PET Imaging of Phosphodiesterase-4 (PDE4) in Brain and Peripheral Organs of Mccune-Albright Syndrome
Phase: PHASE1
Conditions: Nervous System Disease
Sponsor: National Institute of Mental Health (NIMH)

CNS IRB Protocol Template (3.11.15)
Protocol Title: PET imaging of phosphodiesterase-4 (PDE4) in brain and peripheral 
organs of McCune-Albright syndrome
Abbreviated Title: PDE4 in McCune-Albright syndrome
Protocol Number:  16-M-0093
Date of This Submission/Version:   5/14/2019
Principal Investigator
Name, Degree
Branch/Institute
Bldg/Rm
Phone
E-mail
Robert B. Innis, MD, PhD
MIB/NIMH
10/B1D43
301.594.1368
robert.innis@nih.gov
Human Research Protections Program Investigator and Staff Training:
For this protocol, the following “Just in time” human subjects protection training courses 
are required for investigators and staff:

NIAID GCP course or CITI GCP modules

Unanticipated Problems and Reporting Requirements in Biomedical Research
Total requested accrual (separately specify planned accrual for each subject group)  
 (20) Patients   
 (15) Healthy Volunteers 
 
Project Uses Ionizing Radiation:   No 
 Yes (attach RSC/RDRC documentation)
 Medically-indicated only
 Research-related only
 Both
IND/IDE
 No 
Yes (attach FDA documentation)
Drug/Device/# [11C](R)-rolipram/IND #73,149
Sponsor: Robert B. Innis, MD, PhD
Durable Power of Attorney 
 No 
 Yes 
Multi-institutional Project
 No 
 Yes
Data and Safety Monitoring Board 
 No 
 Yes 
Technology Transfer Agreement 
 No 
 Yes 
Samples are being stored 
 No  
 Yes 
Flesch-Kincaid reading level of consent form: 8.4 MAS 
(exclude boilerplate in assessing reading level)   8.4 HV

CNS IRB Protocol Template (3.11.15)
page 2 of 34
Précis:
Objective: McCune-Albright syndrome (MAS) is a mosaic disease arising from early 
embryonic somatic activating mutations of GNAS, which encodes the 3′, 5′-cyclic 
adenosine monophosphate (cAMP) pathway-associated G-protein, Gsα. Constitutive 
activation of Gsα leads to increased cAMP signaling in brain, as well as in peripheral 
organs, particularly bones. Although subjects with MAS show psychiatric and 
neurological symptoms, few studies have attempted to assess brain changes in these 
individuals. This protocol seeks to study changes in the cAMP cascade both in brain and 
peripheral organs of individuals with MAS using [11C](R)-rolipram PET, which binds to 
phosphodiesterase 4 (PDE4) and reflects cAMP cascade activity.
Study population: Participants will include 20 subjects with MAS and 15 healthy subjects 
group-matched to MAS subjects for age and gender. Both MAS subjects and healthy 
controls will have one or two PET scans: one whole body and one brain scan. We expect 
~10 brain and ~10 whole body scan to be performed in each group. In addition, subjects 
who have undergone a baseline whole body scan will be given the option to have a 
second blocked whole body scan after administration of the PDE4 inhibitor roflumilast.  
Design: Subjects with MAS will be recruited from participants in 98-D-0145 “Screening 
and Natural History of Patients with Polyostotic Fibrous Dysplasia and the McCune-
Albright Syndrome” (PI: Alison Boyce, MD). Only participants in protocol (98-D-0145) 
who provided self-consent without a legally authorized representative will be recruited. 
Brain PET scans will be performed by measuring metabolite-corrected arterial input 
function. Venous blood sampling will be performed for whole body scans. Subjects 
willing to undergo a second whole body scan will take 500 µg roflumilast P.O. prior to 
the scan. Blood sampling will be performed for the second whole body scan. 
Outcome measures: The primary outcome measure will be obtained in brain scans as the 
amount of radioligand binding quantified as distribution volume (VT). Calculated from 
both brain and plasma data, VT reflects rolipram binding to PDE4, corrected for any 
individual differences in metabolism of the radioligand or regional blood flow in brain. 
The secondary outcome measure will be obtained in whole body scans as area under the 
curve (AUC) of radioactivity expressed as standard uptake value (SUV). SUV is 
calculated by normalizing radioactivity in PET images to injection activity and body 
weight. VT in brain will be compared between subjects with MAS and healthy controls. 
AUC will be compared within-subjects with MAS between areas of craniofacial fibrous 
dysplasia and adjacent unaffected bone. AUC of whole body scans will also be compared 
between subjects with MAS and healthy controls. We hypothesize that subjects with 
MAS will show greater rolipram binding than healthy controls in brain regions, as well as 
greater rolipram uptake in bones affected by fibrous dysplasia than in unaffected bones. 
The third outcome measure will be the difference in SUV between the baseline whole 
body scan and the blocked (after roflumilast) whole body scan. This will help determine 
the amount of specific [11C](R)-rolipram binding in the periphery. We hypothesize that 
the [11C](R)-rolipram signal in the blocked scan will be greatly reduced, indicating that a 
majority of the signal in the baseline scan is specific

CNS IRB Protocol Template (3.11.15)
page 3 of 34
Table of Contents
1.
Introduction...............................................................................................................6
2.
Study Objectives.....................................................................................................10
3.
Subjects...................................................................................................................11
4.
Study Design and Methods.....................................................................................12
5.
Management of Data and Samples .........................................................................16
6.
Additional Considerations ......................................................................................17
7.
Risks and Discomforts............................................................................................17
8.
Subject Safety Monitoring......................................................................................19
9.
Outcome Measures .................................................................................................20
10.
Statistical Analysis..................................................................................................21
11.
Human Subjects Protection.....................................................................................21
12.
Anticipated Benefit.................................................................................................24
13.
Classification of Risk..............................................................................................24
14.
Consent Documents and Process............................................................................24
15.
Data and Safety Monitoring....................................................................................25
16.
Quality Assurance...................................................................................................25
17.
Reporting of Unanticipated Problems, Adverse Events and Protocol Deviations..26
18.
Alternatives to Participation ...................................................................................26
19.
Privacy ....................................................................................................................26
20.
Confidentiality ........................................................................................................26
21.
Conflict of Interest..................................................................................................27
22.
Technology Transfer...............................................................................................27
23.
Research and Travel Compensation .......................................................................27
24.
References...............................................................................................................29
25.
Attachments/ Appendices .......................................................................................31
List of Abbreviations 
AUC
area under the curve
cAMP
3′, 5′-cyclic adenosine monophosphate
MAS
McCune-Albright syndrome 
MDD
major depressive disorder
MRI
magnetic resonance imaging
PDE4
phosphodiesterase type 4
PET
positron emission tomography

CNS IRB Protocol Template (3.11.15)
page 4 of 34
SUV
standard uptake value
VT 
total (specific plus non-displaceable) distribution volume

CNS IRB Protocol Template (3.11.15)
page 5 of 34
1. Introduction
McCune-Albright syndrome and the cAMP cascade
McCune-Albright syndrome (MAS) is a mosaic disease arising from early 
embryonic somatic activating mutations of GNAS, which encodes the 3′, 5′-cyclic 
adenosine monophosphate (cAMP) pathway-associated G-protein, Gsα.  Constitutive 
activation of Gsα leads to ligand-independent cAMP signaling in affected cells. Clinical 
manifestations of MAS in a given individual are determined by the timing of the GNAS 
mutation during embryogenesis, the tissues involved, and the role of Gsα in affected 
tissues. Affected tissues can include those derived from ectoderm, mesoderm, or 
endoderm, and commonly include skin, skeleton, and endocrine organs. Café-au-lait skin 
macules result from localized hyperpigmentation, and occur in a characteristic mosaic 
distribution. Fibrous dysplasia of bone arises from proliferation of undifferentiated 
skeletal progenitor cells leading to abnormal bone formation. Bone lesions may involve 
any part and combination of the skeleton, ranging from an isolated monostotic lesion to 
severe polyostotic disease with progressive scoliosis, facial deformity, and functional 
impairment. Hyperfunctioning endocrinopathies include any combination of 
hyperthyroidism, autonomous sex steroid production, growth hormone excess, FGF23-
mediated hypophosphatemia, and hypercortisolism. 
The mechanism by which Gsα mutations result in the manifestations of MAS is 
thought to be related to dysregulation of the cAMP signaling cascade.  Numerous 
experimental systems have observed increased cAMP production as a downstream effect 
of activating Gsα mutations (2, 3). In an in vitro model of fibrous dysplasia, skeletal 
progenitor cells stably transfected with the MAS-causing Gsα mutation were shown to 
selectively upregulate specific isoforms of phosphodiesterase, an enzyme superfamily 
that catalyzes hydrolysis of cAMP (1)(Fig. 1). Among these was phosphodiesterase 4 
(PDE4), an isoform previously shown to play a role in modulating the differentiation of 
skeletal progenitor cells into bone-forming osteoblasts (4). This supports a model in 
which skeletal progenitor cell proliferation in fibrous dysplasia may be due in part to 
upregulation of PDE4 in response to Gsα-dependent cAMP production.
While skin, bone, and endocrine disease are the most clearly defined features of 
MAS, multiple additional tissues may be affected due to the ubiquitous nature of Gsα 
signaling. Observations from a long-standing NIDCR natural history protocol (98-D-
0145, “Screening and Natural History of Patients with Polyostotic Fibrous Dysplasia and 
the McCune-Albright Syndrome”, PI: Alison Boyce,MD) have led to the recognition of a 
neuropsychiatric phenotype potentially related to effects of the mutated Gsα in the central 
Fig.1. qPCR analysis of PDE isoform expression in skeletal 
progenitor cells stably transfected with the MAS-causing R201C 
mutation.  Significantly increased expression was detected for 
PDE-3b, -4b, -4d, and -7b isoforms in in comparison with mock- 
and vector-treated cells. From (1).

