Clinical trial NCT05405244
Conditions: Overweight and Obesity Eating Behavior
Keywords: bromocriptine dopamine agonist TaqIA polymorphism eating behavior food intake overweight obesity
Sponsor: University of North Carolina, Chapel Hill
Start date: 2017-09-19; Completion date: 2019-09-19
Enrollment: 55 participants; Allocation: RANDOMIZED; Intervention model: CROSSOVER

Brief summary:
The current project applies an integrative three-prong approach to investigate the potential of the dopamine D2 receptor (DRD2) agonist bromocriptine to: 1) increase homeostatic satiation signaling, 2) alter neural circuitry to reduce hedonically motivated food intake, and 3) examines a genetic predisposition that may markedly impact the effectiveness of this medication in those at high risk for T2DM.

Detailed description:
Twenty-nine million Americans have diagnosed type 2 diabetes (T2DM), with fewer than half able to meet treatment goals and considerably more are at risk for development of T2DM. Implementation of healthy eating behavior has been identified as a barrier to T2DM treatment and efficacy. The determinants of eating behavior and weight regulation involve a complex interaction among individual-level homeostatic, hedonic, and genetic systems, and the external food environment. The high prevalence of obesity and T2DM suggests hedonic motivation to consume food overrides homeostatic satiation signaling, resulting in excess food intake. Elevated intake increases body mass and promotes T2DM incidence via dysregulation of glucagon-like peptide 1 (GLP-1), amylin, and adiponectin, which in turn can negatively impact T2DM treatment options.

Use of a pharmaceutical, such as bromocriptine, to aid in behavioral change is a novel method for treating and ameliorating T2DM and warrants investigation given that previous work has shown reward response to food images mediates T2DM control. Use of functional magnetic resonance imaging (fMRI) techniques to predict and evaluate hedonically-motivated eating behavior can be used to measure sensitivity to reward, and the role it plays in developing obesity, and is therefore an excellent tool to examine the associations among bromocriptine, satiety hormones, reward sensitivity and eating behavior.

Moreover, since 20-35% of the population carries the DRD2 TaqIA A1 allele, and 65% of the population is overweight or obese and at high risk for T2DM development or currently diagnosed, as much as 23% of the population may greatly benefit from dopamine agonist treatment. Despite the possibility that bromocriptine may have robust impact on T2DM treatment or as prevention therapy in those that are genetically predisposed, few data are available that directly examine the three systems (homeostatic, hedonic, genetic) available to assess whether a genetically-informed, personalized T2DM treatment is viable.

Arms: arm 1: During the first intervention visit, participants receive a single dose of 1.6mg of bromocriptine (2 0.8mg capsules). Following a 2-week washout period, participants return for the second intervention visit, where they receive 2 capsules of placebo (sugar free calcium supplement) matched in shape (circle) and color (white) to bromocriptine. Both bromocriptine and placebo are administered orally. arm 2: During the first intervention visit, participants receive 2 capsules of placebo (sugar free calcium supplement). Following a 2-week washout period, participants return for the second intervention visit, where they receive a single dose of 1.6mg of bromocriptine (2 0.8mg capsules). Both bromocriptine and placebo are administered orally.
Interventions: intervention 1: Placebo intervention 2: Bromocriptine-QR
Intervention descriptions: intervention 1: 2 capsules, orally administered once intervention 2: 1.6mg (2 0.8mg capsules), orally administered once

Protocol documents:
Prot_000.pdf: 
Cover Page 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Official Title: A Multimodel Examination of Bromocriptine on 
Homeostatic and Hedonic Mechanisms of Food Intake in Individuals 
at High Risk for Type 2 Diabetes 
 
