Influence of the A2A antagonist KW6002 on dopaminergic system, motor functioning and neuroinflammation in a rodent model of Parkinson's disease

Introduction. Parkinson’s disease (PD) is characterized by a progressive loss of dopaminergic neurons followed by neuroinflammation. While chronic treatments with dopamine replacement drugs such as L-DOPA are effective in ameliorating the symptoms of PD, such treatments are associated with complications such as dyskinesia. A non-dopaminergic treatment targeting adenosine A2A receptors could be promising for the treatment of motor fluctuations and underlying neuroinflammatory processes.
Methods. Rats bearing unilateral intra-striatal 6-hydroxydopamine lesions were treated chronically [20 days] with vehicle [n=8] or KW6002 [3mg/kg, n=8]. Rats were scanned (60-min dynamic) on days 7,8 and 21,22 post-surgery with [11C]PBR28 and [11C]raclopride to measure neuroinflammation and D2R availability respectively. A peak in neuroinflammation was observed with [11C]PBR28-PET on day 7, thus, on that day, 16 other rats were sacrificed for post-mortem analysis. [11C]PBR28 uptake was calculated as a SUVRatio [ipsilateral/contralateral] whereas [11C]raclopride binding potential [BPND] was calculated via SRTM2 , using cerebellum as a reference. BPND values in left and right striatum were calculated from [11C]raclopride scans. Post-mortem analyses were used to quantify microglia (Iba-1 immunofluorescence) and dopaminergic loss (tyrosine hydroxylase [TH] western blot). Motor and asymmetric paw use were evaluated by rotarod and cylinder tests. The generalized estimated equation (GEE) model was applied to longitudinal datasets (PET, behavioral tests), and the independent samples t-test to post-mortem studies. Significance was measured at p<0.05.
Results: Chronic KW6002 treatment significantly decreased the [11C]PBR28 SUVRatio compared with vehicle [vehicle=1.59±0.26, KW6002=1.26±0.06 on day 7; p=0.001 vehicle=1.35±0.21, KW6002=1.19±0.08 on day 21; p=0.045]. KW6002 also significantly decreased the BPND of [11C]raclopride in the left and right-striatum compared with vehicle (vehicle1.80±0.17 and 1.73±0.124; KW6002=1.56±0.194 and 1.50±0.201 on day 8, p=0.03 and 0.049 respectively; vehicle=1.83±0.28 and 1.72±0.24, KW6002=1.56±0.25 and 1.43±0.26 on day 22, p=0.03 and 0.014 respectively]. An 84% increase of DAPI+/Iba-1+ cells was observed in right-striatum [relative to left-striatum] of vehicle-treated compared to KW6002-treated rats on day 22 [p-value=0.007]. Preliminary results of TH immunoblot suggested smaller dopaminergic loss in left -striatum of KW6002 animals compared with vehicle on day 7[p=0.010]. KW6002 caused a significant improvement in rotarod performance [p<0.05] and contralateral forelimb use in the cylinder test on day 4 compared with vehicle [p-<0.05].
Conclusion: The reduced [11C]PBR28 uptake ratio and reduced number of Iba-1+ cells indicate amelioration of neuroinflammation. Reduced [11C]raclopride binding and improved TH levels indicate improved dopamine synthesis. Thus, KW6002 improves the motor symptoms of PD by reducing dopaminergic neuron loss and enhancing dopaminergic functions.