Adenosine A2A Receptor Occupancy by Caffeine After Coffee Intake in Parkinson's Disease

ABSTRACT Background Coffee intake can decrease the risk for Parkinson's disease (PD). Its beneficial effects are allegedly mediated by caffeine through adenosine A2A receptor (A2AR) antagonist action. Objective We aimed to calculate occupancy rates of striatal A2ARs by caffeine after coffee intake in PD. Methods Five patients with PD underwent 11C‐preladenant positron emission tomography scanning at baseline and after intake of coffee containing 129.5 mg (n = 3) or 259 mg (n = 2) of caffeine. Concurrently, serum caffeine levels were measured. Results The mean serum caffeine level (μg/mL) was 0.374 at baseline and increased to 4.48 and 8.92 by 129.5 and 259 mg of caffeine, respectively. The mean occupancy rates of striatal A2ARs by 129.5 and 259 mg of caffeine were 54.2% and 65.1%, respectively. Conclusions A sufficient A2AR occupancy can be obtained by drinking a cup of coffee, which is equivalent to approximately 100 mg of caffeine. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

1 Research Team for Neuroimaging, Tokyo Metropolitan Institute of Gerontology, Tokyo, Japan 2 Department of Neurology, Tokyo Metropolitan Cancer and Infectious Diseases Center Komagome Hospital, Tokyo, Japan 3 School of Allied Health Science, Kitasato University, Sagamihara, Japan 4 Institute of Cyclotron and Drug Discovery Research, Southern Tohoku Research Institute for Neuroscience, Koriyama, Japan 5 Department of Biofunctional Imaging, Fukushima Medical University, Fukushima, Japan AB S T R A CT : Background: Coffee intake can decrease the risk for Parkinson's disease (PD). Its beneficial effects are allegedly mediated by caffeine through adenosine A 2A receptor (A 2A R) antagonist action. Objective: We aimed to calculate occupancy rates of striatal A 2A Rs by caffeine after coffee intake in PD. Methods: Five patients with PD underwent 11 Cpreladenant positron emission tomography scanning at baseline and after intake of coffee containing 129.5 mg (n = 3) or 259 mg (n = 2) of caffeine. Concurrently, serum caffeine levels were measured. Results: The mean serum caffeine level (μg/mL) was 0.374 at baseline and increased to 4.48 and 8.92  caffeine; Parkinson's disease;  Epidemiological studies have consistently demonstrated that coffee intake can decrease the risk for development of Parkinson's disease (PD). 1,2 Because decaffeinated coffee is not protective against PD, 1 caffeine in coffee is believed to be the essential pharmacological factor contributing to its beneficial effects in PD. Caffeine mainly works as a nonselective blocker of all four adenosine receptor subtypes: A 1 (K D = 12 μM), A 2A (K D = 2.4 μM), A 2B (K D = 13 μM), and A 3 (K D = 80 μM). 3 Of these subtypes, adenosine A 2A receptors (A 2A Rs) are believed to underlie most of the beneficial effects of caffeine in PD, 4 although its mechanism is unclear.
A 2A Rs are predominantly distributed in the putamen, caudate, nucleus accumbens, and external globus pallidus, and they interact with dopamine D 2 receptors in the indirect basal ganglia pathway. [5][6][7] Because of these features, A 2A Rs have been recognized as a therapeutic target to modulate motor symptoms in PD. Istradefylline, a selective A 2A R antagonist (K i = 12.4 nM), 8

was then launched in
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Full financial disclosures and author roles may be found in the online version of this article. Published online 9 January 2022 in Wiley Online Library (wileyonlinelibrary.com). DOI: 10.1002/mds.28897 Japan in 2013 as an adjunct to levodopa to alleviate off episodes in PD and was subsequently approved by the US Food and Drug Administration in 2019. 9,10 Currently, once-daily oral administration of istradefylline 20 or 40 mg is recommended. We found that occupancy rates of striatal A 2A Rs after single administration of istradefylline 20 and 40 mg were 39.5% and 52.1%, respectively, 11 and that the corresponding rates after long-term administration of istradefylline 20 and 40 mg increased to 72.1% and 86.5%, respectively. 12 We hypothesized that if the beneficial effects of coffee in PD are mediated by caffeine through A 2A R antagonist action, a substantial amount of A 2A R should be occupied by caffeine after coffee intake, similar to that observed after administration of istradefylline. 11,12 This study aimed to test the hypothesis by calculating occupancy rates of striatal A 2A Rs after coffee intake in patients with PD using 11 C-preladenant positron emission tomography (PET) for measurement of A 2A R availability. Concurrently, the amount of caffeine in coffee was verified, and the serum caffeine levels were measured.

