAH) gene mutations in Han ethnic children with phenylketonuria of Xinjiang region.Methods The mutations in the promoter,exons 1-13 and flanking introns of PAH genes from 71 Han ethnic PKU children and their parents of Xinjiang region were detected by PCR,DNA sequencing and high-throughput targeted sequencing,and the obtained results were compared with those from other four provinces in northwest of China,Japan and Europe.Re suits A total of 37 kinds of mutations,including missense mutation,splice site mutation,nonsense mutation,deletion mutation and frameshift mutation,were detected in 90.1％ (128/142) of PAH alleles from 71 Han ethnic PKU children of Xinjiang region.Most mutations existed in exons 7,6,3,12,2 and 11 and intron 4 of PAH gene.The most common missense mutations were R243Q (21.8％) and R53H (7.7％).The most common splicing sites were EX6-96A ＞ G(6.3％),IVS4-1G ＞ A(4.9％) and V399V (4.2％).Moreover,The most common nonsense mutations were R111X(4.9％) and Y356X(4.9％).The detection rate of R53H mutation (7.7％) in Han ethnic PKU children of Xinjiang region was significantly higher than that in other 4 provinces of northwest of China,and a novel PAH gene nutation P225S(c.673C ＞ T) was found.Conclusion The mutation spectrum of PAH gene in Han ethnic PKU children of Xinjiang region is similar to that in other 4 provinces of northwest of China,but significantly different from that of Japanese and European population,which displays a distinct and conservative characteristic.

Fenilcetonuria de diagnóstico tardío y mutaciones asociadas en una familia ecuatoriana / Phenylketonuria of late diagnosis and associated mutations in an Ecuadorian family

La fenilcetonuria es un error innato del metabolismo, producido por mutaciones en el gen de la fenilalanina hidroxilasa. Se describe el caso de un adolescente de 15 años con diagnóstico tardío de fenilcetonuria, quien presenta retardo mental severo, convulsiones e hipopigmentación. En este estudio se realizó el diagnóstico molecular de fenilcetonuria y se detectó la mutación p.R252W en homocigosis en el gen que codifica para la fenilalanina hidroxilasa. La presencia de esta variante nos permitió inferir la falta de respuesta a la terapia con sapropterina, medicamento que actúa como cofactor de la enzima, por la ausencia de actividad enzimática residual reportada para esta variante. Debido al retraso psicomotor del paciente, se decidió aplicar terapia lúdica y fortalecimiento muscular a través de la intervención fisioterapéutica; sin embargo, no se observó una mejoría permanente al aplicar este tratamiento, motivado por la falta de continuidad.

Phenylketonuria is an inborn error of metabolism due to mutations on the phenylalanine hydroxylase gene. We described the case of a 15 years old-adolescent with late diagnosis of phenylketonuria, who presents severe mental retardation, convulsions and hypopigmentation. In this study, the molecular diagnosis of phenylketonuria was performed, detecting p.R252W mutation in homozygous state on the phenylalanine hydroxylase gene. The presence of this variant allowed us to infer the lack of response to drug therapy with sapropterina which works as an enzyme cofactor, due to the absence of residual enzymatic activity reported for the p.R252W variant. Physical therapy was applied through playful therapy and muscular strengthening, because of the psychomotor retardation present in the patient. The failing in continuing with the physical therapy program stopped the patient´s improvement.

Strategies for Successful Long-Term Engagement of Adults With Phenylalanine Hydroxylase Deficiency Returning to the Clinic

Abstract Nearly half of all patients diagnosed with phenylalanine hydroxylase (PAH) deficiency, also known as phenylketonuria, are lost to follow-up (LTFU); most are adults who stopped attending clinic after the age of 18 years. To understand why adult patients with PAH deficiency disengage from their clinic, a focus group of 8 adults with PAH deficiency who had been LTFU for 2 or more years was held in March 2016. Ten clinicians observed the focus group and discussed strategies for successfully reengaging adult patients and encouraging lifelong management of PAH deficiency. Four strategies were proposed: (1) create a safe, supportive environment, (2) acknowledge patients as partners in their care, (3) develop individualized management plans, and (4) provide patients with additional resources. These strategies provide a framework to motivate change in clinical practice to meet the unique needs of adults with PAH deficiency.

