Anthony G. DiLella

Correlation between polymorphic DNA haplotypes at phenylalanine hydroxylase locus and clinical phenotypes of phenylketonuria

Eight polymorphic sites for seven restriction endonucleases have been reported at the human phenylalanine hydroxylase locus. The composite profile of the presence or absence for each of the eight polymorphic sites within an allele defines the haplotype of the corresponding allele. Twelve such haplotypes associated with normal and mutant phenylalanine hydroxylase alleles have been identified in 33 Danish families with children with phenylketonuria. Of the 66 mutant alleles analyzed, 59 (89%) were associated with only four haplotypes. The identification of individual phenylalanine hydroxylase alleles by haplotype analysis enables correlation of the hyperphenylalaninemic phenotypes of the patients with their genotypes. Patients who were either homozygous or heterozygous for the mutant alleles of haplotypes 2 and 3 had a severe clinical course. Patients who had a mutant allele of either haplotype 1 or 4 usually had a less severe clinical phenotype. The recent demonstration that the mutation responsible for classic phenylketonuria associated with haplotype 3 is not present in mutant alleles of other haplotypes provides unambiguous evidence that there are multiple mutations in the phenylalanine hydroxylase gene and supports the hypothesis that different combinations of mutant alleles may be responsible for the clinical diversity of phenylketonuria.

Molecular biology of -" phenylalanine hydroxylase and phenylketonuria

Abstract The cloning of cDNAs for phenylalanine hydroxylase (PAH) from rat and human livers has let to major advances in understanding the structure and function of this metabolically important enzyme and its deficiency state, phenylketonuria (PKU). The nucleic acid and amino acid sequences of human PAH have been deduced, its genomic locus has been cloned, and mutations at the locus in PKU patients have been examined. Gene transfer experiments result in enzyme activity in cells that do not normally produce PAH, suggesting that somatic gene therapy for PKU may be feasible.

Molecular basis of phenylketonuria and recombinant DNA strategies for its therapy.

Mutations in the human phenylalanine hydroxylase gene associated with two prevalent mutant alleles have been identified and shown to be in linkage disequilibrium with the corresponding mutant restriction fragment length polymorphism haplotypes. These results suggest the possibility of carrier detection in the population without a prior family history of phenylketonuria (PKU). Furthermore, recombinant retroviruses containing the full-length human phenylalanine hydroxylase cDNA have been constructed and used to transduce functional enzymatic activity into cultured hepatoma cells. Together with the recent success in retroviral infection of primary mouse hepatocytes, it will be possible to use the mouse model to investigate somatic gene therapy for PKU.