CNS IRB Protocol Template (3.11.15)
page 6 of 34
nervous system. The clinical features of MAS-associated neuropsychiatric disease are 
currently being investigated under 98-D-0145 and appear to fall along a spectrum that 
includes developmental delay, intellectual disability, anxiety, and impairments in 
attention and activity. Our hypothesis is that similar to the physical manifestations of 
MAS, these neuropsychiatric features result from Gsα mutation-bearing cells in the 
central nervous system, and are influenced by the location and function of cAMP 
signaling in these affected areas of the brain.
PET imaging of the cAMP cascade
PET imaging of phosphodiesterase type 4 (PDE4) using [11C](R)-rolipram has 
been successfully used to study the in vivo activity of the cAMP cascade. PDEs 
hydrolyze the second messengers cAMP and cyclic guanosine monophosphate (cGMP) to 
terminate signal transduction. Eleven PDEs exist in the human body. PDE4 is selective to 
cAMP and is present in both brain and peripheral organs such as heart, lungs, kidney (5), 
immune cells (6), osteoclast (7), and osteoblast (8). Rolipram is a selective inhibitor of 
PDE4; PDE4 has four isozymes, and rolipram is not selective among these isozymes.
Our studies—described below—demonstrated that [11C](R)-rolipram PET is a 
valuable method of detecting cAMP cascade activity in living human and animal 
subjects. Because PDE4 is present in almost all brain regions and no regions without 
PDE4 are available to use as a reference to measure rolipram binding, we used [11C](R)-
rolipram concentrations in arterial plasma (i.e., metabolite-corrected arterial input 
function) to measure rolipram binding in brain (9, 10). We also showed that rolipram 
binding can be accurately measured using only four arterial samples (9).
Broadly, the cAMP theory of depression posits that depression is caused by low 
cAMP signaling. The corresponding theory regarding the relevant mechanism of 
treatment is that chronic, but not acute, administration of antidepressants upregulates 
cAMP signaling. Although the cAMP theory of depression has limited supporting data, 
evidence for the mechanism of antidepressants has repeatedly been confirmed in animal 
studies. Chronic, but not acute, administration of all classes of antidepressants, as well as 
electroconvulsive therapy, upregulates several components of the cAMP pathway, 
including PDE4.
PDE4, an important component of the cAMP cascade, selectively metabolizes 
cAMP in the brain to the inactive monophosphate. Rolipram is a reversible inhibitor of 
PDE4, and binding of [11C](R)-rolipram provides a measure of the activity of this enzyme 
in brain. Due to a feedback mechanism, in vivo binding of [11C](R)-rolipram reflects the 
activity of the cAMP cascade; essentially, increased cAMP stimulates protein kinase A 
(PKA), which phosphorylates PDE4 that, in turn, increases rolipram binding.

CNS IRB Protocol Template (3.11.15)
page 7 of 34
We confirmed in 
animals that increased 
[11C](R)-rolipram 
binding reflects the 
phosphorylated / active 
state of PDE4. Using 
this radioligand, we 
found that PDE4 
binding is decreased in 
unmedicated patients 
with MDD, consistent 
with the cAMP theory 
of depression. Finally, 
we found that two 
months of treatment 
with an SSRI increased 
(normalized) PDE4 
binding, consistent with 
the cAMP theory of the 
mechanism of 
antidepressants.
In Vivo Density 
and Affinity. Our initial 
experiments sought to 
measure in vivo both 
binding site density and 
radioligand affinity of [11C](R)-rolipram in the rat brain (11). We also studied two critical 
factors in small-animal PET scans: the influence of anesthesia and the difference in 
binding under in vivo and in vitro conditions. Binding site density and radioligand affinity 
of conscious rats were significantly greater than those of anesthetized rats, by 29% and 
59%, respectively. In addition, in vitro affinity was five-fold greater than in vivo affinity, 
although density was similar in both conditions (Fig 2). The findings were consistent with 
rapid dephosphorylation of PDE4, and established that [11C](R)-rolipram binding in vivo 
reflects cAMP cascade activity in rats. 
In Vivo Phosphorylation Status. We further examined the effects of PKA 
modulators in conscious rats on [11C](R)-rolipram binding compared to the much less 
active enantiomer [11C](S)-rolipram (Fig 3) (12). Two drugs were studied. db-cAMP, 
which is a cAMP analogue, was used to activate PDE4, thus increasing activity of the 
cAMP-dependent PKA. PKA then phosphorylated the PDE4 enzyme and increased 
PDE4 activity. We also studied Rp-cAMP, a PKA inhibitor that directly inhibited PKA 
function, thereby decreasing PDE4 activity. Unilateral injection of the PKA activator (db-
cAMP) and the PKA inhibitor (Rp-cAMP) into the striatum significantly increased and 
decreased [11C](R)-rolipram binding, respectively. These effects were not caused by 
changes in blood flow or delivery of radioligand to brain, because these agents did not 
affect [11C](S)-rolipram binding. These results supported the importance of measuring in 
Fig 2. In vivo Scatchard plot of PDE4 binding using [11C](R)-rolipram 
in awake rats. PET scans were performed with increasing 
concentrations of radioligand in order to saturate binding to PDE4 
(bottom left), and analyzed as an in vivo Scatchard plot (bottom right). 
The brains were subsequently harvested for in vitro Scatchard analysis. 
Binding density (Bmax) was unchanged, but binding affinity was 
decreased 5-6 fold after death compared to the awake condition. The 
decreased affinity after death is consistent with the rapid 
dephosphorylation (within minutes) of PDE4. The phosphorylation 
state of PDE4 would not be possible in human postmortem tissue; PET 
imaging is the only technique currently available to measure the 
phosphorylated and active state of this enzyme.

CNS IRB Protocol Template (3.11.15)
page 8 of 34
vivo [11C](R)-rolipram binding in brain to assess response to physiological or 
pharmacological challenges to the cAMP second messenger system.
PDE4 Binding in 
MDD. Based on these 
preclinical findings, we 
hypothesized that 
[11C](R)-rolipram PET in 
humans would provide a 
unique in vivo measure of 
PDE4 density and affinity 
not possible in 
postmortem tissue. 
Expanding our work, we 
sought to quantify the 
binding of [11C](R)-
rolipram as an indirect 
measure of this enzyme’s 
activity in the brain of 
individuals with MDD 
compared with healthy 
controls (13). This is 
particularly important 
because animal studies 
had suggested that 
upregulation of the cAMP 
cascade, including PDE4, 
was a mechanism of 
action common to several 
antidepressant treatments. 
To avoid the misleading results that can be obtained from small sample sizes, we have 
now scanned a total of 43 unmedicated, moderately depressed patients with MDD and 25 
age- and gender-matched healthy controls, which is about twice the size of most PET 
studies in psychiatry. Notably, about half the patients were treatment-naïve. [11C](R)-
rolipram binding in brain was measured using arterial [11C](R)-rolipram levels to correct 
for the influence of cerebral blood flow. MDD subjects showed a widespread, 18% 
reduction in [11C](R)-rolipram binding (p=0.0001) that was not caused by different gray 
matter volumes (Fig 4). Decreased rolipram binding of similar magnitude was observed 
in most brain areas. Rolipram binding did not correlate with the severity of depressive or 
anxiety symptoms. These results were the first to demonstrate that brain levels of PDE4, 
a critical enzyme that regulates cAMP, are decreased in unmedicated individuals with 
MDD in vivo. Furthermore, the results are in line with human postmortem and rodent 
studies demonstrating downregulation of the cAMP cascade in MDD and support the 
hypothesis that PDE4 inhibitors—which increase cAMP cascade activity—may have 
antidepressant effects. 
Fig 3. [11C](R)-rolipram binding in rat brain depends on the 
phosphorylation state of PDE4. Rats were unilaterally injected in 
striatum with either a PKA activator (db-cAMP, upper panels) or a 
PKA inhibitor (Rp-cAMPS, lower panels). PKA-mediated 
phosphorylation of PDE4 increased both enzyme activity and the 
affinity of [11C](R)-rolipram binding about 10-fold. Both the PKA 
activator and inhibitor had the expected effect on radioligand uptake 
in striatum injected with drug compared to the side injected with 
saline. These effects were not merely caused by altered blood flow, 
given that they were not found using the inactive enantiomer 
[11C](S)-rolipram (right panels).

