Conditions: Multiple System Atrophy
Allocation: RANDOMIZED | Intervention model: SINGLE_GROUP
Enrollment: 47
Healthy volunteers: no

Brief summary:
Single-center, randomized, placebo-controlled, phase-II, futility clinical trial to determine if oral sirolimus is of sufficient promise to slow disease progression in MSA, prior to embarking on a large-scale and costly phase III study to assess its efficacy. A futility design under the null hypothesis assumes that sirolimus will slow the progression of the disease, whereas the alternative hypothesis assumes no benefit of sirolimus. If the null hypothesis is rejected (i.e., futility of sirolimus to slow progression of MSA), a major phase III study will be discouraged, whereas non-futility will offer strong support for a phase III trial to detect clinical efficacy.

Detailed description:

Arms:
arm 1: 2 mg/day (one 2-mg tablet/day). The dose of sirolimus will be adjusted throughout the trial based on sirolimus plasma levels and the presence of drug-related adverse events. The maximum dose of sirolimus will be6 mg/day (three 2-mg tablets/day). arm 2: Patients receiving placebo will undergo analog sham level measurements and the number of tablets will be also adjusted to maintain the blinding of the trial.

Interventions: intervention 1: Sirolimus 2 MG intervention 2: Placebo
Intervention descriptions: intervention 1: Dose will be adjusted throughout this trial based on sirolimus plasma levels and the presence of drug-related adverse events. The maximum dose will be 6mg/day. intervention 2: Patients receiving placebo will undergo analog sham level measurements and the number of tablets will be also adjusted to maintain the blinding of the trial.

Protocol document:
Prot_SAP_000.pdf: 
STUDY PROTOCOL 
 
A futility trial of sirolimus in multiple system atrophy 
A single center, randomized, double blind, placebo-controlled futility trial to determine if sirolimus is of 
sufficient promise to slow the progression of multiple system atrophy 
 
Protocol Number: 17-01392 
 
National Clinical Trial (NCT) Identification Number:  NCT03589976 
 
 
Study Chairs: 
Horacio Kaufmann MD  
Jose-Alberto Palma MD PhD 
New York University School of Medicine 
 
IND #: 136871 
Sponsor of the IND: 
Horacio Kaufmann MD 
 
Support from: 
National Institute of Neurological Disorders and Stroke (NINDS) 
 
Version Number:  v.9 (4-JUN-2020) 
 
 
 


 
 
Summary of Changes from Previous Version: 
Affected 
Section(s) 
Summary of Revisions Made 
Rationale 
1.3. Schedule of 
activities and 
4.1, 5.2. 
Patients will undergo PCR nasal swab to screen 
for COVID-19 
Sirolimus treatment can cause 
immunosuppression and, therefore, 
potentially increase the risk of severe 
infection. We will ensure that patients with 
COVID-19 (+) are not enrolled in the study, 
regardless of their symptoms or severity. 
 
 
 
 
 
 
 


  
1 
TABLE OF CONTENTS 
STATEMENT OF COMPLIANCE ....................................................................................... 1 
1 
PROTOCOL SUMMARY .............................................................................................. 2 
1.1 
Synopsis ............................................................................................................................2 
1.2 
Schema ..............................................................................................................................6 
1.3 
Schedule of Activities (SoA) ...........................................................................................9 
2 
INTRODUCTION ......................................................................................................... 11 
2.1 
Study Rationale............................................................................................................... 11 
2.2 
Background ..................................................................................................................... 11 
2.3 
Risk/Benefit Assessment .............................................................................................. 14 
2.3.1 
Known Potential Risks ........................................................................................................ 14 
2.3.2 
Known Potential Benefits ................................................................................................... 15 
2.3.3 
Assessment of Potential Risks and Benefits .................................................................. 16 
3 
OBJECTIVES AND ENDPOINTS .............................................................................. 17 
4 
STUDY DESIGN.......................................................................................................... 19 
4.1 
Overall Design................................................................................................................. 19 
4.2 
Scientific Rationale for Study Design .......................................................................... 23 
4.3 
Justification for Dose ..................................................................................................... 24 
4.4 
End of Study Definition .................................................................................................. 24 
5 
STUDY POPULATION ............................................................................................... 24 
5.1 
Inclusion Criteria ............................................................................................................ 25 
5.2 
Exclusion Criteria ........................................................................................................... 25 
5.3 
Lifestyle Considerations ................................................................................................ 26 
5.4 
Screen Failures ............................................................................................................... 26 
5.5 
Strategies for Recruitment and Retention ................................................................... 27 
5.5.1 
Use of DataCore/Epic Information for Recruitment Purposes ..................................... 28 
6 
STUDY INTERVENTION ............................................................................................ 28 
6.1 
Study Intervention Administration ............................................................................... 28 
6.1.1 
Study Intervention Description .......................................................................................... 28 
6.1.2 
Dosing and Administration ................................................................................................. 29 
6.2 
Preparation/Handling/Storage/Accountability ............................................................ 31 
6.2.1 
Acquisition and accountability ........................................................................................... 31 
6.2.2 
Formulation, Appearance, Packaging, and Labeling ..................................................... 31 
6.2.3 
Product Storage and Stability ............................................................................................ 32 
6.3 
Measures to Minimize Bias: Randomization and Blinding ........................................ 32 
6.4 
Study Intervention Compliance .................................................................................... 34 
6.5 
Concomitant Therapy..................................................................................................... 34 
Prohibited medications ...................................................................................................................... 34 


  
6.5.1 
Rescue Medicine ................................................................................................................. 35 
7 
STUDY INTERVENTION DISCONTINUATION AND PARTICIPANT 
DISCONTINUATION/WITHDRAWAL .............................................................................. 36 
7.1 
Discontinuation of Study Intervention ......................................................................... 36 
7.2 
Participant Discontinuation/Withdrawal from the Study ........................................... 36 
7.3 
Lost to Follow-Up ........................................................................................................... 37 
8 
STUDY ASSESSMENTS AND PROCEDURES ........................................................ 37 
8.1 
Efficacy Assessments ................................................................................................... 37 
8.2 
Safety and Other Assessments ............................................................................................ 40 
8.3 
Adverse Events and Serious Adverse Events ............................................................ 41 
8.3.1 
Definition of Adverse Events (AE) .................................................................................... 41 
8.3.2 
Definition of Serious Adverse Events (SAE) ................................................................... 41 
8.3.3 
Classification of an Adverse Event ................................................................................... 41 
8.3.4 
Time Period and Frequency for Event Assessment and Follow-Up ........................... 42 
8.3.5 
NON-SERIOUS Adverse Event Reporting ...................................................................... 43 
8.3.6 
Serious Adverse Event Reporting .................................................................................... 43 
8.4 
Unanticipated Problems ................................................................................................ 44 
8.4.1 
Definition of Unanticipated Problems (UP)...................................................................... 44 
8.4.2 
Unanticipated Problem Reporting ..................................................................................... 44 
9 
STATISTICAL CONSIDERATIONS ........................................................................... 45 
9.1 
Statistical Hypotheses ................................................................................................... 45 
9.2 
Sample Size Determination ........................................................................................... 46 
9.3 
Populations for Analyses .............................................................................................. 46 
9.4 
Statistical Analyses ........................................................................................................ 47 
9.4.1 
General Approach ............................................................................................................... 47 
9.4.2 
Analysis of the Primary Efficacy Endpoint ....................................................................... 47 
9.4.3 
Analysis of the Secondary Endpoint ................................................................................ 47 
9.4.4 
Safety and tolerability analyses ........................................................................................ 48 
9.4.5 
Planned Interim Analyses .................................................................................................. 48 
9.4.6 
Sub-Group Analyses ........................................................................................................... 49 
10 
SUPPORTING DOCUMENTATION AND OPERATIONAL CONSIDERATIONS. 49 
10.1 
Regulatory, Ethical, and Study Oversight Considerations .................................... 49 
10.1.1 Informed Consent Process ................................................................................................ 49 
10.1.2 Study Discontinuation and Closure .................................................................................. 50 
10.1.3 Confidentiality and Privacy ................................................................................................ 50 
10.1.4 Future Use of Stored Specimens and Data .................................................................... 51 
10.1.5 Key Roles and Study Governance ................................................................................... 51 
10.1.6 Safety Oversight and clinical monitoring ......................................................................... 52 
10.1.7 Quality Assurance and Quality Control ............................................................................ 52 
10.1.8 Data Handling and Record Keeping ................................................................................. 53 


  
10.1.9 Protocol Deviations ............................................................................................................. 55 
10.1.10 
Publication and Data Sharing Policy ............................................................................ 55 
10.1.11 
Conflict of Interest Policy ................................................................................................ 56 
10.2 
Abbreviations .............................................................................................................. 57 
10.3 
Protocol Amendment History .................................................................................... 59 
11 
REFERENCES ......................................................................................................... 61 
 


A futility trial of sirolimus in MSA 
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Protocol # 17-01392 
15-Oct- 2018 
 
1
STATEMENT OF COMPLIANCE 
 
The trial will be conducted in accordance with International Conference on Harmonisation Good Clinical 
Practice (ICH GCP), applicable United States (US) Code of Federal Regulations (CFR), and the NINDS 
Terms and Conditions of Award. The Principal Investigator will assure that no deviation from, or changes 
to the protocol will take place without prior agreement from the Investigational New Drug (IND) 
sponsor, funding agency and documented approval from the Institutional Review Board (IRB), except 
where necessary to eliminate an immediate hazard(s) to the trial participants. All personnel involved in 
the conduct of this study have completed Human Subjects Protection and ICH GCP Training. 
 
The protocol, informed consent form, recruitment materials, and all participant materials will be 
submitted to the IRB for review and approval.  Approval of both the protocol and the consent form must 
be obtained before any participant is enrolled.  Any amendment to the protocol will require review and 
approval by the IRB before the changes are implemented to the study.  All changes to the consent form 
will be IRB approved; a determination will be made regarding whether a new consent needs to be 
obtained from participants who provided consent, using a previously approved consent form. 
 
 


A futility trial of sirolimus in MSA 
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1 
 
PROTOCOL SUMMARY 
1.1 
SYNOPSIS  
 
Title: 
A futility trial of sirolimus in multiple system atrophy (MSA) 
Study Description: 
Single-center, randomized, placebo-controlled, phase-II, futility clinical trial 
to determine if oral sirolimus is of sufficient promise to slow disease 
progression in MSA, prior to embarking on a large-scale and costly phase III 
study to assess its efficacy. A futility design under the null hypothesis 
assumes that sirolimus will slow the progression of the disease, whereas the 
alternative hypothesis assumes no benefit of sirolimus. If the null 
hypothesis is rejected (i.e., futility of sirolimus to slow progression of MSA), 
a major phase III study will be discouraged, whereas non-futility will offer 
strong support for a phase III trial to detect clinical efficacy.  
Objectives: 
 
Primary objective: To determine if oral sirolimus is of sufficient promise or 
futile to slow disease progression in MSA. 
Secondary objectives: To assess the potential effects of sirolimus on retinal 
and brain neuroimaging in MSA 
Exploratory objective: To explore the potential effects of sirolimus on 
blood biomarkers (exosomal alpha-synuclein particles) in MSA. 
Endpoints: 
Primary Endpoint: Change from baseline to 48 weeks (or last available date) 
in United Multiple System Atrophy Rating Score (UMSARS) total score 
compared with described progression rates in historical controls. We will 
assume that patients receiving sirolimus will have a change of 4 points less 
than those reported in natural history studies (see statistical section). 
 
Secondary Endpoints:  
1. Change from baseline to 48 weeks (or last available date) in 
UMSARS-2 subscale score. We will assume that patients receiving 
sirolimus will have a change of 2 points less than those reported in 
historical controls. 
2. Change from baseline to 48 weeks (or last available date) in 
UMSARS-1 subscale score. We will assume that patients receiving 
sirolimus will have a change of 2 points less than those reported in 
historical controls. 
3. Magnetic resonance changes using dedicated algorithms to 
evaluate rate of change in diffusivity of defined areas of brain from 
baseline to 48 weeks. 
4. Changes in retinal nerve fiber layer using optical coherence 
tomography from baseline to 48 weeks.  
 
Exploratory outcome 
1. Change in plasma biomarkers (exosomal alpha-synuclein particle 
concentration) from baseline to 48 months. 


A futility trial of sirolimus in MSA 
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Protocol # 17-01392 
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Study Population: 
A total of 56 patients with multiple system atrophy will be randomized (42 
to active agent and 14 to placebo) 
Inclusion criteria: 
1. Participants aged 30-80 years old with a diagnosis of MSA. Patients 
have to fulfill current consensus criteria (1) for probable MSA of 
the parkinsonian subtype (MSA-P) or cerebellar subtype (MSA-C). 
2. Participants who are less than 4 years from the time of 
documented MSA diagnosis. 
3. Participants who are still able to walk with or without assistance. 
4. Participants with an anticipated survival of at least 3 years in the 
opinion of the investigator. 
5. Participants who are willing and able to give informed consent. 
6. Montreal Cognitive Assessment (MoCA) > 20. 
7. Ability to take oral medication and be willing to adhere to the 
study drug regimen 
8. For females of reproductive potential: use of highly effective 
contraception during study participation and for an additional 8 
weeks after the end of study drug administration  
9. For males of reproductive potential: use of condoms or other 
methods to ensure effective contraception with partner 
10. Agreement to adhere to Lifestyle Considerations (see section 5.3) 
throughout study duration 
Exclusion criteria:  
1. Women of childbearing potential who do not practice an 
acceptable method of birth control. Acceptable methods of birth 
control in this study are: surgical sterilization, intrauterine devices, 
partner’s vasectomy, a double-protection method (condom or 
diaphragm with spermicide), hormonal contraceptive drug (i.e., 
oral contraceptive, contraceptive patch, long-acting injectable 
contraceptive) with a required second mode of contraception. 
2. Participants with a clinically significant or unstable medical or 
surgical condition that, in the opinion of the investigator, might 
preclude safe completion of the study or might affect the results 
of the study. These include but are not limited to severe, 
uncontrolled heart failure, recent (<6 months) myocardial infarct, 
severe, uncontrolled cardiopulmonary disease, severe, 
uncontrolled hypertension (sustained systolic blood pressure 
above 200 mmHg in any position), thrombocytopenia (< 50 x 
10(9)/L), severe anemia (< 8g/dl), immunocompromised state, 
liver or kidney disease (creatinine > 1.5 mg/dl or proteinuria > 20 
mg/dl), uncontrolled diabetes mellitus (HbA1c >10g%), alcoholism, 
amyloidosis, uncontrolled hypothyroidism, sympathectomy, 
unstable peripheral neuropathies, concurrent infections, severe 
orthopedic problems that compromise mobility and activity of 
daily living, recent (< 6 month) acute cerebrovascular accidents, 
neurotoxin or neuroactive drug exposure, parkinsonism due to 
drugs (including neuroleptics, alpha-methyldopa, reserpine, 
metoclopramide). 
3. Participants with high LDL cholesterol levels (LDL > 190 mg/dL) 
and/or high triglycerides levels (> 500 mg/dL).  Patients receiving 


A futility trial of sirolimus in MSA 
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15-Oct- 2018 
 
4
medications for elevated cholesterol or triglycerides are eligible to 
participate as long as their LDL cholesterol and/or triglyceride 
levels are below the abovementioned limits. 
4. Participants with positive interferon-gamma release-assay 
(QUANTIferon®) indicating latent tuberculosis infection. 
Participants with a positive Quantiferon® may be enrolled after 
the patient has been evaluated by an infectious diseases specialist 
that has cleared the patient for this trial. 
5. Participants with history of untreated tuberculosis. Participants 
with a history of treated tuberculosis may be enrolled after an 
infectious diseases specialist evaluates patient and has cleared the 
patient for this trial. 
6. Participants with a history of active, acute or chronic, or latent 
hepatitis B or hepatitis C. 
7. Participants with a nasal swab PCR positive for COVID-19. If 
positive, participant may be re-screened after 1 month. If 
the second nasal swab PCR is negative for COVID-19, 
participant may be enrolled.  
8. Participants with human immunodeficiency virus (HIV) infection, 
or other congenital or acquired causes of immunosuppression. 
9. Participants with active malignant neoplasms or history of 
malignant neoplasm in the last 5 years. 
10. Movement disorders other than MSA; e.g., Parkinson disease, 
dementia with Lewy bodies, essential tremor, progressive 
supranuclear palsy, spinocerebellar ataxia, spastic paraparesis, 
corticobasal degeneration, or vascular, pharmacological or post-
encephalitic parkinsonism. 
11. Dementia (DSM-V criteria).  
12. History of electroconvulsive therapy. 
13. History of deep brain stimulation surgery. 
14. Patients with contraindication for MRI scanning, including those 
with MRI-incompatible pacemaker 
15. History of organ transplant 
16. Participants who have taken any investigational products within 60 
days prior to baseline. 
17. Treatment with cyclosporine, oral or intravenous corticosteroids, 
methotrexate, rituximab within 3 months prior to baseline.  
18. Treatment with inhibitors of CYP3A4 (which may decrease the 
metabolism of sirolimus and increase sirolimus levels): nicardipine, 
verapamil, clotrimazole, fluconazole, itraconazole, clarithromycin, 
erythromycin, troleandomycin, cisapride, metoclopramide, 
bromocriptine, cimetidine, danazol, HIV-protease inhibitors (e.g., 
ritonavir, indinavir); grapefruit. 
19. Treatment with inducers of CYP3A4 (which may increase the 
metabolism of sirolimus and decrease sirolimus levels): 
carbamazepine, phenobarbital, phenytoin, rifabutin, rifapentine. 
20. Inability or unwillingness of subject to give written informed 
consent. 


A futility trial of sirolimus in MSA 
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Protocol # 17-01392 
15-Oct- 2018 
 
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Phase: 
Phase II 
Description of Sites: 
Single-center clinical trial performed at the New York University Langone 
Medical Center 
Description of Study 
Intervention: 
Oral sirolimus capsules at a dosage of 2 mg (one 2 mg capsule), 4 mg (two 
2 mg capsules) or 6 mg (three 2 mg capsules) taken once a day. Dosage will 
be adjusted based on determination of sirolimus levels in blood performed 
throughout the trial and on the frequency of adverse events related to the 
study drug.  
Study Duration: 
3 years 
Participant Duration: 
48 weeks plus a safety follow-up phone call 2 weeks after the end of the 
trial. 
 
