PAH deficient pathology in humanized c.1066-11G>A phenylketonuria mice

Area of Science:

• Genetics and Genomics
• Neuroscience
• Metabolic Diseases

Background:

• Phenylketonuria (PKU) is a neurometabolic disorder caused by mutations in the phenylalanine hydroxylase (PAH) gene.
• A common human PAH variant, c.1066-11G>A, leads to aberrant splicing and reduced enzyme activity.
• Existing models may not fully recapitulate the human PKU phenotype.

Purpose of the Study:

• To generate and characterize a novel, partially humanized mouse model of PKU.
• To utilize CRISPR/Cas9 technology for precise genetic modification.
• To create a tool for studying PKU pathophysiology and therapeutic development.

Main Methods:

• CRISPR/Cas9 gene editing to introduce a humanized intron 10 sequence with the c.1066-11G>A PAH variant.
• Phenotypic analysis of homozygous Pah c.1066-11A mice.
• Biochemical and molecular analyses of hepatic PAH activity, neurotransmitter levels, and brain morphology.

Main Results:

• The mouse model accurately replicates the splicing defect and exhibits negligible hepatic PAH activity.
• Mice display hallmark PKU symptoms: hypopigmentation, reduced growth, decreased survival, elevated blood/brain phenylalanine, and altered neurotransmitter levels.
• Neuropathological changes include glial cell alterations, increased GFAP/Iba1 staining, and reduced myelination, alongside behavioral deficits.

Conclusions:

• The developed mouse model serves as a valuable preclinical tool for PKU research.
• It accurately recapitulates key aspects of human PKU pathophysiology, including splicing defects and neurological consequences.
• This model facilitates the investigation of disease mechanisms and the testing of novel therapeutic strategies for PKU.