NCT Number: NCT05405244 
 
Document Date: June 6, 2022 
 


Study Protocol 
 
A. Study Rationale 
 
Twenty-nine million Americans suffered from type 2 diabetes (T2DM) in 2012 (2014 National 
Diabetes Report), with fewer than half able to meet treatment goals and considerably more are 
at risk for development of T2DM (1). Implementation of healthy eating behavior has been 
identified as a barrier to T2DM treatment and efficacy (2). The determinants of eating behavior 
and weight regulation involve a complex interaction among individual-level homeostatic, 
hedonic, and genetic systems, and the external food environment (3,4). The high prevalence of 
obesity and T2DM suggests hedonic motivation to consume food overrides homeostatic 
satiation signaling, resulting in excess food intake. Elevated intake increases body mass and 
promotes T2DM incidence via dysregulation of GLP1, amylin, and adiponectin, which in turn can 
negatively impact T2DM treatment options. Use of a pharmaceutical, such as bromocriptine, to 
aid in behavioral change is a novel method for treating and ameliorating T2DM and warrants 
investigation given that previous work has shown reward response to food images mediates 
T2DM control (5). Use of fMRI techniques to predict and evaluate hedonically-motivated eating 
behavior can be used to measure sensitivity to reward, and the role it plays in developing 
obesity (6,7), and is therefore an excellent tool to examine the associations among 
bromocriptine, satiety hormones, reward sensitivity and eating behavior. Moreover, since 20 
35% of the population carries the DRD2 TaqIA A1 allele (8), and 65% of the population is 
overweight or obese and at high risk for T2DM development or currently diagnosed, as much as 
23% of the population may greatly benefit from dopamine agonist treatment (9). Despite the 
possibility that bromocriptine may have robust impact on T2DM treatment or as prevention 
therapy in those that are genetically predisposed, few data are available that directly examine 
the three systems (homeostatic, hedonic, genetic) available to assess whether a genetically-
informed, personalized T2DM treatment is viable. 
 
B. Study Objectives 
 
Aim 1: To test the hypothesis that acute bromocriptine administration will alter neural and 
perceptual hedonic drivers of eating behavior (Visual analog scales), both contributing to a 
decrease in palatable food intake in those at high risk for T2DM. We expect that bromocriptine 
will increase BOLD response during palatable food intake in dopaminergic brain regions thought 
to encode reward and increase perceptual hedonic food ratings. We also expect that 
bromocriptine will result in decreased ad lib palatable food intake.  
 
Aim 2: Test whether the A1 variant of the DRD2 TaqIA polymorphism, which is associated with 
reduced dopamine-D2 receptor density and dopamine signaling, moderates the impact of 
bromocriptine on neural and perceptual hedonic drivers of eating behavior. By testing the 
bromocriptine by TaqIA allele status interaction, we expect that the effects described in aim 1 
will be pronounced in those with at least 1 A1 variant of the TaqIA polymorphism. 
 
Exploratory Aim 1: To test the hypothesis that acute bromocriptine administration will alter the 
homeostatic mechanisms of satiation (meal termination). We expect bromocriptine to modify 
endocrine-mediated satiation signaling via suppressed prolactin, augmented adiponectin, 
amylin, glucagon like peptide-1 (GLP1), and decrease postprandial plasma glucose and insulin. 
 
 
 
 


C. Study Design 
 
This within-subject pilot study will look at 180 overweight/obese (25 < BMI < 35) adults ages 18-
35 years of age. All participants will be screened at the first visit for 50 eligible participants who 
will complete the other visits. 25 participants will have the DRD2 TaqIA A1 allele and 25 will 
have the A2/A2 variant. The study will be completed over two testing visits with a two-week 
washout period in between. Following published reports of the effect of acute administration of 
bromocriptine (10), participants will be given a 1.6mg dose of either bromocriptine-QR 
(Cycloset) or a placebo after a 6-8 hour fast, paralleling the recommended dosage and time of 
administration (11). Order of agonist/placebo will be counterbalanced across the two visits. At 
both visits, the participant will have a fasting blood draw, anthropometric measurements, 
complete visual analogue scales (VAS) for hunger, and satiety and surveys. Once 
bromocriptine/placebo is metabolically active (~60 mins) subjects will undergo a functional MRI 
with our milkshake receipt paradigm and a resting state MRI to assess functional connectivity. 
The Biomedical Research Imaging Center (BRIC) is located on the UNC-CH campus and 7-10 
min walk from our laboratory (Neuropsychology of Ingestive Behavior Laboratory). Post-MRI, 
subjects will consume a fixed portion of milkshake (12 fl oz) and perform VAS hedonic ratings; 
will complete a test of reaction time and behavioral impulsivity, and two postprandial blood 
draws (30 & 60mins post milkshake ingestion). Finally, subjects will perform perceptual hedonic 
assessments of palatable foods and given the opportunity for ad lib consumption. 
 