 


A futility trial of sirolimus in MSA 
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Protocol # 17-01392 
15-Oct- 2018 
 
6
 
 
 
1.2 
SCHEMA 
 


A futility trial of sirolimus in MSA 
Version 4 
Protocol # 17-01392 
15-Oct- 2018 
 
7
 
Screening
Month -1
•Total n=56
•Obtain informed consent
•Screen potential participants by inclusion and exclusion criteria
•Obtain history,  document concomitant medications, MSA diagnosis and demographics
•Asessments: EKG, MoCA, Physical examination and UMSARS, vital signs
•Safety assessments: CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel, urine pregnacy test, HIV, hepatitis B and C, QuantiFERON
Randomization and baseline visit
Day 1
•Sirolimus (n=42)
•Placebo (n=14)
•Perform randomization
•EKG, MoCA, Physical examination and UMSARS, vital signs
•Safety assessments: CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel
•Collect blood for exosomes
•Perform brain MRI (+/- 5 days)
•Perform retinal OCT (+/- 5 days)
•Administer initial dose of study intervention
•Dispense study drug
Study drug levels
Day 8, Day 15, Day 22
•Measure sirolimus plasma levels (+3 days) and adjust study drug dose accordingly
Safety labs (can be performed locally; adverse event recording performed over the phone)
Month 1 and Month 2
•CBC, CMP, CPK, urinalysis, and 24-hour urine collection, lipid panel, and plasma sirolimus levels.
•Adverse event recording
Follow-up assessments of study endpoints and safety
Month 3
•Concomitant medications
•EKG, physical exam, vital signs, height and weight and UMSARS
•CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel, and plasma sirolimus levels.
•Adverse event recording
•Retrieve study drug
•Dispense study drug
Safety labs (can be performed locally; adverse event recording performed over the phone)
Month 4 and Month 5
•CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel, and plasma sirolimus levels.
Adverse event recording
Follow-up assessments of study endpoints and safety
Month 6
•EKG, physical exam, vital signs, height and weight and UMSARS
•Adverse event recording
•CBC, CMP, CPK, urinalysis, and 24-hour urine collection lipid panel, and plasma sirolimus levels.
•Obtain blood for exosomes
•Retrieve study drug
•Dispense study drug 
Safety labs (can be performed locally; adverse event recording performed over the phone)
Month 7 and Month 8
•CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel, and plasma sirolimus levels.
Adverse event recording
Follow-up assessment
Month 9
•EKG, physical exam, vital signs, height and weight and UMSARS
•Adverse event recording
•CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel, and plasma sirolimus levels.
•Retrieve study drug
•Dispense study drug
Safety labs (can be performed locally)
Month 10 and Month 11
•CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel, and plasma sirolimus levels.
•Adverse evente recording
End of study visit
Month 12
•MoCA, Physical examination and UMSARS, vital signs
•Safety assessments: CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel
•Collect blood for exosomes
•Perform brain MRI (+/- 5 days)
•Perform retinal OCT (+/- 5 days)
•Retrieve study drug
•End of study 
Follow-up phone call
Month 12.5
•Adverse event recording
Final safety assessment (can be performed locally; adverse event recoring performed over the phone)
Month 13
•Safety assessments: CBC, CMP, CPK, urinalysis and 24-hour urine collection, lipid panel
•Adverse event recording


A futility trial of sirolimus in MSA 
Version 4 
Protocol # 17-01392 
15-Oct- 2018 
 
8


A futility trial of sirolimus in MSA 
Version 4 
Protocol # 17-01392 
15-Oct- 2018 
 
9
  
1.3 
SCHEDULE OF ACTIVITIES (SOA) 
 
 
Procedures 
Screening (V1) 
Day -42 to -1 
Baseline / Random. 
Visit 2, Day 1
Study Visit 3  
Day 7 +3 day
Study Visit 4 
Day 14 + 3 day
Study Visit 5 
Day 21 +3 day
Study Visit 6 
Day 28 +3 day
Study Visit 7 
Day 56 +15 day
Study Visit 8 
Day 84 +15 day
Study Visit 9 
Day 112 +15 day
Study Visit 10 
Day 140 +15 day
Study Visit 11 
Day 168 +15 day
Study Visit 12 
Day 196 +15 day
Study Visit 13 
Day 224 +15 day
Study Visit 14 
Day 252 +15 day
Study Visit 15 
Day 280 +15 day
Study Visit 16 
Day 308 +15 day
End of study V17 
Day 336 +15 day
Phone followup V18 
Day 350 +/-5 day
Safety assess. V19 
Day 365 +/-5 day
Visit type 
S 
S 
SL 
SL 
SL 
SL 
SL 
S 
SL 
SL 
S 
SL 
SL 
S 
SL 
SL 
S 
P 
SL 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Randomization 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Administer study 
intervention 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Dispense study drug 
 
X 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
 
 
 
Retrieve study drug 
 
 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
Conclusion of study  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Conc. Medication review 
X 
X 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
Physical exam 
X 
X 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
Vital signs, height & 
weight 
X 
X 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
MSA criteria 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
MoCA 
X 
Xe 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
UMSARS 
X 
Xe 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
Sirolimus levels in blood d 
 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
Complete blood count  
X 
Xe 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
Comprehensive metabolic 
a 
X 
Xe 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
CPK 
X 
Xe 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
Lipid panel 
X 
Xe 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
Urinalysis 
X 
Xe 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
24-hour urine collection 
X 
X C 
 
 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
Pregnancy test b 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
HIV testing, hepatitis B 
and C testing, 
QuantiFERON® 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Nasal swab PCR COVID-
19 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
EKG 
X 
Xe 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
Adverse event review  
 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Brain MRI 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
Retinal OCT 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
Blood sample for 
exosomes 
 
X 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
X 
 
 
Complete Case Report 
Forms  
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
a: 
Albumin, alkaline phosphatase, total bilirubin, bicarbonate, BUN, calcium, chloride, creatinine, glucose,potassium, total 
protein, AST, ALT, sodium. 
B: 
Serum pregnancy test (women of childbearing potential). 
c.      24-hour urine collection at baseline will be performed only if the baseline visit occurs > 30 days from the 24-hour urine 
assessment performed as part of the screening visit.  
d.     Sirolimus blood level should be drawn 7 days after previous dose adjustment has taken place. 
e.     If baseline visit (Visit 2) occurs within 15 days of the screening visit, the selected procedures can be foregone and the 
results of the screening day will be used.  
S: In-site visit at NYU (denoted in grey) 
SL: Safety labs visit, this can be done either at NYU or locally. If done locally, a phone call will be performed to inquire about 
adverse events. Safety labs should be drawn after a light (low fat) breakfast. 
P: Phone call or video-conference call 


A futility trial of sirolimus in MSA 
Version 4 
Protocol # 17-01392 
15-Oct- 2018 
 
10
Procedures 
Screening (V1) 
Day -42 to -1 
Baseline / Random. 
Visit 2, Day 1
Study Visit 3  
Day 7 +3 day
Study Visit 4 
Day 14 + 3 day
Study Visit 5 
Day 21 +3 day
Study Visit 6 
Day 28 +3 day
Study Visit 7 
Day 56 +15 day
Study Visit 8 
Day 84 +15 day
Study Visit 9 
Day 112 +15 day
Study Visit 10 
Day 140 +15 day
Study Visit 11 
Day 168 +15 day
Study Visit 12 
Day 196 +15 day
Study Visit 13 
Day 224 +15 day
Study Visit 14 
Day 252 +15 day
Study Visit 15 
Day 280 +15 day
Study Visit 16 
Day 308 +15 day
End of study V17 
Day 336 +15 day
Phone followup V18 
Day 350 +/-5 day
Safety assess. V19 
Day 365 +/-5 day
CPK: Creatine Phosphokinase 
 
 
 
 
 
 


A futility trial of sirolimus in MSA 
Version 4 
Protocol # 17-01392 
15-Oct- 2018 
 
11
 
2 
 
INTRODUCTION 
 
2.1 
STUDY RATIONALE  
 
Multiple system atrophy (MSA) is an adult-onset, fatal neurodegenerative disease characterized by 
progressive autonomic failure, parkinsonism, and cerebellar and pyramidal features in various 
combinations (2). Depending on their presenting predominant motor deficits, MSA is sub-classified into 
parkinsonian (MSA-P) or cerebellar (MSA-C) variant (3). MSA is an orphan disease (i.e., less than 200,000 
cases in the U.S.). The estimated mean incidence is 0.6 to 0.7 cases per 100,000 person-years, with a 
range of 0.1 to 2.4 cases per 100,000 person-years (4). The estimated point prevalence is 3.4 to 4.9 cases 
per 100,000 inhabitants, increasing to 7.8 per 100,000 among persons older than 40 years of age (5). 
Cases of the parkinsonian subtype outnumber cases of the cerebellar subtype in most countries by 2:1 
to 4:1, although the cerebellar subtype is more frequent in Japan (2). Disease onset is usually in the sixth 
decade of life, with both sexes equally affected (6). MSA is a relentlessly progressive disease, and the 
mean survival from the onset of symptoms is ~8 years (3, 6), with few patients with autopsy-proven 
MSA surviving more than 15 years (7). 
 
MSA is the most fatally progressive of synucleinopathies, disorders characterized by the abnormal 
accumulation of the protein α-Synuclein (αSyn) in the nervous system (3, 8). In contrast to Parkinson 
disease (PD), in which αSyn predominantly accumulates in neurons forming Lewy bodies, in MSA it 
accumulates predominantly in glia forming glial cytoplasmic inclusions (GCI) (9). Immunohistochemical 
analysis of the inclusions shows that they stain with antibodies to a variety of proteins in addition to α-
syn. These include ubiquitin, tubulin, synphilin-1, and DJ-1. Although GCI are the primary 
neuropathological hallmark of MSA, neuronal cytoplasmic and nuclear inclusions of αSyn have also been 
reported. Neuronal loss occurs in the striatum, cerebellum, brainstem, and cortex; and is accompanied 
by astrogliosis, microgliosis, and myelin loss (10, 11). Neurodegeneration in MSA also affects the retinal 
ganglion cells, a process that can be non-invasively quantified in-vivo with optical coherence 
tomography (OCT) (12-14).  
 
Currently there are no disease-modifying treatments for MSA. Therefore, there is an urgent unmet 
medical need for therapies than can halt or slow the progression of this devastating disease. Many lines 
of evidence highlight the pathogenic importance of αSyn aggregation and spread in MSA (15-18). 
Although the mechanisms that lead to this aggregation remain to be elucidated, there is evidence to 
support roles for neuroinflammation (19), oxidative stress (20), and defective lysosome-mediated 
clearance of protein aggregates. Therefore, potential therapeutic options for MSA are aimed at 
correcting these mechanisms (21). 
 
2.2 
BACKGROUND  
 
Autophagy is the major cellular digestion process that removes damaged macromolecules, organelles 
and abnormally misfolded proteins, including αSyn (22). Under normal conditions, autophagy is 
regulated by the protein mammalian target of rapamycin (mTOR): there is a tight, direct coupling of 
autophagy induction and mTOR inhibition (23). In synucleinopathies such as MSA, the presence of αSyn 
aggregates is associated with accumulation of autophagosomes and reduction of lysosomes in neurons, 
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with PD and MSA, misfolded αSyn fails to be processed and αSyn clearance is blocked (24, 25). 
Pharmacological activation of the autophagy function by inhibition of mTOR with rapamycin (a.k.a. 
sirolimus) produces neuroprotective effects in animal and cellular models of synucleinopathies (26-28). 
Thus, inhibition of the mTOR pathway with sirolimus has been proposed to be a highly efficient strategy 
to block αSyn-induced neurodegeneration (28, 29). 
 
Sirolimus is an orally active macrolide compound and a potent immunosuppressant agent and is 
currently approved by the U.S. Food and Drug Administration in 1999 to prevent organ transplant 
rejection and, since 2015, for the treatment of lymphangioleiomiomatosis (LAM), a rare progressive lung 
disease (30). Thus, sirolimus has been widely used for almost 20 years and its adverse events profile is 
relatively safe and well known.  
 
Sirolimus is also one of the few pharmacological molecules known to effectively induce autophagy (23).  
 
Of interest to this protocol, there is ample evidence that sirolimus is lipophilic and crosses the brain-
blood barrier of murine animals and humans (31-35), the levels of sirolimus in CSF and blood correlate in 
humans (31), and that sirolimus effectively inhibits mTOR activity in CNS cells of humans (31). 
In addition to a potentially promising drug, two developments enhance the probability of success of this 
study. The first is that NYU is the main site of the international NIH-sponsored Natural History Study of 
Multiple System Atrophy (PI: Dr. Horacio Kaufmann), which is currently being conducted within the NIH-
sponsored Rare Disease Autonomic Disorders Clinical Research Consortium (RDCRC; U54 NS065736; PI: 
Dr. David Robertson). This Natural History Study includes a cohort of ~200 patients with MSA fully 
characterized neurologically, being followed closely with validated instruments to fully document and 
define neurologic and autonomic symptoms and deficits. The second development is the use of 
instruments and objective outcome measures which have been validated in longitudinal studies of MSA 
patients. These include the Unified Multiple System Atrophy Rating Scale (UMSARS), measurements of 
putaminal diffusivity by brain magnetic resonance imaging (MRI), and measurement of retinal nerve 
fiber layer thickness by optical coherence tomography (OCT).    
 
The UMSARS is a scale that comprises 4 parts. UMSARS-1 scores symptoms of neurological and 
autonomic dysfunction. UMSARS-2 is a motor examination. UMSARS-3 is a cardiovascular autonomic 
examination; and UMSARS-4, a global disability scale (36). The UMSARS in the only validated 
investigator-administered rating scale to reliably measure semi-quantitatively symptoms and signs in 
MSA. The instrument has high inter-rater reliability, and internal consistency, for instance the UMSARS-1 
has a Crohnbach’s alpha of 0.84 and the UMSARS-2 of 0.90 (36). The instrument is sensitive in measuring 
disease progression (37). In a study to assess rates of disease progression in MSA and validating UMSARS 
for sensitivity to change over time, MSA patients were assessed at two time points 12 months apart 
using the UMSARS 1-4. Fifty patients were assessed twice with an interval of 12.3 months. UMSARS-2 
scores progressed by 57.3% (P<0.0001) and the UMSARS-1 scores by 35.6% (P<0.0001) in relation to the 
respective baseline scores with no differences between motor subtypes, diagnostic categories and 
gender (37). More recently, the minimally clinically important difference (MCID) for the UMSARS scores 
were determined (38): the MCID for the UMSARS-1 and the UMSARS-2 was 1.5 points, whereas it was of 
3.5 points for the UMSARS total score (UMSARS-1 + UMSARS-2). Already completed natural history 
studies of MSA (8, 39) have reported the annual rates of change of UMSARS. This has also been reported 
in the placebo group of a previous clinical trial with rifampicin for MSA (40). These developments are 
relevant because appreciation of the MCID as well as the described annual changes in UMSARS are 
crucial for statistically significant effects and power analysis calculation (see statistical section). 
 


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The rate of progression of objective markers in MSA has been reported in longitudinal studies measuring 
the thickness of the retinal nerve fiber layer using retinal OCT (12-14), as well as in studies measuring 
changes in diffusion to water molecules in the pons and the putamen with specific sequences of brain 
MRI (41, 42). These markers have been specifically measured at baseline and after 12 months, are able 
to detect change, correlate with disease progression, and appear to be good candidates to be used as 
objective outcome measures in clinical trials of MSA.  
 
2.2.1. Mechanism of action of sirolimus  
Sirolimus inhibits T lymphocyte activation and proliferation that occurs in response to antigenic and 
cytokine (interleukin [IL]-2, IL-4, and IL-15) stimulation by a mechanism that is distinct from that of other 
immunosuppressants.  Sirolimus also inhibits antibody production. In cells, sirolimus binds to the 
immunophilin, FK binding protein-12 (FKBP-12), to generate an immunosuppressive complex.  The 
sirolimus:FKBP-12 complex binds to and inhibits the activation of the mTOR. This inhibition suppresses 
cytokine-driven T-cell proliferation, inhibiting the progression from the G1 to the S phase of the cell 
cycle. Studies in experimental models show that sirolimus prolongs allograft (kidney, heart, skin, islet, 
small bowel, pancreatico-duodenal, and bone marrow) survival and reverses acute rejection of heart 
and kidney allografts in animal models.  In some studies, the immunosuppressive effect of sirolimus 
lasted up to 6 months after discontinuation of therapy. In animal models of autoimmune disease, 
sirolimus suppresses immune-mediated events associated with systemic lupus erythematosus, collagen-
induced arthritis, autoimmune type I diabetes, autoimmune myocarditis, experimental allergic 
encephalomyelitis, graft-versus-host disease, and autoimmune uveo-retinitis. 
 
2.2.2. mTOR signaling pathway 
mTOR is a serine/threonine protein kinase that oversees multiple functions in cells that involve gene 
transcription, protein synthesis, cytoskeletal organization, cell metabolism, proliferation and survival. In 
mammalian cells, there are two mTOR complexes: mTOR complex 1 (mTORC1) and mTOR complex 2 
(mTORC2). mTORC1 controls protein homeostasis and autophagy, whereas mTORC2 regulates cell 
survival and proliferation. mTORC1 and mTORC2 share three regulatory components in common, 
including Deptor, mLST8 and Tti1/Tel2. Despite the presence of a catalytic mTOR subunit in both 
mTORC1 and mTORC2, mTORC1 (involved in autophagy) is more sensitive than mTORC2 (involved in cell 
survival) to the inhibition by sirolimus (25). 
 
2.2.3. Pre-clinical data on efficacy of sirolimus in synucleinopathy models 
Overall, there is accumulating evidence that treatment with sirolimus exerts neuroprotective effects and 
improves motor function in synucleinopathy mouse models by increasing αSyn autophagy and, possibly, 
reducing neuroinflammatory processes. Masliah’s group (43) delivered a lentivirus expressing beclin-1, 
an activator of autophagy, into the brain of a synucleinopathy mouse model. This approach reduced 
αSyn pathology in the limbic system without deleterious effects. Rapamycin (i.e., sirolimus) enhanced 
the beneficial effects of beclin-1 (43). Malagelada and colleagues (44) found that treatment with 
rapamycin (i.e., sirolimus) at a dose of 7.5 mg/kg protected neurons from death in MPTP-induced mice 
models of synucleinopathy, by sparing phosphorylation of the survival kinase Akt. In an MPTP-induced 
synucleinopathy mouse model, treatment with rapamycin (i.e., sirolimus) at a dosage of 5 mg/kg/day for 
up to 4 days restored lysosomal levels, and attenuated αSyn accumulation, which was associated with 
an attenuation of neurodegeneration in the animals (29). Rapamycin (i.e., sirolimus) at a dosage of 5 
mg/kg/day for 7 days protected against neuronal loss, and alleviated mitochondrial ultrastructural 
damage in a 6-hydroxydopamine rat model of synucleinopathy (45). Moreover, antioxidant activity and 
expression of anti-apoptotic markers was significantly higher in the synucleinopathy rats treated with 
rapamycin compared to those treated with placebo (45). Wills and colleagues (46) performed a dual 


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immunohistochemical staining with co-localization of αSyn with mTOR in striatal and midbrain sections 
of paraquat-induced synucleinopathy mice or control mice. Dual staining of αSyn and mTOR in striatal 
and midbrain sections showed increased levels of both proteins that were co-localized with one another 
in the synucleinopathy mice but not in the control mice. Western blot analysis confirmed increased 
levels of mTOR only in the synucleinopathy mice (46). To expand their results, the same group analyzed 
postmortem brain samples of patients with synucleinopathies finding similar results: mTOR levels were 
increased in the striatum of patients compared to age-matched controls (46). Liu and colleagues (47) 
showed that the number of dopaminergic neurons in the substantia nigra of MPTP-induced mice with 
synucleinopathy was significantly higher in animals treated with rapamycin compared to saline-treated 
controls. αSyn aggregates were significantly decreased in rapamycin-treated mice compared to controls 
(47). Bai and colleagues (48) studied whether a 24-week course of rapamycin at a dose of 2.25 
mg/kg/day had any behavioral or motor effect in αSyn transgenic mice.  Rapamycin-treated mice 
showed improved performance on the forepaw stepping adjustment test, accelerating rotarod and pole 
test (48). Siracusa and colleagues (49) showed that treatment with temsirolimus (an intravenous analog 
of sirolimus) at a dosage of 5 mg/kg/day for 8 days significantly ameliorated behavioral deficits, 
increased tyrosine hydroxylase and dopamine transporter expression, and decreased αSyn in the 
substantia nigra of MPTP-induced synucleinopathy mice. Western blot and immunohistochemistry 
showed that temsirolimus increased autophagy significantly. Temsirolimus also had anti-inflammatory 
properties as assessed by significant inhibition of the expression of mitogen-activated protein kinases 
such as p-JNK, p-p38, and p-ERK. Collectively, these findings suggest that inhibition of mTOR with 
sirolimus in MSA may be beneficial (26). 
 
mTOR inhibition with sirolimus or analogs also has demonstrated neuroprotective effects in models of 
other disorders caused by abnormal accumulation of misfolded proteins such as Huntington’s disease or 
Alzheimer’s disease  (35, 50-53). 
 