Progress in molecular genetics of hyperphenylalanemia / 国际儿科学杂志

Hyperphenylalaninemia is a common group of inherited metabolic diseases.It is characterized by the increased concentration of plasma phenylalanine.The metabolism of phenylalanine requires phenylalanine hydroxylase and coenzyme tetrahydrobiopterin.These enzymes cannot function normally if there is any mutation in their encoding genes.Children suffering from hyperphenylalaninemia without promptly treatment may present mental development delay and other serious nervous system sequelae.With the technical improvement of molecular genetics,there have been many progresses in the study of genetic diagnosis,genotype-phenotype correlation and gene therapy of hyperphenylalaninemia.This article reviews the history and classification of hyperphenylalaninemia,the characteristics of gene mutation,the methods of genetic diagnosis,the genotype-phenotype correlation,and the progress of new therapy.

Caracterización fenotípica y molecular de una familia colombiana con fenilcetonuria / Phenotypic and molecular characterization of a Colombian family with phenylketonuria

Introducción. La fenilcetonuria es un trastorno metabólico caracterizado por un compromiso neurológico grave y por alteraciones del comportamiento. Su diagnóstico temprano permite establecer un tratamiento efectivo que evita las secuelas y modifica el pronóstico. Objetivo. Caracterizar a una familia con fenilcetonuria en Colombia, a nivel clínico, bioquímico y molecular. Materiales y métodos. Se estudió una población de siete individuos de una familia consanguínea en la que cuatro hijos presentaban signos clínicos sugestivos de fenilcetonuria. Una vez firmado el consentimiento informado, se tomaron muestras de sangre y orina para las pruebas colorimétricas, la cromatografía de capa fina y la cromatografía líquida de alta eficacia. Se extrajo el ADN y se secuenciaron los 13 exones del gen PAH de todos los sujetos estudiados. Se diseñaron iniciadores para cada exón con el programa Primer 3; la secuenciación automática se hizo con el equipo Abiprism 3100 Avant y, el análisis de las secuencias, con el programa SeqScape v2.0. Resultados. Se describieron las características clínicas y moleculares de una familia colombiana con fenilcetonuria en la que se identificó la mutación c.398_401delATCA; se presentó una correlación fenotipo-genotipo con una interesante variabilidad clínica entre los afectados, a pesar de tener la misma mutación. Conclusiones. Es importante el reconocimiento temprano de esta enfermedad para evitar sus secuelas neurológicas y psicológicas, pues los pacientes llegan a edades avanzadas sin diagnóstico ni tratamiento adecuados.

Introduction: Phenylketonuria is a metabolic disorder characterized by severe neurological involvement and behavioral disorder, whose early diagnosis enables an effective treatment to avoid disease sequelae, thus changing the prognosis. Objective: To characterize a family with phenylketonuria in Colombia at clinical, biochemical and molecular levels. Materials and methods: The population consisted of seven individuals of a consanguineous family with four children with suggestive symptoms of phenylketonuria. After signing an informed consent, blood and urine samples were taken for colorimetric tests and high performance liquid and thin layer chromatographies. DNA extraction and sequencing of the 13 exons of the PAH gene were performed in all subjects. We designed primers for each exon with the Primer 3 software using automatic sequencing equipment Abiprism 3100 Avant. Sequences were analyzed using the SeqScape, v2.0, software. Results: We described the clinical and molecular characteristics of a Colombian family with phenylketonuria and confirmed the presence of the mutation c.398_401delATCA. We established a genotype-phenotype correlation, highlighting the interesting clinical variability found among the affected patients despite having the same mutation in all of them. Conclusions: Early recognition of this disease is very important to prevent its neurological and psychological sequelae, given that patients reach old age without diagnosis or proper management.

Conventional Phenylketonuria Treatment

Abstract Phenylketonuria (PKU) is caused by a deficient activity of enzyme phenylalanine (Phe) hydroxylase, which results in high Phe blood concentration, which is toxic to the central nervous system. The fundamental purpose of nutritional treatment is to reduce and maintain blood Phe between 2 mg/dL (120 µmol/L) and 6 mg/dL (360 µmol/L) in order to prevent neuropathogenic complications. At the same time, nutrition support must provide enough energy and nutrients to promote normal growth and development and also to avoid vitamin and mineral deficiencies. Phenylketonuria treatment must be maintained long-life and its adherence must be frequently assessed. The amount of Phe required by patients with PKU varies throughout life and must be adjusted according to individual tolerance, residual phenylalanine hydroxylase enzymatic activity, age, sex, growth rate, protein intake, and nutritional and biochemical status among others. Treatment must be done by trained personnel. It is necessary to unify treatment criteria and further research must be done.