CNS IRB Protocol Template (3.11.15)
page 9 of 34
The Effect of Chronic Antidepressants on PDE4 Binding Building on this work, 
we sought to determine if antidepressant treatment upregulates PDE4 in humans as it 
does in animals. In addition to the rolipram PET scans without medication reported 
above, 22 of the 43 unmedicated MDD patients had a follow up rolipram scan after 
starting treatment with SSRIs. In this ongoing study, our pre-determined sample size is 
25 patients to be scanned after SSRI treatment.  These patients showed an increase of 13 
± 36% in rolipram binding after SSRI treatment across all brain regions (p = 0.001 when 
age was used as a covariate). In contrast, 11 healthy controls who had a repeat scan 
without SSRI showed similar binding on repeat scans with changes of only -2  ± 13%. 
The change in rolipram binding after SSRI varied markedly among patients as indicated 
by the large standard deviation of 36%. Those patients with lower rolipram binding 
before SSRI showed significantly greater increases after therapy (p < 0.007 in all regions) 
indicating normalization of rolipram binding by treatment. Older patients also showed 
greater increases in rolipram binding after SSRI (p ≤ 0.001) while young patients with 
higher baseline rolipram binding tended to show decreased rolipram binding after SSRI. 
According to the interim analysis, the changes in rolipram binding did not correlate with 
symptom improvement in all 
patients. Nevertheless, posthoc 
analyses after the full sample size 
has been accrued (i.e., 25 patients) 
may show correlations in subgroups 
of patients and/or specific regions of 
brain.
Taken together, these results 
elucidate two important and related 
points. First, the cAMP cascade, as 
indirectly measured with PDE4 
binding, was downregulated in 
unmedicated patients with MDD. 
Second, antidepressant treatment 
normalized this [11C](R)-rolipram 
downregulation. These studies 
suggest that PDE4 inhibition, 
perhaps via subtype selective agents, 
might again be assessed for efficacy 
in MDD; the results also broadly 
support the cAMP theory of 
depression and of antidepressant 
action.
PDE4 inhibitor roflumilast
There are currently two FDA approved PDE4 inhibitors on the market: 
roflumilast (Daliresp) and apremilast (Otezla). Both are marketed as anti-inflammatory, 
but the former is indicated for chronic obstructive pulmonary disease, while the latter is 
used to treat psoriasis/ psoriatic arthritis. The side effect profiles for both drugs are 
similar in that the primary complaint is gastrointestinal (Table 1). While both drugs could 
Fig 4.  [11C](R)-rolipram binding was globally 
decreased by 18% in brains of unmedicated MDD 
patients during a major depressive episode (n = 43) 
compared to matched controls (n = 35; p < 0.001). 
These mean parametric images of the two groups 
represent the “absolute” quantitation of radioligand 
binding calculated on an individual voxel level and 
using the concentration of radioligand in serial arterial 
blood samples. 

CNS IRB Protocol Template (3.11.15)
page 10 of 34
be suitable for this study, roflumilast has been investigated more thoroughly in rodents 
and humans than apremilast. Therefore, we can better estimate an appropriate dose of 
roflumilast for this study based on previous experiments and clinical trials. A study 
conducted in non-human primates found that 200 µg roflumilast resulted in a plasma 
concentration similar to peak concentration in humans (approximately 5 – 9 ng/ml) (14). 
This study hypothesized that a 500µg dose in humans would result in a PDE4 occupancy 
of 30-40%. The recommended dose of roflumilast is 500 µg per day, with or without 
food. Maximum plasma concentrations occur approximately 1-2 hours after dosing 
depending on food consumption. The half-life of roflumilast and its primary metabolite, 
roflumilast N-oxide, are approximately 17 and 30 hours, respectively. 
Table 1. Adverse reactions reported in ≥2% of patients
Roflumilast 
(500 mcg daily over 6 – 12 
months in N=4438)
Placebo
Apremilast
(30 mg BID over 1 -5 days in 
N=497). 
Placebo
Diarrhea
9.5%
2.7%
9.3%
1.2%
Weight decreased
7.5%
2.1%
NR
NR
Nausea
4.7%
1.4%
7.4%
1.4%
Headache
4.4%
2.1%
4.8%
1.8%
Back pain
3.2%
2.2%
NR
NR
Influenza
2.8%
2.7%
0.6%
0.6%
Dizziness
2.1%
1.1%
NR
NR
Decreased appetite
2.1%
0.4%
NR
NR
Vomiting
NR
NR
0.8%
0.4%
BID, twice a day; NR, not reported
Table modified from (15) and (16) 
The purpose of this study is to assess whether uptake in dysplastic bone reflects 
parent radioligand binding to the enzyme or merely accumulation of radiometabolites, 
which are expected to have no affinity for the enzyme. For this purpose, we need not 
administer a receptor saturating dose of roflumilast, as the Lassen occupancy plot has 
great sensitivity to measure the percentage occupancy and then to extrapolate to 100% 
occupancy (Cunningham et al., J. Cereb. Blood Flow & Metab., 30: 46-50, 2010). The 
sensitivity derives from the fact that it is a within subject comparison of multiple regions 
(in brain or periphery) at baseline and after blockade. We have used the Lassen plot 
numerous times, including to measure the occupancy of PDE4D subtype by the 
therapeutic candidate BPN14470. The figure below shows that that the occupancy (i.e., 
slope) is ~65% and the nondisplaceable uptake (VND or x-intercept) is ~4.9 mL ·cm-3.

CNS IRB Protocol Template (3.11.15)
page 11 of 34
0
5
10
15
0
1
2
3
4
Frontal Cortex
Cingulate Cortex
Striatum
Insula
Temporal Lobe
Amygdala
Hippcampus
Thalamus
Paracentral Lobule
Occipital Lobe
Cerebellum
VT (Baseline)
VT (Baseline) - VT (Blocked)
2. Study Objectives
This study will investigate cAMP signaling in the brains of individuals with MAS 
using [11C](R)-rolipram PET imaging.
a. Primary objectives
We will study PDE4 levels in patients with MAS by comparing [11C](R)-rolipram 
binding in various brain regions between patients and age-matched healthy controls. In 
addition, in whole body imaging of patients with MAS, we will compare [11C](R)-
rolipram uptake between areas of fibrous dysplasia and adjacent unaffected bones. In 
whole body scans, we will also compare [11C](R)-rolipram uptake in peripheral organs 
between subjects with MAS and healthy controls. The reason for doing the latter 
comparison is that some tissues of MAS may contain the GNAS mutations even if there 
are no obvious structural changes.
b. Secondary objective
In order to quantify the amount of [11C](R)-rolipram binding in patient whole body 
scans, we will perform a blocking scan with 500 µg roflumilast, a FDA approved PDE4 
inhibitor. Because of potential differences in metabolism of [11C](R)-rolipram in in the 
periphery of healthy controls and patients, it is difficult to interpret whether or not the 
[11C](R)-rolipram uptake is specific. By performing a blocking scan, we can quantify the 
amount of specific [11C](R)-rolipram binding to determine if there is a difference between 
healthy controls and patients.
3. Subjects
a. Description of study populations
This study will have two populations: subjects with MAS and healthy control 
subjects. MAS subjects will be recruited from participants in 98-D-0145 “Screening and 
Natural History of Patients with Polyostotic Fibrous Dysplasia and the McCune-Albright 

CNS IRB Protocol Template (3.11.15)
page 12 of 34
Syndrome” (PI: Alison Boyce, MD). Participation in our protocol will not delay or 
interfere with the patient's treatment. For all patients, the diagnosis of MAS will have 
been made under 98-D-0145.
Healthy subjects will be recruited and screened by the Molecular Imaging Branch 
(MIB) under a separate protocol (01-M-0254 or 17-M-0181), and group-matched for age 
to the MAS subjects.
Our accrual number will be 20 for subjects with MAS and 15 for healthy controls. 
Assuming a withdrawal / dropout rate of ~20%, our target is to complete the study with 
16 MAS subjects and 12 healthy controls. Both MAS and healthy subjects have one or 
two PET scans: one whole body and one brain scan. We expect that healthy subjects are 
more likely to have both brain and whole body scans and MAS subjects are less likely to 
do so. We expect ~10 brain and ~10 whole body scan to be performed in each group.
b. Inclusion criteria
Subjects with MAS
1. At least 18 years of age
2. Able to provide self-consent
3. Diagnosed with MAS under 98-D-0145.
4. Have craniofacial fibrous dysplasia
Healthy Subjects
1. At least 18 years of age.
2. Healthy based on medical history and physical examination.
c. Exclusion criteria
Subjects with MAS
1. Serious medical conditions, which may interfere with study procedures. Such 
conditions include but not limited to significant bone abnormalities in wrist 
areas of both arms, which makes it difficult to place a radial arterial line, 
clinically marked dysfunction of liver or kidney, which may delay clearance 
of [11C](R)-rolipram. 
2. Clinically significant laboratory abnormalities not linked to endocrine 
abnormalities but that may interfere with the PET measurement or affect 
safety of the participant during this study.
3. Positive HIV test.
4. Head trauma resulting in a period of unconsciousness lasting longer than one 
hour.
5. Metallic foreign bodies that would be affected by the magnetic resonance 
imaging (MRI) magnet, or fear of enclosed spaces likely to make the subject 
unable to undergo an MRI scan.
6. Recent research-related exposure to radiation (i.e., PET from other research) 
that, when combined with this study, would be above the allowable limits. 
7. Inability to lie flat on camera bed for about two and a half hours.
8. Pregnancy or breastfeeding.
9. NIMH employees/staff and their immediate family members will be excluded 
from the study per NIMH policy.