Thus, this clinical trial is based on i) the unmet medical need of a disease-modifying treatment for 
patients with MSA; ii) the plausible biological mechanism of sirolimus increasing phagocytosis and 
clearance of αSyn; and iii) robust preclinical data from rigorously performed studies on the effect of 
sirolimus in cellular and murine models of synucleinopathy demonstrating that there is an adequate 
scientific foundation to justify this futility trial of sirolimus in patients with MSA. 
 
2.3 
RISK/BENEFIT ASSESSMENT   
 
2.3.1 KNOWN POTENTIAL RISKS  
 
Specific adverse reactions associated with the administration of sirolimus 2 mg once a day occurred at a 
significantly higher frequency than in the respective control group. These include hypercholesterolemia, 
hyperlipemia, hypertension, rash, acne, anemia, arthralgia, diarrhea, hypokalemia, and 
thrombocytopenia. The elevations of triglycerides and cholesterol and decreases in platelets and 
hemoglobin occurred in a dose related manner in patients receiving sirolimus (see FDA-approved 
package insert information (54)).  Combination of sirolimus with statins and other medications for 
dyslipidemia has been shown to increase the risk of rhabdomyolysis. To evaluate the risk of 
rhabdomyolysis, CPK will be checked at each study visit including safety laboratory assessments, and 
patients will be specifically asked about new-onset myalgia at each study visit.    
  


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Risk involving non-contrast brain magnetic resonance imaging (MRI): There are no known risks or 
adverse effects resulting directly from exposure to non-contrast MRI.  However, subjects who have a 
pacemaker or metal objects in their body such as shrapnel or metal in the eye should not have the scan 
performed. Every care will be taken to prevent this, including a thorough interview of each patient and 
review of his/her medical history to ensure that no metallic implants are present in patient’s body. 
There is a potential risk of burns in the event that patients are inadvertently carrying metallic 
ferromagnetic objects or particles in or on the patient. These include but are not limited to pulse 
15ximeter, EKG leads, or skin tattoos. Every care will be taken to prevent this, including a thorough 
screening of each patient to ensure that no metallic materials are present in or on the patient’s body. 
Additionally, the power limit of the brain MRI will be adjusted as necessary. The introduction of metallic 
ferromagnetic objects in the scanner room may result in magnetic displacement of the metallic object 
causing injuries to the patient. To reduce this risk, patients and every medical personnel near the MRI 
magnet will remove metal contents from them.  
Some people may feel confined and experience anxiety in the MR scanner. In the patient experiences 
this during the scanning procedure, the scanner will be stopped immediately. Discussion with the PI and 
the radiology team will follow, and measures to prevent this could be implemented, such as 
administering a patient a low dose of anxiolytic medication prior to the scanning.  
The MR scanner produces tapping sounds during operation, which may reach very loud levels.  To 
minimize any discomfort from this noise, patients will be given disposable earplugs to reduce the noise 
levels but will still allow voice communication with the scanner operator. 
In extremely rare cases, a magnet can lose its magnetism, in which case cooling fluids may be released 
noisily through escape valves and may collect in gas form in the scan room.  The gas is not harmful in 
itself as long as fresh air is available.  In this very remote event, the patient will immediately be brought 
out of the magnet room. 
Risk involving retinal optical coherence tomography (OCT):  The retinal OCT is a non-invasive procedure 
and there are no known risks or side effects from this procedure. Patients may experience some 
discomfort due to holding their head still for several minutes in the specific chin-piece while the OCT 
acquires the images. No pupil dilatation with eye drops is required.  
Risks involving those of a blood draw. The most common complication of this procedure is a small 
bruise. Rarely infection can occur. Standing up can result in a drop in blood pressure.  
 
Risk of orthostatic hypotension. Because selected participants with MSA may have symptomatic 
orthostatic hypotension, orthostatic syncope is a possibility, but we can almost invariably prevent such 
an event by having the participant sit back down or lie down. This risk should be no greater than that 
encountered during a standard clinical visit. Every care will be taken to prevent this occurrence. 
 
 
2.3.2 KNOWN POTENTIAL BENEFITS  
 
The potential benefits of this study are: The drug could potentially slow, halt or reverse the progression 
of the neurological abnormalities in MSA, although this is unproven. These include difficulties with 
speech, walking and autonomic function. There is also the possibility that the drug may be ineffective. 
 


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2.3.3 ASSESSMENT OF POTENTIAL RISKS AND BENEFITS  
 
Given the rapidly progressive and fatal nature of multiple system atrophy, there is an urgent need for 
therapies that can halt or slow the progression of the disease. Thus, performing well-designed clinical 
trials with medications that have the potential of modifying the natural history of multiple system 
atrophy is a research priority. Because sirolimus is an FDA-approved medication with a well-known side 
effects profile, the potential benefits for patients outweigh the potential risks of the study drug.  
 
 


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3 
OBJECTIVES AND ENDPOINTS 
   
OBJECTIVES 
ENDPOINTS 
JUSTIFICATION FOR 
ENDPOINTS 
Primary 
 
 
To determine if oral sirolimus is 
of sufficient promise or futile to 
slow disease progression in 
patients with MSA  
 
Primary endpoint: Change from 
baseline to 48 weeks in United 
Multiple System Atrophy Rating 
Score (UMSARS) total score in 
patients taking sirolimus 
compared with described 
progression rates in historical 
controls. We will assume that 
patients receiving sirolimus will 
have a change of 4 points less 
than those reported in natural 
history studies (see statistical 
section). 
 
Secondary endpoints: Change 
from baseline to 48 weeks in 
United Multiple System Atrophy 
Rating Score (UMSARS) 
subscores (UMSARS-1 and 
UMSARS-2) in patients taking 
sirolimus compared with 
described progression rates in 
historical controls. We will 
assume that patients receiving 
sirolimus will have a change of 
2.5 points less than those 
reported in natural history 
studies (see statistical section). 
 
 
UMSARS is a patient-reported, 
semi-quantitative, validated, 
disease-specific scale 
representing the diverse signs 
and symptoms in MSA (36).  
The UMSARS in the only 
validated investigator-
administered rating scale to 
reliably measure semi-
quantitatively symptoms and 
signs in MSA. The instrument 
has high inter-rater reliability, 
and internal consistency, for 
instance the UMSARS-1 has a 
Crohnbach’s alpha of 0.84 and 
the UMSARS-2 of 0.90 (36). The 
instrument is sensitive in 
measuring disease progression 
(37). It has an Activities of Daily 
Living score (UMSARS-1, 12 
questions) that evaluates motor 
including autonomic activities 
and the Motor Examination 
score (UMSARS-2, 14 
questions). UMSARS-3 
measures supine/standing BP 
and UMSARS-4 is a disability 
scale (1-5). Higher scores on the 
UMSARS scales mean poorer 
health.  
Secondary 
 
 
To assess the potential effects 
of sirolimus on brain 
neuroimaging and retinal 
biomarkers in patients with 
MSA 
Brain magnetic resonance 
changes using dedicated 
algorithms to evaluate rate of 
change in diffusivity of defined 
areas of brain from baseline to 
48 weeks in patients receiving 
sirolimus compared to the 
progression of putaminal 
diffusivity reported in 
longitudinal studies of MSA. 
Increased putaminal diffusivity 
has been consistently reported, 
even in early disease stages of 
MSA (42, 55-59). Increase in 
putaminal diffusivity has been 
reported to be a sensitive 
neuroimaging measures of 
disease progression in patients 
with MSA and has been used 
previously as neuroimaging 


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OBJECTIVES 
ENDPOINTS 
JUSTIFICATION FOR 
ENDPOINTS 
 
 
 
 
Changes in retinal nerve fiber 
layer (RNFL) using optical 
coherence tomography from 
baseline to 48 weeks in patients 
receiving sirolimus compared to 
the rate of progression in 
longitudinal studies of MSA.  
outcome measure in previous 
clinical trials of MSA (41, 42).   
 
 
The use of optical coherence 
tomography (OCT) is emerging 
as a non-invasive fast technique 
to measure the thickness of the 
retinal layers. Even though 
patients with MSA only rarely 
have visual complaints, recent 
studies by our group using OCT 
showed that the retina of 
patients with MSA have 
consistent atrophy of the 
peripapillary retinal nerve fiber 
layer (RNFL) and to a lesser 
extent the macular ganglion cell 
layer (GCL) complex (13). These 
abnormalities are progressive 
overtime and have been 
recently confirmed by 
pathological examination in the 
retinas from three patients with 
MSA (12, 14). 
Tertiary/Exploratory  
 
 
To explore the effects of 
sirolimus on blood biomarkers 
(exosomal alpha-synuclein 
particles) in patients with MSA 
Change from baseline to week-
48 in exosomal alpha-synuclein 
particle concentration in plasma 
in patients taking sirolimus 
versus the placebo group 
 
The concentration of exosomal 
alpha-synuclein particles in 
plasma has been proposed as a 
biomarker of diagnosis of 
synucleinopathies. We 
hypothesize that sirolimus will 
increase autophagy and, 
therefore, the degradation of 
these exosomal alpha-synuclein 
particles. 
 
 
 
 
 
 
 
 
 


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4 
STUDY DESIGN  
 
4.1 
OVERALL DESIGN 
 
To determine whether to test sirolimus in a large phase 3 trial, we propose to conduct a clinical trial of 
sirolimus in MSA using a futility design. A futility design under the null hypothesis assumes that 
sirolimus will slow the progression of the disease, whereas the alternative hypothesis assumes no 
benefit of sirolimus. If the null hypothesis is rejected (i.e., futility of sirolimus to slow progression of 
MSA), major phase 3 studies will be discouraged, whereas non-futility will offer strong support for a 
phase III trial to detect clinical efficacy.  
 
To test this hypothesis we will perform a single-center, randomized, double blind, placebo-controlled, 
phase-II, futility trial, chosen as an efficient approach to determine if sirolimus would be worthy of 
future investigation in larger, more definite trials. This trial was designed to compare the mean value for 
the primary outcome measure, change in total Unified Multiple System Atrophy Rating Scale (UMSARS) 
score from baseline to week 48, between the sirolimus arm and a pre-specified fixed value, representing 
a 4-point reducing in the expected mean decline without treatment; this fixed value was obtained using 
data from previously completed a large natural history study(39) and the placebo arm of a recently 
completed clinical trial in MSA (40). The 4-point reduction was based on the recently reported minimally 
clinically important decline (MICD) in the total UMSARS score of 3.5-points (38).  
 
The null hypothesis is that sirolimus will reduce the mean decline in the total UMSARS score over 12 
month by, at least 4-points, compared with the expected (fixed) value obtained from the historical 
controls. If the null hypothesis is rejected, then the drug would be considered futile for further 
investigation in MSA. If the null hypothesis is not rejected, sirolimus would be considered for a phase 3 
clinical trial.  
 
Thus, the primary goal is to evaluate the effect of sirolimus on progression of MSA, including autonomic, 
cerebellar and parkinsonian features as measured by the UMSARS, a relevant clinical measure of disease 
progression. Secondary goals are to determine if there is any effect on neuroimaging (brain MRI) and 
ophthalmological (retinal OCT) biomarkers of the disease. As exploratory aim we will determine if there 
are any changes in plasma αSyn exosomes, as surrogate markers of autophagy. 
 
This study will include a placebo group in 3:1 proportion. Our reason for including a placebo arm is to 
facilitate blinding to treatment assignment and also to permit a descriptive assessment of the validity of 
the assumed change in UMSARS over time in the historical control group; the placebo group will not be 
used for primary outcome purposes with the active treatment arm. 
 
This will be single-center study performed at the NYU Dysautonomia Center, one of the leading centers 
in the diagnosis and management of MSA. Dr. Horacio Kaufmann, the director of the Center, is the PI of 
the international Natural History Study of MSA, within the NIH-funded Rare Diseases Autonomic 
Disorders Consortium. NYU has the infrastructure and personnel in place to undertake this study and is 
experienced in the conduct of clinical trials. NYU is in a position to rapidly commence the study and 
conduct the study efficiently. We will review the option of adding additional sites if recruitment in the 
first 15 months is slow. Our aim is to complete recruitment within 24 months from commencement of 
the study.  


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Eligible participants, who signed the approved informed consent form at the screening visit, will initially 
receive sirolimus 2 mg/day (one 2-mg capsule/day) or matching placebo in a 3:1 fashion. The dose of 
sirolimus will be adjusted throughout the trial based on sirolimus blood levels and the presence of drug-
related adverse events. The maximum dose of sirolimus will be 6 mg/day (three 2-mg capsules/day) or 
matching placebo. Patients receiving placebo will undergo analog sham level measurements and the 
number of capsules will be also adjusted to maintain the blinding of the trial. Patients will receive active 
agent or placebo for 48-weeks. 
 
Participants will be evaluated at study clinic by the investigator at the screening visit and at baseline visit 
and will return to the clinic at Day 84 (month 3), Day 168 (month 6), Day 252 (month 9), and Day 363 
(month 12) (+/- 10 days) of beginning the study treatment.  
 
Detailed study plan 
 
Screening visit (Visit -1, Day -42 to -1): Prior to performing any study activities/evaluations, the potential 
participant must be thoroughly informed about all aspects of the study, including scheduled study visits 
and activities, and must sign the informed consent. A signed copy of the approved informed consent will 
be given to the participant.  Screening procedures will include demography, medical history, inclusion 
and exclusion criteria assessment, MSA diagnosis confirmation, physical examination, vital signs & 
height/weight, EKG, concomitant medication, adverse events, laboratory and safety testing (serum 
pregnancy test, if applicable, CBC, CMP, CPK, urinalysis and 24-hour urine collection (to quantify the 
protein/creatinine ratio), lipid panel, HIV, hepatitis B and C serologies, and QuantiFERON®), and 
UMSARS scale.    
COVID-19 nasal swab PCR will be performed at screening. If negative, enroll patient. If positive, patient 
should be quarantined for at least 2 weeks and consult with their PCP to determine if any treatment is 
necessary. Participants can be re-screened after 4 weeks with a second nasal swab PCR for COVID-19. If 
negative, the patient can be enrolled. If still positive, patient will be definitely excluded and 
recommended continued follow-up with an infectious disease specialist. 
If the subject has laboratory tests and EKG results available performed within 90 days before the 
screening date, it will be allowable to use these for the screening visit.  
 
Safety labs should be performed at least 3 hours after a light (low fat) breakfast. 
 
There will be a screening period of up to 42 days.  
 
Baseline visit (Visit 1, Day 1): The following tasks/procedures will be performed at the clinic during the 
baseline visit: 
o EKG 
o MoCA 
o Concomitant medications 
o Physical examination 
o Vitals signs 
o CBC, CMP, CPK (8 ml of blood) urinalysis  
o Lipid panel 
o Dispensing of study medication 


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o UMSARS scale 
o Blood sample for exosomes (2 ml of blood) 
o Retinal optical coherence tomography 
o 24-hour urine collection. This test should be performed only if the baseline visit occurs > 30 days 
from the screening visit. 
 
If the baseline visit is performed less than 14 days from the screening visit the following procedures can 
be foregone, and the results will be assumed to be similar to those obtained at the screening visit: 
 
o EKG 
o MoCA 
o CBC, CMP, CPK (8 ml of blood), urinalysis  
o Lipid panel 
o UMSARS scale 
 
 
Participants will be dispensed study medication during the baseline visit. In case this is not possible, 
participants will be mailed the study medication within 30 days of the baseline visit. 
 
The case report forms will note the date when the subject will start the study medication.  
 
Participants will also undergo a non-contrast brain MRI with specific sequences to evaluate the diffusion 
at the putamen and pons. The brain MRI will be performed at the NYU Center for Biomedical Imaging. 
There will be a brain MRI period of up to 7 days after the baseline visit (i.e., subjects must be scanned 
within 7 days after the baseline visit). 
 
Scheduled site visits: assessment of the primary outcome measure and safety labs will be conducted on-
site at study visits at Visit 8 (month 3 after initiation of study drug), Visit 11 (month 6 after initiation of 
study drug), Visit 14 (month 9 after initiation of study drug), and Visit 17 (month 12 after initiation of 
study drug) (+ 15 days). We will accept a visit window of 15 days after the target date. The following 
tasks will be performed: 
o EKG 
o Concomitant medications 
o Adverse event recording 
o Physical exam 
o Vital signs 
o UMSARS scale 
o CBC, CMP, CPK (8 ml of blood) and urinalysis with 24-hour urine collection  
o Lipid panel 
o Sirolimus levels in blood (5 ml of blood) 
o Retrieve study drug 
o Dispense study drug. If dispensation of study drug is not possible during the visit, the study drug 
will be mailed within 15 days.  
 
Safety Laboratory Visits: Assessment of safety will be conducted on-site or locally at study visits 7 days 
(with a window of 3 days after the target date) after initiation of study drug (Study Visit 3), and 7 days 
after the patient has increased dose of study drug (i.e., Study Visit 4, Study Visit 5 and Study Visit 6, with 


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a window of 3 days after the target date) during the first month of study drug treatment. We will accept 
a visit window of 3 days after the target date of these visits.  
 
Assessment of safety visits will also occur on-site or locally on Month 2 (Study Visit 7, this visit may be 
skipped in previous visits were delayed, to ensure that Visit 8 occurs at Month 3), Month 4 (Study Visit 
9), Month 5 (Study Visit 10), Month 7 (Study Visit 12), Month 8 (Study Visit 13), Month 10 (Study Visit 
15), and Month 11 (Study Visit 16). We will accept a visit window of 15 days before or after the target 
date of these visits.  
 