D. Study Duration, Enrollment and Number of Subjects 
 
For each participant, the study will include three visits on three separate days. 
Total study duration will be approximately two years.  
We will recruit and screen 180, 18-35-year-old overweight/obese individuals in order to enroll 50 
participants with and without the presence of the DRD2 TaqIA A1 allele, and at high risk for 
T2DM by virtue of HbA1c. The n=42 represents the number of participants who will provide 
usable data and accounts for possible attrition and negative side effects of the bromocriptine 
(based on previous repeated measures fMRI studies and reports of bromocriptine, expected 
attrition is 4%-7%). 
 
E. Study Population 
 
50 overweight/obese (25 < BMI < 35) adults ages 18-35 years of age. We will screen for 
enrollment such that 25 participants will have the DRD2 TaqIA A1 allele and 25 will have the 
A2/A2 variant. We will exclude participants who report current: a) contraindications of fMRI (e.g., 
metal implants, braces, or serious medical problems [e.g., known diabetes, CVD, recent 
stroke]), b) any previous habitual use of cigarettes or illicit drugs, or c) current major psychiatric 
disorders (depression, bipolar disorder, panic disorder, generalized anxiety disorder), d) 
pregnancy or breastfeeding e) current weight loss dieting and/or weight fluctuations great than 
10 lbs in the previous 6 weeks. f) do not consume dairy e) allergy to bromocriptine, dairy, and 
nuts. Diabetes status will be assessed via self-report. 
 
F. Study Procedures 
 
1. Screening Visit 
At the screening visit, following consent, the potential participant will be weighed and height will 
be measured to ensure they are overweight (25 < BMI < 35). A brief screener for identifying MRI 
contraindications will be administered before inviting subjects to join the MRI portion of the 


study, as done in prior fMRI studies. If participants satisfy inclusion and exclusion criteria during 
the screen, they will be scheduled for the testing assessments. Female participants will be 
asked to self-report the date of their last menstruation to ensure they are in the follicular phase 
and are not pregnant (pregnancy test will be offered). Saliva (genotyping) and blood glucose 
values (mg/dL) will be collected from consented participants at the initial screening appointment. 
(30 minutes) 
 
2. Testing Visits – Outline 
• 
At the Neuropsychology of Ingestive Behavior Lab 
Baseline: Fasting blood draw, fasting drug or placebo administration 
Baseline-60 minutes: Visual analogues scales (VAS) hunger and satiety, height and weight 
measurement, demographic and psychological surveys, waiting for drug to become active 
60 minutes: Walk to Biomedical Research Imaging Center 
• 
At the Biomedical Research Imaging Center 
90 minutes - 150 minutes: VAS hunger and satiety, MRI assessment, taste administration, 
resting state, anatomical scans 
• 
At the Neuropsychology of Ingestive Behavior Lab 
150 minutes-180 minutes: VAS hunger and satiety, fixed milkshake consumption 
180 minutes: Blood draw 
180 minutes-210 minutes: Behavioral impulsivity, surveys 
210 minutes: Blood draw 
210-240 minutes: VAS ratings, hedonic ratings, ad libitum food intake 
 
The testing visit will last approximately 4 hours. 
Testing visit occurs twice for each participant. 
 
 
G. Study Evaluations and Measurements 
 
Bromocriptine/placebo administration 
Participants will be given either bromocriptine (1.6mg Cycloset) or placebo at each testing visit. 
Order of bromocriptine/placebo will be counterbalanced across the participants. The researcher 
and participant will be blinded to the treatment given. Dr. Burger, Dr. Klett, and IDS will have the 
randomization key. The research assistant will pick up the Cycloset or placebo from IDS. He or 
she will be blinded to the contents (whether placebo or Cycloset) and will give the participant the 
capsule to take orally. To allow the drug to reach maximum plasma concentration, participants 
will be scanned approximately 90 minutes following drug administration. The placebo will be 
developed by IDS and will come in a capsule which will look identical to the capsule the 
Cycloset is in. 
  