Research Participants
This study was conducted in accordance with the Declaration of Helsinki and approved by the Ethics Committee of the Tokyo Metropolitan Institute of Gerontology (R19-19). Written informed consent was obtained from all five patients with PD (three men and two women) aged 61 to 76 years (Table 1). All patients were taking at least one antiparkinsonian drug other than istradefylline. Two patients regularly consumed at least one cup of coffee per day. Two other patients occasionally drank coffee. The remaining one patient rarely drank coffee. None of the patients had a history of smoking after middle age.

Study Protocol, Coffee Intake, and Serum Caffeine Level
Commercially available "canned coffee" (Suntory Premium Boss; Suntory Holdings Limited, Osaka, Japan) was used for caffeine loading by drinking coffee. One can of coffee (185 g) contained 129.5 mg of caffeine (70 mg/100 g). The five patients with PD were classified into either the low-caffeine-loading group or the highcaffeine-loading group according to their requests. The two patients in the high-caffeine-loading group consumed two cans of coffee containing 259 mg of caffeine. The other three patients in the low-caffeine-loading group consumed one can of coffee containing 129.5 mg of caffeine. To calculate A 2A R occupancy rates by caffeine after coffee intake, each patient underwent a total of two 11 C-preladenant PET scans in two conditions:  Fig. 1A-E, respectively. L-Dopa, levodopa. a Binding potential (BP ND ) was calculated using the volume-of-interest-based method. 11 caffeine restricted (ie, baseline) and caffeine loading. The interval between the two PET scans was less than 3 months. In both caffeine-restricted and caffeine-loading conditions, all patients were instructed to avoid consuming caffeine-containing products such as coffee, tea, energy drinks, or chocolate from the evening before undergoing PET. We also instructed them not to withhold any antiparkinsonian drugs before the PET scan. On the day of the examination, all patients visited the PET center after taking breakfast and their antiparkinsonian drugs. In the caffeine-restricted condition, the PET scan began at about 13:00 after blood samples were collected immediately before the injection of 11 C-preladenant. In the caffeine-loading condition, each patient drank one can or two cans of coffee at about 12:15. The PET scan was then started at about 13:00 immediately after the collection of blood samples.
The collected blood samples were transferred to the Tsukuba Research Institute (BoZo Research Center, Tokyo, Japan), and serum caffeine levels were measured using liquid chromatography mass spectrometry. The pharmacokinetics of caffeine have been established. 13 After oral administration of a single 250-mg dose of caffeine, the peak plasma caffeine level reaches approximately 10 μg/mL in an hour, and the plasma elimination half-lives (t 1/2 ) range from 3 to 7 hours. Therefore, the 11 C-preladenant PET scan in the caffeine-loading condition was performed approximately 45 minutes after coffee intake; as such, serum caffeine level reached a peak during PET scanning.

11
C-preladenant PET scanning and image processing were conducted basically as described previously. 11,12 In brief, after a bolus injection of approximately 500 MBq of 11 C-preladenant, emission data were acquired for 60 minutes. Binding potential (BP ND ) in the whole striatum was calculated to measure A 2A R availability using the Simplified Reference Tissue Model, 14 after the cerebellum was set as a reference region. In addition, BP ND maps were generated using the Simplified Reference Tissue Model 2. 15 The A 2A R occupancy was calculated using the following equation: Occupancy (%) = 100 Â [(BP ND in caffeine restricted) À (BP ND in caffeine loading)]/(BP ND in caffeine restricted). The relationships between A 2A R occupancy and serum caffeine levels (μg/mL) or dose of caffeine (mg) were modeled using the following equation: occupancy (%) = α Â [D/(D + ED 50 )], where α refers to the maximal receptor occupancy, D refers to serum caffeine levels or dose of caffeine, and ED 50 refers to the level resulting in 50% of maximal receptor occupancy. [16][17][18] The two parameters, α and ED 50 , were estimated with a nonlinear regression analysis, using SPSS Statistics version 25 (IBM Corporation, Armonk, NY, USA).