The following safety laboratory assessments will be performed: 
o CBC, CMP, CPK (8 ml of blood) and urinalysis with 24-hour urine collection  
o Lipid panel (2 ml of blood)     
o Sirolimus levels in blood (5 ml of blood) 
o Adverse event reporting 
 
In case the safety lab assessments are performed locally (e.g., at a local laboratory near the subjects’ 
home), the PI or a licensed research member of the team will provide the patient with a prescription for 
the required tests. A research nurse, investigator or study manager will perform a phone call to assess 
adverse events within 7 days of the study visit.  
 
To account for delays during the fist month of treatment, Visit 7 (2-month) may be skipped to ensure 
that Visit 8 occurs 3 months (+/- 15 days) after the initiation of the study drug.  
 
To maintain the blinding, the results of the sirolimus levels in blood will remain blinded to the 
investigators and study coordinators and delivered to the independent pharmacist in charge of 
determining if the sirolimus dosage should be increased or decreased.  
 
End of study visit/early termination: he following tasks/procedures will be performed at the clinic during 
the termination visit (Study Visit 17): 
o EKG 
o MoCA 
o Concomitant medications 
o Physical examination 
o Vitals signs 
o CBC, CMP, CPK (8 ml of blood) and urinalysis with 24-hour urine collection 
o Lipid panel (2 ml of blood) 
o Sirolimus levels in blood (5 ml of blood) 
o Retrieve study medication 
o Recording of AE 
o UMSARS scale 
o Blood sample for exosomes (2 ml of blood) 
o Retinal optical coherence tomography 
o Brain MRI 
 
We will accept a visit window of 15 days before or after the target date of this visit.  
 
The reason for discontinuation of therapy will be documented in the source documents and captured on 
the CRF. In the event that the study drug is discontinued early, an early termination visit should be 


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completed as close to the last dose date as possible. Participants will be followed for the duration of 12 
months.  
 
If a participant is withdrawn because of an adverse event, the appropriate “Withdrawal Section” of the 
CRFs should be fully completed in addition to the Adverse Event module. The Independent Medical 
Monitor should be informed of all participants who are withdrawn for this reason. Participants will be 
followed for 12 months.  
 
Follow-up phone call: Patients will be contacted 14 days (+/- 5 days) after the End of Study visit to 
follow up on and record any Aes that were ongoing at the end of their previous visit (Study Visit 18). The 
follow-up visit may be conducted over the phone; however a response from the patient should be 
obtained. This response may be obtained by a phone call with the patient and appropriately 
documented in the source notes. In the event that the patient is unreachable by phone, all reasonable 
efforts should be made and documented before considering the patient lost to follow-up. 
 
Safety assessment: Assessment of safety will be conducted on-site 30 days (+/- 5 days) after the End of 
Study visit (Study Visit 19). We accept a visit window of 3 days before or after the target date. The 
following safety laboratory assessments will be performed: 
o CBC, CMP, CPK (8 ml of blood) and urinalysis with 24-hour urine collection  
o Lipid panel (2 ml of blood)     
o Adverse event reporting 
Alternatively, these safety lab assessments can be performed locally (e.g., at a local laboratory near the 
subjects’ home). In this case, the PI or a licensed research member of the team will provide the patient 
with a prescription for the required tests. Also, a research nurse or study manager will perform a phone 
call for adverse event reporting.  
 
Unscheduled visit: An un-scheduled visit may be performed in-office or locally, should the investigators 
determine the need for so.  The following safety laboratory assessments may be performed: 
o CBC, CMP, CPK (8 ml of blood) and urinalysis with 24-hour urine collection  
o Lipid panel (2 ml of blood)     
o Sirolimus levels in blood (5 ml of blood) 
o Adverse event reporting 
 
 
4.2 
SCIENTIFIC RATIONALE FOR STUDY DESIGN 
 
To determine whether to test sirolimus in a large phase 3 trial, we propose to conduct a clinical trial of 
sirolimus in MSA using a futility design. This approaches minimizes the number of subjects and resources 
required for the study and provides an indication of whether or not to pursue larger and more definite 
studies.  A futility design (60) is highly appropriate for MSA given: a) the low prevalence of the disease; 
b) the availability of historical controls from recent large studies (8, 39); and c) the availability of 
sensitive outcome measures. A futility design under the null hypothesis assumes that sirolimus will slow 
the progression of the disease, whereas the alternative hypothesis assumes no benefit of sirolimus. If 
the null hypothesis is rejected (i.e., futility of sirolimus to slow progression of MSA), a major phase III 
study will be discouraged, whereas non-futility will offer strong support for a phase III trial to detect 
clinical efficacy. 
 


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This study will also include a placebo group in 3:1 proportion. Our reason for including a placebo arm is 
to facilitate blinding to treatment assignment and also to permit a descriptive assessment of the validity 
of the assumed change in UMSARS over time in the historical control group; it will not be used as a basis 
of comparison for the active treatment arm. 
 
This placebo-controlled futility design comparing outcome measures to historical controls has been used 
in other neurodegenerative disorders (61-63). 
The study population (n=42 active agent, n=14 placebo) was selected based on power analysis 
calculation using the known annual progression of the UMSARS total score from published natural 
history studies (8, 39) and the progression in the placebo group of previous clinical trials of MSA(40), as 
well as from the minimally clinically significant difference in UMSARS (38) (see statistical section).  
 
4.3 
JUSTIFICATION FOR DOSE 
We will use sirolimus dosages of 2 mg, 4 mg or 6 mg, once a day based on therapeutic drug monitoring 
in order to maintain sirolimus drug levels within a FDA-recommended target-range: 5-20 ng/ml. This 
daily dosage is within the range that is used clinically, and we expect this dose to be well tolerated. This 
is well below the currently FDA-approved maximum sirolimus dose administered per day, which is 40 mg 
(54). The use of sirolimus for 1 year is known to be safe, based on long experience in the prevention of 
organ transplant rejection, without significant complications (64, 65). Moreover, there is ample evidence 
that sirolimus at these dosages crosses the blood-brain barrier (31-35). 
 
4.4 
END OF STUDY DEFINITION 
 
A participant is considered to have completed the study if he or she has completed all phases of the 
study including the last visit or the last scheduled procedure shown in the Schedule of Activities (SoA), 
Section 1.3. 
 
The end of the study occurs when the following circumstance have ALL occurred: 
1. Completion of the last visit or procedure shown in the SoA in the trial globally. 
2. Completion of data analysis 
3. Completion of final study report 
 
5 
STUDY POPULATION 
 
The study will include 56 participants, randomized in a 3:1 ratio to receive oral sirolimus or matching 
placebo. All participants will fulfill the diagnostic criteria for probable MSA outlined by the MSA 
Consensus Criteria (1). Participants with MSA-P and MSA-C diagnosed less than 4 years ago will be 
accepted into the study. This is because natural history studies have found that later stage MSA patients 
have only modest rates of progression while patients with milder symptoms have a more rapid rate of 
progression over 12 months. We hypothesize that these milder cases also have a greater probability to 
respond to effective treatment. We aim to study 56 participants 30-80 years of age. Sex will be 
approximately evenly represented. It is expected that approximately 10% of participants will be from 
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5.1 
INCLUSION CRITERIA 
 
1. Participants aged 30-80 years old with a diagnosis of MSA. Patients have to fulfill current 
consensus criteria (1) for probable MSA of the parkinsonian subtype (MSA-P) or cerebellar 
subtype (MSA-C). 
2. Participants who are less than 4 years from the time of documented MSA diagnosis. 
3. Participants who are still able to walk with or without assistance. 
4. Participants with an anticipated survival of at least 3 years in the opinion of the investigator. 
5. Participants who are willing and able to give informed consent. 
6. Montreal Cognitive Asessment (MoCA) > 20. 
7. Ability to take oral medication and be willing to adhere to the study drug regimen 
8. For females of reproductive potential: agreement to use highly effective contraception method 
during study participation and for an additional 8 weeks after the end of study drug 
administration  
9. For males of reproductive potential: use of condoms or other methods to ensure effective 
contraception with partner 
10. Agreement to adhere to Lifestyle Considerations (see section 5.3) throughout study duration 
 
5.2 
EXCLUSION CRITERIA 
 
1. Women of childbearing potential who do not practice an acceptable method of birth control. 
Acceptable methods of birth control in this study are: surgical sterilization, intrauterine devices, 
partner’s vasectomy, a double-protection method (condom or diaphragm with spermicide), 
hormonal contraceptive drug (i.e., oral contraceptive, contraceptive patch, long-acting 
injectable contraceptive) with a required second mode of contraception. 
2. Participants with a clinically significant or unstable medical or surgical condition that, in the 
opinion of the investigator, might preclude safe completion of the study or might affect the 
results of the study. These include severe uncontrolled congestive heart failure, recent (<6 
months) myocardial infarct, severe uncontrolled cardiopulmonary disease, severe, uncontrolled 
hypertension (sustained systolic blood pressure above 200 mmHg in any position), 
thrombocytopenia (< 50 x 10(9)/L), severe anemia (< 8g/dl), immunocompromised state, liver or 
kidney disease (creatinine > 1.5 mg/dl or proteinuria > 20 mg/dl), uncontrolled diabetes mellitus 
(HbA1c >10g%), alcoholism, amyloidosis, uncontrolled hypothyroidism, sympathectomy, 
unstable peripheral neuropathies, concurrent infections, severe orthopedic problems that 
compromise mobility and activity of daily living, recent (< 6 months) cerebrovascular accidents, 
neurotoxin or neuroactive drug exposure, parkinsonism due to drugs (including neuroleptics, 
alpha-methyldopa, reserpine, metoclopramide). 
3. Participants with high LDL cholesterol levels (LDL > 190 mg/dL) and/or high triglycerides levels (> 
500 mg/dL) under fasting conditions. Patients receiving medications for elevated cholesterol or 
triglycerides are eligible to participate as long as their LDL cholesterol and/or triglyceride levels 
are below the abovementioned limits.   
4. Participants with positive interferon-gamma release-assay (QUANTIferon®) indicating latent 
tuberculosis infection. Participants with a positive Quantiferon® may be enrolled after the 
patient has been evaluated by an infectious diseases specialist that has cleared the patient for 
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5. Participants with history of untreated tuberculosis. Participants with a history of treated 
tuberculosis may be enrolled after an infectious diseases specialist evaluates patient and has 
cleared the patient for this trial. 
6. Participants with a history of active, acute or chronic, or latent hepatitis B or hepatitis C. 
7. Participants with human immunodeficiency virus (HIV) infection, or other congenital or acquired 
causes of immunosuppression. 
8. Participants with a nasal swab PCR positive for COVID-19. If positive, participant may 
be re-screened after 1 month. If the second nasal swab PCR is negative for COVID-19, 
participant may be enrolled.  
9. Participants with active malignant neoplasms or history of malignant neoplasm in the last 5 
years. 
10. Movement disorders other than MSA; e.g., Parkinson disease, dementia with Lewy bodies, 
essential tremor, progressive supranuclear palsy, spinocerebellar ataxia, spastic paraparesis, 
corticobasal degeneration, or vascular, pharmacological or post-encephalitic parkinsonism. 
11. Dementia (DSM-V criteria). 
12. History of electroconvulsive therapy. 
13. History of deep brain stimulation surgery. 
14. Patients with contraindication for MRI scanning, including those with MRI-incompatible 
pacemaker 
15. History of organ transplant 
16. Participants who have taken any investigational products within 60 days prior to baseline. 
17. Treatment with cyclosporine, oral or intravenous corticosteroids, methotrexate, rituximab 
within 3 months prior to baseline.  
18. Treatment with inhibitors of CYP3A4 (which may decrease the metabolism of sirolimus and 
increase sirolimus levels): nicardipine, verapamil, clotrimazole, fluconazole, itraconazole, 
clarithromycin, erythromycin, troleandomycin, cisapride, metoclopramide, bromocriptine, 
cimetidine, danazol, HIV-protease inhibitors (e.g., ritonavir, indinavir); grapefruit. 
19. Treatment with inducers of CYP3A4 (which may increase the metabolism of sirolimus and 
decrease sirolimus levels): carbamazepine, phenobarbital, phenytoin, rifabutin, rifapentine. 
20. Inability or unwillingness of subject to give written informed consent. 
 
5.3 
LIFESTYLE CONSIDERATIONS 
 
During this study, participants are asked to: 
• 
Refrain from consumption of red wine, Seville oranges, grapefruit or grapefruit juice from 30 
days before the start of study drug until after the final dose. 
• 
Participants who use tobacco products will be instructed that use of nicotine-containing 
products (including nicotine patches) will not be permitted while they are in the clinical unit.  
• 
Minimize interactions with household contacts that may be immunocompromised. 
 
5.4 
SCREEN FAILURES 
 
Screen failures are defined as participants who consent to participate in the clinical trial but are not 
subsequently randomly assigned to the study intervention or entered in the study. A minimal set of 
screen failure information is required to ensure transparent reporting of screen failure participants, to 
meet the Consolidated Standards of Reporting Trials (CONSORT) publishing requirements and to 


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respond to queries from regulatory authorities. Minimal information includes demography, screen 
failure details, eligibility criteria, and any serious adverse event (SAE).  
 
A screen failure log will be maintained in the study binders.  
 
Individuals who do not meet the criteria for participation in this trial (screen failure) because of an 
unallowable medication may be rescreened after discontinuation of such medication. Individuals who 
do not meet the criteria for participation in this trial (screen failure) because of an acute infection may 
be rescreened after resolution of such infection. Rescreened participants should be assigned the same 
participant number as for the initial screening. 
 
Individuals with a positive Quantiferon® will be recommended to see an infectious diseases specialist. In 
the event that the infectious diseases specialist considers that the positive Quantiferon® was a false 
positive, or that there is negligible safety risks for a patient with positive Quantiferon® to be enrolled in 
the study, the patient will be re-screened.  
 
5.5 
STRATEGIES FOR RECRUITMENT AND RETENTION 
 
Patients with MSA will be recruited from those that we are currently following at the NYU Dysautonomia 
Center. Our Center currently follows approximately 100 patients with MSA. In addition, we are in close 
contact with MSA patients advocacy groups (e.g. Multiple System Atrophy Coalition) and will collaborate 
closely with them in order to disseminate information about the trial and increase the likelihood of 
recruitment, by means of e-mail blast or letter. The recruitment e-mail/letter to be sent to patients with 
MSA is attached as an appendix. Any recruitment e-mail or letter to be used in this trial will be 
submitted to the IRB for approval prior to use. 
 
We expect to screen an average of 8 patients per month and accruing an average of 3 patients per 
month. At this predicted rate, we expect to complete enrollment of the subjects (n=56) within 24 
months with patients recruited only at the NYU Dysautonomia Center. A special effort will be made to 
include women and minorities. 
 
A screening and enrollment log to record the consent and screening of all subjects and the outcome of 
each screening will be maintained. The log will provide a comprehensive list of all subjects who were 
screened for eligibility as well as consented, including the screened failures, the reasons for ineligibility, 
and reasons for nonparticipation of eligible subjects. This information will be collected centrally, will be 
included in the Essential Documents Binder, and will be used to enhance subject recruitment efforts.  
 
The principal investigators or co-investigators will approach patients with MSA during a routine clinic 
visit to inquire whether they would be interested in being enrolled in this clinical trial. If so, patients will 
be given the consent form. The subject and family will be given enough time to read the informed 
consent in a quiet room to maintain privacy. The principal investigators, co-investigator or clinical study 
manager will explain the design of the study, discuss it with the patient and answer any questions or 
concerns. If the patient is satisfied by the responses of the investigators, the principal investigator will 
ask the patient to sign the consent form before entering the study. The original signed form will be kept 
within the study files, and the patient and family will be given a copy. 
 
Screening procedures will include demography, medical history, inclusion and exclusion criteria 


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assessment, disease diagnosis confirmation, physical examination, vital signs & height/weight, EKG, 
concomitant medication, adverse events, laboratory testing (serum pregnancy test, if applicable, CBC, 
metabolic panel, CPK, urinalysis and 24-hour urine collection, QuantiFERON®, hepatitis B and C and HIV 
testing), and UMSARS scale. There will be a screening period of up to 42 days.  
 
To encourage retention, participants will be compensated for the time/travel/meals with $75 per visit. 
This will be done with a check payable to the subject, mailed to the subject’s mail address within 30 days 
after the study visit.  
 
5.5.1 USE OF DATACORE/EPIC INFORMATION FOR RECRUITMENT PURPOSES 
This study will utilize EPIC to identify participants.  
Any recruitment information sent by email will utilize Send Safe email.  
Once potential participants have been identified, the study team will notify the treating physician (TP) 
that they have patients eligible to participate as follow:  
• 
TP agrees to permit study team to directly contact potential participants on behalf of TP.  
• 
TP has been notified that the study team will contact potential participants directly, phone, email, 
and/or the MyChart portal. 
Once contact is made, approved recruitment language will be used to communicate the reason they are 
being contacted and participants will be asked if they are interested in participating in this specific 
study. Should the potential participants agree, the study team will provide the participants with 
information regarding the next steps for participation.  
If a participant requests information regarding opting out of further recruitment for all research, 
participants will be directed to contact research-contact-optout@nyumc.org or 1-855-777-7858. 
 
6 
STUDY INTERVENTION 
 
6.1 
STUDY INTERVENTION ADMINISTRATION 
 
6.1.1 STUDY INTERVENTION DESCRIPTION 
 
Sirolimus (a.k.a. rapamycin, a.k.a. Rapamune) is an immunosuppressive agent.  Sirolimus is a macrocyclic 
lactone produced by Streptomyces hygroscopicus.  The chemical name of sirolimus (also known as 
rapamycin) is (3S,6R,7E,9R,10R,12R,14S,15E,17E,19E,21S,23S,26R,27R,34aS)-
9,10,12,13,14,21,22,23,24,25,26,27,32,33,34,34a-hexadecahydro-9,27-dihydroxy-3-[(1R)-2-[(1S,3R,4R)-
4-hydroxy-3-methoxycyclohexyl]-1-methylethyl]-10,21-dimethoxy-6,8,12,14,20,26-hexamethyl-23,27-
epoxy-3H-pyrido[2,1-c][1,4] oxaazacyclohentriacontine-1,5,11,28,29 (4H,6H,31H)-pentone.  
 


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Its molecular formula is C51H79NO13 and its molecular weight is 914.2. Sirolimus is a white to off-white 
powder and is insoluble in water, but freely soluble in benzyl alcohol, chloroform, acetone, and 
acetonitrile. 
 