Blood Draws 
Each blood draw will occur in either the antecubital vein in the arm or in the cephalic vein in the 
hand. 6 mL of blood will be drawn three times. For each visit, the first blood draw (fasting) will 
occur at the start of the visit and the second and third will occur 30mins and 60mins after the 
fixed milkshake portion (after fMRI scan; see Fig 1). This will allow us to assess fasted 


circulating hormone levels and gives us the ability to examine satiation hormone fluctuation over 
two time points that encompass the entirety of the previously characterized GLP1 peak in 
serum. A trained phlebotomist will perform the blood draws. The phlebotomist will try no more 
than 3 times unless the participant consents to further attempt. Blood samples for satiety 
hormones will be assayed using Bio-Plex Pro Human Diabetes 7-plex Assay (Bio-rad, Hercules, 
CA). This kit analyzes adiponectin, amylin, and GLP1. Additionally, a custom Human Cancer 
Panel 1 with only prolactin will be used to analyze prolactin concentrations (Bio-rad, Hercules, 
CA) using a Millipore Luminex (Darmstadt, Germany). All samples will be kept at -80° until 
processed. Our team is well versed in blood processing procedures and Luminex operation. 
  
Height and Weight 
Height and weight will be measured without shoes or heavy clothing. 
  
Visual Analogue Scale 
Visual Analogue Scales will be used to assess hunger, fullness, thirst, and nausea periodically 
throughout the testing visits. These will be distributed via iPad on Qualtrics software. 
  
Neuroimaging Paradigms 
Participants will complete 2 functional MRI paradigms: milkshake receipt paradigm and resting 
state paradigm, in addition to an anatomical scan. These paradigms examine different aspects 
of neural functioning. First, the milkshake receipt paradigm allows us to examine how the brain 
responds during exposure to a primary reward (receipt of highly palatable taste) and a tasteless 
control. Second, the steady state paradigm will allow for functional connectivity analyses to 
examine reward and behavioral differences in the brain at ‘rest’. 
  
Milkshake receipt fMRI paradigm 
The paradigm is controlled by in-house Python scripts. Each trial in the paradigm starts with the 
presentation of a cue (duration: 1s) signaling the impending delivery of either 3 mL of highly 
palatable milkshake or a control water solution over a period of 6s. Taste delivery is followed by 
a 1s wait period and 2s rinse (tasteless solution). The next trial begins after a 1-9s jitter. Order 
of milkshake and water trials is pseudo randomized. In total, there are 32 trials per run for a total 
of 2 8-minute runs. This paradigm allows us to assess brain response to a receipt of primary 
reward (milkshake) and control (water solution) as well as in vivo neuroadaptations over 
repeated exposures.  
 
Resting state functional connectivity 
We will use resting state fMRI data for functional connectivity analyses to provide data about 
brain functioning and connectivity. Functional connectivity is assessed by examining the 
correlation of fluctuations of BOLD signals in different regions of the “resting” brain and is 
thought to provide a measure of its functional organization Steady-state fMRI data will be 
acquired in one run of 5 minutes; participants will be asked to remain still and relaxed and stay 
awake during the scanning. This paradigm allows us to assess the brain’s functional 
connectivity. We hypothesize that connectivity from the hypothalamus will be significantly 
altered by bromocriptine. 
Total scan time will be approximately 50 minutes. 
  


Behavioral impulsivity task 
We will use a basic stop-signal task that assesses behavioral inhibitory control in response to 
palatable food images, adapted from Batterink and colleagues (12) and in current use in our lab. 
This task requires subjects to respond to “go” signals and occasionally inhibit responses to “no-
go” signals. It measures the ability to inhibit a pre-potent tendency to respond. The task consists 
of 80 trials, evenly split between left-button responses (the letter A) and right-button responses 
(the letter Z). 16 total stop trials are presented (the stop signal is the letter X). The stop signal 
delay begins at 200ms, and increments by one screen refresh period depending on trial 
performance. The number of responses to stop signal stimuli (commission errors) will be used 
to provide a behavioral measure of inhibition failure. 
  
