Triple Transduction with Adeno-Associated Virus Vectors Expressing Tyrosine Hydroxylase, Aromatic-L-Amino-Acid Decarboxylase, and GTP Cyclohydrolase I for Gene Therapy of Parkinson's Disease

Abstract

Parkinson's disease (PD), a neurological disease suited to gene therapy, is biochemically characterized by a severe decrease in the dopamine content of the striatum. One current strategy for gene therapy of PD involves local production of dopamine in the striatum achieved by inducing the expression of enzymes involved in the biosynthetic pathway for dopamine. We previously showed that the coexpression of tyrosine hydroxylase (TH) and aromatic-L-amino-acid decarboxylase (AADC), using two separate adeno-associated virus (AAV) vectors, resulted in more effective dopamine production and more remarkable behavioral recovery in 6-hydroxydopamine-lesioned parkinsonian rats, compared with the expression of TH alone. Not only levels of TH and AADC but also levels of tetrahydrobiopterin (BH4), a cofactor of TH, and GTP cyclohydrolase I (GCH), a rate-limiting enzymes for BH4 biosynthesis, are reduced in parkinsonian striatum. In the present study, we investigated whether transduction with separate AAV vectors expressing TH, AADC, and GCH was effective for gene therapy of PD. In vitro experiments showed that triple transduction with AAV-TH, AAV-AADC, and AAV-GCH resulted in greater dopamine production than double transduction with AAV-TH and AAV-AADC in 293 cells. Furthermore, triple transduction enhanced BH4 and dopamine production in denervated striatum of parkinsonian rats and improved the rotational behavior of the rats more efficiently than did double transduction. Behavioral recovery persisted for at least 12 months after stereotaxic intrastriatal injection. These results suggest that GCH, in addition to TH and AADC, is important for effective gene therapy of PD.

Delayed delivery of AAV-GDNF prevents nigral neurodegeneration and promotes functional recovery in a rat model of Parkinson's disease

“…14 Previous studies, including our own, have demonstrated that AAV vector-mediated delivery of dopamine-synthesizing enzymes results in efficient long-term expression of the transgene in the striatum. [14][15][16] The present study used the Sauer and Oertel partial PD model. 17 In this model, intrastriatal injection of 6-OHDA induces progressive retrograde degeneration of DA neurons that starts between 1 to 2 weeks after lesioning and continues over 8 to 16 weeks.…”

“…The total number of complete body turns was counted during an observation period of Ն60 min, as described previously. 16 Spontaneous limb use was scored every 4 weeks using the cylinder test method. 10 Rats were placed in a clear glass cylinder large enough for free movement.…”

“…Levels of dopamine and its metabolites, HVA and DOPAC, were estimated using high-performance liquid chromatography (HPLC) as described previously. 16 Immunohistochemistry Rats were perfused with PBS under deep anesthesia followed by ice-cold 4% PFA. Brains were dissected, and then post-fixed for 4 h in 4% PFA.…”

Neurotransmitter release: vacuolar ATPase V0 sector c-subunits in possible gene or cell therapies for Parkinson’s, Alzheimer’s, and psychiatric diseases

“…ATP6V0C plays a central role in H + transport as one part of the multi-subunit complex of ATPase [30][31][32][33][34][35][36][37][38][39][40][41][42][43][44]. Israel and his group initially proposed that medatophore forms a proteinaceous pore by the six transmemebrane c subunits of ATPase [10,11].…”

Gene therapy progress and prospects: Parkinson's disease

“…Various vector systems have been used in recent preclinical studies to deliver different combinations of these enzymes. These include multicistronic lentiviruses simultaneously encoding GCH1, TH and AADC; 20 combinations of AAV vectors separately encoding GCH1 and TH 21 or GCH1, TH and AADC; 22,23 an HSV vector coexpressing AADC and TH. 24 In all cases, coexpression of the enzymes and functional recovery of the experimentally lesioned animals was observed.…”