DOUBLE TRANSDUCTION WITH GTP CYCLOHYDROLASE-I AND TYROSINE-HYDROXYLASE IS NECESSARY FOR SPONTANEOUS SYNTHESIS OF L-DOPA BY PRIMARY FIBROBLASTS

Gene transfer of tyrosine hydroxylase (TH) in animal models of Parkins on's disease (PD), using either genetically modified cells or recombin ant virus vectors, has produced partial restoration of behavioral and biochemical deficits. The limited success of this approach may be rela ted to the availability of the cofactor, tetrahydrobiopterin (BH4), be cause neither the dopamine-depleted striatum nor the cells used for ge ne transfer possess a sufficient amount of BH4 to support TH activity. To determine the role of BH4 in gene therapy, fibroblast cells transd uced with the gene for TH were additionally modified with the gene for GTP cyclohydrolase I, an enzyme critical for BH4 synthesis. In contra st to cells transduced with only TH, doubly transduced fibroblasts spo ntaneously produced both BH4 and 3,4-dihydroxy-L-phenylalanine. To exa mine further the importance of GTP cyclohydrolase I in gene therapy fo r PD, in vivo micro-dialysis was used to assess the biochemical change s in the dopamine-denervated striatum containing grafts of genetically modified fibroblasts. Only denervated striata grafted with fibroblast s possessing both TH and GTP cyclohydrolase I genes displayed biochemi cal restoration. However, no significant differences from controls wer e observed in apomorphine-induced rotation. This is partly attributabl e to a limited duration of gene expression in vivo. These differences between fibroblasts transduced with TH alone and those additionally mo dified with the GTP cyclohydrolase I gene indicate that BH4 is critica l for biochemical restoration in a rat model of PD and that GTP cycloh ydrolase I is sufficient for production of BH4.