Ad lib taste test 
After the fMRI scan and fixed consumption of milkshake, participants undergo the postprandial 
blood draw and perform a taste test which includes perceptual hedonic ratings of a variety of 
foods and an ad lib intake challenge. Participants first perform a taste test of 1g of each of the 4 
foods (M&Ms, Skittles, Doritos, cheddar popcorn). These foods were selected as they are 
generally considered highly palatable and represent a variety of flavors, mouth feels and 
nutrient contents (e.g., sugar, fat, salt). Participants complete perceptual hedonic ratings of the 
pleasantness, sweetness, taste intensity, and desire to consume on generalized, labeled 
magnitude scales of each of the foods. After the taste test, participants will be told that they are 
free to eat as many snacks as they like, as we have to discard them after each participant (they 
will not be allowed to have backpacks, purses, or bags, so they do not take snacks from the 
room). Participants will be left alone during the tasting to minimize demand characteristics. Food 
will be pre- and post-weighed to determine ad lib intake. Total caloric intake will be calculated 
and translated to % of calorie needs using gender-specific formulas (NAS, 2005). We currently 
use these techniques (NORC5-34387, HHSN275201300015C). 
  
Eating Attitudes 
The Dutch Eating Behavior Questionnaire (DEBQ; 13) is a 46-item questionnaire with three 
subscales measuring restrained, external, and emotional eating. Responses are recorded on a 
5-point Likert scale. These three scales demonstrated strong internal consistency (Cronbach’s 
alpha = 0.80 – 0.95), interrelationships between scales, and factorial validity. 
  
Food Craving 
The Food Craving Inventory (FCI) will assess the degree of craving for high- and low-fat/sugar 
foods (14). This scale has shown internal consistency (α= .93), 2-week test-retest reliability (r = 
.86), and sensitivity to detecting change. 
  
Self-report Impulsivity 
The Barratt Impulsivity Scale (BIS-15) is a 15-item self-report assessment that will measure 
attentional, motor, and non-planning impulsivity (15). This scale has shown internal consistency 
(α=.81), 2-week test-retest reliability (r =.88), and discriminates between psychiatric patients and 
controls (56). 
  
Acute and Habitual Dietary Intake 
Participants will complete our adapted version of the Block Food Frequency Questionnaire 
(Block FFQ) which inquiries about the frequency of consumption of 60 specific foods over the 


past 2 weeks (16) to measure habitual dietary intake. Food frequency questionnaires are the 
most practical and economical method for collecting dietary intake data in large studies (17). 
Block FFQ values correlated (r = .57) with 4-day food record estimates for energy and most 
nutrients (18). We have successfully used this version in previous studies (NORC5-34387, 
HHSN275201300015C) and published results. 
  
Additional Surveys 
We will administer the Short-Form International Physical Activity Questionnaire (IPAQ), which 
queries about physical activity over the previous days and has shown strong reliability (r = .80; 
(19)). Participants will also complete a modified version of the Pittsburgh Sleep Quality Index, 
which will ask about sleep quality over the past week (20), which we can determine the previous 
night’s sleep. Lastly, participants will complete the Edinburgh Handedness Inventory, which is a 
continuous measure of hand dominance (21). 
  
All Surveys will be administered via an iPad using Qualtrics software and applications. 
 
H. Study Intervention  
 
- 
Bromocriptine-QR cycloset; 1.6mg; once; oral 
- 
Placebo; 2 capsules; once; oral 
Participants will be administered two placebo pills at one testing visit and a two 0.8mg dose of 
bromocriptine at the other visit. Bromocriptine dose agrees with optimal dose according to 
clinical trials (22,10). Adverse effects related to bromocriptine include nausea, dizziness, 
fatigue, headache, diarrhea, and constipation (11). However, adverse events (AEs) are 
generally not severe; according to Scranton & Cincotta, 17% subjects on bromocriptine reported 
adverse drugs experiences as severe, while 14% on placebo reported AEs as severe (22). 
Moreover, the most commonly reported adverse event, nausea, was not reported as severe 
(22). To prevent any potential adverse interactions with bromocriptine, participants will be asked 
to abstain from any drugs for 12 hours before the assessment including over the counter 
pharmaceuticals such as (ex. Advil, aspirin, Benadryl), and for the remainder of the day after the 
session. Participants will be asked to avoid consuming alcohol for 24 hours after bromocriptine 
administration to prevent increased risk of drowsiness. Bromocriptine may decrease the 
effectiveness of oral contraceptives, female participants taking oral contraceptives will be asked 
to refrain from intercourse for 24 hours or to use an additional form of birth control. Because the 
half-life for bromocriptine is 6 hours after 24 hours the amount of active drug in the participant’s 
system will have decreased 98%. 
 