Analysis of neonatal hyperphenylalaninemia screening and therapeutic effect in Xuzhou from 2003 to 2015 / 中华围产医学杂志

Objective To investigate the prevalence,clinical classification,treatment and prognosis of neonatal hyperphenylalaninemia(HPA) in Xuzhou area,China.Methods Infants born between July 1,2003 and July 1,2015 in Xuzhou area were investigated.Heel blood samples of neonates were collected at 72 hours after birth,and the concentration of blood phenylalanine(Phe) was determined by fluorescent quantitative method in Xuzhou Maternity and Child Health Care Station Neonatal Disease Screening Center.Differential diagnosis was performed in all 265 cases diagnosed as HPA by urinary pterin analysis and dihydropteridine reductase activity determination.The blood Phe concentration and mental development were followed up regularly in infants with HPA.Mutations of phenylalanine hydroxylase (PAH) gene were analyzed by gene sequencing.The relationship between blood Phe concentration and mental development was analyzed by Bivariate correlation analysis.Results (1) The prevalence of HPA in neonates in Xuzhou was 1/4 635.Among the 265 confirmed HPA cases,260 cases(98.11％) had PAH deficiency,including 90(33.96％) classical phenylketonuria(PKU),84(31.70％) mild PKU and 86(32.45％) mild HPA.The other five patients(1.89％) diagnosed with tetrahydrobiopterin (BH4) deficiency all had 6-pyruvoyl tetrahydropteim synthase(PTPS) deficiency.(2) Among the 265 HPA cases,26 cases refused any treatment,including five cases of PTPS deficiency and 21 cases of PKU.Of the five patients with PTPS deficiency,two died and the other three had normal mental and physical development.Twenty-one PKU patients who refused treatment had mental retardation of various degrees.Among 153 PKU patients who received medical treatment,three died and 12 were lost to follow-up.(3) For 138 PKU patients who received dietary treatment and follow-up,the ages at the last visit were two months to 12 years,116 of them had normal mental development,the remaining 22 patients had mental retardation,and a negative correlation was observed between mental development and the average Phe concentration.(4) Thirty-five patients with PAH deficiency underwent gene sequencing,and 22 kinds of mutations of PAH gene were detected.Conclusions The prevalence of HPA in Xuzhou area is higher than the average national level.With early diagnosis and standard treatment,most of PKU neonates can have normal mental development.Phe level control is an important factor for mental development.

Thermal stability improvement for phenylalanine hydroxylase by site-directed mutagenesis / 生物工程学报

In proteins of thermophilic bacteria, Gly is tend to be replaced by Ala and Lys is tend to be replaced by Arg to adapt the high temperature. In order to improve the thermal stability of phenylalanine hydroxylase (PAH) from Chromobacterium violaceum, all the Gly on PAH were mutated to Ala and Lys to Arg. Positive mutant enzymes with improved thermal stability were selected, followed by combined mutation and characterization. The results revealed that half-lives of K94R and G221A mutants at 50 °C were 26.2 min and 16.8 min, which were increased by 1.9-times and 0.9-times than the parent enzyme (9.0 min). The residual activity of K94R/G221A mutant was improved to 65.6% after keeping at 50 °C for 1 h, which was 6.6 time higher than the parent enzyme (8.6%). Circular dichroism (CD) spectroscopy revealed that Tm values of the parent enzyme, K94R, G221A and K94R/G221A were 51.5 ℃, 53.8 ℃, 53.1 ℃ and 54.8 ℃, respectively. According to the protein structure simulation, the two mutations were located on flexible loop. In the K94R mutant, the mutated Arg94 on the surface of the enzyme formed an extra hydrogen bond with Ile95, which stabilized the located loop. In the G221A mutant, the mutated Ala221 formed hydrophobic interaction with Leu281, which could stabilize both the loop and flexible area of the C-terminus of G221A. The results not only provided a reference for protein modification on thermal stability, but also laid the foundation for application of phenylalanine hydroxylase in the field of functional foods.