Focal striatal dopamine may potentiate dyskinesias in parkinsonian monkeys

Abstract

Striatal neurons convert L-dopa to dopamine (DA) following gene transfer of aromatic L-amino acid decarboxylase (AADC) via adeno-associated virus (AAV) in parkinsonian monkeys. We investigated whether AAV-AADC could reduce or eliminate L-dopa-induced dyskinesias (LIDs) and side effects in MPTP-treated monkeys. Five monkeys were made parkinsonian by bilateral MPTP lesions. The optimal therapeutic dose of L-dopa was determined using an acute dose response regimen. After 3 weeks of chronic L-dopa treatment, AAV-AADC or control vector was bilaterally injected into the striatum. Animals were assessed for 6 months with the same L-dopa dosing as presurgery as well as chronic oral L-dopa treatment. Presurgery LID was observed at doses greater than 5 mg/kg. The AAV-AADC-treated animals displayed an average 7.3-fold decrease in the therapeutic dose of L-dopa throughout the 6-month follow-up period. Only AAV-AADC-treated monkeys were susceptible to dyskinesias even at sub-clinical doses. Immunohistochemical analysis revealed well-delineated foci of AADC within the striatum. These results suggest that high levels of focal DA were generated in response to L-dopa administration and may be responsible for the exacerbation of dyskinesias. This may be similar to focal dopaminergic activity in PD patients that developed off-drug or "runaway" dyskinesias following fetal mesencephalic grafts.

Fig. 1

Immunostaining and PET imaging of AAV-AADC-treated monkey (RQ 1065).

Fig. 2

Immunostaining and PET imaging of AAV-AADC-treated monkey (RQ 1077).

Fig. 3

Immunostaining and PET imaging of AAV-AADC-treated monkey (TO11). (A, B) Low power photomicrographs of AADC-IR with focal AADC expression. One site in the caudate nucleus and 2 sites in the putamen were targeted bilaterally. Note very low levels of AADC in the striatum with exception of nucleus accumbens and focal regions (as indicated by arrows) of AADC gene transfer 6 months following AAV-AADC administration. (C–F) High power photomicrographs of the AADC-transduced regions in the caudate as shown in panels A and B. AADC expression is restricted to medium spiny neurons (expressing D1 and D2 receptors). Also note the very short distance of AADC-IR fibers extending from the focus of AADC transduction. Local DA production by AADC-expressing striatal neurons in response to l-dopa administration most likely represents focal DA production. l-dopa administration in these monkeys resulted in significant induction of dyskinesias (see clinical rating figures). (G–H) 6-[¹⁸F]fluoro-l-m-tyrosine (FMT) positron emission tomography (PET) before and approximately 4 weeks after AAV-AADC gene transfer. Three brain levels (anterior to posterior) are shown for baseline and post-gene transfer. Low levels of FMT activity were seen following AADC gene transfer due to limited expression of AADC in the striatum. Scale bar: 5 mm.

Fig. 4

PET imaging-percent change from baseline. Quantification of AADC activity. Data represent %change in FMT striatal uptake ratios between baseline and post-AAV scan for each monkey in right and left hemispheres. The two AAV-NULL animals showed little change in PET ratios from baseline. Two of the AAV-AADC-treated animals showed small bilateral increases and the third animal showed a more prominent increase bilaterally.

Fig. 5

Response of AAV-AADC-treated monkeys to acute l-dopa. (A) Clinical responses to acute administration of l-dopa (0, 5, 10, 20 mg/kg) were rated at 45 min. A 3-day wash-out was implemented between each dose. Improved l-dopa response was seen by reduction of CRS scores after AADC gene transfer. However, it was associated with significantly elevated dyskinesias as shown in panel B. (B) The animals received weekly injections of increasing doses of l-dopa: 1.5, 2.5, 5, 10, 20, 30 mg/ml combined with the decarboxylase inhibitor benserazide (2 mg/kg) and dyskinesias were scored. The dyskinesia scores of animals treated with AAV-AADC were averaged for each l-dopa dose before and after surgery and are reported as mean ± SEM.

Fig. 6

Response of AAV-AADC-treated and control monkeys to chronic l-dopa. Sinemet CR was given BID for 5 weeks. The therapeutic dose of l-dopa was determined for each monkey and adjusted to one of the following doses: 50/200 mg, 25/100 mg, 12.5/50 mg (l-dopa/Carbidopa) to avoid or minimize dyskinesias or other adverse effects of l-dopa administration. (A and B) Clinical improvement in response to l-dopa administration was achieved in 2 NHP treated with AAV-NULL without inducing dyskinesias. However, the AADC-treated group developed dyskinesias in the first 5 days of treatment that persisted despite lowering the dose of Sinemet. Both groups had some degree of other adverse effects of l-dopa treatment such as hyperactivity, aggression, occasional apparent hallucinations and self-mutilation. (C and D) Eight monkeys, in which diffuse transgene expression (NULL or AADC; n = 4 in each group) was achieved, were subjected to a similar regimen of Sinemet CR for 7 weeks. No dyskinesias were observed in response to either acute or chronic l-dopa.