Validation and noninvasive kinetic modeling of [11C]UCB-J PET imaging in mice

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Dopamine D2 receptor pharmacology : In vitro analyses and in vivo pet imaging

The goals of the present thesis were to study high affinity agonist binding at the D2 dopamine receptor and to explore the role of agonist induced internalization of the receptor in positron emission tomography (PET) imaging studies. For this purpose, we combined in vitro studies of radioligand binding and receptor internalization with in vivo studies in rodents using PET. A pharmacological characterization of the novel PET radioligand MNPA in vitro demonstrated that it is a potent and full agonist at the D2 receptor. In membrane homogenates, MNPA bound to both a high and low affinity site of the D2 receptor, of which the high affinity site was sensitive to guanosine triphosphate. In intact cells, however, MNPA bound to only one site of low affinity. As a typical agonist at G-protein coupled receptors, MNPA also induced receptor internalization, which was further augmented by arrestin proteins. [11C]MNPA was evaluated as an agonist radioligand and used to measure the baseline occupancy of the D2 dopamine receptors by dopamine. In vivo, [11C]MNPA was a D2 selective radioligand. Comparison of [11C]MNPA binding in dopamine depleted rats and control rats suggested that about 50% of the D2 receptors are occupied by dopamine at baseline. To study high affinity agonist binding in vivo, we compared the binding of [11C]MNPA in control mice to that of dopamine-beta-hydroxylase (DBH) knockout mice, which have been reported to exhibit an increased percentage of D2 receptors in the high affinity state. The in vivo studies were accompanied by in vitro binding experiments in striatal membrane homogenates. We did not observe any differences in [11C]MNPA BPND between DBH knockout and control mice. Nor did we find any differences in density or percentage of D2 receptors in high affinity state with in vitro binding experiments. Combined, this study suggests that DBH knockout mice have normal densities of D2 receptors in the high affinity state. Agonist induced D2 receptor internalization was evaluated using immunohistochemistry in striatal tissue slices from arrestin3 knockout and wild-type mice. Both dopamine and MNPA induced D2 receptor internalization in wild-type tissue but not in knockout tissue. These results demonstrated that D2 receptor internalization is mediated by arrestin3 and that the arrestin3 knockout mice can be used to study D2 receptor internalization in vivo. To determine whether the prolonged decrease of radioligand binding after amphetamine is caused by receptor internalization, we imaged wild-type and arrestin3 knockout mice, which are incapable of internalizing D2 receptors. The mice were imaged with both the D2 agonist [11C]MNPA and the D2 antagonist [18F]fallypride. The effect of amphetamine on radioligand binding was examined at two time points; 30 min and 4 hours post-amphetamine. At 30 min, [11C]MNPA showed greater displacement than [18F]fallypride, but each radioligand gave similar displacement in knockout and wild-type mice. At 4 hours, the binding of both radioligands returned to baseline in knockout mice but remained decreased in wild-type mice. Our results suggest that the prolonged decrease of radioligand binding after amphetamine is primarily due to D2 receptor internalization rather than dopamine displacement. In conclusion, the present thesis demonstrates that MNPA is a potent and full agonist at the D2 dopamine receptor and binds in vitro to both a high and low affinity state of the D2 receptor. We also demonstrated that arrestin3 mediates internalization of the D2 dopamine receptor and that the prolonged in vivo decrease of radioligand binding after amphetamine is likely due to receptor internalization rather than dopamine displacement

Longitudinal preclinical evaluation of the novel radioligand [\ub9\ub9C]CHDI-626 for PET imaging of mutant huntingtin aggregates in Huntington's disease

PURPOSE: As several therapies aimed at lowering mutant huntingtin (mHTT) brain levels in Huntington’s disease (HD) are currently being investigated, noninvasive positron emission tomography (PET) imaging of mHTT could be utilized to directly evaluate therapeutic efficacy and monitor disease progression. Here we characterized and longitudinally assessed the novel radioligand [(11)C]CHDI-626 for mHTT PET imaging in the zQ175DN mouse model of HD. METHODS: After evaluating radiometabolites and radioligand kinetics, we conducted longitudinal dynamic PET imaging at 3, 6, 9, and 13 months of age (M) in wild-type (WT, n = 17) and heterozygous (HET, n = 23) zQ175DN mice. Statistical analysis was performed to evaluate temporal and genotypic differences. Cross-sectional cohorts at each longitudinal time point were included for post-mortem [(3)H]CHDI-626 autoradiography. RESULTS: Despite fast metabolism and kinetics, the radioligand was suitable for PET imaging of mHTT. Longitudinal quantification could discriminate between genotypes already at premanifest stage (3 M), showing an age-associated increase in signal in HET mice in parallel with mHTT aggregate load progression, as supported by the post-mortem [(3)H]CHDI-626 autoradiography. CONCLUSION: With clinical evaluation underway, [(11)C]CHDI-626 PET imaging appears to be a suitable preclinical candidate marker to monitor natural HD progression and for the evaluation of mHTT-lowering therapies. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00259-021-05578-8