Analiza genov za fenilalanin - hidroksilazo pri bolnikih s hiperfenilalanemijo : magistrska naloga

Matuc, Vanda (author), Trebušak Podkrajšek, Katarina (mentor)

Blaga hiperfenilalaninemija (HFA) je avtosomno recesivno podedovana bolezen, ki nastane kot posledica prirojene motnje v hidroksilaciji fenilalanina (Phe) zaradi znižane aktivnosti encima fenilalanin-hidroksilaze. Posledično pride do motenj pri presnovi fenilalanina v tirozin, kar privede do povišanih vrednosti fenilalanina v krvi. Blaga HFA ima enako genetsko osnovo kot fenilketonurija (PKU), vendar gre v primeru blage HFA za dosti milejšo obliko bolezni z zmerno do blago povišanimi vrednostmi Phe v krvi. Vzročnega zdravljenja bolezni ne poznamo. Pri hujših oblikah bolezni, oziroma pri močno povišanih vrednostih Phe, je potrebna čim hitrejša uvedba dietnega zdravljenja z omejitvijo vnosa Phe. Zato je ključnega pomena presejalno testiranje novorojencev na PKU, ki se v Sloveniji izvaja od leta 1979. Namen magistrske naloge je bila opredelitev povezave med koncentracijo Phe pri preiskovancih z blago HFA in genotipom gena za fenilalan-hidroksilazo (PAH). V analizo je bilo vključenih 97 preiskovancev z blago HFA, ki so imeli koncentracijo Phe višjo od 200 μmol/L. Opredelili smo nabor 22 genetskih sprememb na skupno 194 neodvisnih alelih. Po dve genetski spremembi sta bili prisotni pri 21 preiskovancih, po ena sprememba pri 39 preiskovancih, pri preostalih preiskovancih nismo opredelili sprememb. Najpogosteje je bila zastopana genetska sprememba p.R408W v eksonu 12. Druge pogostejše genetske spremembe so bile p.A403V, p.R158Q, p.A300S in p.E390G. Pri 21 bolnikih, ki so imeli po dve spremembi, smo opredelili 19 različnih genotipov. Pri tem sta se ponavljali dve kombinaciji genotipa, in sicer: p.A403V v kombinaciji s p.D415N in p.A403V v kombinaciji s p.Phe55fs. Opredelili smo tudi dve novi, še neopisani spremembi (p.W326X in p.V45I). Primerjava nabora opredeljenih genetskih sprememb pri bolnikih s HFA, s spremembami predhodno opredeljenimi pri slovenskih bolnikih s PKU, je pričakovano pokazala razlike v naboru in frekvenci sprememb. Najpomembnejša razlika med skupinama je, da v skupini z blago HFA nismo določili nobene genetske spremembe v homozigotni obliki. Povprečna vrednost fenilalanina preiskovancev z blago HFA, ki niso imeli opredeljene vzročne spremembe v genu PAH, je bila 129 μmol/L, preiskovancev z eno vzročno spremembo 157 μmol/L in preiskovancev z dvema vzročnima spremembama 300 μmol/L. Dokazali smo statistično značilno povezavo med genotipom in presnovnim fenotipom bolnikov z blago HFA.

• Language: Slovenian
• Keywords: hiperfenilalaninemija fenilketonurija značilnosti bolezni fenotipske značilnosti zdravljenje bolezni nabor genotipov
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: FFA - Faculty of Pharmacy
• Place of publishing: Ljubljana
• Publisher: [V. Matuc]
• Year: 2016
• Number of pages: IV, 54 f.
• PID: 20.500.12556/RUL-120983
• UDC: 615.2:612.015.3(043.3)
• COBISS.SI-ID: 4149105
• Publication date in RUL: 28.09.2020

Secondary language

• Language: English
• Title: Analysis of phenylalanine hydroxylase gene in patients with hyperphenylalaninemia

Abstract

Mild hyperphenylalaninaemia (HPA) is an autosomal recessive inherited disease that occurs as a consequence of an inborn aberration of hydroxylation of phenylalanine due to deficiency of phenylalanine hydroxylase. Consequently, a disturbance in the metabolism of phenylalanine to tyrosine results in elevated blood levels of phenylalanine (Phe). Mild HPA has the same etiology as phenylketonuria (PKU), but in the case of mild HPA the disease presents itself in milder form with moderately to mildly elevated blood levels of Phe. There is no causal treatment of the disease. In the case of severe forms of the disease or heavily increased Phe levels, introduction of dietary treatment limiting the Phe intake is necessary. Newborn screening of PKU in Slovenia was implemented in 1979. The aim of this master's thesis was to evaluate the link between Phe concentrations in patients with mild HPA and the genotype of the PAH gene. The analysis was carried out on 97 patients with mild HPA who had the Phe concentration higher than 200 μmol/L. We have detected 22 genetic variants on 194 independent alleles. Two variants per individual were detected in 21 patients, one per individual in 39 patients while there were no variants in the remaining ones. The genetic variant p.R408W in exon 12 was the most frequent variant detected in this study. Other frequent variants were p.A403V, p.R158Q, p.A300S and p.E390G. Among 21 patients with two variants, 19 different genotypes were identified, where two combinations were more common, namely: p.A403V in combination with p.D415N and p.A403V in combination with p.Phe55fs. We have also identified two novel previously unidentified variants (p.W326X and p.V45I). We compared defined set of genetic variants in patients with HPA to genetic variants previously identified among Slovenian PKU patients who, as expected, showed differences in spectrum and frequency. The most important difference was that there were no causal genetic variants in homozygous form present in the mild HPA group. The average value of phenylalanine in subjects with mild HPA and without any causal genetic variant in the PAH gene was 129 μmol/L, among subjects with one identified causal variant 157 μmol/L and among subjects with two identified causal variants 300 μmol/L. We demonstrated statistically significant association between genotype and metabolic phenotype of patients with mild HPA.