STUDY POPULATION
Eleven patients with PD, Hoehn and Yahr (H&Y) stage < 3, were recruited via advertisement on the Swedish Parkinson Foundation website and via two specialist outpatient clinics in Stockholm (Academic Specialist Centre, Karolinska University Hospital). None of the subjects had clinically relevant somatic comorbidities, cognitive decline, history of psychiatric disease, illicit drug abuse or alcoholism, as assessed by a structured interview. Physical examination, electrocardiography, and routine blood tests were normal. One patient had to be excluded from the PET analysis because the cerebellum was partly out of the PET axial field of view. Demographic details are shown in Table 1.
DATA COLLECTION
Activity monitor and disease severity assessment
An activity monitor (Actigraph GT3X+) was worn on the hip for 5—7 days before each PET measurement. Average amount of steps and magnitude of movement per day were measured as a supportive measure of clinical motor stability [18,19]. Only days with minimal 540 minutes wear time were included in the calculation [20]. As measure of disease severity, the Movement Disorder Society Unified Parkinson’s Disease Rating Scale part 3 motor function (MDS-UPDRS-III) was done, including H&Y staging. All MDS-UPDRS-III assessments were performed on the same time of day by the same physician (VSK) in practically defined “OFF” (see below). Symptom duration was defined as the time from reported onset of first motor symptoms.
TABLE 1 Demographic and clinical characteristics of the patients
Subject
|
Age (y) / sex
|
Symptom duration (y)
|
MDS-UPDRS-III
|
H&Y
|
LEDD (mg)*
|
Days
between
PET 1 - 2
|
Avg. steps/day**
|
Avg. magnitude counts/day**
|
|
|
|
PET1
|
PET2
|
|
|
|
PET1
|
PET2
|
PET1
|
PET2
|
1
|
67 / F
|
8
|
16
|
16
|
1
|
425
|
7
|
5133
|
6925
|
296027
|
390733
|
2
|
68 / M
|
7
|
10
|
9
|
1
|
500
|
7
|
11904
|
7254
|
584009
|
369687
|
3
|
56 / F
|
14
|
16
|
21
|
1
|
680
|
7
|
6496
|
4429
|
345169
|
335207
|
4
|
71 / M
|
5
|
18
|
28
|
1
|
560
|
7
|
4361
|
4770
|
261553
|
275948
|
5
|
69 / M
|
8
|
30
|
30
|
2
|
675
|
20
|
5428
|
6854
|
340436
|
403469
|
6
|
54 / M
|
3
|
10
|
7
|
2
|
650
|
28
|
7232
|
6930
|
422193
|
368935
|
7
|
74 / M
|
8
|
25
|
20
|
2
|
400
|
7
|
3976
|
6152
|
267269
|
350235
|
8
|
47 / M
|
2.5
|
3
|
6
|
1
|
320
|
17
|
7084
|
7166
|
501065
|
524408
|
9
|
67 / F
|
5
|
40
|
36
|
2.5
|
300
|
7
|
3091
|
5971
|
248016
|
323042
|
10
|
61 / M
|
2.5
|
16
|
16
|
2
|
150
|
14
|
12020
|
11801
|
676188
|
857889
|
Mean ± SD
|
63.4 ± 8.6
|
6.3 ±
3.5
|
18.2 ± 10.8
|
18.7 ± 10.2
|
|
465 ± 180
|
-
|
-
|
-
|
Median
|
|
|
|
1.5
|
|
7
|
5962
|
6889
|
342803
|
369311
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
*No changes in LEDD has occurred between PET assessments. **Assessed with the activity monitor (Actigraph GT3X+)
MDS-UPDRS-III: Movement Disorder Society Unified Parkinson’s Disease Rating Scale, part 3 motor function (range 0-72). Assessed before PET1 and PET2, respectively. H&Y: Hoehn & Yahr (range 1-5); LEDD: Levodopa equivalent daily dose.
MRI acquisition
Using a 3 Tesla MRI system (General Electric, Discovery MR750), T2-weighted images were acquired to exclude clinically significant pathology, and 3D T1-weighted images were acquired for co-registration with PET and delineation of the regions of interest (ROI). This last sequence has 176 slices of 1 mm thickness, field of view 256 x 256 mm, resolution 1x1x1 mm, inversion time 450 ms, echo time 3.18 ms, repetition time 8.16 ms.
