Retrovirus-mediated double transduction of the GTPCH and PTPS genes allows 6-pyruvoyltetrahydropterin synthase-deficient human fibroblasts to synthesize and release tetrahydrobiopterin.

The tetrahydrobiopterin (BH4) cofactor is essential for the aromatic amino acid hydroxylases that are involved in phenylalanine degradation and catecholamine and serotonin biosynthesis. Furthermore, BH4 is an essential and limiting cofactor for all types of nitric oxide synthases. BH4 deficiency results in hyperphenylalaninemia and monoamine neurotransmitter depletion associated with progressive mental retardation and is most commonly due to autosomal recessive mutations in 6-pyruvoyltetrahydropterin synthase (PTPS), the second enzyme for cofactor biosynthesis. Due to the relatively poor blood-brain barrier penetration of the cofactor, conventional therapy requires, besides oral doses of synthetic BH4, administration of neurotransmitter precursors and an aromatic amino acid decarboxylase inhibitor. The outcome of this therapy is not always beneficial. In this study we transduced into primary patient fibroblasts the human cDNAs for the BH4 biosynthetic enzymes GTP cyclohydrolase I and PTPS, expressed from different retroviral vectors. This allowed BH4 biosynthesis in originally PTPS-deficient cells. Moreover, the double-transduced fibroblasts released between 200 and 800 pmol of BH4/10(6) cells/day. Such engineered fibroblasts may be grafted into the central nervous system and used as depository cells for constitutive delivery of BH4.