I. 
Study Intervention Administration 
 
This is a within subjects designed experiment, so all subjects will receive the Cycloset and 
placebo. The administration will be blinded to subject and the research. Dr. Burger will oversee 
the blinding procedure. The order of the drug and placebo will be counterbalanced across the 
two visits, and Dr. Burger will see to the drug/placebo administration schedule. Because the 
screening visit is before the testing visit and randomization we are confident in the participant's 
eligibility. 
 
J. Safety Management 
It is possible that the questionnaires may cause distress or embarrassment to the participants. 
One potential risk to participants is that it may be distressing to disclose information about 


psychiatric difficulties. In our estimation, there is a low risk of this possibility and the effects 
would probably be short-lived. We have conducted several thousand of these types of 
interviews with no adverse events. We will take steps to minimize this risk by informing them 
that their data will be confidential. Additionally, it is possible someone maybe embarrassed to be 
weighed. The research staff will be trained to be highly professional for the entire study visit. 
Data will be obtained from participants through surveys, taste assessments, height and weight 
measurements, fMRI scans, blood draws, and saliva. There is a slight risk that these research 
records might be obtained by persons not authorized to do so. There is also a risk that 
participants may not understand the limits of confidentiality (i.e. that we will not keep certain 
information confidential, such as spontaneous disclosures on their part regarding suicidal 
ideation and child abuse [no measures inquire about sensitive topics such as these]). Data for 
all participants will be kept strictly confidential, except as mandated by law. All research files will 
be kept in locked file cabinets in a locked room in a locked building. Participants will be 
assigned a numerical code for identification in the files. Names and other identifiers will be kept 
in separate locked files and will never be recorded on any data forms. Statistical analyses will 
be performed on aggregate-level data; participants are never individually named. All entered 
data will be kept on the password protected, secured computers or networks at the research 
site. These data will be accessible only to research staff, using confidential usernames and 
passwords. All staff on this project will receive training in the ethical conduct of research with 
human participants prior to contact with data or research participants. Participants will be 
informed in detail about the limits of confidentiality, including disclosure of intentions to inflict 
harm to self or others and the possibility that we might break confidentiality to report suspected 
child or elder abuse. All participants will be told that participation is voluntary and that they may 
terminate their involvement at any time without any consequences. If, during any assessment, 
the project coordinator determines that a participant has developed a serious health problem, 
efforts will be made to help the participant access appropriate treatment as quickly as possible. 
In addition, any scans that suggest possible problems are sent to a radiologist for a consult. In 
the event of a potential problem, Dr. Burger will meet with the participant and the radiologist to 
discuss the findings, per protocol from the Biomedical Research Imaging Center. 
 
K. Data Collection and Management 
 
All paper data will be stored in a locked filing cabinet in a locked room. All digital data will be 
stored on servers which require passwords to access. Data will be saved as confidential, coded 
files. All blood data and genetic data will be de-identified and will be kept in secure freezers until 
it is assayed. 
 
Only authorized project staff will have access to the information gathered for this project. We will 
follow established data handling procedures as follows: (a) data will be reported in aggregate 
form only; (b) all data will be kept in locked file cabinets and/or in password-protected computer 
files; and (c) all personnel interacting with participants and having access to data are trained in 
confidentiality procedures. 
 
Any paper documents will be stored in locked files, behind locked doors at the laboratory on 
campus at UNC-CH. The key that links participant names to the identification numbers will be 
kept separate from the data, also in a locked file. 
 
fMRI data collected at the BRIC is sent over a secured server (PACS), this data is attached to 
coded numbers. fMRI data is accessed through this password protected served and transferred 
to the lab's protected databases. Unprotected personal information of subjects is never sent 
over the web. 


 
L. Recruitment Strategy 
 
Recruitment posters will be placed on the UNC campus, in community centers, and restaurants 
in local Chapel Hill/Durham area. Additionally, we will use the ‘UNC-Chapel Hill Biomedical 
Research Imaging Center (BRIC) Database’ (UNC-CH IRB #13-0959) and the ‘Join the 
Conquest’ (UNC-CH IRB #13-1768) recruitment databases. These and similar approaches have 
been successfully used for recruitment in this sampling frame (e.g., R01DK092468, NORC5-
34387). Those interested in participating will be directed to the screening website. Information 
needed to determine participant eligibility will be obtained via web, phone or in person (see 
attached prescreening material). The subjects will be contacted by research staff via the 
preferred method of the subject (phone, email, in person) to provide the consent for review and 
schedule the first assessment (if applicable). 
 