PET acquisition
[18F]FE-PE2I was prepared as previously described [21]. Two 93-minute [18F]FE-PE2I PET measurements were acquired in each subject within an interval of 7–28 days (see PET injection characteristics, Supplementary Table S1). PET measurements were done on the same time of day, around 1:30 pm. Patients were asked to be practically defined ”OFF”, meaning a withdrawal of levodopa-medication for at least 12 hours and other dopaminergic medication for at least 24 hours. Also, abstinence of caffeine 3 hours before PET, nicotine on day of PET, alcohol 48 hours before PET, and cardiovascular training 96 hours before PET were requested. An individually made plaster helmet was used for head fixation in the PET camera.
PET-measurements were acquired with a high-resolution research tomograph (HRRT, Siemens Molecular Imaging) after an intravenous bolus injection of [18F]FE-PE2I. Details can be found in Supplementary Table S1. A 6-minute transmission scan with a Caesium-137 source was obtained for attenuation correction. Due to technical reasons, the transmission scan for one patient could not be acquired on the day of first PET measurement, so the transmission scan acquired before the second PET measurement was used for attenuation correction of the first PET measurement.
List mode PET data were reconstructed into 37 frames (8x10, 5x20, 4x30, 4x60, 4x180, 12x360 seconds) using 3D OP OSEM with 10 iterations and 16 subsets, including modeling of the PSF [22]. Frame-to-frame realignment was performed as previously described [23], with the only difference that the first two minutes instead of the first minute were used as reference frame for PET-realignment.
PET motion correction
Head motion was evaluated by patient observation during data acquisition as well as during image analysis by reviewing the realignment plots and brain time activity curves (TACs). Translation of more than 3 mm on the realignment plots led to additional motion correction using an in-house developed automatic procedure. Description of the method is given in Supplementary Text 1.
IMAGE ANALYSIS
Using SPM12, the T1-weighted 3D MRI sequence was first realigned to the AC-PC-plane (anterior commissure-posterior commissure), after which the PET was realigned and co-registered to the realigned MRI. The following regions of interest were then delineated automatically on the T1-weighted images with Freesurfer version 6.0.0 (http://surfer.nmr.mgh.harvard.edu/): whole striatum (STR), caudate (CAU), putamen (PUT), ventral striatum (VS), and cerebellum. For substantia nigra (SN), the functional molecular template, as created in the research group [9], was used. As exploratory outcome, three functionally subdivided striatal areas [14] were added to the analysis (http://fsl.fmrib.ox.ac.uk/fsl/fslwiki/Atlases/striatumconn). For all regions, regional non-specific binding potentials (BPND) of [18F]FE-PE2I were generated using wavelet-aided parametric imaging (WAPI) [24] with t* = 27 min and cerebellum as reference region.
As described above, for one subject, only one transmission scan could be acquired, which had to be used for attenuation correction of both PETs. This technical issue introduced a bias in BPND due to misaligned attenuation correction in the reference region. See Supplementary Text 2 for analysis and explanation. It was, therefore, decided to exclude the subject for the main analysis and report the results including the outlier as supplementary material. We believe that the test-retest metrics calculated without the outlier are more representative of the study sample.
STATISTICAL ANALYSIS
For statistical analysis, R version 3.4.3, was used with the package relfeas (https://github.com/mathesong/relfeas). [18F]FE-PE2I measurement reproducibility was determined with calculation of repeatability (absolute intrasubject variability, AbsVar; and the minimum detectable difference, MDD) and reliability (intraclass correlation coefficient, ICC), as per recommendation of Weir, Baumgartner, and Matheson [25–27]. Absolute variability was calculated as: (test-retest)/(mean test and retest)*100. For ICC, the two-way random effects, absolute agreement, single rater/measurement was used, corresponding to:
See formula 1 in the supplementary files.
The ICC represents the proportion of the variability not attributable to measurement error. As such, an ICC of 1 indicates perfect measurement reliability with all observed variability being due to true (biological) differences and none to measurement variability (error), while an ICC of 0.5 indicates that the variability is comprised of true differences and measurement error in equal measure. Different interpretations of the ICC exist: as proposed by Portney & Watkins [28] and suggested by Matheson [27], we regard an ICC < 0.5 as low, 0.5—0.75 moderate, 0.75—0.9 good, and > 0.9 excellent. Measurement reliability with an ICC > 0.9 is recommended as a lowest acceptable standard for measurements from which diagnostic decisions are made, ICC > 0.7 for research purposes, with 0.95 and 0.8 considered as adequate, respectively [29].
The agreements between measurements in each region were plotted with the Bland Altman plots. Power plots were generated with the jamovi software (https://www.jamovi.org/). The results of study variables are expressed as mean ± standard deviation unless otherwise stated.