Clinical trial NCT02925650
Conditions: Alzheimer's Disease
Keywords: Alzheimer's Disease SILK™ Pharmacodynamics Safety Pharmacokinetics
Sponsor: Annovis Bio Inc.
Start date: 2017-03-02; Completion date: 2021-12-31
Enrollment: 18 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
This study evaluates the safety and pharmacological effects of 3 different doses of Posiphen® when compared to a placebo, in adult male and female patients with early Alzheimer's disease (AD).

Detailed description:
Posiphen®, which was discovered by the US National Institute on Aging (NIA) is a small, orally active, experimental drug that specifically inhibits the synthesis of amyloid precursor protein (APP), Tau and α-Synuclein. It is distinct from other Alzheimer's disease drugs currently in development, because it inhibits the formation of several toxic proteins, rather than removing individual toxic protein after they are produced. Posiphen has potential utility as a disease modifying treatment for AD. The present study will confirm the pharmacokinetics (PK) of Posiphen and its metabolites in plasma and cerebral spinal fluid (CSF). It will also measure the effects of a 23-25 day treatment period with Posiphen on the CSF and plasma levels of a number of biomarkers, inflammatory factors and control proteins. It will also expand the safety data in humans by extending the treatment period from 10 days to a treatment period from 23-25 days. In addition, this study will measure concentrations of various soluble biomarkers in CSF and use the SILK™ assay methods to directly measure the effect of Posiphen on the fractional synthesis rate of Aβ40 in CSF, which will help guide the further development of Posiphen and determine the feasibility of SILK™ in a multicenter trial.

Arms: arm 1: The study drug Posiphen dosage of 60mg is to be taken orally in divided doses, three times per day for a total 23-25 days. arm 2: The study drug Posiphen dosage of 120mg is to be taken orally in divided doses, three times per day for a total 23-25 days. arm 3: The study drug Posiphen dosage of 180mg is to be taken orally in divided doses, three times per day for a total 23-25 days. arm 4: The Placebo comparator is to be taken orally in divided doses, three times per day for a total 23-25 days.
Interventions: intervention 1: Posiphen intervention 2: Placebo
Intervention descriptions: intervention 1: oral solid dosage form capsule intervention 2: oral solid dosage form capsule

Protocol documents:
ICF_003.pdf: 
 
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Informed Consent to Act as a Research Subject In: 
A Safety and Pharmacokinetics- 
Pharmacodynamics Study in Subjects 
with Early Alzheimer’s Disease 
This consent form describes a research study and your role as 
a research subject.  This document is intended to inform you 
about the possible risks and benefits of the research study, 
other options that may be available to you and your rights as 
a research subject.  Please read this consent form carefully 
and do not hesitate to ask the study doctor any questions you 
may have about the study or the information provided below. 
Your participation in this study is entirely voluntary.  Please 
take as much time as you need to discuss the study with your 
doctors, family and friends.  The decision to participate or not 
is yours.  If you choose to participate, you have the right to 
withdraw from the study at any time.   
A description of this research study will be available on 
http://www.ClinicalTrials.gov.  This website will not include 
information that can identify you.  At most, the website will 
include a summary of the results.  You can search this website 
at any time.  
This study is being conducted by the Alzheimer’s Disease 
Cooperative Study (ADCS), through a grant from the National 
Institute on Aging (NIA).  Drug will be provided by QR, Inc. 
 
 
 
 
 
 
 
 
 
 
 
Investigator(s) 
Sample Doctor, MD 
Sample Doctor, PhD 
 
Location 
Site Address 
Site Address 
Site Address 
Site Address 
 
Protocol Title 
A Multicenter, Randomized, 
Double-Blind, Placebo-Controlled 
Ascending Dose Study to 
Evaluate the Safety, Tolerability, 
Pharmacokinetics (PK) and 
Pharmacodynamic (PD) Effects of 
Posiphen® in Subjects with Early 
Alzheimer’s Disease (AD) 
 
Protocol Number ADC-043-DISC 
(
)