Results
The serum caffeine levels and striatal BP ND values are shown in Table 1. The mean serum caffeine level (μg/mL) was 0.374 in the caffeine-restricted condition (n = 5) and increased to 4.48 and 8.92 in the low-caffeine-loading (129.5 mg: n = 3) and high-caffeine-loading (259 mg: n = 2) conditions, respectively. The mean A 2A R occupancy rates in the low-caffeine and high-caffeine groups were 54.2% and 65.1%, respectively. The changes in the striatal BP ND values after coffee intake are displayed in BP ND maps (Fig. 1A-E).
The relationships between A 2A R occupancy and serum caffeine levels or doses of caffeine are depicted (Fig. 1F,G). The two estimated parameters, maximal receptor occupancy and ED 50 , in the relationship between A 2A R occupancy and serum caffeine levels were 81.1% (standard error [SE], 10.0%) and 2.2 μg/ mL (SE, 1.0 μg/mL), respectively. The corresponding values in the relationship between A 2A R occupancy and dose of caffeine were 81.3% (SE, 9.4%) and 64.7 mg (SE, 27.2 mg), respectively.

Discussion
This study found that the mean occupancy rates of striatal A 2A Rs by 129.5 and 259 mg of caffeine were 54.2% and 65.1%, respectively. Meanwhile, the corresponding rates after single administration of istradefylline 20 and 40 mg are 39.5% and 52.1%, respectively, 11 and those after long-term administration of istradefylline 20 and 40 mg are 72.1% and 86.5%, respectively. 12 These findings suggest that striatal A 2A R occupancy by caffeine after coffee intake is comparable with that by the administration of the approved dose of istradefylline (20-40 mg) and strongly support the hypothesis that a substantial amount of A 2A R is occupied by caffeine after coffee intake. Zhou et al. 19 assessed the suitability of 11 Cpreladenant PET for the quantification of striatal A 2A Rs and calculated occupancy rates of striatal A 2A R by caffeine in conscious monkeys. They demonstrated that occupancy rates after intravenous injections of caffeine at doses of 2.5, 5.0, and 10.0 mg/kg were 64%, 74%, and 81%, respectively. Meanwhile, the average body weight of five patients in our study was 54 kg. Applying this mean value to 129.5 and 259 mg of caffeine, our results restated that occupancy rates of striatal A 2A Rs after administration of 2.4 and 4.8 mg/kg caffeine were 54.2% and 65.1%, respectively. Given the methodological differences, our results from human patients seem to agree well with those carried out by Zhou et al. 19 involving animals. The ED 50 values were estimated to be 2.2 μg/mL for serum caffeine levels and 64.7 mg for doses of caffeine. A cup of coffee generally contains approximately 100 mg of caffeine. Therefore, after drinking a cup of coffee (ie, intake of 100 mg of caffeine), serum caffeine level can exceed its ED 50 value (ie, 2.2 μg/mL). According to the National Coffee Association, USA, the average American coffee drinker drinks about three cups per day, which is equivalent to approximately 300 mg of caffeine per day. Considering the ED 50 (64.7 mg) and t 1/2 (3-7 hours) of caffeine, it is quite possible that at least approximately 50% of striatal A 2A Rs is constantly blocked by caffeine in coffee drinkers. This estimated occupancy rate is novel and is an important finding to help understand the impact of caffeine on health and disease in coffee drinkers. 4,20 Recently, a randomized controlled trial reported that compared with administration of placebo, consumption of caffeine-containing capsules 200 mg twice daily did not improve motor manifestations in PD. 21 This result appears to conflict with epidemiological links between coffee intake and lower PD risk. 1,2 One possible reason for this contradiction is that in the placebo group, daily intake of 92.17 AE 50.30 mg of caffeine was allowed during the trial. 21 Considering the ED 50 (64.7 mg) and t 1/2 (3-7 hours) of caffeine, 92.17 mg of caffeine can occupy a substantial amount of A 2A Rs, and it is possible that daily caffeine intake in the placebo group 21 might already exert some symptomatic effects in PD. Therefore, further studies are required to investigate the symptomatic effects of caffeine in patients with PD who have no or less caffeine consumption. In addition, this study recommends measuring blood caffeine concentration or at least avoiding the consumption of caffeine-containing products when investigating human A 2A Rs.
In conclusion, this study shows that caffeine binds to striatal A 2A Rs in a dose-dependent manner. A sufficient A 2A Rs occupancy can be obtained by drinking one cup of coffee, which is equivalent to approximately 100 mg of caffeine.