CNS IRB Protocol Template (3.11.15)
page 13 of 34
10. Currently taking a PDE4 inhibitor (such as roflumilast or apremilast), a drug 
that inhibits or induces cytochrome P450 (such as rifampicin or 
erythromycin), or oral contraceptives containing gestodene and ethinyl 
estradiol (such as Minulet and Femodene). 
Healthy Subjects
1. Serious medical conditions, which may interfere with study procedures. Such 
conditions include but not limited to clinically marked dysfunction of liver or 
kidney, which may delay clearance of [11C](R)-rolipram.
2. Clinically significant laboratory abnormalities that may interfere with the 
PET measurement or affect safety of the participant during this study.
3. Personal history of any DSM  Axis I disorder.
4. Positive HIV test.
5. Head trauma resulting in a period of unconsciousness lasting longer than one 
hour.
6. Metallic foreign bodies that would be affected by the MRI magnet, or fear of 
enclosed spaces likely to make the subject unable to undergo an MRI scan.
7. Recent research-related exposure to radiation (i.e., PET from other research) 
that, when combined with this study, would be above the allowable limits.
8. Inability to lie flat on camera bed for about two and a half hours.
9. Pregnancy or breastfeeding.
10. Current substance use disorder based on DSM.
11. NIMH employees/staff and their immediate family members will be 
excluded from the study per NIMH policy.
12. Currently taking a PDE4 inhibitor (such as roflumilast or apremilast), a drug 
that inhibits or induces cytochrome P450 (such as rifampicin or 
erythromycin), or oral contraceptives containing gestodene and ethinyl 
estradiol (such as Minulet and Femodene). 
4. Study Design and Methods
a.
Study overview 
This study will have two populations: subjects with MAS and healthy control subjects. 
Both MAS subjects and healthy controls will have up to three PET scans. All subjects 
will be given the option to have the following: one brain scan, one baseline whole-body 
scan, and one blocked whole-body scan after roflumilast administration. The two whole 
body scans can be performed on the same day or on separate days. The blocked whole 
body scan will be performed approximately 1-2 hours after the administration of 500 µg 
roflumilast P.O. 
 
All MAS subjects will also have a brain MRI scan and will administered the WAIS and 
SCID neuropsychiatric evaluations.  Each of these scans requires one outpatient visit 
lasting up to several hours; therefore, subjects in this study will have between one and 
five study visits depending on the number of scans they participate in. In addition, under 

CNS IRB Protocol Template (3.11.15)
page 14 of 34
this protocol MAS subjects will have their blood tested to rule out serious medical 
conditions.  Women of child-bearing potential will be tested for pregnancy within 24 
hours before each PET scan. Resting EKG will also be obtained.  
All PET and MRI scans will be performed at the NIH Clinical Center.
b. Recruitment
MAS patients will be recruited from the existing cohort of subjects enrolled in 
protocol 98-D-0145. Referrals will be made by a study team member of 98-D-0145. 
Criteria will be based on the inclusion and exclusion criteria of this PET protocol. 
Eligible subjects will be contacted by a member of the study team of this PET protocol 
who has not been directly involved in their clinical care under protocol 98-D-0145. 
Healthy volunteers will be recruited by the MIB and undergo screening under a 
separate protocol (01-M-0254 or 17-M-0181). We expect to recruit approximately five 
MAS subjects and five healthy controls per year.
c.
Screening
Written, informed consent will be obtained before any study procedures, 
including screening procedures, are done. For MAS subjects, eligibility will be 
determined based on a previous diagnosis of MAS as documented in 98-D-0145. The 
procedures to determine eligibility will not be repeated under this PET protocol if the 
same information has already been obtained under 98-D-0145 within six months. Tests 
done solely for the purpose of diagnosis of MAS will not be repeated even if not done 
within the last six months.  With participant permission, identifiable data will be shared 
between 98-D-0145 in order to confirm the diagnosis of MAS. If the information on 
medical history or examination is more than six months old, the updated information will 
be obtained under the current PET protocol. If some of laboratory testing described below 
in subsection ii is more than six months old or missing, the laboratory testing will be 
performed under this PET protocol. 
Healthy volunteers will be screened under a separate protocol (01-M-0254 or 
17-M-0181) to determine eligibility for the study procedures in this protocol. For healthy 
subjects, SCID-NP will be performed as a part of the screening under 01-M-0254 or 
17-M-0181 to determine eligibility.
Patients with MAS who have completed a baseline whole body scan and return to 
undergo a blocked whole body scan will be re-screened if it has been more than 6 months 
since their initial screening. Healthy volunteers who are returning to do a blocked whole 
body scan will be rescreened if it has been more than 12 months since their initial 
screening.
i. Medical history and examination (about one hour)
Under this PET protocol, MAS subjects will undergo a comprehensive medical 
history and examination by a clinically credentialed study investigator if those procedures 
were not done within 6 months under 98-D-0145. The purpose of the medical history and 
examination is to rule out serious medical conditions that may be contraindications to 
safe protocol participation. If H&P was done within 6 months under 98-D-0145, H&P 

CNS IRB Protocol Template (3.11.15)
page 15 of 34
will not be repeated in this PET protocol. A copy of the medical evaluation will be placed 
in the subject’s NIH chart.
For healthy subjects, H&P will be done under 01-M-0254 or 17-M-0181 but not 
in this PET protocol.
ii. Blood draw for laboratory testing
Under this PET protocol, for MAS subjects, blood testing will be performed to 
rule out serious medical conditions. Clinical laboratory testing will include complete 
blood count, PT/PTT, chemistry panel, liver function tests, lipid panel, HIV status, 
hepatitis serologies, and urine drug screen.  Resting EKG will also be obtained.  If any of 
these lab tests have been done in the last 6 months, they will not be repeated. With 
participant permission, identifiable data will be shared between 98-D-0145 in order to 
prevent unnecessary repeat of blood tests.
For healthy subjects, laboratory testing including urine drug screen will be done 
under 01-M-0254 or 17-M-0181 but not in this PET protocol.
iii Pregnancy testing: Women of child-bearing potential will be tested for pregnancy with 
a urine test at the time of screening.
d. Study procedures
Study procedures, including the PET scan, are for research purposes only.
The entire study procedures are summarized below.
Subjects with MAS:
1. Neuropsychiatric testing: Three hours.
2. Brain MRI: One hour.
3. Brain PET scan: Five hours.
4. Baseline whole body PET: Four hours.
5. Roflumilast administration and blocked whole body PET: Five hours
As long as the schedule of the participant allows, some of these procedures are 
combined in a single visit.  MRI, brain and whole body PET scans do not need to be 
performed in a specific order.
6. Pregnancy tests:  For women able to become pregnant, urine pregnancy testing will 
be done within the 24 hours prior to any MRI or PET scan.  PET and MRI will not be 
done if the pregnancy test is positive. 
Healthy controls:
1. Brain MRI: One hour.
2. Brain PET scan: Five hours.
3. Baseline whole-body PET: Four hours.
4.  Roflumilast administration and blocked whole body PET: Two hours
5. Pregnancy tests:  For women able to become pregnant, urine pregnancy testing will 
be done within the 24 hours prior to any MRI or PET scan.  PET and MRI will not be 
done if the pregnancy test is positive.

CNS IRB Protocol Template (3.11.15)
page 16 of 34
As long as the schedule of the participant allows, some of these procedures are combined 
in a single visit. The MRI, brain scan, and whole-body PET scans do not need to be 
performed in a specific order, however if the subject agrees to doing both whole body 
scans, the baseline scan will be performed before the blocked scan. The subject cannot do 
a blocked whole body scan without doing a baseline scan first. The baseline and blocked 
whole body scans can be performed in the same day, or on separate days. Subjects who 
have previously had a baseline whole body scan will be given the option to return for the 
blocked whole-body scan. For MAS patients, the goal will be to perform the blocked 
whole body scan within 6 months of the baseline whole body scan. If, however this is not 
possible due to scheduling restraints, screening procedures will be repeated to ensure that 
there has not been significant disease progression. Please note that MAS is a relatively 
stable condition in adulthood, and little change is expected over 6 months. For healthy 
controls, the time limit between the baseline and blocked whole body scans is 1 year. 
All subjects with MAS will be asked to have up to three [11C](R)-rolipram PET scans: 
one brain, one baseline whole body, and a blocked (with roflumilast) whole body scan. 
The subject can choose whether to undergo a brain scan or a baseline whole body scan, or 
both, but the subject is only eligible for the blocked whole body scan if they have the 
baseline whole body scan first. For brain scans, one arterial line will be placed for blood 
sampling, and one venous line will be placed for radioligand injection. For whole body 
scans, two venous lines will be placed, one for radioligand injection and another to 
measure [11C](R)-rolipram concentrations. In the case that a patient is only willing to do 
one of the scans, either a brain or a whole body scan will be performed based on patient 
preference. If the patient has no preference, one whole body scan will be performed. If 
patients are willing to have two PET scans (one whole body and one brain), the scans will 
not be performed in any particular order. All subjects with MAS will undergo the WAIS-
or KBIT-2, Edinburgh Handedness Inventory, and Structured Clinical Interview for Non-
Patient (SCID-NP).
Healthy controls will similarly have the option to undergo either one brain scan 
and/or a baseline whole body scan. The blocked whole body scan will be offered only if 
the subject has agreed to the baseline whole body scan. 
 