 
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Why is this study being done?  
Alzheimer’s disease (AD) is a brain disease that results in a loss of mental function. 
 A protein called “amyloid” or “beta-amyloid” forms plaques in the brains of people with AD, and 
scientists believe the build-up of amyloid may play a key role in the developing the disease. The drugs 
currently approved by the United States Food and Drug Administration (FDA) for AD treat symptoms 
such as memory loss but do not prevent the disease, delay its onset or slow its progression.  
As an alternative to being in the study, you may ask your doctor about receiving Aduhelm™ (an 
antibody also known as aducanumab) which has received accelerated approval by the FDA for the 
treatment of AD. This treatment Aduhelm™  is not permitted within this study. 
You are being invited to take part in a research study that will look at an experimental drug called 
“Posiphen”.  This means it has not been approved by the United States Food and Drug Administration 
(FDA) for the treatment of AD. You are being asked to take part in this study because you are 55-89 
years old (inclusive), in good health with no frailty,  and have a clinical profile consistent with early AD 
including Mild Cognitive Impairment due to Alzheimer’s disease (MCI-AD) or Mild AD.   
Posiphen is being developed as an anti-amyloid drug that may delay the onset or slow the 
progression of possible AD-related brain damage due to amyloid build-up. 
The purpose of this study is to: 
• test the safety and tolerability of the study drug. 
• measure the amount of the study drug and drug-related material  in the cerebrospinal fluid (CSF, 
the fluid surrounding the brain and spinal cord) and blood. 
• measure the effect of Posiphen on the levels of certain proteins (biomarkers) associated with AD 
in the blood and CSF.  
In this study, participants will be given either active study drug (Posiphen 60 mg taken either once, 
twice or three times per day) or placebo (capsules that looks like the study drug but do not contain 
any active drug). 
How many people will take part in this study?  
This study is being conducted at 3 to 6 sites across the US.  If you are eligible and agree to participate, 
you will be one of approximately 24 individuals enrolled in this study. 
What will happen if I take part in this research study? 
In this study, participants will take one capsule of Posiphen or placebo, up to three times a day by 
mouth for up to 25 days. 
This is a randomized, double-blind, placebo-controlled study.  This means that  if you choose to 
participate, it will be determined by chance (for example, flipping a coin) which of the two groups 
(active or placebo) you will be assigned to at each dose level (60 mg once per day,  60 mg twice per 


 
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day or 60 mg three times per day of active study drug, or placebo).   Since approximately two-thirds 
of subjects will receive active study drug, there is a greater chance that you will receive Posiphen 
rather than the placebo.   
Neither you nor the study personnel will know whether you are receiving the active study drug or 
placebo.  However, if it becomes medically necessary, Dr. [INSERT PI NAME] may obtain this 
information.   
Because this is a research study, Posiphen will be given only during this study and not after the study 
is over.  
If you are to qualify and agree to participate in the study, your participation will last up to 68 days and 
will require 5 study visits - including an approximate 3-day stay (Confinement visit) at the clinical 
research unit.  One of those 5 “visits” will be done over the telephone.  Study staff will explain this to 
you and work out the best schedule for you to complete all of the study procedures. 
To participate in this study, you must have a spouse, friend or relative (called a “Study Partner”), who 
is willing to: 
• Attend the entire screening and baseline visits with you. Your study partner should also stay for 
the first three hours of the confinement visit and return to drive you home.  
• Answer questions about your memory, behavior and daily functioning. 
• Make sure that you are taking the study drug as instructed. 
• Tell the study staff of any changes in your health status over the period of this study. 
While you are in the study, you must: 
• Follow the instructions you are given. 
• Come to the study site for all visits.  
• Tell the study staff about any changes in your health. 
• Tell the study doctor about all medications you are taking and check with the study doctor 
before you begin taking a new medication.  
• Tell the study doctor or study staff if you want to stop being in the study at any time. 
During this study, Dr. [INSERT PI NAME] and [his/her] staff will be monitoring your condition.   
Description of Visit Activities 
Screening Visit (Visit 1) 
The first part of the study is called the Screening Period.  During this time, the study team will decide 
if you qualify to be in the study.  The screening activities could take place over 2 days at the clinic over 
a 6 week period.  Each visit may take several hours to complete. 
At the beginning of your first clinic visit, the study staff will explain all of the study procedures and 
answer any questions that you and your study partner may have.   


