Hepatocytes from wild-type or heterozygous donors are equally effective in achieving successful therapeutic liver repopulation in murine phenylketonuria (PKU)

• Published in: Molecular genetics and metabolism Vol. 104; no. 3; pp. 235 - 240
• Main Authors: Hamman, Kelly J., Winn, Shelley R., Harding, Cary O.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.11.2011
• Subjects: Animals; DNA Primers - genetics; Genetic Therapy - methods; Hepatocyte transplantation; Hepatocytes - transplantation; Hydrolases - deficiency; Liver - cytology; Liver - pathology; Liver Regeneration - genetics; Mice; Mice, Mutant Strains; Models, Biological; Mouse model; Phenotype; Phenylalanine; Phenylalanine - metabolism; Phenylalanine Hydroxylase - genetics; Phenylalanine Hydroxylase - metabolism; Phenylalanine hydroxylase deficiency; Phenylketonuria; Phenylketonurias - therapy; Polymerase Chain Reaction; Therapeutic liver repopulation; Phenylketonuria; NTBC; Hepatocyte transplantation; Phenylalanine; PAH; Therapeutic liver repopulation; Mouse model; PKU; Phenylalanine hydroxylase deficiency; Phe; Tyr; FAH
• ISSN: 1096-7192; 1096-7206
• DOI: 10.1016/j.ymgme.2011.07.027

Successful restoration of phenylalanine (Phe) clearance following liver-directed gene therapy in murine phenylketonuria (PKU) is likely dependent upon both the number of cells successfully transduced and the amount of phenylalanine hydroxylase (PAH) activity expressed per cell. At low levels of transduction, Phe clearance could be limited by the low absolute number of PAH-expressing cells rather than the total amount of PAH activity produced in the liver. We have evaluated the interrelationship between the number of PAH positive cells, the amount of PAH activity produced and Phe clearance through experiments with hepatocyte-mediated therapeutic liver repopulation in the Pahenu2 mouse, a model of PKU. We compared the therapeutic efficacy of transplantation with either wild-type hepatocytes or hepatocytes from heterozygous Pahenu2/+ donors into PAH deficient, hyperphenylalaninemic Pahenu2/Pahenu2 mice. The recipient mice were also homozygous for fumarylacetoacetate hydrolase (FAH) deficiency. In this model system, FAH positive donor hepatocytes enjoy a selective growth advantage in the FAH-deficient recipient. If Phe clearance is governed predominantly by the total PAH activity, then more heterozygous cells, which express lower PAH activity than wild-type cells, should be required to correct Phe clearance. If the absolute donor cell number is more important, then wild-type hepatocytes should have no advantage over heterozygous cells. We successfully carried out therapeutic liver repopulation with heterozygous donor cells in fifteen mice and an additional thirteen transplants with wild-type cells. Blood Phe was successfully reduced in both transplant groups, and the relationship between the final blood Phe level and the extent of liver repopulation with donor cells did not differ between the two donor groups. Regardless of the type of donor cell, liver repopulation of approximately 3–10% was sufficient to at least partially reduce blood phenylalanine, and blood Phe levels were completely corrected in mice that had attained greater than approximately 10% liver repopulation. We conclude from our study that the absolute number of PAH-expressing cells likely governs Phe clearance at least at the levels of repopulation reported here and that the amount of PAH activity per donor cell is a less critical variable. The implication for liver-directed gene therapy of PKU is that only partial correction of cellular PAH deficiency may yet improve Phe clearance as long as a sufficient number of hepatocytes is successfully transduced. ► Therapeutic liver repopulation corrects hyperphenylalaninemia in murine PKU. ► Liver repopulation requires a selective growth advantage for donor hepatocytes. ► We transplanted either wild type or PAH deficient hepatocytes into PKU mice. ► Wild type or PAH deficient hepatocytes are equally effective in treating PKU. ► Phe clearance is influenced both by PAH activity and transmembrane Phe transport.