i. Brain [11C](R)-rolipram PET
Healthy controls will undergo PET scanning as outpatients, while MAS patients 
may be either inpatient or outpatient. An arterial line will be placed for blood sampling. 
A venous line will be placed for radioligand injection. To minimize discomfort, an 
anesthesiologist will place the arterial catheter in the subject’s wrist after numbing the 
area with a local anesthetic prior to the placement.
PET scanning will be performed in the NIH Clinical Center and will begin with a 
transmission scan by either CT, 68Ge- or 137Cs-source to correct for attenuation in the 
subsequent emission images. The PET radioligand [11C](R)-rolipram (maximum activity 
of 20 mCi) will be injected intravenously over about one minute. The subject will be 
scanned over approximately two hours. To quantify the density of PDE4 in the body, we 
will obtain arterial blood samples to correct for individual differences in clearance of the 

CNS IRB Protocol Template (3.11.15)
page 17 of 34
radioligand.  The volume of blood drawn will not exceed 200 mL per scan. Following the 
PET scan, the arterial and venous catheters will be removed.
ii. Whole body [11C](R)-rolipram PET
Healthy controls will undergo PET scanning as outpatients, while MAS patients 
may be either inpatient or outpatient. Two intravenous catheters will be placed, one for 
radioligand injection and another to measure [11C](R)-rolipram concentrations. Total 
volume of blood sampling is about 30 mL.
PET scanning will be performed in the Clinical Center and will begin with a 
transmission scan by either CT or 68Ge or 137Cs source to correct for attenuation in the 
subsequent emission images. The PET radioligand [11C](R)-rolipram (maximum activity 
of 20 mCi) will be injected intravenously over about one minute. The subject will be 
scanned over approximately two hours. Following the PET scan, the venous catheter will 
be removed. 
If the brain or whole body PET scan is cancelled or incomplete for any reason—
eg, synthesis failure of [11C](R)-rolipram, camera issues, or other problems—and if the 
study participant did not receive any radioactive material, the scan will be repeated on the 
same or on another day as long as the participant is willing. The transmission scan may 
be repeated as well.
iii. Blocked Whole body scan [11C](R)-rolipram PET with roflumilast
Healthy controls will undergo PET scanning as outpatients, while MAS patients 
may be either inpatient or outpatient. Two intravenous catheters will be placed, one for 
radioligand injection and another to measure [11C](R)-rolipram concentrations. Total 
volume of blood sampling is about 30 mL per scan.
PET scanning will be performed in the Clinical Center and will begin with a 
transmission scan by either CT or 68Ge or 137Cs source to correct for attenuation in the 
subsequent emission images. The PET radioligand [11C](R)-rolipram (maximum activity 
of 20 mCi) will be injected intravenously over about one minute at the start of each scan. 
The subject will be scanned over approximately two hours. Subjects will be administered 
500 µg roflumilast P.O. approximately 1-2 hours prior to [11C](R)-rolipram 
administration. Because food consumption can affect the absorption of roflumilast, 
subjects will be asked to have a light breakfast (and light lunch if scan occurs in the 
afternoon) prior to the scan. Following the PET scan, the venous catheter will be 
removed. 
iv. Brain MRI
All subjects, including those who receive only a whole body scan, will undergo a 
dedicated MRI of the brain for co-registration of PET images for analysis and to estimate 
brain morphology. Portions of the whole body PET scans image the brain. However, both 
volunteers and patients need not repeat the MRI scan if they had one of appropriate 
sequence within the prior year.
The MRI will be performed on a 3 Tesla magnet without the use of gadolinium 
contrast. Subjects will undergo safety screening prior to the MRI to rule out 
contraindications. The MRI will take place at the NIH Clinical Center and requires about 

CNS IRB Protocol Template (3.11.15)
page 18 of 34
thirty to forty minutes. If subjects become anxious during a scan, lorazepam (0.5–1 mg) 
or another benzodiazepine may be administered orally, if not clinically contraindicated. 
Subjects can also request that the operator stop the scan at any time. Women of child-
bearing potential without a history of sterilization will be tested for pregnancy with a 
urine test prior to the MRI. 
If the scan is cancelled or incomplete due to a problem with the scanner, the scan 
will be repeated on the same or on another day if the participant is willing.
v. Neuropsychiatric testing
Subjects with MAS will have the following examinations:
1.  Wechsler Adult Intelligence Scale, Fourth Edition (WAIS-IV) or the Kaufman 
Brief Intelligence Test, Second Edition (KBIT-2) (17, 18)
2. Structured Clinical Interview for Non-Patient (SCID-NP) or SCID Patient Edition 
(SCID-P) 
3. Edinburgh Handedness Inventory
Both the WAIS-IV and KBIT-2 provide an overall assessment of intelligence. 
SCID-NP characterizes neuropsychiatric symptoms. The Edinburgh Handedness 
Inventory assesses the dominance of a person’s right or left hand. A decision to perform 
either the SCID-NP or SCID-P will be based on the patient’s psychiatric history. 
The WAIS-IV, KBIT-2, or the Edinburgh Handedness Inventory will not be 
repeated if the subject has already undergone these assessments under protocol 98-D-
0145. The SCID-NP (or SCID-P) will not be repeated if performed within the past month. 
e.
End of participation
Study participation will end with the last PET or MRI scan the participant 
agreed to do. Patients will return for follow up under 98-D-0145. No follow up 
care will be needed for healthy volunteers.
5. Management of Data and Samples
a. Storage
Demographic and clinical data will be archived on a password-protected server. 
Imaging and blood data will be offloaded from the scanner/blood sampling device to a 
password-protected server. Clinical Safety Monitoring data will be archived together with 
other data. Laboratory test results will be stored on CRIS. Only study investigators will 
have access to stored data.
Blood samples for measurement of radioligand will be discarded after analysis, 
which will be performed on the day of the PET scan.
b. Data (if applicable: including genomic data) and sample sharing plan
Data may be shared. Data may be shared with collaborating laboratories 
at NIH or outside of NIH and/or submitted to NIH-designated 
repositories and databases if consent for sharing was obtained. 
Repositories receiving data from this protocol may be open-access or 
restricted access. 

CNS IRB Protocol Template (3.11.15)
page 19 of 34
Data will be stripped of identifiers and may be coded (“de-identified”) 
or unlinked from an identifying code (“anonymized”). When coded 
data is shared, the key to the code will not be provided to collaborators, 
but will remain at NIH.  Data may be shared with investigators and 
institutions with an FWA or operating under the Declaration of 
Helsinki (DoH) and reported at the time of continuing review.  Sharing 
with investigators without an FWA or not operating under the DoH will 
be submitted for prospective IRB approval.  Submissions to NIH-
sponsored or supported databases and repositories will be reported at 
the time of Continuing Review.  Submission to non-NIH sponsored or 
supported databases and repositories will be submitted for prospective 
IRB approval.
Required approvals from the collaborating institution will be obtained 
and materials will be shipped in accordance with NIH and federal 
regulations.   
6. Additional Considerations
a.
Research with investigational drugs or devices
[11C](R)-rolipram will be administered under IND #73,149, with Robert Innis, 
MD as the Sponsor and Investigator. The radioligand will be synthesized by the NIMH 
radiochemistry laboratory directed by Victor Pike, PhD.
Roflumilast is an FDA approved drug and meets the criteria of an IND exempt 
drug:
1. The drug product is lawfully marketed in the United States.
2. The investigation is not intended to be reported to FDA as a well-controlled
study in support of a new indication and there is no intent to use it to support any
other significant change in the labeling of the drug.
3. In the case of a prescription drug, the investigation is not intended to support a
significant change in the advertising for the drug.
4. The investigation does not involve a route of administration, dose, patient
population or other factor that significantly increases the risk (or decreases the
acceptability of the risk) associated with the use of the drug product (21 CFR
312.2 (b)(1)(iii).
5. The investigation is conducted in compliance with the requirements for review
by an IRB (21CFR 56) and with the requirements for informed consent (21 CFR
part 50).
6. The investigation is conducted in compliance with the requirements of 312.7
(i.e. the investigation is not intended to promote or commercialize the drug 
product).
b.
Gene therapy
None.