Sirolimus is available for administration as an oral solution containing 1 mg/mL sirolimus. Sirolimus is 
also available as a tan, triangular-shaped tablet containing 0.5 mg sirolimus, as a white, triangular-
shaped tablet containing 1 mg sirolimus, and as a yellow-to-beige triangular-shaped tablet containing 2 
mg sirolimus. The inactive ingredients in sirolimus tablets include sucrose, lactose, polyethylene glycol 
8000, calcium sulfate, microcrystalline cellulose, pharmaceutical glaze, talc, titanium dioxide, 
magnesium stearate, povidone, poloxamer 188, polyethylene glycol 20,000, glyceryl monooleate, 
carnauba wax, dl-alpha tocopherol, and other ingredients. The 0.5 mg and 2 mg dosage strengths also 
contain yellow iron (ferric) oxide and brown iron (ferric) oxide. 
 
In this trial, sirolimus capsules (2 mg) and matching placebo will be manufactured in compliance with 
current GMP standards and guidelines applicable to study drugs. Sirolimus capsules (2 mg) and their 
matching placebo will be of identical appearance to maintain study blinding. 
 
This trial will use sirolimus (Rapamune®) 2 mg tablets manufactured by Pfizer (NDC #: 0008-1032-05). 
Matching placebo will be compounded by Joseph Navarra, R.Ph., FACA, Town Total Compounding 
Center. 
 
The matching placebo will be composed of lactose and encapsulated in an orange gelatin capsule. 
6.1.2 DOSING AND ADMINISTRATION 
 
During the baseline visit, participants will be allocated to a treatment group, based on a randomization 
procedure employing a 3:1 assignment ratio, and a scheme using blocks stratified by MSA type 
(cerebellar or parkinsonian) and sex. The randomization sequence will be performed by the independent 
medical monitor (IMM) and delivered, in a closed envelope, to Town Total Compounding Pharmacy  
(415 Crossways Park Drive, Suite B Woodbury NY 11797). The Town Total Compounding Pharmacy 
pharmacist will use the randomization order to send the proper medication to the study coordinator. 
The patient will be then instructed to take one capsule (i.e., 2 mg of sirolimus or matching placebo) daily 
and have sirolimus levels in blood 7 days (with a window of 3 days after the target date) after start of 
study drug.  
 
Participants will then undergo a dose-selecting procedure based on therapeutic level measurements. 
The results of these measurements will be revealed only to an independent study pharmacist 
(independent from the Town Total Compounding pharmacy) who will make dosing recommendations to 
maintain sirolimus trough levels between 5 and 20 ng per milliliter, as well as corresponding sham dose 
adjustments in the placebo group. 
 
Until the dose selection period has been completed, subjects will undergo sirolimus measurements in 
blood frequently during the first month of study drug treatment. Sirolimus levels will be measured again 
7 days (with a window of 3 days after the target date) after a subject has been instructed to increase the 
dose of study drug. Once the initial dose selection has been achieved, subjects will then undergo 
measurements of sirolimus in blood levels monthly since the start of the study drug (with a window of 
15 days after the target date). 


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The initial dose selection period is as follows: from day 1 to day 7, all subjects will receive sirolimus 2 mg 
one capsule daily or matching placebo. Seven days later (+ 3 days), subjects will undergo measurement 
of sirolimus levels. Once the results of the levels are back, the independent pharmacist will assess and 
determine if an increase in study drug is required. 
 
o If sirolimus levels are < 5 ng/mL, the participant will be instructed to take sirolimus 2 mg two 
capsules daily (4 mg total dose) or matching placebo; sirolimus levels will be then measured 
again 7 days after the patient began taking 4 mg/day  (+ 3 days). If sirolimus levels are between 
5 and 20 ng/ml, the subject will continue with that dose until the next measurement of sirolimus 
levels on Visit  (+ 3 days).  
o If sirolimus levels are > 30 ng/ml with one capsule/day, the subject will be excluded from the 
trial due to the high risk of adverse events. 
 
Subjects who were instructed to increase their dose to 4 mg will undergo another measurement of 
sirolimus levels 7 days after the previous dose adjustment (+ 3 days): 
 
o If sirolimus levels are < 5 ng/mL, the participant will be instructed to increase sirolimus to 2 mg 
three capsules daily (6 mg total dose) or matching placebo. Sirolimus levels will be then 
measured 7 days after the patient began taking 6 mg/day (+ 3 days).  
o If sirolimus levels are between 5 and 20 ng/ml, the subject will continue with that dose until the 
next measurement of sirolimus. If the sirolimus levels are > 20 ng/ml, the subject will be 
instructed to go back to the 2 mg/day dose and continue with that dose until the next 
measurement of sirolimus levels.  
 
From Visit 3 to Visit 6, sirolimus level blood draw will take place 7 days (+3 days) after the previous dose 
adjustment has taken place (i.e., 7 days after the patient started taking the adjusted dose). 
 
From Visit 7 onwards, subjects will be instructed to increase or decrease their sirolimus dose (between a 
min dose of 2 mg/day to a max dose of 6 mg/day) depending on blood levels (target: 5-20 ng/ml) and 
the emergence of adverse events.  
 
Expected dose-limiting effects requiring sirolimus dose reduction are:  
 
• 
Grade 2 stomatitis or mucositis 
• 
Grade 2 lymphedema 
• 
Grade 2 proteinuria 
• 
Grade 2 renal failure 
• 
Grade 3 pneumonitis 
• 
Grade 2 diarrhea 
 
Every attempt will be made to maintain the participant on the full dose. If the participant is unable to 
maintain the full dose, the minimum acceptable dose is 2 mg once a day. An attempt will be made to 
increase the dose with a re-challenge at a later date.  Capsules should be swallowed whole with a glass 
of water or may be taken with food to minimize gastrointestinal side effects.  
 
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measurements in blood will be revealed only to an independent study pharmacist who will make dosing 
recommendations, and a sham therapeutic drug level will be simulated for those patients receiving 
placebo (66) (see Measures to Minimize Bias section). 
 
 
6.2 
PREPARATION/HANDLING/STORAGE/ACCOUNTABILITY 
 
6.2.1 ACQUISITION AND ACCOUNTABILITY 
 
Sirolimus capsules (2 mg) and matching placebo will be acquired in randomized per patient bottles from 
the Town Total Compounding Pharmacy and delivered to the study coordinator/ PI for dispensing to 
subjects 
 
Town TOTAL Compounding Center 
415 Crossways Park Dr., Suite B 
Woodbury, N.Y. 11797 
516-249-7436 
 
The Town Total Compounding Center pharmacy will randomize, store and dispense study drug to our 
center. 
 
The study coordinator will maintain drug accountability with a drug accountability log. 
 
6.2.2 FORMULATION, APPEARANCE, PACKAGING, AND LABELING 
 
Sirolimus capsules (2 mg) and matching placebo capsules will be packaged in tight, light-resistant bottles 
as defined in the US Pharmacopeia. Sufficient bottles for 3 months of study will be packed for each 
participant, taking into consideration the potential increases in sirolimus dose after measuring sirolimus 
levels in blood. Each bottle label will bear the site number, investigator’s name, participant number and 
the bottle number which comprises the participant number followed by a sequential number (01, 02, 
03, 04, etc.).  
 
The label on the bottles will include a fixed information section (to include the product name, storage 
conditions, instructions etc.) and variable information section. The variable information section will 
include 3 parts: 2 detachable parts, which will be attached to the participant’s file upon bottle 
dispensing and 1 part to remain on the bottle. The variable information section will include the following 
variable data: site number, investigator name, participant number, bottle number, batch code and 
expiration date (all to be pre-printed on the label) and blank field for visit number to be manually filled 
in by the Investigator or designee upon provision of bottle to the participant.  
 
Study drug supplies will be kept at the Total Town Compounding pharmacy, in a secure, limited-access, 
temperature/humidity-controlled area and a temperature/humidity log will be maintained. Only 
authorized personnel will have access to the study drug. The Total Town Compounding pharmacy 
personnel will be responsible for correct storage and handling of the study drug.  The study drug bottles 
will be dispensed to the participant at the study site at each study visit starting at baseline visit (except 
for termination visit). The site number and investigator’s name will be pre-printed on the bottle label. 
Since the participant number is pre-printed on the bottle label, assigning the right bottles to the 


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participants is straightforward. After assigning a bottle to the participant, the site personnel should fill in 
the visit number on the label of the dispensed bottle as well as on the detachable parts that would be 
attached to the participant’s file.  
 
At each dispensing visit, the participant will be dispensed with the study drug with the participant’s 
number printed on the label. Any of the bottles designated for a participant can be dispensed in each of 
the dispensing visits. The bottle with the bottle number “participant # -R” is designated to replace 
damaged or lost bottles and should be used only for replacement.  
 
In case a replacement bottle is required, the Total Town compounding pharmacist will be notified.  
 
Participants will be instructed to return all used empty bottles and unused study drug at each visit.  
 
The study coordinator is responsible for performing study drug accountability at the site and the 
accountability of the returned study drug.  
 
6.2.3 PRODUCT STORAGE AND STABILITY 
 
Capsules will be stored at 20 to 25C (US Pharmacopeia Controlled Room Temperature) (68 - 77F). If 
the study drug supplies appear to be damaged/missing upon arrival at the NYU Dysautonomia Center, 
the study manager should be contacted immediately. Each shipment of study drug supplies for the study 
will contain a shipment form describing the content of shipment. This form will assist in maintaining 
current and accurate inventory records. When a shipment is received, the NYU Dysautonomia Center 
will acknowledge receipt of the study drug supply by signing the acknowledgment of receipt section. 
 
 
6.3 
MEASURES TO MINIMIZE BIAS: RANDOMIZATION AND BLINDING 
 
After a participant meets the eligibility criteria, During the baseline visit, participants will be allocated to 
a treatment group, based on a randomization procedure employing a 3:1 assignment ratio, and a 
scheme using blocks stratified by MSA type (cerebellar or parkinsonian) and sex. The randomization 
sequence will be performed by the independent medical monitor (IMM) and delivered, in a closed 
envelope, to the Total Town compounding pharmacy. The compounding pharmacist will use the 
randomization order to send the proper medication to the study coordinator. The patient will be then 
instructed to take one capsule (i.e., 2 mg of sirolimus or matching placebo) daily and have sirolimus 
levels in blood at specific times throughout the study. Depending on the sirolimus levels, the sirolimus 
dose may be increased or decreased throughout the duration of the study.  
 
A participant ID will be assigned by the center and kept in a written log. The participant IDs will be used 
to code the data obtained during the study visits, including brain MRI, OCT, and blood for exosomes. The 
participant IDs will follow the following format: three letters followed by three numbers (i.e., NYU001). 
The three letters correspond to center (i.e., New York University).  
 
Drug Kit numbers will be assigned by the pharmacy and provided to the statistician who will then assign 
to the randomization block scheme. There will be an electronic validation of the participant’s eligibility. 
If they are found to be eligible the system will send the drug kit number based on the known 
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by the pharmacist to send the proper medication. The Drug Kit number will not replace the Participant 
ID.  
 
Study drug will be stored and dispensed by the study PI or research nurses delegated by the PI, located 
at: 
 
NYULMC 
530 First Ave. 
Dysautonomia Center, suite 9Q 
New York, NY 10016 
 
 
Blinding  
The participant, as well as investigators and any personnel involved in the participant’s recruitment, 
assessment, monitoring, analysis and data management, will be blinded to the participant’s treatment 
assignment.  
 
The only personnel unblinded to the intervention will be the compounding pharmacist, the independent 
medical monitor (IMM), the independent pharmacist (independent from the compounding pharmacy 
and any other study members) for the purposes of sirolimus therapeutic levels monitoring. The 
independent pharmacist will maintain the blinding of the sirolimus levels and dose escalation during the 
clinical trial by make dosing recommendations for those patients receiving active agent, and by 
generating a sham therapeutic drug level for those patients receiving placebo. The independent study 
pharmacist will then make dosing recommendations to maintain sirolimus trough levels between 5 and 
20 ng per milliliter in those patients taking active agent, as well as corresponding sham dose 
adjustments in patients taking placebo, in order to maintain investigator and subject blinding during the 
clinical trial. 
 
Sirolimus therapeutic levels monitoring 
An independent pharmacist will remain unblinded to each patient allocation and to the sirolimus blood 
level of each patient to allow the adjustment of such levels. The independent pharmacist will generate a 
simulated/sham value for patients receiving placebo in order to maintain the blinding of the 
investigators and the patients (66).  
 
Emergency Unblinding  
Unblinding of the participant’s drug assignment will occur under any of the following circumstances: 
1. Pregnancy. 
2. Serious adverse event when the study drug assignment is required to make treatment 
decisions for the individual participant. 
3. Death of participant when the death has been considered to be probably or possibly related 
to the study drug. 
4. Death of participant when the relationship to study drug has been undetermined (e.g., after 
multiple efforts, the investigator team was unable to obtain medical records and/or death 
certificate). 
5. Other circumstances that, under the opinion of the PI or the overseeing agencies, require 
emergency unblinding.  
 


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Unblinding of the participant’s drug assignment will be done by the compounding pharmacist, the 
independent medical monitor, or the independent pharmacist, after the PI request. The IRB should be 
notified of the event prior to breaking of the code, if possible. If this is not possible, the IRB should be 
notified immediately afterwards, and the participant’s drug assignment should be revealed. The 
circumstances leading to the breaking of the code should be fully documented, in the investigator’s 
study files and in the participant’s source documentation. The independent medical monitor and the 
compounding pharmacist will maintain the randomization lists and will be available to break the blind in 
the event of an emergency. A randomization list will be sent to the compounding pharmacist at the 
beginning of the study. A backup copy of the complete set of randomization assignments will also be 
given to the IMM and the PIs in a sealed envelope. We do not anticipate that it will ever be necessary 
that the PI will have to open this envelope during the trial. We will carefully document the reasons for 
any emergency unblinding should it ever become necessary to do this.  
 
 
6.4 
STUDY INTERVENTION COMPLIANCE 
 
At each study visit, the investigator/site coordinator will assess the participant’s compliance with the 
prescribed regimen for the study medication. This will include checks of protocol compliance and use of 
study drug in order to assess the reliability of participant-generated data. Participants who fail to comply 
with the study requirements may be withdrawn from the study, following consultation with the PI.  
 
Compliance with the dosing regimen will be determined by performing study drug accountability of 
returned capsules and bottles of the study drug used and unused. The number of used, unused and lost 
capsules will be recorded on the CRF by the study coordinator.  
 
Participants with less than 75% overall compliance during the entire study will be considered non-
compliant. These participants will be included in the intention to treat analysis set and complete analysis 
set as defined in sections 12.3.1 and 12.3.2 respectively. However, these participants will be excluded as 
in the per-protocol analysis set.  
 
Accountability records must be maintained at the site at all times. Accountability of the returned study 
drug should be performed and recorded at the site by the study coordinator. The participant number, 
the date, batch code, bottle number and quantity of study drug returned by the participant will be 
checked for correctness and recorded on the appropriate accountability forms.  
 
6.5 
CONCOMITANT THERAPY 
 
Prohibited medications 
• 
Intravenous immunoglobulin within 3 months prior to baseline visit  
• 
Rituximab within 3 months prior to baseline visit 
• 
Oral or intravenous corticosteroids within 3 months prior to baseline visit (topical or inhaled 
corticosteroids are allowable) 
• 
Cyclosporine within 3 months prior to baseline visit 
• 
Methotrexate within 3 months prior to baseline visit 
• 
Azathioprine, mycophenolate or any other immunosuppressants or immunomodulatory drugs 
within 3 months prior to baseline visit 
• 
Any anti-neoplastic or immunomodulatory agent including but not limited to nilotinib, 


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natalizumab, infliximab and bevacizumab within 3 months prior to baseline visit  
• 
Any other investigational drug within 3 months prior to baseline visit 
 
Allowed medications 
• 
All anti-parkinsonian medications including but not limited to L-dopa, rasagiline, selegiline, 
amantadine. 
• 
Fludrocortisone, pyridostigmine and vasopressin and analogs. 
• 
Beta-blockers 
• 
ACE inhibitors 
• 
Alpha adrenergic agonists including but not limited to midodrine 
• 
Norepinephrine precursors including but not limited to droxidopa 
• 
Norepinephrine reuptake inhibitors including but not limited to atomoxetine or venlafaxine 
• 
Serotonin reuptake inhibitors and other antidepressants 
• 
Benzodiazepines 
• 
Baclofen 
• 
CoQ10 
• 
Metformin and other anti-diabetic medications 
• 
Topical or inhaled corticosteroids 
• 
Non-steroidal anti-inflammatory drugs 
• 
Topical anesthetic (e.g., lidocaine patch, lidocaine mouthwash) 
• 
Statins 
• 
Bile sequestrants 
• 
Fibrates 
• 
Trimethoprim/sulfamethoxazole 
• 
Clindamycin 
• 
Primaquine 
 
Efforts will be made to maintain medications and dosages stable until the final study visit.  
 
Standard non-pharmacologic measures to treat orthostatic hypotension are encouraged. These include 
the use of elastic stockings/abdominal binder, elevation of the head of the bed, increased fluid intake, 
and liberal salt intake. The participant is encouraged to maintain stable conditions of these measures.  
 
Disallowed concomitant medications during the study: 
• 
Treatment with inhibitors of CYP3A4 (which may decrease the metabolism of sirolimus and 
increase sirolimus levels) within 1 month prior to baseline visit: nicardipine, verapamil, 
clotrimazole, fluconazole, itraconazole, clarithromycin, erythromycin, troleandomycin, cisapride, 
metoclopramide, bromocriptine, cimetidine, danazol, HIV-protease inhibitors (e.g., ritonavir, 
indinavir); grapefruit. 
• 
Treatment with inducers of CYP3A4 (which may increase the metabolism of sirolimus and 
decrease sirolimus levels) within 1 month prior to baseline visit: carbamazepine, phenobarbital, 
phenytoin, rifabutin, rifapentine. 
 
6.5.1 RESCUE MEDICINE 
 
Not applicable 
 


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7 
STUDY INTERVENTION DISCONTINUATION AND PARTICIPANT 
DISCONTINUATION/WITHDRAWAL 
 
7.1 
DISCONTINUATION OF STUDY INTERVENTION 
 
Discontinuation from study drug does not mean discontinuation from the study, and remaining study 
procedures should be completed as indicated by the study protocol. If a clinically significant finding is 
identified (including, but not limited to changes from baseline) after enrollment, the investigator or 
qualified designee will determine if any change in participant management is needed. Any new clinically 
relevant finding will be reported as an adverse event (AE). 
 
The data to be collected at the time of study intervention discontinuation will include the following: 
o EKG 
o Concomitant medications 
o Physical examination 
o Vitals signs 
o CBC, CMP, CPK, urinalysis 
o 24-hour urine collection 
o Lipid panel 
o Sirolimus levels in blood 
o Retrieve study medication 
o Recording of AE 
o UMSARS scale 
 
7.2 
PARTICIPANT DISCONTINUATION/WITHDRAWAL FROM THE STUDY 
 
Participants are free to withdraw from participation in the study at any time upon request. An 
investigator may discontinue or withdraw a participant from the study for the following reasons: 
 
o Pregnancy 
o Significant study intervention non-compliance  
o If any clinical adverse event (AE), laboratory abnormality, or other medical condition or situation 
occurs such that continued participation in the study would not be in the best interest of the 
participant 
o Disease progression which, in the opinion of the investigator, requires discontinuation of the 
study intervention 
o If the participant meets an exclusion criterion (either newly developed or not previously 
recognized) that precludes further study participation 
o Participant unable to receive study drug for 15 days or more. 
 
