Genomic profiling, implications for genotype-based treatment of 131 patients with phenylketonuria and characterization of novel p.Pro416Leu PAH variant.

Phenylketonuria (PKU) is the most common inborn disorder of amino acid metabolism caused by biallelic pathogenic variants in phenylalanine hydroxylase (PAH) gene. This study comprised genomic profiling and phenotypic characterization of 131 Serbian PKU patients along with implications for BH4 therapy. By combining Sanger sequencing, MLPA and WES re-analysis of previously unsolved cases, we identified 38 different disease causing variants in the PAH gene and classified them using ACMG guidelines. The most frequent variant was p.Leu48Ser (30.92%), followed by p.Arg408Trp (12.21%) and p.Ile306Val (8%). We detected one novel variant, p.Pro416Leu, which was classified as pathogenic, based on computational algorithms prediction, with destabilization as the mechanism of the effect upon PAH protein. For patients' phenotypic classification, we used pre-treatment serum Phe level and Phe tolerance, indicating that 42% of patients had classic PKU, 22% had mild PKU and 32% were classified as MHP (4% remained unclassified). Furthermore, we performed genotype-phenotype correlation study which emphasized the inconsistency of p.Leu48Ser variant. Given that not all PKU patients benefit from genotype-based therapy (Kuvan and sepiapterin), we assessed the potential responsiveness in our patients. We categorized all detected PAH genotypes accordingly and with 39.1% responsive and 44.5% probably responsive found that majority of patients may respond to BH4 therapy. Our study brings the updated spectrum of molecular genetic data, variant classification, characterization of novel p.Pro416Leu variant, detailed phenotypic characteristics and BH4 responsiveness for PKU patients from Serbia, therefore contributing to better understanding of molecular landscape of PKU.