CNS IRB Protocol Template (3.11.15)
page 20 of 34
7. Risks and Discomforts
Potential risks from this study include those associated with: a) laboratory testing 
including urine toxicology testing, b) placement of an arterial and venous line, c) arterial 
and venous blood sampling, d) MRI, possibly including benzodiazepine drug 
administration, e) radiation exposure from PET scans, and f) PET scanning.
a. Blood Draw and laboratory testing
Laboratory testing is associated with minimal risks. We will first explain and 
familiarize subjects with laboratory testing to minimize discomfort, if any, during testing. 
During the screening it will be necessary to test urine for illegal drugs (urine toxicology). 
Subjects will be informed that if the drug test is positive, they will be told promptly. The 
results of the drug testing will be noted in their NIH medical record. There is minimal 
medical risk to urine collection for pregnancy testing.
b. Arterial/Venous line placement
Arterial catheterization has been shown to be a generally safe and reliable method 
of obtaining arterial blood samples (19). Placing a radial arterial catheter may cause 
bruising or infection. There is also a risk of occlusion and microemboli. In the past, over 
3,000 arterial catheters have been placed at NIH Clinical Center. Of these, two 
complications requiring physician’s care were reported. In the first case, a small radial 
artery aneurysm developed several months later, which was successfully repaired 
surgically. In the second case, a radial artery thrombosis developed 28 days later, which 
was also successfully repaired surgically. The arterial line will be placed by a member of 
the anesthesiology staff after confirming normal double circulation (both radial and ulnar 
arteries).
Venous catheter insertion, which is less invasive than arterial catheterization, can 
be associated with bruising, infection, or clot formation. Using proper placement 
techniques will minimize these risks.
c. Arterial and venous sampling
Blood draw volumes will not exceed the limits allowed by the NIH Clinical 
Center (Medical Administrative Policy M95-9: Guidelines for Blood Drawn for Research 
Purposes in the Clinical Center); in this study, the volume of blood drawn will not exceed 
230 mL per scan. Blood sampling may lead to the formation of a small subcutaneous 
hematomas caused by blood leaking from a punctured blood vessel. Such hematomas 
cause only minor discomfort. They are not dangerous and require no treatment other than 
reassuring the patient. There is also a small risk of infection at the site of the needle 
puncture, which can be readily treated with antibiotic therapy. Subjects will be asked not 
to donate blood for a period of eight weeks after participation is completed.
d. MRI
Three-Tesla MRI is widely used as a clinical imaging tool. Subjects will be 
screened and excluded for the presence of any metallic prostheses both at the time of 
recruitment and just prior to the MRI. Subjects will wear earplugs to minimize exposure 
to excessively loud noises. Occasionally subjects become anxious during the scan. In that 
case, lorazepam (0.5–1 mg) or a comparable benzodiazepine anti-anxiety agent may be 

CNS IRB Protocol Template (3.11.15)
page 21 of 34
given orally before the MRI as requested by subjects, and if not clinically 
contraindicated. Subjects can also request that the operator stop the scan at any time. Side 
effects of anti-anxiety medications may include prolonged sedation (usually less than 
eight hours), lightheadedness, lassitude, motor incoordination, and difficulties with 
balance. These effects are transient and go away after several hours. Subjects will be 
instructed to not drive, handle machinery, or drink alcohol until the next day. Subjects 
who receive a sedative will be observed in clinic until the sedation wears off and will be 
required to bring a driver or will be provided a taxi home.
Severely claustrophobic subjects find it difficult to undergo MRI scans; thus, such 
subjects will be excluded at the time of recruitment.
e. Radiation exposure
Radiation exposure in this protocol will be from [11C](R)-rolipram and transmission scans 
for PET using 68Ge or 137Cs source or CT transmission for PET/CT. Among these three 
types of transmission scans, CT transmission causes the highest radiation as calculated by 
Craig Barker, PhD, CC PET Dept., July 22 2015; according to ICRP103. Therefore, the 
radiation exposure from CT transmission is used to calculate total radiation exposure in 
88-23(a). Radiation dose estimates for [11C](R)-rolipram are based upon biodistribution 
data in healthy human subjects performed by the MIB, NIMH under protocol 06-M-0002 
“PET whole body biodistribution and test retest brain imaging studies using a 
phosphodiesterase 4 inhibitor (R)-[11C]rolipram” (PI: M Fujita, MD, PhD). That study 
showed that the highest effective dose in this protocol is about four rem for patients with 
MAS and healthy controls who would undergo one brain scan and two whole-body scans 
with two CT transmission scans for each brain and whole-body scan, respectively. 
Although CT transmission scans have higher radiation exposure than 68Ge or 137Cs 
transmission scans, CT transmission scans are far more commonly performed in both 
clinical and research settings because they are the only choice for PET/CT scanners. This 
amount is within NIH RSC guidelines. Detailed results of this dosimetry study are 
included in the attached NIH 88-23(a). In case the subject moves between the 
transmission scan and the injection of [11C](R)-rolipram, the MIB routinely includes two 
transmission scans for each PET scan to allow a repeat of transmission. Note: the 
designation of radiation dose of “about four rem” was recently approved by the Radiation 
Safety Committee as an alternative way to present risk to patients using rounded 
numbers, which better reflects the actual dose to an individual subject.
All subjects will be asked about any prior research participation involving 
radiation exposure so that the total exposure, in combination with the present study, will 
not exceed an effective dose of five rem per 12 months. Examples may include 
mammograms, x-rays taken in radiology departments, cardiac catheterization, and 
fluoroscopy as well as nuclear medicine scans in which radioactive materials were 
injected into your body.
f. PET scan
PET scanning, which detects injected radioactivity within the body, has been used 
as a clinical imaging tool. We routinely use a series of procedures to minimize the risk of 
discomfort during scanning sessions. Namely, the procedures are conducted in the 
presence of trained health professionals to whom subjects will have ready access should 

CNS IRB Protocol Template (3.11.15)
page 22 of 34
they experience any problems. Subjects can communicate with the trained health 
professionals while in the scanner and can be removed from the scanner or withdraw 
from the study at any time if they wish to do so.
Occasionally subjects become anxious during the scan. In that case, subjects can 
request that the operator stop the PET scan.
g. Roflumilast administration
     Roflumilast is an FDA approved PDE4 inhibitor, used commonly to treat COPD. In 
this study, subjects may receive a single dose of 500 µg roflumilast P.O. For treatment of 
COPD, roflumilast is normally given at a dose of 500 µg over a period of weeks or 
months (15). Therefore, we feel that the risks of serious reactions are low given that we 
are administering only a single dose. Adverse reactions to roflumilast administration 
reported by ≥ 2% of patients (n = 4438) include: diarrhea, weight loss, nausea, headache, 
back pain, influenza, dizziness, and decreased appetite. Psychiatric disorders (anxiety, 
depression) occurred at a frequency of 1 to 2% greater than placebo. We will contact all 
patients who receive roflumilast 24-48 hours after its administration to confirm they are 
feeling well.
8. Subject Safety Monitoring
Participants will be carefully monitored by clinical staff throughout the study. 
Participants will be asked about the presence of adverse events throughout the procedure. 
To confirm the absence of the pharmacological effects of the PET ligand, for each PET 
scan, a clinically credentialed investigator will monitor vital signs within three hours 
before tracer injection, and again at approximately 30 and 90 minutes after radioligand 
injection. The site of the arterial and venous catheter insertion will be carefully monitored 
for signs of bleeding during the PET scans and also after removal of the catheter.
All adverse events (AEs) will be recorded on the case report form. All AEs, 
including clinically significant abnormal findings on laboratory evaluations, regardless of 
severity, will be evaluated until satisfactory resolution. After removal of venous lines, the 
site of the catheter insertion will be carefully monitored for signs of bleeding. 
Toxicity tables/criteria to be used
Toxicity will be assessed according to the NCI Common Terminology Criteria for 
Adverse Events (CTCAE) v.4.03   http://evs.nci.nih.gov/ftp1/CTCAE/About.html.
Criteria for individual subject withdrawal
We will stop study procedures for any subject with any Grade 2, Grade 3, or 
greater non-laboratory toxicity that is intolerable, persistent, or uncorrectable for more 
than several minutes. In addition, the Medical Advisory Investigator or a clinically 
credentialed investigator may terminate study procedures based on clinical judgments. 
Likely causes are pain at the site of catheter insertion and difficulties to stay in MRI and 
PET scanners, and the judgement of withdrawal by study investigators will be done as 
described above. In addition, study participants can voluntarily withdraw at any time for 
any reason. 

CNS IRB Protocol Template (3.11.15)
page 23 of 34
9. Outcome Measures
a. Primary outcome measures
The primary outcome measure will be obtained in brain scans as the amount of 
radioligand binding quantified as distribution volume (VT). Calculated from both brain 
and plasma data, VT reflects the density of PDE4, corrected for any individual differences 
in metabolism of the radioligand or regional blood flow in brain. 
b. Secondary outcome measures
The secondary outcome measure will be obtained in whole body scans as area under 
the curve (AUC) of radioactivity expressed as standard uptake value (SUV). SUVs are 
calculated by normalizing radioactivity in PET images to injection activity and body 
weight. Although AUC of SUV is not as fully quantitative as VT, the AUC is widely used 
in clinical studies where arterial data are not obtained. If patients and healthy controls 
have markedly different clearance of [11C](R)-rolipram, group comparison using AUC of 
SUV may include errors. To correct group difference in clearance, in whole body 
imaging, venous concentrations of [11C](R)-rolipram will be measured at several time 
points. MRI measures of brain morphology including gray matter, white matter and CSF 
volume and folding will be estimated from the MRI images. 
c.
Tertiary outcome measure
The third outcome measure will be the difference in SUV between the baseline and the 
blocked whole-body scans. SUV is calculated as stated above. 
10. Statistical Analysis
a. Analysis of data/ study outcomes
PET and MRI images will be co-registered for anatomic definition of brain 
regions. Brain and plasma data will be analyzed with compartmental and graphical 
modeling to calculate VT, which is proportional to the density of receptor or enzyme. VT 
will be calculated both for regions of the brain and on a voxel basis. PET whole body 
images will be analyzed by drawing organs and by calculating AUC of SUV. For 
comparisons between subjects with MAS and healthy controls in brain and peripheral 
organs, an unpaired test, such as t- and Mann-Whitney tests will be used. In addition, in 
peripheral organs of MAS, for comparisons of PET data between affected and unaffected 
organs, a paired test will be applied if the contralateral side is unaffected. If contralateral 
side is also affected or not available, an unpaired test will be applied to compare the same 
tissue/organ between patients and healthy controls. For the statistical analyses, SPSS, 
PMOD, SPM, Freesurfer or equivalent programs will be used.
b.
Power analysis
This is an exploratory study focusing on PET imaging. Clinical data obtained under 
98-D-0145 may be used to interpret the relationship between PET findings and clinical 
manifestations of the disease. Although constitutive activation of Gsα is expected to 
increase cAMP signaling, which increases PDE4 levels, PDE4 levels in MAS are not 
known in postmortem tissue either in brain or in peripheral organs such as bone. 
Therefore, we cannot do a realistic power calculation.