The reason for participant discontinuation or withdrawal from the study will be recorded on the 
electronic Case Report Form (CRF). Subjects who sign the informed consent form and are randomized 
but do not receive the study intervention may be replaced.  Subjects who sign the informed consent 
form, and are randomized and receive the study intervention, and subsequently withdraw, or are 
withdrawn or discontinued from the study, will not be replaced. 
 


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7.3 
LOST TO FOLLOW-UP 
 
A participant will be considered lost to follow-up if he or she fails to return for 2 scheduled visits and is 
unable to be contacted by the study site staff.  
 
The following actions must be taken if a participant fails to return to the clinic for a required study visit: 
o The site will attempt to contact the participant and reschedule the missed visit within 10 days 
and counsel the participant on the importance of maintaining the assigned visit schedule and 
ascertain if the participant wishes to and/or should continue in the study. 
o Before a participant is deemed lost to follow-up, the investigator or designee will make every 
effort to regain contact with the participant (where possible, 3 telephone calls, 3 e-mail 
messages and, if necessary, a certified letter to the participant’s last known mailing address or 
local equivalent methods). These contact attempts should be documented in the participant’s 
medical record or study file.  
o Should the participant continue to be unreachable, he or she will be considered to have 
withdrawn from the study with a primary reason of lost to follow-up. 
 
 
8 
STUDY ASSESSMENTS AND PROCEDURES 
 
8.1 
EFFICACY ASSESSMENTS  
 
Primary efficacy outcome 
 
The Unified Multiple System Atrophy Rating Scale (UMSARS) total score. The primary outcome measure 
will be the total UMSARS score progression at week 48 compared to baseline. UMSARS is a validated, 
disease-specific scale representing the diverse signs and symptoms in MSA (36). It can assess rates 
of progression and is sensitive to change over time (37). It has an Activities of Daily Living score 
(UMSARS-1, 12 questions) that evaluates motor including autonomic activities and the Motor 
Examination score (UMSARS-2, 14 questions). UMSARS-3 measures supine/standing BP and 
UMSARS-4 is a disability scale (1-5). Higher scores on the UMSARS scales mean poorer health. 
The UMSARS will be determined by the same investigator at each study visit to ensure consistency. This 
investigator has undergone specific UMSARS training and has extensive experience in performing this 
scale. The investigator rating the UMSARS will be blinded to the previous UMSARS score of the patient 
as well as to the treatment group assignment of the subject.   
 
Secondary efficacy outcomes 
 
The Unified Multiple System Atrophy Rating Scale (UMSARS) part 1 and part 2. Secondary outcome 
measures include the UMSARS-2 score (neurological examination) progression at week 48 compared to 
baseline; the UMSARS-1 score (activity of daily living) progression at week 48 compared to baseline. 
 
Neuroimaging: We will compare several diffusion parameters of the individual and combined putamen 
in subjects using brain magnetic resonance imaging (MRI) obtained at baseline and after 48 weeks. 
Diffusion-based MRI techniques measure how the random movement of water is altered in normal and 
disease nervous tissue. Diffusion MRI data can be analyzed with a mathematical tensor that provides 


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model parameters for the mean diffusion of water (the apparent diffusion coefficient, or “ADC”), the 
degree of diffusion orientation dependence (fractional anisotropy, or “FA”) and the dominant 
orientation of water diffusion in an individual image voxel (first Eigen vector). In most 
neurodegenerative conditions, the tissue environment sampled by water becomes less organized over 
time and this is reflected by increasing ADC and decreasing FA. Compared to PD, increased putaminal 
ADC in MSA-P has been consistently reported, even in early disease stages (42, 55-59). Putaminal ADC 
changes have been reported to be a highly sensitive MRI marker of disease progression in patients with 
MSA and has been used previously as neuroimaging outcome measure in previous clinical trials of MSA 
(41, 42).   
 
MRI will be carried out with a state-of-the-art 1.5-T MRI system and 24-channel head coil (Aera, Siemens 
Healthcare, Erlangen, Germany) – note, while 3-T has some signal-to-noise advantages, many MSA 
patients have cardiac pacer wires that make 3-T imaging difficult or impossible to obtain routinely. First 
a volumetric 3D T1-weighted MR-RAGE (TR/TE/TI 10/2/600 ms) and volumetric 3D SPACE FLAIR  MRI 
sequences will both be obtained with 1-mm isotropic resolution (10 minutes total). Routine 3D 
susceptibility-weighted imaging will be obtained (5 min). Finally axial high-angular resolution diffusion 
MRI will be obtained of the whole brain using 3-mm isotropic resolution using echoplanar imaging, 
parallel imaging and 2-slice acceleration with simultaneous multi-slice acquisitions (TR/TE = 7000/100 
ms, b = 0 and 1500 s/mm2, 30 gradient directions).  These data will be analyzed with inline scanner 
diffusion tensor model and parameter maps of ADC, FA and fiber orientation generated. These will be 
co-registered with the volumetric data and manual regions-of-interest drawn to include the entire left 
and right putamen to extract mean ADC and FA. A secondary neuroimaging aim in this study will be to 
determine whole brain, cortical, white matter and individual structure volumes at baseline and after 48 
weeks using automated FreeSurfer analysis as described in the literature (67). 
 
Dr. Timothy Shepherd, Assistant Professor of Radiology at the NYU Center for Advanced Imaging 
Innovation and Research and Director of the NYU Brain Mapping Service (which performs fMRI and 
diffusion tractography for neurosurgical planning) will supervise creation of the MRI protocol, data 
acquisition, diffusion analysis and interpretation of the neuroimaging data. Dr. Shepherd is an academic 
neuroradiologist with MD/Ph.D. training in diffusion MRI and 6 years of experience performing 
advanced diffusion acquisitions in patients with MSA, Parkinson disease and other neurodegenerative 
diseases, brain tumors and epilepsy. 
 
Retinal optical coherence tomography: Retinal nerve fiber layer thickness progression at week 48 
compared to baseline. The use of optical coherence tomography (OCT) is emerging as a non-invasive fast 
technique to measure the thickness of the retinal layers. Even though patients with MSA only rarely 
have visual complaints, recent studies by our group using OCT showed that the retina of patients with 
MSA have consistent atrophy of the peripapillary retinal nerve fiber layer (RNFL) and to a lesser extent 
the macular ganglion cell layer (GCL) complex (13). These abnormalities are progressive overtime and 
have been recently confirmed by pathological examination in the retinas from three patients with MSA 
(12, 14). In brief, high definition OCT will be performed in a darkened room by the same trained 
personnel using standardized Food and Drug Administration (FDA)-approved equipment to evaluate the 
structure of the eye (Cirrus 4000, Carl Zeiss, CA, USA). Images will be acquired in the seated position 
with the subjects facing the OCT equipment. Subjects were instructed to fixate their gaze at a green 
target during the scan. Each subject had both eyes scanned at least three times using two standard 
acquisition protocols: macular cube (512x128 line scans) and optic disc cube (200x200 line scans). In 
both protocols the scanned area was a 6-mm cube with no signal averaging. The quality of the obtained 
images was assessed by evaluation of the signal strength (a value from 0-10 in arbitrary units) 


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automatically provided by the system. Only scans with signal strength above 6 units were included in the 
analysis. In some patients with MSA, involuntary eye movements will make the test difficult to obtain. In 
such cases, repeated scans will be performed to obtain at least three scans without eye movement 
artifacts. The scan with the best resolution will be used for analysis. For OCT quantification, two 
automatic segmentation algorithms will be used to determine the RNFL and ganglion cell complex (GCC) 
thicknesses (Cirrus 4000, Carl Zeiss, CA, USA). For the RNFL analysis, an optic disc cube of data (200x200-
line scans) centered in the optic nerve head will be acquired. Subsequently, a recognition algorithm 
detected the inner (vitreoretinal interface) and outer (inner plexiform layer) border of the RNFL, from a 
1.73-mm diameter circle will extract from the optic nerve cube and centered in the optic nerve head. 
The distance between the two lines will be measured as RNFL thickness at specific quadrants around the 
optic nerve: temporal, superior, nasal, inferior and global. For the GCC analysis the macular cube 
512x128 centered in the foveal pit will be used. A different recognition algorithm will be applied to 
detect the outer border of the RNFL and the inner plexiform layer. The GCC analysis will evaluate the 
thickness of the ganglion cell plus inner plexiform layers. The average, minimum and sectorial 
thicknesses of the GCC will be measured in an elliptical annulus (vertical inner and outer radius of 0.5 
mm and 2.0 mm respectively; horizontal inner and outer radius of 0.6 and 2.4 mm, respectively) around 
the fovea. The GCC thickness will be measured in different locations in the macula, around the foveal 
center: temporal-superior, superior, nasal-superior, nasal inferior, inferior, temporal-inferior, and global. 
The global RNFL and GCC thickness values, as well as those at each specific quadrant, expressed in 
micrometers (μm), will be used for analysis, although only the total RNFL will be used as secondary 
outcome measure.  
 
Exploratory outcomes 
 
Measurement of αSyn in CNS-derived extracellular vesicles (Evs): As exploratory outcome measure we 
will measure the concentration of αSyn in neuronal and oligodendroglial Evs isolated from patients’ 
serum at baseline, on week 24, and on week 48. Recently, exosomes have been implicated in the 
dissemination of misfolded proteins in a variety of neurodegenerative disorders, including Parkinson 
disease.(68) Exosomes are Evs of 30–200 nm diameters that are released from various cells including 
neurons.(69) Exosomal release and transport of αSyn followed by intracellular uptake and induction of 
toxicity and cell death has been reported by several groups in vivo and in vitro.(70, 71) Several groups 
also have measured exosomal αSyn particles in cerebrospinal fluid (CSF) of patients with 
synucleinopathies.(72) However, because the lumbar puncture required for obtaining CSF is rather 
invasive and patients often refuse it, we opt to use a new methodology in which brain-derived Evs will 
be isolated from serum. The methodology has been introduced originally in 2014 in studies of 
Alzheimer’s disease and frontotemporal dementia(73-75) and in two recent studies was applied also to 
patients with Parkinson disease.(76, 77) This methodology has been improved by Professor Gal Bitan’s 
team at UCLA, who has been able to distinguish among healthy controls, patients with Parkinson 
disease, and patients with MSA with high sensitivity and specificity (see preliminary data section). Our 
group has been collaborating with Dr. Bitan and contributed part of the samples included in the 
preliminary data. Using the same protocol, we will obtain samples from the patients included in the 
study proposed here and ship them to the Bitan laboratory for analysis. In brief, 10 ml of blood will be 
extracted from each patient and centrifuged immediately to extract the serum. Because αSyn is one of 
the major biomarkers in our analysis, and in addition to neurons it exists in high levels in erythrocytes, it 
is crucial to separate the serum within a 20-minute window from drawing the blood, before hemolysis 
begins. The serum samples will be flash frozen immediately and stored at -80C. They will then be 
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will be performed by operators blinded to the study intervention. Dr. Bitan will receive only de-
identified samples. He will have no way to link the samples to the subject’s identities or PHI. 
 
 
8.2 
SAFETY AND OTHER ASSESSMENTS 
 
The primary safety outcome is number of participants (%) who discontinue the study due to AE.  
 
Secondary safety outcomes include active monitoring for severe adverse events (SAE’s) throughout the 
trial. Specific SAE’s are anticipated to be related to sirolimus. These selected SAE’s include mucositis, 
anemia, thrombocytopenia, hyperglycemia, hyperlipidemia, proteinuria, renal failure, pneumonitis, and 
infection. The occurrences of these safety outcomes by treatment arm will be reported in bi-annual 
safety reports to the IMM. We will also report counts and proportions of mortality for each treatment 
arm (by rhythm), along with the number of SAEs that are probably or definitely related to mortality. 
 
Adverse events will be recorded from the time a participant has signed the Informed Consent Form and 
throughout the 12 months of the study, even if they prematurely discontinue the study drug. They 
should be reviewed and updated at each subsequent visit and during any phone contact with the 
participant. Participants will be instructed to call a provided number if they experience AE between 
scheduled visits. An unscheduled visit may be needed so that the participant can be assessed.  
 
Safety laboratory evaluations. Safety laboratory evaluations will be performed at screening, baseline, 
during each study visit and at termination visit. In addition, laboratory evaluations at months 2, 4, 5, 7, 
8, 10 and 11 will be performed either at the NYU Dysautonomia Center or locally if the patient is unable 
to come to NYU. HIV testing, hepatitis B and C, QuantiFeron and serum pregnancy testing (women) will 
be performed only at screening. In addition, the following tests will be performed during all monthly 
safety assessments (Visit 7 and onwards).  
o Complete blood count with differential – approximately 1 tablespoon of blood will be 
required. 
o Comprehensive metabolic panel and CPK– approximately 1 tablespoon of blood will be 
required. 
o Lipid panel – approximately 1 tablespoon of blood will be required. 
o Urinalysis – approximately a cup of urine will be required. 
o 24-hour urine collection – approximately 4 cups of urine, collected throughout a day, will 
be required 
 
Physical examination including the cardiovascular, dermatological, respiratory and gastrointestinal and 
neurological organ systems and vital signs, height and weight will be performed at each scheduled and 
unscheduled visits.  
 
EKG: EKG will be performed locally at Screening and Termination visits. Twelve-lead ECG should be 
performed following the participant being in a supine position for 5 minutes. The investigator will 
evaluate the ECG at time of performance (signed and dated) and the printout should be kept in the 
source documentation file. If the investigator detects potentially clinically significant findings, a 
cardiologist should be consulted for a definitive interpretation. All communications and diagnoses 
should be filed in the source documentation file.  
 


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8.3 
ADVERSE EVENTS AND SERIOUS ADVERSE EVENTS 
 
8.3.1 DEFINITION OF ADVERSE EVENTS (AE) 
 
Adverse event means any untoward medical occurrence associated with the use of an intervention in 
humans, whether or not considered intervention-related (21 CFR 312.32 (a)). 
 
8.3.2 DEFINITION OF SERIOUS ADVERSE EVENTS (SAE)  
 
An adverse event (AE) or suspected adverse reaction is considered “serious” if, in the view of either the 
investigator or sponsor, it results in any of the following outcomes: death, a life-threatening adverse 
event, inpatient hospitalization or prolongation of existing hospitalization, a persistent or significant 
incapacity or substantial disruption of the ability to conduct normal life functions, or a congenital 
anomaly/birth defect. Important medical events that may not result in death, be life-threatening, or 
require hospitalization may be considered serious when, based upon appropriate medical judgment, 
they may jeopardize the participant and may require medical or surgical intervention to prevent one of 
the outcomes listed in this definition. Examples of such medical events include allergic bronchospasm 
requiring intensive treatment in an emergency room or at home, blood dyscrasias or convulsions that do 
not result in inpatient hospitalization, or the development of drug dependency or drug abuse. 
 
8.3.3 CLASSIFICATION OF AN ADVERSE EVENT 
8.3.3.1 
SEVERITY OF EVENT 
 
All reported adverse events will be classified using the Common Terminology Criteria for Adverse Events 
(CTCAE) developed and maintained by CTEP at National Cancer Institute. The intensity or severity of the 
AE will be graded as follows: 
 
• 
Grade 1: Mild AE 
• 
Grade 2: Moderate AE 
• 
Grade 3: Severe AE 
• 
Grade 4: Life-threatening or disabling AE 
• 
Grade 5: Death related to AE 
 
8.3.3.2 
RELATIONSHIP TO STUDY INTERVENTION 
 
All adverse events (AE) must have their relationship to study intervention assessed by the clinician who 
examines and evaluates the participant based on temporal relationship and his/her clinical judgment. 
The degree of certainty about causality will be graded using the categories below. In a clinical trial, the 
study product must always be suspect.  
 
• 
Related – The AE is known to occur with the study intervention, there is a reasonable possibility 
that the study intervention caused the AE, or there is a temporal relationship between the study 
intervention and event. Reasonable possibility means that there is evidence to suggest a causal 
relationship between the study intervention and the AE. 


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• 
Not Related – There is not a reasonable possibility that the administration of the study 
intervention caused the event, there is no temporal relationship between the study intervention 
and event onset, or an alternate etiology has been established. 
 
8.3.3.3 EXPECTEDNESS  
 
The Pis will be responsible for determining whether an adverse event (AE) is expected or unexpected.  
An AE will be considered unexpected if the nature, severity, or frequency of the event is not consistent 
with the risk information previously described for the study intervention. 
 
Common adverse events of sirolimus that have been described in previous clinical trials (> 10% of 
patients) include: 
o Blood and lymphatic disorders (anemia, leucopenia, thrombocytopenia) 
o Dermatological disorders (acneiform rash, oral mucositis) 
o Dyslipidemia (high cholesterol, high lipids) 
o Hyperglycemia/glucose intolerance/new onset diabetes mellitus 
o Hypertension 
o Lymphocele 
o Pneumonitis 
o Renal disorders (proteinuria/increased creatinine/increased blood urea) 
o Wound complications (healing problems) 
o Infections (urinary tract infection, pneumonia) 
 
In addition, patients with MSA have increased morbidity associated to the motor and autonomic 
dysfunction inherent to the disease, regardless of any study drug or intervention. Expected events that 
are expected to occur as part of the natural history of MSA are: 
 
o Urinary tract infection 
o Dizziness, lightheadedness or feeling about to faint 
o Syncope due to orthostatic hypotension 
o Shortness of breath upon standing 
o Head or neck pain/discomfort upon standing 
o Blurry vision or other vision changes upon standing 
o Falls 
o Aspiration pneumonia 
  
8.3.4 TIME PERIOD AND FREQUENCY FOR EVENT ASSESSMENT AND FOLLOW-UP 
 
The occurrence of an adverse event (AE) or serious adverse event (SAE) may come to the attention of 
study personnel during study visits and interviews of a study participant presenting for medical care, or 
upon review by a study monitor. 
 
All Aes including local and systemic reactions not meeting the criteria for SAEs will be captured on the 
appropriate case report form (CRF). Information to be collected includes event description, time of 
onset, clinician’s assessment of severity, relationship to study product (assessed only by those with the 
training and authority to make a diagnosis), and time of resolution/stabilization of the event. All Aes 


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occurring while on study must be documented appropriately regardless of relationship. All Aes will be 
followed to adequate resolution. 
 
Any medical condition that is present at the time that the participant is screened will be considered as 
baseline and not reported as an AE. However, if the study participant’s condition deteriorates at any 
time during the study, it will be recorded as an AE.  
 
Changes in the severity of an AE will be documented to allow an assessment of the duration of the event 
at each level of severity to be performed. AEs characterized as intermittent require documentation of 
onset and duration of each episode. 
 