M. Consent Process 
 
If the potential participant is interested in the study they will be scheduled for a screening visit 
and a consent form will be mailed or emailed to them. At the screening visit, before an 
assessments, the consent form will be reviewed, all answers regarding study expectations 
will be answered, and consent will be confirmed by trained study staff (listed in the personnel 
section of the application). Consenting will be performed in a private room. 
 
 
 
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http://informahealthcare.com/doi/full/10.1517/14656560903501544 

SAP_001.pdf: 
Cover Page 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Official Title: A Multimodel Examination of Bromocriptine on 
Homeostatic and Hedonic Mechanisms of Food Intake in Individuals 
at High Risk for Type 2 Diabetes 
 
NCT Number: NCT05405244 
 
Document Date: June 9, 2022 
 
 
 


Statistical Analysis Plan  
 
 
A. Sample Size and Power 
We will recruit and screen 180, 18-35 year old overweight/obese individuals in order to enroll 50 
participants with and without the presence of the DRD2 TaqIA A1 allele, and at high risk for 
T2DM by virtue of HbA1c. The n=42 represents the number of participants who will provide 
usable data and accounts for possible attrition and negative side effects of the bromocriptine 
(based on previous repeated measures fMRI studies and reports of bromocriptine, expected 
attrition is 4%-7%). In evaluating our inclusion/exclusion criteria (see below) conservatively we 
anticipate 180 participants per year will be eligible for the present investigation. Of those, based 
on previous work, 36 are expected to have one A1 variant of the TaqIA polymorphism and 144 
to have A2/A2 (per each recruitment year). Therefore, over the three-year duration of the 
present study, we will have ample opportunity to meet our recruitment targets (Y1: n= 17; Y2: n= 
18; Y3: n= 15). Power analyses were conducted using fMRIpower (for fMRI contrasts described 
in Aim 1b) and G*Power (for lab measures) to determine sample size based on previous work 
(25). Sample sizes were determined by the between-group (TaqIA allele status; Aim 2) 
difference, which is the least sensitive of the proposed tests. Based on a small to medium sized 
effect (r = .30), we determined that a similarly sized strategy-by-assignment interaction would be 
detectable with 0.8 power with at least n=21 per group, yielding a total sample size of 42. To 
account for the small possibility of complications with blood draws, excessive movement in the 
scanner, side effects from the medication or other protocol deviations we plan on recruiting a 
total of 50 (25 per cell). 
 
 
B. Analysis Plan  
 
1. Primary and Secondary Outcomes 
 
Behavioral data analysis (food intake + hedonic ratings of food). Continuous variables will be 
summarized by descriptive statistics (e.g., mean, SD, range, median), examined via graphical 
analyses (e.g. histogram, Q-Q plot) and analyzed to assess the normality of the data. Data 
transformation of continuous variables will be used as necessary. Model assumptions and 
potential outliers will be analyzed with various modeling diagnostics and plotting techniques. 
Visual analog scale (VAS) measures will be analyzed using PROC MIXED implemented in SAS 
(Version 9.4, Cary, NC). VAS scales for milkshake, snack food pleasure and desire, ad lib 
intake of those foods as well as changes in hunger (fasted to fed) will be entered into 2x2 
repeated measures mixed models including drug-by-gene interactions which addresses Aim 1a 
and Aim 2. Control variables of no interest to be included are BMI, gender, visit number and 
nausea (after scan assessments only). In addition to main effects of the drug and the TaqIA 
allele status, interactions with p < 0.10 will be probed with least squared mean comparisons of a 
priori contrasts of interest including tests of the: 1) impact of TaqIA allele status with 
bromocriptine, 2) impact of TaqIA allele status without bromocriptine 3) impact of bromocriptine 
on those with the TaqIA A2/A2 variant, and 4) impact of bromocriptine on those with at least one 
TaqIA A1 variant. 
 