CNS IRB Protocol Template (3.11.15)
page 24 of 34
To achieve the goals of this exploratory study, we request a total of 35 subjects: ~10 
MAS subjects to undergo brain PET scan, ~10 MAS subjects to undergo whole body 
PET, and 15 healthy controls to undergo a brain scan and/or a whole body PET scan.  By 
limiting subjects with MAS to those who have craniofacial fibrous dysplasia and 
neuropsychological symptoms, we expect to increase the likelihood of detecting changes 
in PET in this small sample size. To limit the number of study participants, all MAS and 
healthy subjects will be asked if they would be willing to have both brain and whole body 
PET scans. A smaller number of 15 healthy subjects are requested because healthy 
subjects are more likely to have both brain and whole body scans. Our previous study of 
[11C](R)-rolipram brain scans in healthy subjects found that the coefficient of variation 
(SD/mean) of VT was 19%. Based on the activation of Gsα, we hypothesize that MAS 
patients will show increased rolipram binding for whichever one-sided test is appropriate. 
Assuming a 20% dropout from 10 subjects (thus, eight subjects per group) plus a 19% 
coefficient of variation (SD/mean) will provide a power of 0.80 to detect a 19% increase. 
11. Human Subjects Protection
a.
Subject selection
The study will be open to all racial and ethnic groups and will not target any 
particular group. Healthy volunteers will be recruited to match the patient group for age. 
Those who are HIV positive are excluded because the possible viral effects on the brain 
could interfere with interpretation of the outcome of the study.
b.
Justification for exclusion of children
We will exclude children younger than 18 years of age to avoid radiation exposure to 
minors because the risks of radiation exposure outweighs the benefits.
c.
Justification for inclusion of other vulnerable subjects
No population of vulnerable subjects will be included. Pregnant and breastfeeding 
females will be excluded because of greater risks of radiation to fetus and babies.   
Subjects without consent capacity will be excluded because this protocol has more than 
minimal risks and provides no direct benefits.  Protections for NIH employees and staff 
participating in this study include 1) assuring that the participation or refusal to 
participate will have no effect, either beneficial or adverse, on the subject’s employment 
or position at the NIH, 2) giving employees and staff who are interested in participating 
the “NIH Information Sheet on Employee Research Participation” prior to obtaining 
consent, and 3) assuring that there will be no direct solicitation of employees or staff.
d.
Justification for sensitive procedures
This protocol does not include sensitive procedures such as use of placebo, 
medication withdrawal, provocative testing, or deception.
e.
Safeguards for vulnerable populations and sensitive procedures
Pregnancy testing and interview will be performed for females with childbearing 
potential to prevent the enrollment of pregnant females.  This protocol may enroll NIH 
employees. Protections for NIH employees and staff participating in this study include 1) 

CNS IRB Protocol Template (3.11.15)
page 25 of 34
assuring that the participation or refusal to participate will have no effect, either 
beneficial or adverse, on the subject’s employment or position at the NIH, 2) giving 
employees and staff who are interested in participating the “NIH Information Sheet on 
Employee Research Participation” prior to obtaining consent, and 3) assuring that there 
will be no direct solicitation of employees or staff.
When reviewing responses to the SCID, experienced clinical interviewers will 
supervise interviewers. Although rare, should the study staff identify symptoms of 
suicidality or suicidal ideations during the interview procedures, all necessary steps will 
be taken. The on-call clinician will be contacted to provide further evaluation, the 
subjects’ own clinician may be notified after permission is granted by the subject. If not, 
other referral sources are provided and followed up, and necessary treatment steps (e.g., 
hospitalization, referral to a care provider, etc.) will be offered. However, NIH does not 
pay for outside hospitalization if that is required.
12. Anticipated Benefit
This study offers no direct benefit to individual subjects, but will lead to generalizable 
knowledge about MAS.
13. Classification of Risk
a. For adults
This study has more than minimal risk.
b. For adults without consent capacity
Not applicable.
c.
For children
Not applicable.
d. Overall risk and benefit consideration
The risks are reasonable in relation to anticipated benefit.
14. Consent Documents and Process
a. Designation of those obtaining consent
Study investigators designated as able to obtain consent in section 11f above will 
obtain informed consent. All study investigators obtaining informed consent have 
completed the NIMH HSPU ‘Elements of Successful Informed Consent’ training. 
b. Consent procedures 
All participants will receive a verbal explanation in terms suited to their 
comprehension of the purposes, procedures, and potential risks of the study and of their 
rights as research participants. Participants will have the opportunity to carefully review 
the written consent form and ask questions regarding this study prior to signing. NIMH 
Human Subject Protection Unit (HSPU) will be consulted if there is any question about 
the consent capacity of an individual.
c.
Consent documents

CNS IRB Protocol Template (3.11.15)
page 26 of 34
Two separate consent forms have been submitted: one for healthy volunteers, one for 
subjects with MAS. Both of these groups will be asked if they are willing to have one 
brain and one whole body PET scans. In addition, there will be two separate consent 
forms (one for healthy volunteers and one for subjects with MAS) giving subjects who 
have previously had a baseline whole-body scan the option to return for a blocked whole-
body scan with roflumilast administration.  All of these consent forms contain all 
required elements.
15. Data and Safety Monitoring
a. Data and safety monitor
This study will be monitored by the following independent safety monitor: 
Kenneth Towbin, MD. Dr. Towbin is a Senior Research Physician at NIMH, board-
certified in adult psychiatry, and in child and adolescent psychiatry and Chief of Clinical 
Child and Adolescent Psychiatry in the Emotion and Development Branch. Dr. Towbin 
has been the primary physician overseeing care for inpatients with major depression 
treated in the Mood and Brain Disorders Unit. He also assists in overseeing care, as 
needed, in the Section on Developmental Affective Neuroscience (SDAN).   
b. Data and safety monitoring plan
The PI will prepare a report on data and safety parameters for the 
Independent Monitor for the first ten subjects or annually, whichever 
comes first.  The Independent monitor will provide a written monitoring 
report to be submitted to the IRB at the time of continuing review. 
c.
Criteria for stopping the study or suspending enrollment or procedures
The Independent Monitor and IRB will determine if changes are needed for 
the research to continue or if it will be closed. Any changes required as 
conditions for resuming the research must be submitted as an amendment 
and IRB-approved before the changes can be implemented. The 
PI/Independent Monitor and IRB will determine if changes are needed for 
the research to continue or if it will be closed. Any changes required as 
conditions for resuming the research must be submitted as an amendment 
and IRB-approved before the changes can be implemented. 
16. Quality Assurance
a. Quality assurance monitor
Quality assurance will be monitored by the PI, The research team and the NIMH Office 
of Regulatory Compliance (ORO).
b. Quality assurance plan
ORO monitors intramural research studies to ensure compliance with GCP, 
organizational policies and regulations.  Audit frequency is determined by the ORO SOP 
based on the study level of risk.  Results of ORO audits are provided to the PI, The 
Clinical Director and the CNS IRB. As an IND study, this protocol will be subject to 
GCP audits at study initiation and after the first enrolled subject.  Timing of subsequent 
review will be established by the ORO but no less frequent than every other year.