The PI will record all reportable events with start dates occurring any time after informed consent is 
obtained until 7 (for non-serious AEs) or 30 days (for SAEs) after the last day of study participation.  At 
each study visit, the investigator will inquire about the occurrence of AE/SAEs since the last visit.  Events 
will be followed for outcome information until resolution or stabilization. 
 
 
8.3.5 NON-SERIOUS ADVERSE EVENT REPORTING  
 
Non-serious expected adverse events: non-serious expected adverse events that are reported to or 
observed by the investigator or a member of his research will be presented in tabular form and given to 
the IMM on a bi-annual basis or as requested.  
 
Non-serious Aes that are expected or unexpected but do not increase the risk of the study drug will be 
reported in annual reports to the IRB. 
 
8.3.6 SERIOUS ADVERSE EVENT REPORTING  
 
The research study team member will immediately report to the PI any serious adverse event, whether 
or not considered study intervention related, including those listed in the protocol or investigator 
brochure and must include an assessment of whether there is a reasonable possibility that the study 
intervention caused the event.  
 
Serious adverse events (e.g., death, hospitalizations) with evidence suggesting a probable or possible 
causal relationship between the study intervention and the event (e.g., death from anaphylaxis) will be 
reported by the PI or study coordinator to the IRB and the IIM within 7 working days after the PI’s initial 
receipt of the information. If required, emergency unblinding will follow the criteria specified in section 
6.3.  
 
Serious adverse events (e.g., death, hospitalizations) with no evidence of a causal relationship between 
study intervention and event need to be communicated to the IMM within 14 working days after the 
PI’s initial receipt of the information. 
 
All serious adverse events (SAEs) will be followed until satisfactory resolution or until the site 
investigator deems the event to be chronic or the participant is stable. Other supporting documentation 
of the event may be requested by any regulatory agency and should be provided as soon as possible. 
 


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The PI will be responsible for notifying the IRB, IMM and Food and Drug Administration (FDA) of any 
unexpected fatal or life-threatening suspected adverse reaction as soon as possible, but in no case later 
than 7 calendar days after the PI’s initial receipt of the information. Emergency unblinding will follow 
the criteria specified in section 6.3.  
 
 
8.4 
UNANTICIPATED PROBLEMS 
8.4.1 DEFINITION OF UNANTICIPATED PROBLEMS (UP) 
 
The Office for Human Research Protections (OHRP) considers unanticipated problems involving risks to 
participants or others to include, in general, any incident, experience, or outcome that meets all of the 
following criteria: 
 
• 
Unexpected in terms of nature, severity, or frequency given (a) the research procedures that are 
described in the protocol-related documents, such as the Institutional Review Board (IRB)-
approved research protocol and informed consent document; and (b) the characteristics of the 
participant population being studied; 
• 
Related or possibly related to participation in the research (“possibly related” means there is a 
reasonable possibility that the incident, experience, or outcome may have been caused by the 
procedures involved in the research); and 
• 
Suggests that the research places participants or others at a greater risk of harm (including 
physical, psychological, economic, or social harm) than was previously known or recognized. 
 
 
8.4.2  UNANTICIPATED PROBLEM REPORTING  
 
The investigator will report unanticipated problems (Ups) to the reviewing Institutional Review Board 
(IRB) and to the principal investigator (PI). The UP report will include the following information: 
 
• 
Protocol identifying information: protocol title and number, PI’s name, and the IRB project 
number; 
• 
A detailed description of the event, incident, experience, or outcome;  
• 
An explanation of the basis for determining that the event, incident, experience, or outcome 
represents an UP;  
• 
A description of any changes to the protocol or other corrective actions that have been taken or 
are proposed in response to the UP. 
 
To satisfy the requirement for prompt reporting, Ups will be reported using the following timeline:   
 
• 
Ups that are serious adverse events (SAEs) will be reported to the IRB and to the IMM within 7 
working days of the investigator becoming aware of the event.  
• 
Any other UP that significantly increases the risk of the study will be reported to the IRB and to 
the IMM within 7 working days of the investigator becoming aware of the problem.  
• 
All UP that do not increase the risk of the study will be reported to the IMM in the bi-annual 
safety report. 
 
 


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9 
STATISTICAL CONSIDERATIONS  
 
9.1 
STATISTICAL HYPOTHESES 
 
Primary Efficacy Endpoint:  We hypothesize that treatment with sirolimus will show promise to slow the 
progression of the disease of patients with MSA. Specifically, our null hypothesis is that patients 
receiving sirolimus will have a UMSARS total score progression of only 4.6 points at week 48 higher than 
baseline, compared to a progression of 8.6 points at week 48 from baseline reported in historical 
controls from natural history studies and placebo-controlled trials of MSA. Our alternate hypothesis is 
that sirolimus will be futile to slow the progression of the disease in patients with MSA. The statistical 
definitions are as follows:  
 
H0:  θE – θp ≤ 4 where θp = 8.6 points increase at week 48 from baseline described in historical controls 
H0: θE ≤ 4.6 points increase at week 48 from baseline 
H1: θE > 4.6 points increase at week 48 from baseline 
 
Secondary Efficacy Endpoints: 
 
Brain MRI: We hypothesize that treatment with sirolimus will slow the increase of putaminal diffusivity 
in patients with MSA. Specifically, our null hypothesis states that patients receiving sirolimus will have a 
putaminal Trace (D) total score progression of 0.030 x 10-3 mm2/sec or less at week 48 versus baseline, 
compared to a progression of 0.070 x 10-3 mm2/sec at week 48 from baseline reported in specific 
longitudinal studies of MSA (41). 
 
H0:  θE – θp ≤ 0.04 where θp = 0.07 (x 10-3 mm2/sec) increase in putaminal diffusivity at week 48 from 
baseline described in specific studies 
H0: θE ≤ 0.03 points increase at week 48 from baseline 
H1: θE > 0.03 points increase at week 48 from baseline 
 
Optical coherence tomography: We hypothesize that treatment with sirolimus will slow the thinning of 
the retinal nerve fiber layer (RNFL) in patients with MSA. Specifically, our null hypothesis states that 
patients receiving sirolimus will have a RNFL thickness thinning of 1.45 μm at week 48 versus baseline, 
compared to a decrease of 3.7 μm at week 48 from baseline reported in specific longitudinal studies of 
MSA (12).  
 
H0:  θE – θp ≤ 2.25  where θp =  3.7 μm decrease in RNFL at week 48 from baseline described in specific 
studies 
H0: θE ≤ 1.45 μm decrease at week 48 from baseline 
H1: θE > 1.45 μm decrease at week 48 from baseline 
 
Concentration of exosomal αSyn particles: We hypothesize that treatment with sirolimus will reduce 
the concentration of exosomal αSyn particles in patients with MSA. Specifically, our null hypothesis is 
that patients receiving sirolimus will have statistically lower concentration of exosomal αSyn particles at 
week 48 compared to baseline: 
  
H0:  θE < θp  
H1:  θE ≥ θp  


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9.2 
SAMPLE SIZE DETERMINATION 
 
This is study is a randomized, double blind placebo-controlled futility trial in which the primary outcome 
measure value (UMSARS total score) in a cohort of patients with MSA taking active agent will be compared 
to a known value from historical controls previously published in the literature. The primary outcome 
measure was used to calculate the sample size.  
 
The primary statistical analysis will involve a comparison of the mean absolute difference in the active 
sirolimus group with a pre-specified fixed value, constituting a 4-point reduction in the expected mean 
increase in the UMSARS total score over 48 weeks without treatment. The expected mean increase in 
total UMSARS outcomes were obtained from the U.S. Natural History Study of MSA (n=96) (39), that 
reported that the total UMSARS score at 12 months increased a mean of 7.5 (SD: 8.4) absolute points 
from baseline. Also data from a phase III double-blind placebo-controlled 12-month clinical trial with 
rifampicin in MSA performed in the U.S. (40) (which was found to be ineffective to slow the progression 
of the disease) showed that in patients taking placebo (n=50) the total UMSARS score at 12 months 
increased a mean of 10.8 (SD: 10.7) absolute points from baseline. A weighted analysis of these two 
populations showd he pooled total UMSARS score at 12-months in patients with MSA in large studies 
performed in the U.S. had increased a mean of 8.6 (SD: 9.2) points from baseline. 
 
Also, recent studies (38) have determined that the minimally clinically important difference (MCID) in the 
total UMSARS score is 3.5 points after 48-weeks from baseline.  
 
Taking into consideration this MCID, for this trial, we will assume a promising difference in the UMSARS 
total score of 4 points between those taking active agent and the historical controls. Specifically, our null 
hypothesis is that patients receiving sirolimus will have a UMSARS total score progression of only 4.6 
points at week 48 higher than baseline, compared to a progression of 8.6 points at week 48 from 
baseline reported in historical controls from natural history studies and placebo-controlled trials of MSA. 
Our alternate hypothesis is that sirolimus will be futile to slow the progression of the disease in patients 
with MSA. The statistical definitions are as follows:  
 
H0:  θE – θp ≤ 4 where θp = 8.6 points increase at week 48 from baseline described in historical controls 
H0: θE ≤ 4.6 points increase at week 48 from baseline 
H1: θE > 4.6 points increase at week 48 from baseline 
 
Therefore, assuming an alpha of 0.05, and a power of 0.8 (one sided, as patients with MSA have not been 
reported to improve spontaneously) to detect a progression of 4.6 points (i.e., 8.6 minus 4) in the UMSARS 
total score after 48-weeks from baseline we will need to enroll 38 patients in the active agent arm. 
Anticipating a 10% for dropouts rate, we will need to enroll 42 patients in the active agent arm. 
 
This trial will include a placebo group to facilitate blinding in a 3:1 proportion. However, statistical 
comparisons for the primary and secondary efficacy outcome measures will be performed between the 
active agent arm and the historical controls. We will therefore enroll 14 patients in the placebo arm.  
 
9.3 
POPULATIONS FOR ANALYSES 
 


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Analyses will be performed according to the intention-to-treat principle and will include all randomized 
subjects who have completed, at least, 3 months of study drug.  
 
9.4 
STATISTICAL ANALYSES 
 
9.4.1 GENERAL APPROACH 
 
The two treatment groups (sirolimus and placebo) will be compared for baseline characteristics. This 
analysis will include demographic data and MSA history. The continuous variables will be examined 
using the two-sample t-test or Wilcoxon rank sum test when appropriate. For testing the assumption of 
normality prior to applying parametric approaches (i.e., two sample t-test), visual inspection of the 
normality plots as well as the Kolmogorov Smirnov test of normality will be used. We will use 5% level of 
significance for the test of normality. If the underlying data were to be non-normal, we will use non-
parametric approaches for analyzing the data (i.e., Wilcoxon rank sum test). Categorical variables will be 
examined for differences between groups using the Chi-Square test or the Fisher’s exact test when 
appropriate.  
 
 
9.4.2 ANALYSIS OF THE PRIMARY EFFICACY ENDPOINT 
 
The primary outcome endpoint (UMSARS total score) will be analyzed by comparing the mean decline in 
total UMSARS score in the active sirolimus group (UMSARS total score at week 48 minus UMSARS total 
score at baseline) with the pre-specified fixed value of 8.6 (SD: 9.2) using a one-sample t-test. If the 
difference between the fixed value of 8.6 minus the UMSARS total score in the active sirolimus group is 
4 or less, the null hypothesis will be rejected, indicating futility.  
 
Missing data will be addressed by using a regression-based multiple imputation model (78) 
 
 
9.4.3 ANALYSIS OF THE SECONDARY ENDPOINT 
 
In order to maintain the overall type I error at the 0.05 level, an hierarchy will be employed as follows: If 
the primary endpoint is found to be significant at a significance level of 0.05, then the first key 
secondary endpoint will be tested; if this endpoint is found to be significant in a significance level of 
0.05, then the second key secondary endpoint will be tested and so on. The key secondary endpoints 
and their order in the hierarchy are: 
 
UMSARS sub-scores: The secondary outcome endpoint (UMSARS-1 and UMSARS-2 scores) will be 
analyzed by comparing the mean decline in total UMSARS-1 and UMSARS-2 score in the active sirolimus 
group (UMSARS-1 and UMSARS-2 at week 48 minus UMSARS-1 and UMSARS-2 at baseline) with the pre-
specified fixed value of 4.5 (SD: 5.5) in each score using a one-sample t-test. 
 
Brain magnetic resonance imaging: The secondary outcome endpoint (putaminal diffusivity) will be 
analyzed by comparing the mean increase in Trace(D) score in the active sirolimus group (Trace(D) score 
at week 48 minus Trace(D) score at baseline) with the pre-specified fixed value of 0.07 (x 10-3 mm2/sec) 
using a one-sample t-test. 
 
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thickness) will be analyzed by comparing the mean decline in RNFL thickness in the active sirolimus 
group (RNFL at baseline minus RNFL at week 48) with the pre-specified fixed value of 3.7 μm using a 
one-sample t-test. 
 
Concentration of exosomal αSyn particles: This exploratory outcome endpoint will be analyzed by 
comparing the mean change in αSyn exosomal particles concentration (week 48 minus baseline) in the 
active sirolimus group with the placebo group. 
 
 
In all cases missing data will be addressed by using a regression-based multiple imputation model (78) 
 
9.4.4 SAFETY AND TOLERABILITY ANALYSES 
 
Adverse events: The incidence and frequency of adverse events will be presented by System Organ Class 
and preferred terminology according to NCI and CTCAE reporting standard. Aes will also be presented by 
System Organ Class, High Level Term and preferred terminology. Data will be tabulated by treatment 
group, sex, age, maximal severity, maximal outcome, maximal action taken and maximal relationship to 
the tested drug. Serious adverse events and seriousness criteria will be listed and discussed on a case by 
case basis.  
 
Laboratory tests: The incidence of laboratory tests outside the normal range and the incidence of 
measurements of potential clinical significance will be presented by treatment group. Shift analysis of 
these counts from baseline will also be provided. Descriptive statistics as well as their changes from 
baseline will also be presented by study group.  
 
Vital Signs & Height/Weight: Incidence of measurements of potential clinical significance will be 
presented by study group. Shift analysis from baseline will be provided as well. Descriptive statistics of 
vital signs & height/weight, as well as, their changes from baseline, will be presented by study group.  
 
EKG: The incidence of abnormal measurements will be presented by treatment group. Shift analysis 
from baseline will be provided as well. Descriptive statistics, as well as, their changes from baseline, will 
also be presented by study group.  
 
Tolerability assessments: Tolerability analysis will be based on the number (%) of participants who 
failed to complete the study, the number (%) of participants who failed to complete the study due to 
adverse events. Time to withdrawal will be presented by Kaplan-Meier curves. Significance testing of 
time to withdrawal will be done using Cox’s proportional hazards model.  
 
 
9.4.5 PLANNED INTERIM ANALYSES  
 
Interim efficacy analysis will be performed when 24 patients have completed the trial.  
 
If the efficacy pre-specified criteria are fulfilled, the trial will be stopped for overwhelming efficacy. 
These pre-specified criteria are: 
o The average increase in the total UMSARS score is 4 points or less from baseline to week 48 in 
patients taking active agent – OR – 


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o The average increase in the total UMSARS score from baseline to week 48 in patients taking 
active agent is significantly lower than in patients taking placebo with a P<0.001. 
 
If the futility pre-specified criteria are fulfilled the trial will be stopped for futility. These pre-specified 
criteria are: 
o The average increase in the total UMSARS score is at least 7 points or more from baseline to 
week 48 in patients taking active agent  - OR – 
o The average increase in the total UMSARS score from baseline to week 48 in patients taking 
active agent is significantly higher than in patients taking placebo with a P<0.001.   
 
 
9.4.6 SUB-GROUP ANALYSES 
 
Not applicable 
 
10 
SUPPORTING DOCUMENTATION AND OPERATIONAL CONSIDERATIONS 
 
10.1 
REGULATORY, ETHICAL, AND STUDY OVERSIGHT CONSIDERATIONS  
 
10.1.1 INFORMED CONSENT PROCESS 
 
10.1.1.1 
CONSENT/ASSENT AND OTHER INFORMATIONAL DOCUMENTS PROVIDED TO 
PARTICIPANTS 
 
Consent forms describing in detail the study intervention, study procedures, and risks are given to the 
participant and written documentation of informed consent is required prior to starting 
intervention/administering study intervention.  The following consent materials are submitted with this 
protocol: 
 
o Consent form 
 
10.1.1.2 
CONSENT PROCEDURES AND DOCUMENTATION 
 
Informed consent is a process that is initiated prior to the individual’s agreeing to participate in the 
study and continues throughout the individual’s study participation. Consent forms will be Institutional 
Review Board (IRB)-approved and the participant will be asked to read and review the document.  
 
All subjects will have the capacity to provide informed consent. The subject’s capacity to provide 
informed consent will be confirmed by the subject’s treating physician in the course of receiving clinical 
care. 
 
The investigator or research coordinator will explain the research study to the participant and answer 
any questions that may arise. A verbal explanation will be provided in terms suited to the participant’s 
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 prior to signing. The participants should have the opportunity to discuss the 


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study with their family or think about it prior to agreeing to participate. The participant will sign the 
informed consent document prior to any procedures being done specifically for the study. Participants 
must be informed that participation is voluntary and that they may withdraw from the study at any 
time, without prejudice. A copy of the informed consent document will be given to the participants for 
their records. The informed consent process will be conducted and documented in the source document 
(including the date), and the form signed, before the participant undergoes any study-specific 
procedures. The rights and welfare of the participants will be protected by emphasizing to them that the 
quality of their medical care will not be adversely affected if they decline to participate in this study. 
10.1.2 STUDY DISCONTINUATION AND CLOSURE 
 
This study may be temporarily suspended or prematurely terminated if there is sufficient reasonable 
cause.  Written notification, documenting the reason for study suspension or termination, will be 
provided by the suspending or terminating party to study participants, investigator, funding agency, the 
Investigational New Drug (IND) sponsor and regulatory authorities.  If the study is prematurely 
terminated or suspended, the Principal Investigator (PI) will promptly inform study participants, the 
Institutional Review Board (IRB), and sponsor and will provide the reason(s) for the termination or 
suspension.  Study participants will be contacted, as applicable, and be informed of changes to study 
visit schedule. 
 
After consultation with the PI, the IRB and the IMM the entire trial may be stopped if, at least, 40% of 
enrolled patients develop either frequent (5 or more events of Grade 2 or above) or severe (at least one 
event of Grade 3 or above) adverse effects. 
  
Additional circumstances that may warrant termination or suspension include, but are not limited to: 
o Determination of unexpected, significant, or unacceptable risk to participants 
o Insufficient compliance to protocol requirements 
o Data that are not sufficiently complete and/or evaluable 
 
Study may resume once concerns about safety, protocol compliance, and data quality are addressed, 
and satisfy the NIH, IRB and/or Food and Drug Administration (FDA). 
 
