New approaches for the treatment of Phenylketonuria: Evaluation of human phenylalanine hydroxylase (hPAH) formulations in cellular models  

Investigador: Paula Peralta
Instituição: Met&Gen, iMed.UL / FFUL – Lisboa

The current universal established PKU treatment relies on a highly restrictive diet of difficult compliance which may compromise the patient’s neurologic outcome and decrease their quality of life. The use of the pharmacological form of the natural hPAH cofactor BH4 has been recently approved by American and European authorities and a pegylated (PEG) form of a recombinant non-mammalian L-Phe degrading enzyme (phenylalanine ammonia lyase; PAL) already reached Phase III clinical trials. However, BH4 supplementation only allow diet liberalization in the restrict group of PKU patients presenting the mild form and PEG-PAL administration has been reported as potentially immunogenic. Therefore, a safety and efficacious treatment for all forms of PKU is still needed.

Abstract

The current universal established PKU treatment relies on a highly restrictive diet of difficult compliance which may compromise the patient’s neurologic outcome and decrease their quality of life. The use of the pharmacological form of the natural hPAH cofactor BH4 has been recently approved by American and European authorities and a pegylated (PEG) form of a recombinant non-mammalian L-Phe degrading enzyme (phenylalanine ammonia lyase; PAL) already reached Phase III clinical trials. However, BH4 supplementation only allow diet liberalization in the restrict group of PKU patients presenting the mild form and PEG-PAL administration has been reported as potentially immunogenic. Therefore, a safety and efficacious treatment for all forms of PKU is still needed.

Reposition of the deficient human enzyme, hPAH, is a potential approach to treat the full spectrum of PKU clinical phenotypes. However, this approach has never been truly investigated due to the difficulty to obtain a stable formulation of the recombinant hPAH protein mainly due to its high in vitro instability. The research team successfully obtained recombinant hPAH-loaded NP. The physicochemical and functional attributes of the obtained NP as well as their blood compatibility and absence of cytotoxicity demonstrate that the research team holds now relevant data indicating that enzyme reposition therapy for PKU is feasible. Consequently, we propose to investigate the capacity of the hPAH-loaded NP to render human cells lacking PAH activity (e.g. HEK293) capable of L-Phe metabolism. The uptake and cellular localization of formulated protein will be additionally studied in hepatocytes derived cells (HepG2), the cellular target for the enzyme reposition therapy here investigated. The obtained data will indicate if these studies could be extended in a second stage to an in vivo model of PKU or if a chemical/structural refinement of NP will be necessary in order to optimize cellular targeting and trafficking.

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