CNS IRB Protocol Template (3.11.15)
page 27 of 34
17. Reporting of Unanticipated Problems, Adverse Events and Protocol Deviations
Reportable events for this protocol will be tracked and reported in compliance with 
Policy 801.
18. Alternatives to Participation 
Subjects do not receive any treatment in this study or forego any treatment in 
order to participate in this study. The alternative, therefore, is not to participate.
19. Privacy
All research activities will be conducted in as private a setting as possible.
20. Confidentiality
a. For research data and investigator medical records
Data will be stored using codes that we assign. Data will be kept in password-
protected computers. Only study investigators will have access to the data. The results of 
urine drug screen will be in the NIH Medical Record.
This study collects sensitive information on drug and alcohol use. The PI will 
train study staff regarding obtaining and handling potentially sensitive and private 
information about co-workers through staff discussions and written branch/section 
procedures.  
b. For stored samples
Samples are not stored.
c. Special precautions
Samples and data will be stored using codes that we assign. Data will be kept in 
password-protected computers.  Samples will be kept in locked storage. Only study 
investigators will have access to the samples and data. De-identified results from clinical 
trials will be posted on cctrials.gov.
Any clinically relevant results will be returned to the participant.
21. Conflict of Interest
a. Distribution of NIH guidelines
NIH guidelines on conflict of interest have been distributed to all investigators.
b. Conflict of interest 
There are no conflicts of interest to report.
c. Role of a commercial company or sponsor
Commercial company or sponsor is not involved.
22. Technology Transfer
No Technology Transfer is involved in this study.
23. Research and Travel Compensation
Subjects will be compensated for time and research-related inconveniences. 
Reimbursement is based on NIH standards for time devoted to the research project. This 
study will also provide travel compensation based on NIH standards. Subjects will be 
paid for each portion of the study they have completed whether or not they opt for early 

CNS IRB Protocol Template (3.11.15)
page 28 of 34
withdrawal from participation. MAS patients and controls may receive a maximum of 
$1190 and $930, respectively. Subjects will be paid for each portion of the study they 
have completed whether or not they opt for early withdrawal from participation. Payment 
will be sent after participation has been completed. Employees and staff who participate 
during work hours must have permission from their supervisor. NIH employees must 
either participate outside of work hours or take leave in order to receive compensation.

CNS IRB Protocol Template (3.11.15)
page 29 of 34
Schedule of Visits and Remuneration
 
Inconvenience 
Units
Inconvenience 
Remuneration
Time 
(hours)
Time 
Remuneration
Total 
Remuneration
Visit 1: screening 
(patient only)*
 
 
 
 
 
Medical history, 
physical exam, EKG, 
and lab tests if not 
obtained in prior six 
months
2
$20
2
$30 
$50 
Neuropsych Testing
4
$40
3
$40
$80
Visit 2**
 
 
 
 
 
Pregnancy test***
1
$10 
 
 
$10 
MRI
8
$80 
2
$30 
$110 
Visit 3**
 
 
 
 
 
Pregnancy test***
1
$10 
 
 
$10 
Brain PET scan
15
$150
5
$60
$210
Venous catheter
3
$30 
 
 
$30 
Arterial catheter
7
$70
$70
Movement restriction
1
$10 
 
 
$10 
Visit 4**
 
 
 
 
 
Pregnancy test***
1
$10 
 
 
$10 
Whole body PET scan
15
$150
4
$50
$200
Venous catheter x2
6
$60 
 
 
$60 
Movement restriction
1
$10 
 
 
$10 
Visit 5**
 
 
 
 
 
Roflumilast admin
1
$50
$50
Pregnancy test*** †
1
$10 
 
 
$10 
Whole body PET scan
15
$150
4
$50
$200
Venous catheter x2 ††
6
$60 
 
 
$60 
Movement restriction
1
$10 
 
 
$10 
Total
89
$930
20
$260
$1190
*Screening will be repeated for subjects with MAS if no information is available within 6 
months under 98-D-0145. Healthy volunteers will be screened under protocol 01-M-0254 
or 17-M-0181.
**The order of visits 2, 3, 4 and 5 may change depending on the availability of the 
scanners.
***Pregnancy testing for women of childbearing potential
†The pregnancy test for the second whole body PET scan will not be conducted if the 
scan takes place the same day as the baseline whole body scan. 

CNS IRB Protocol Template (3.11.15)
page 30 of 34
†† Subjects will not receive compensation for placement of catheters if blocked scan 
occurs on the same day as baseline scan and catheters are already in place.
 Note: Some visits may be combined.

CNS IRB Protocol Template (3.11.15)
page 31 of 34
24. References
1.  Piersanti S, Remoli C, Saggio I, Funari A, Michienzi S, Sacchetti B, Robey PG, 
Riminucci M, Bianco P. Transfer, analysis, and reversion of the fibrous dysplasia cellular 
phenotype in human skeletal progenitors. Journal of bone and mineral research : the 
official journal of the American Society for Bone and Mineral Research. 2010;25:1103-
1116.
2.  Lania A, Persani L, Ballare E, Mantovani S, Losa M, Spada A. Constitutively active 
Gs alpha is associated with an increased phosphodiesterase activity in human growth 
hormone-secreting adenomas. The Journal of clinical endocrinology and metabolism. 
1998;83:1624-1628.
3.  Lader AS, Xiao YF, Ishikawa Y, Cui Y, Vatner DE, Vatner SF, Homcy CJ, Cantiello 
HF. Cardiac Gsalpha overexpression enhances L-type calcium channels through an 
adenylyl cyclase independent pathway. Proceedings of the National Academy of Sciences 
of the United States of America. 1998;95:9669-9674.
4.  Wakabayashi S, Tsutsumimoto T, Kawasaki S, Kinoshita T, Horiuchi H, Takaoka K. 
Involvement of phosphodiesterase isozymes in osteoblastic differentiation. Journal of 
bone and mineral research : the official journal of the American Society for Bone and 
Mineral Research. 2002;17:249-256.
5.  Engels P, Fichtel K, Lubbert H. Expression and regulation of human and rat 
phosphodiesterase type IV isogenes. FEBS Lett. 1994;350:291-295.
6.  Beghe B, Rabe KF, Fabbri LM. Phosphodiesterase-4 inhibitor therapy for lung 
diseases. American journal of respiratory and critical care medicine. 2013;188:271-278.
7.  Park H, No AL, Lee JM, Chen L, Lee SY, Lee DS, Yim M. PDE4 inhibitor 
upregulates PTH-induced osteoclast formation via CRE-mediated COX-2 expression in 
osteoblasts. FEBS letters. 2010;584:173-180.
8.  Nomura-Furuwatari C, Wakitani S, Hashimoto Y, Imai Y, Ohta Y, Nakagawa K, 
Nakao Y, Takayama K, Manaka T, Takaoka K. Expression profiles of phosphodiesterase 
4D splicing variants in osteoblastic cells. Journal of bone and mineral metabolism. 
2008;26:152-158.
9.  Zanotti-Fregonara P, Zoghbi SS, Liow JS, Luong E, Boellaard R, Gladding RL, Pike 
VW, Innis RB, Fujita M. Kinetic analysis in human brain of [11C](R)-rolipram, a positron 
emission tomographic radioligand to image phosphodiesterase 4: a retest study and use of 
an image-derived input function. Neuroimage. 2011;54:1903-1909.
10.  Fujita M, Zoghbi SS, Crescenzo MS, Hong J, Musachio JL, Lu J-Q, Liow J-S, 
Seneca N, Tipre DN, Cropley VL, Imaizumi M, Gee AD, Seidel J, Green MV, Pike VW, 
Innis RB. Quantification of brain phosphodiesterase 4 in rat with (R)-[11C]rolipram-PET. 
NeuroImage. 2005;26:1201-1210.
11.  Itoh T, Abe K, Zoghbi SS, Inoue O, Hong J, Imaizumi M, Pike VW, Innis RB, Fujita 
M. Positron emission tomographic measurement of the in vivo affinity of 11C-(R)-
rolipram and the density of its target, phosphodiesterase-4, in brain of conscious and 
anesthetized rats. Journal of nuclear medicine : official publication, Society of Nuclear 
Medicine. 2009;50:749-756.
12.  Itoh T, Abe K, Hong J, Inoue O, Pike VW, Innis RB, Fujita M. Effects of cAMP 
dependent protein kinase activator and inhibitor on in vivo PET rolipram binding to 
phosphodiesterase 4 in conscious rats. Synapse. 2010;64:172-176.

CNS IRB Protocol Template (3.11.15)
page 32 of 34
13.  Fujita M, Hines CS, Zoghbi SS, Mallinger AG, Dickstein LP, Liow J-S, Zhang Y, 
Pike VW, Drevets WC, Innis RB, Zarate CA. Downregulation of brain phosphodiesterase 
type IV measured with 11C-(R)-rolipram positron emission tomography in major 
depressive disorder. Biol Psychiatry. 2012;72:548-554.
14.  Takano A, Uz T, Garcia-Segovia J, Tsai M, Lahu G, Amini N, Nakao R, Jia Z, 
Halldin C. A Nonhuman Primate PET Study: Measurement of Brain PDE4 Occupancy by 
Roflumilast Using (R)-[(11)C]Rolipram. Mol Imaging Biol. 2018;20:615-622.
15.  Daliresp (roflumilast) [package insert].  Wilmington, DE: AstraZeneca 
Pharmaceuticals LP. 2011.
16.  Otezla (apremilast) [package insert]. Summit, NJ: Celgene Corporation. 2014.
17.  Wechsler D: Wechsler Adult Intelligence Scale–Fourth Edition. San Antonio, TX, 
Pearson; 2008.
18.  Kaufman AS, Kaufman NL: Kaufman Brief Intelligence Test, Second Edition. San 
Antionio, TX, Pearson; 2004.
19.  Lockwood AH. Invasiveness in studies of brain function by positron emission 
tomography. Journal of cerebral blood flow and metabolism : official journal of the 
International Society of Cerebral Blood Flow and Metabolism. 1985;5:487-489.

CNS IRB Protocol Template (3.11.15)
page 33 of 34
25. Attachments/ Appendices

Eligibility checklist

Case report forms (CRFs)

CNS IRB Protocol Template (3.11.15)
page 34 of 34
 Consent Forms

Patient

Healthy adult volunteer