fMRI data preprocessing: Neuroimaging data will be preprocessed using the fMRIPrep pipeline 
[45]. DICOMS will be converted to the Brain Imaging Data Structure (BIDS file structure) [46], 
then preprocessed using fMRIPrep. fMRIPrep preprocessing includes skull stripping using 
Advanced Normalization Tools (ANTs); susceptibility distortion correction using field maps; 
tissue segmentation using FSL’s Automated Segmentation Tool (FAST); and spatial 


normalization to Montreal Neurological Institute (MNI) 152-Asymmetrical space using ANTs’ 
registration option. Functional data will be co-registered to anatomical data using FSL's FMRIB's 
Linear Image Registration Tool (FLIRT) with boundary-based registration. Functional data will 
additionally be motion corrected using FSL’s MCFLIRT. In FSL, final preprocessing will include 
spatial smoothing using a 5mm full width half maximum isotropic Gaussian kernel, high-pass 
filtering, and adjusting for nuisance regressors, including the 6 motion parameters, the 
derivatives of each, and high motion time points (FD>0.9mm). A functional run for a subject will 
be excluded if >25% of total volumes are flagged as high motion points.      
  
fMRI primary analyses: Neuroimaging analyses will be primarily completed in FSL (FMRIB 
Software Library,www.fmrib.ox.ac.uk/fsl). Within-run (level 1), within-subject (level 2) and group-
level (level 3) analyses will be carried out in FSL's FMRI Expert Analysis Tool (FEAT). At the 1st 
(within-run) level, models will assess brain response to the following contrasts of interest: effect 
of milkshake cue (milkshake cue > h2o cue) and effect of milkshake taste (milkshake taste > 
h2o taste). Inverse contrasts (h2o cue > milkshake cue; h2o taste > milkshake taste) will also be 
assessed. 
At the 2nd level analyses, to test Aims 1 and 2, milkshake cue > h2o cue and milkshake 
taste > h2o taste contrasts will be carried forward to a fixed effects within-subject t-test 
estimating individual-level differences as a function of bromocriptine, i.e., active drug 
(bromocriptine) > placebo. Bilateral striatum will be used as the a priori region of interest (ROI). 
Binary masks of caudate, putamen and ventral striatum will be included in the level 2 within-
subject models to extract parameter estimates for the contrasts of interest only within those 
regions.  Extracted parameter estimates will be averaged across the three regions resulting in a 
single striatal parameter estimate value per contrast per subject.  
For group-level analyses, striatal parameter estimates will be entered in 2x2 repeated 
measures mixed models including drug-by-gene interactions. As there are two scans, pre-scan 
covariates will likely vary between the two scans, as such pre-scan thirst and milkshake 
pleasantness ratings from both visits will be included as time-varying covariates. Models will 
also include visit order, BMI, and gender as covariates. 
 
2. Exploratory Outcomes 
 
Appetitive hormone analysis: Participants will have three blood draws per visit. For each visit, 
the first blood draw (fasting) will occur at the start of the visit prior to drug administration. 
Second blood draw will occur 180min after drug administration (and 30min after a fixed 
milkshake consumption). Third blood draw will occur 210min after drug administration (and 
60min after a fixed milkshake consumption). Blood samples for satiety hormones will be 
assayed using Bio-Plex Pro Human Diabetes 7-plex Assay (Bio-rad, Hercules, CA). This kit 
analyzes adiponectin, amylin, and GLP-1. Additionally, a custom Human Cancer Panel 1 with 
only prolactin will be used to analyze prolactin concentrations (Bio-rad, Hercules, CA) using a 
Millipore Luminex (Darmstadt, Germany). Using the 3 timepoints for the blood draws, we will 
create the area under the curve (AUC) for each appetitive hormone (prolactin, adiponectin, 
amylin, GLP-1, insulin) for each visit. AUC values will be entered into 2x2 repeated measures 
mixed models including drug-by-gene interactions. Control variables of no interest to be 
included are BMI, gender, visit number, and baseline (fasting) hormone levels. 
 
Behavioral response inhibition analysis: Reaction time, % commission errors and % omission 
errors from the Stop Signal Task will be used as a behavioral measure of inhibition. The three 
outcome variables will be entered into 2x2 repeated measures mixed models including drug-by-
gene interactions. Control variables of no interest to be included are BMI, gender, visit number, 
and pre-task nausea ratings.