 
10.1.3 CONFIDENTIALITY AND PRIVACY  
 
Participant confidentiality and privacy is strictly held in trust by the participating investigators, their 
staff, and the sponsor(s) and their interventions. This confidentiality is extended to cover testing of 
biological samples in addition to the clinical information relating to participants. Therefore, the study 
protocol, documentation, data, and all other information generated will be held in strict confidence. No 
information concerning the study or the data will be released to any unauthorized third party without 
prior written approval of the sponsor-investigator.  
 
All research activities will be conducted in as private a setting as possible. 
 
The study monitor, other authorized representatives of the sponsor, representatives of the Institutional 
Review Board (IRB), regulatory agencies or pharmaceutical company supplying study product may 
inspect all documents and records required to be maintained by the investigator, including but not 


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limited to, medical records (office, clinic, or hospital) and pharmacy records for the participants in this 
study. The clinical study site will permit access to such records. 
 
The study participant’s contact information will be securely stored at each clinical site for internal use 
during the study. At the end of the study, all records will continue to be kept in a secure location for as 
long a period as dictated by the reviewing IRB, Institutional policies, or sponsor requirements. 
 
Study participant research data, which is for purposes of statistical analysis and scientific reporting, will 
be transmitted to and stored in TrialMaster This will not include the participant’s contact or identifying 
information. Rather, individual participants and their research data will be identified by a randomization 
number. TrialMaster will be secured and password protected. At the end of the study, all study 
databases will be de-identified and archived at the NYU Dysautonomia Center. 
 
10.1.4 FUTURE USE OF STORED SPECIMENS AND DATA  
 
Data collected for this study will be analyzed and stored at the NYU Dysautonomia Center. After the 
study is completed, the de-identified, archived data will be stored at NYU Dysautonomia Center, for use 
by other researchers including those outside of the study. Permission to store data at the NYU 
Dysautonomia Center will be included in the informed consent.  
 
All left over de-identified biological samples will be destroyed upon study completion. No lef over 
samples will be kept for future analyses or investigations.  
 
10.1.5 KEY ROLES AND STUDY GOVERNANCE 
 
Co-Principal Investigator 
Co-Principal Investigator  
Independent Medical Monitor  
Horacio Kaufmann, MD 
Jose-Alberto Palma MD PhD 
Amir Steinberg MD PhD 
New York University School of 
Medicine 
New York University School of 
Medicine 
Mount Sinai School of Medicine 
Professor of Neurology 
Associate Professor of 
Neurology 
Associate Professor of Hematology and 
Oncology 
530 First Ave Suite 9Q 
New York, NY 10016 
530 First Ave Suite 9Q 
New York, NY 10016 
Tisch Cancer Institute 
One Levy Place 
Box 1410 
New York, NY 10029 
 


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212 263 7225 
212 263 7225 
 212 241 6021 
Horacio.Kaufmann@nyumc.org 
JoseAlberto.PalmaCarazo@nyu
mc.org 
amir.steinberg@mountsinai.org 
 
 
 
10.1.6 SAFETY OVERSIGHT AND CLINICAL MONITORING 
 
Because this is a single center study enrolling a small number of subjects and using a study drug that has 
been FDA-approved for over 10 years with a well-known profile of adverse event, an Independent 
Medical Monitor (IMM) will oversee the safety and data monitoring procedures of this study. The 
proposed IMM for this study is Amir Steinberg MD PhD, Associate Professor of Hematology & Oncology 
at the Mount Sinai School of Medicine. Dr. Steinberg has extensive expertise in sirolimus and its 
immunosuppressive side effects and has overseen several clinical trials in the past. Dr. Steinberg will be 
independent from study design and conduct and free from any potential conflict of interest.  
 
The PI, IMM and independent pharmacist will have monthly conference calls to assess AE and the 
overall safety of the study. During the meeting, the PI will offer information on recruitment, screen 
failures, withdrawals and AE. The PI will be responsible for identifying AE. Aggregate report -detailed by 
severity, attribution (expected or unexpected), and relationship to the study drug/study procedures- will 
be available to the IMM for review on a monthly basis. Additional responsibilities of the IMM will 
include: 
 
o Ensure that the rights and well being of trial participants are protected, that the reported trial 
data are accurate, complete, and verifiable. 
o The conduct of the trial is in compliance with the currently approved protocol/amendment(s), 
with International Conference on Harmonisation Good Clinical Practice (ICH GCP), and with 
applicable regulatory requirements. 
o Initial review of the research protocol and ongoing activities including data integrity, protocol 
adherence and study participant safety issues.  
o Monthly review of adverse events and reasons for losses to follow-up. 
o Raising any concern to the sponsor and PI and recommending the sponsor or PI continuation, 
modification or termination of the trial. 
o Protect the confidentiality of the trial data and results of monitoring.  
o Submit an annual report to the PI and the NYU IRB. 
 
 
10.1.7 QUALITY ASSURANCE AND QUALITY CONTROL 
 
The NYU Dysautonomia Center will perform internal quality management of study conduct, data and 
biological specimen collection, documentation and completion.  An individualized quality management 
plan will be developed to describe a site’s quality management. 
 


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Study team weekly meetings will be conducted to review subjects status, enrolment, adverse events and 
other regulatory issues.  
 
Quality control (QC) procedures will be implemented beginning with the data entry system and data QC 
checks that will be run on the database will be generated. Any missing data or data anomalies will be 
communicated to study personnel for clarification/resolution. 
 
Following written Standard Operating Procedures (SOPs), the IMM will verify that the clinical trial is 
conducted and data are generated and biological specimens are collected, documented (recorded), and 
reported in compliance with the protocol, International Conference on Harmonisation Good Clinical 
Practice (ICH GCP), and applicable regulatory requirements (e.g., Good Laboratory Practices (GLP), Good 
Manufacturing Practices (GMP)).  
 
The NYU Dysautonomia Center will provide direct access to all source data/documents, and reports for 
the purpose of monitoring and auditing by the sponsor, and inspection by local and regulatory 
authorities. 
 
10.1.8 DATA HANDLING AND RECORD KEEPING  
 
10.1.8.1 DATA COLLECTION AND MANAGEMENT RESPONSIBILITIES  
Data collection is the responsibility of the clinical trial staff at the site under the supervision of the site 
investigator. The investigator is responsible for ensuring the accuracy, completeness, legibility, and 
timeliness of the data reported. 
 
All source documents should be completed in a neat, legible manner to ensure accurate interpretation 
of data.   
 
Hardcopies of the study visit worksheets will be provided for use as source document worksheets for 
recording data for each participant enrolled in the study.  Data recorded in the electronic case report 
form (eCRF) derived from source documents should be consistent with the data recorded on the source 
documents.  
 
Clinical data (including AEs, concomitant medications, and expected adverse reactions data) and clinical 
laboratory data will be entered into TrialMaster, a 21 CFR Part 11-compliant data capture system 
provided by the NYU CTSI. The data system includes password protection and internal quality checks, 
such as automatic range checks, to identify data that appear inconsistent, incomplete, or inaccurate. 
Clinical data will be entered directly from the source documents. 
 
Data quality will be assessed at the data entry point using intelligent on-line forms using TrialMaster. 
Data element constraints, whether independent range and/or format limitations or ‘relative’ referential 
integrity limitations, can be enforced by all methods employed for data input. QA reports assess data 
quality post-data entry. As we note, data quality begins with the design of the data collection forms and 
procedures and incorporates reasonable checks to minimize transcription and omission errors. Of the 
more important quality assurance measures are the internal validity checks for reasonableness and 
consistency. In addition to those described above, we propose to build these checks into the initial 
tables and cross tabulations that should reveal any remaining data quality issues. All study data will 
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Registration: A system of coded identifiers will be used to protect participant confidentiality and safety. 
Each participant enrolled will be assigned a local identifier by the enrollment site. Local IDs should follow 
the following format: two letters followed by three numbers (i.e., JD001). The two letters correspond to 
the initials of the first name, and the last name of the patient (e.g., John Doe). When the participant is 
registered to participate in the study, using TrialMaster, the system will assign a participant ID number.  
 
Data Entry: Data collection for this study will be accomplished with paper and online electronic case 
report forms by using TrialMaster. Using encrypted communication links, on-line forms will be 
developed that contain the requisite data fields.  
 
Data Quality Control: As much as possible data quality is assessed at the data entry point. Data element 
constraints, whether independent range and/or format limitations or ‘relative’ referential integrity 
limitations, can be enforced by all methods employed for data input. QA reports assess data quality 
post-data entry. As we note, data quality begins with the design of the data collection forms and 
procedures and incorporates reasonable checks to minimize transcription and omission errors. Of the 
more important quality assurance measures are the internal validity checks for reasonableness and 
consistency. In addition to those described above, we propose to build these checks into the initial 
tables and cross tabulations that should reveal any remaining data quality issues.  
o Data Monitoring: The data manager will identify missing or unclear data and generates a data 
query to the study manager. 
o Data Delinquency Tracking: The data manager will monitor data delinquency on an ongoing 
basis.  
 
Laboratory data flow  
On-line forms and/or electronic data exchange mechanisms to enter, update and obtain relevant data 
will be also available.  
Additional Documents and Records  
o Participant Screening and Assignment Log – A listing of all participants who signed the informed 
consent and were screened.  
o Drug Accountability Log – This form documents the total amount of study drug dispensed to and 
returned by each participant throughout the study.  
o Adverse event log 
o Protocol deviation log 
 
10.1.8.2 STUDY RECORDS RETENTION  
 
Study documents should be retained for a minimum of 2 years after the last approval of a marketing 
application in an International Conference on Harmonization (ICH) region and until there are no pending 
or contemplated marketing applications in an ICH region or until at least 2 years have elapsed since the 
formal discontinuation of clinical development of the study intervention. These documents should be 
retained for a longer period, however, if required by local regulations. No records will be destroyed 
without the written consent of the sponsor, if applicable. It is the responsibility of the sponsor to inform 
the investigator when these documents no longer need to be retained. 
 
Primary source documents 
The investigator must maintain primary source documents to support CRF data entries. These 
documents, which are considered “source data”, may include but are not limited to:  


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o Demographic information  
o Evidence supporting the diagnosis/condition for which the participant is being studied  
o General information supporting the participant’s participation in the study  
o Medical history and physical findings  
o Hospitalization or Emergency Room records (if applicable)  
o Each study visit by date, including any relevant findings/notes by the investigator(s), occurrence 
(or lack) of adverse events, and changes in medication usage, including the date the study drug 
was commenced and stopped.  
o Any additional visits during the study  
o Any relevant telephone conversations with the participant regarding the study or possible 
adverse events  
o Original, signed informed consent forms for study participation  
 
The investigator must also retain all participant specific printouts/reports of tests/procedures 
performed as a requirement of the study. During monitoring visits, the monitor will need to verify data 
in the CRFs against these source data. 
 
 
10.1.9 PROTOCOL DEVIATIONS  
A protocol deviation is any noncompliance with the clinical trial protocol, International Conference on 
Harmonisation Good Clinical Practice (ICH GCP), or Manual of Procedures (MOP) requirements. The 
noncompliance may be either on the part of the participant, the investigator, or the study site staff. As a 
result of deviations, corrective actions are to be developed by the site and implemented promptly.  
 
These practices are consistent with ICH GCP:  
• 
4.5 Compliance with Protocol, sections 4.5.1, 4.5.2, and 4.5.3  
• 
5.1 Quality Assurance and Quality Control, section 5.1.1  
• 
5.20 Noncompliance, sections 5.20.1, and 5.20.2.  
 
It is the responsibility of the site investigator to use continuous vigilance to identify and report 
deviations within 30 working days of identification of the protocol deviation, or within 40 working days 
of the scheduled protocol-required activity.   
 
Protocol deviations that compromise patient safety will be reported to the IRB.  
 
A protocol deviation log will be maintained.  
 
10.1.10 
PUBLICATION AND DATA SHARING POLICY 
 
This study will be conducted in accordance with the following publication and data sharing policies and 
regulations: 
 
National Institutes of Health (NIH) Public Access Policy, which ensures that the public has access to the 
published results of NIH funded research. It requires scientists to submit final peer-reviewed journal 
manuscripts that arise from NIH funds to the digital archive PubMed Central upon acceptance for 
publication. 
 


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This study will comply with the NIH Data Sharing Policy and Policy on the Dissemination of NIH-Funded 
Clinical Trial Information and the Clinical Trials Registration and Results Information Submission rule. As 
such, this trial will be registered at ClinicalTrials.gov, and results information from this trial will be 
submitted to ClinicalTrials.gov. In addition, every attempt will be made to publish results in peer-
reviewed journals.  Data from this study may be requested from other researchers 10 years after the 
completion of the primary endpoint by contacting Horacio Kaufmann MD.  
 
 
10.1.11 
CONFLICT OF INTEREST POLICY 
The independence of this study from any actual or perceived influence, such as by the pharmaceutical 
industry, is critical.  Therefore, any actual conflict of interest of persons who have a role in the design, 
conduct, analysis, publication, or any aspect of this trial will be disclosed and managed. Furthermore, 
persons who have a perceived conflict of interest will be required to have such conflicts managed in a 
way that is appropriate to their participation in the design and conduct of this trial.  The study 
leadership in conjunction with the NINDS has established policies and procedures for all study group 
members to disclose all conflicts of interest and will establish a mechanism for the management of all 
reported dualities of interest. 
 
 
 
 


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10.2 ABBREVIATIONS 
 
AE 
Adverse Event 
αSyn 
Alpha-synuclein 
CBC 
Complete blood count 
CFR 
Code of Federal Regulations 
CLIA 
Clinical Laboratory Improvement Amendments 
CMP 
Comprehensive metabolic panel 
CNS 
Central Nervous System 
COC 
Certificate of Confidentiality 
CONSORT 
Consolidated Standards of Reporting Trials 
CSF 
Cerebrospinal Fluid 
CRF 
Case Report Form 
DCC 
Data Coordinating Center 
DHHS 
Department of Health and Human Services 
DSMB 
Data Safety Monitoring Board 
DRE 
Disease-Related Event 
eCRF 
Electronic Case Report Forms 
EKG 
Electrocardiogram 
FDA 
Food and Drug Administration 
FDAAA 
Food and Drug Administration Amendments Act of 2007 
FFR 
Federal Financial Report 
GCP 
Good Clinical Practice 
GCI 
Glial Cytoplasmic Inclusions 
GLP 
Good Laboratory Practices 
GMP 
Good Manufacturing Practices 
HIPAA 
Health Insurance Portability and Accountability Act  
HIV 
Human Immunodeficiency Virus 
ICH 
International Conference on Harmonization  
ICMJE 
International Committee of Medical Journal Editors 
IND 
Investigational New Drug Application 
IRB 
Institutional Review Board 
IMM 
Independent Medical Monitor 
ISO 
International Organization for Standardization 
MCID 
Minimally Clinically Important Difference 
MRI 
Magnetic resonance imaging 
MOP 
Manual of Procedures 
MoCA 
Montreal Cognitive Assessment 
MPTP 
1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine 
MSA 
Multiple System Atrophy 
MSDS 
Material Safety Data Sheet 
mTOR 
Mammalian Target of Rapamycin 
mTORC1 
Mammalian Target of Rapamycin Complex 1 
mTORC2 
Mammalian Target of Rapamycin Complex 2 
NCT 
National Clinical Trial 
NIH  
National Institutes of Health 
NINDS 
National Institutes of Neurological Disorders and Stroke 
OCT 
Optical coherence tomography 
OHRP 
Office for Human Research Protections 
PI 
Principal Investigator 


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PD 
Parkinson disease 
QA 
Quality Assurance 
QC 
Quality Control 
RDCRC 
Rare Diseases Autonomic Disorders Clinical Research Consortium 
SAE 
Serious Adverse Event 
SAP 
Statistical Analysis Plan 
SMC 
Safety Monitoring Committee 
SOA 
Schedule of Activities 
SOC 
System Organ Class 
SOP 
Standard Operating Procedure 
UMSARS 
Unified Multiple System Atrophy Rating Scale 
UP 
Unanticipated Problem 
US 
United States 
 
 
 


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10.3 PROTOCOL AMENDMENT HISTORY 
 
Version 
Date 
Description of Change  
1 
1 June 2018 
 
2 
 
3 September 2018 
Minor changes including clarification of sirolimus levels schedule, 
clarification that sirolimus levels will be measured in whole blood, 
extension of the screening period to 42 days (instead of 30), and 
clarification that sirolimus (Rapamune®) marketed by Pfizer will be 
used (instead of generic sirolimus) 
3 
28 September 2018 Specification that the Town Total Compounding Pharmacy will 
perform the randomization and delivery of study drug, with no 
involvement of the NYU Investigational Pharmacy 
4 
15 October 18 
24-hour urine collection at baseline will be performed only if 
baseline occurs > 30 days from the 24-h urine assessment of the 
screening. 
 
Sirolimus levels in blood during the first month will take place 7 
days after the previous dose adjustment has taken place. 
 
Safety labs to the performed after a light (low fat) breakfast 
 
Clarification that the IMM is unblinded to the study allocation 
5 
19 March 19 
Elimination of the need to perform autonomic testing to diagnose MSA 
as inclusion criteria  
 
Clarification of exclusion criteria including: 
 
-Specification that heart failure and cardiopulmonary disease should be 
severe and uncontrolled. 
 
 
- Specification that patients with positive Quantiferon® may be enrolled 
after an infectious diseases specialist has cleared the patient for this trial 
owing to the low probability of reactivation when patient is 
immunosuppressed with sirolimus. 
 
- Modification of the unhallowed high levels for LDL cholesterol and 
triglycerides 
 
-Clarification that study intervention will be modified based on study 
drug levels and the frequency of adverse events related to the study 
drug.  
 
Clarification that the active drug and placebo are administered in 
capsules, not in tablets  
 
Clarification of windows for study visits.  
 


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Clarification on when sirolimus levels will be performed and how the 
dose adjustment will be performed 
 
Specification of Unscheduled Visits.  
 
Clarification on reporting of serious adverse events, both expected and 
unexpected.  
 
Clarification on when an interim analysis for efficacy will be performed  
 
Modification on how the ID of each subject Specification that a protocol 
deviation log will be created and maintained for this study will be 
obtained 
6 
25 Jul 2019 
Clarification of target sirolimus levels to 5-20 ng/ml (instead of 10-20 
ng/ml)  
Visit 7 may be skipped to account for delays in previous visits, to ensure 
that Visit 8 occurs 3-months after initiation of study drug.  
 
Interim analyses will ensure that there is evidence for the study to 
continue - or to declare it futile earlier than expected.  
 
Specification that the study will terminate if 30% of patients early 
withdraw due to disease progression or side effects 
7 
29 Oct 2019 
The study will be using DataCore/EPIC for recruitment. 
8 
19 Dec 2019 
Clarification of the criteria to unblind the allocation to active agent or 
placebo 
 
Specification that the PI and IMM and the independent pharmacist will 
have monthly conference calls to evaluate the safety of the study. 
 
 


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APPENDICES 
 
Appendix 1. Unified Multiple System Atrophy Rating Scale 
 
Appendix 2. Montreal Cognitive Assessment