 
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You or your legally authorized representative (if applicable), and also your study partner must sign 
this consent form before beginning the screening process.  After you sign consent to participate you 
should report all injuries, reactions to medications or illnesses to the staff at this site so that it can be 
recorded in your study records. 
During the Screening Visit (which includes your Study Partner): 
• Your general medical history will be recorded and your medical records will be reviewed. 
• A list of the vitamins and current medications you are taking will be collected. 
• Your weight and vital signs (blood pressure, pulse, temperature, and respiration rate) will be 
recorded. 
• A physical and neurological exam will be performed. 
• An electrocardiogram (ECG) will also be performed.  An ECG is a tracing test that measures 
electrical signals from your heart. 
• You will be given tests of your memory and thinking and you will be asked questions about your 
daily functioning and your behavior.  You can skip any questions you do not want to answer, and 
you can take breaks if needed.  
• Blood will be drawn from a vein in your arm and you will be asked for a urine sample. The blood 
and urine samples are for routine laboratory tests to make sure you do not have any medical 
conditions that would interfere with your participation in the study. 
You may undergo an MRI (magnetic resonance imaging) scan during the Screening Period if you 
have not had a scan within the last 12 months before the Screening Visit.  An MRI uses a large 
magnet and computer equipment to produce electronic pictures of your brain.  Each MRI will take 
approximately 45 minutes to complete.  You will lie on your back and enter the MR machine for 
the scan, during which time you will hear loud knocking noises.  
People with pacemakers, aneurysm clips, cochlear implants, or metal/foreign objects in their eyes 
are not permitted to undergo MR studies. 
• A lumbar puncture (LP) - also known as a “spinal tap” - will be performed. A lumbar puncture is a 
procedure that involves inserting a needle in the lower back in order to collect a small amount 
(16-20 mL - approximately 1 tablespoon), of the spinal fluid that surrounds the brain and spinal 
cord.  Your body will replace this spinal fluid within 1-2 hours.  If you have had a lumbar puncture  
for another study, within the last 90 days at this site,  the samples may be used in place of the 
screening lumbar puncture. 
You will be asked to fast overnight (a minimum of 8 hours) prior to coming to the clinic for the LP.  
This means no food or drinks such as coffee, tea, milk and juice (water is permitted). 
During the procedure you will lay on your side curled up into a ball or you will sit on the edge of a 
chair or bed and lean forward, whichever is easier for you.  The lower part of your back will be 
cleaned with an antiseptic.  A local anesthetic will be injected into the skin of your lower back at 
the area of the lumbar puncture.  When the area is numb, a very thin needle will be inserted into 
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After the lumbar puncture is complete, you will remain at the site for about 30 minutes. You will 
be given something to eat and drink before you leave. You should avoid any strenuous physical 
activity for the next 24 hours.  This includes lifting, bending, doing housework and gardening, or 
doing exercise such as jogging or bicycle riding. 
Baseline Visit (Visit 2) 
If the screening phase shows that you are eligible to continue and participate in this study, you will 
return to the clinic for the Baseline evaluation. 
During the Baseline Evaluation (which includes your Study Partner): 
• You will be given tests of your memory and thinking and you will be asked questions about 
your daily functioning and your behavior.   
• A physical and neurological exam will be performed. 
• Your weight and vital signs (blood pressure, temperature, etc.) will be recorded. 
• You will be asked about any adverse events (changes in health or any side effects) that you 
may have experienced since your last visit. 
• A list of vitamins and medications you are currently taking will be collected. 
• You will be asked questions about how you feel about being in this study.  
If you continue to meet all protocol inclusion criteria and no exclusion criteria, after completion of all 
baseline procedures, you will be randomly assigned to receive active drug or placebo in each of the 
three treatment groups:  
GROUP 
Posiphen 60 mg Once per day or Placebo 
Posiphen 60 mg Twice per day or Placebo 
Posiphen 60 mg Three times per day or Placebo 
You will be given a supply of study drug and will be instructed to take your first dose of study drug the 
morning after you have completed the Baseline visit.  You will take 3 capsules per day of either study 
drug or placebo, with or without food, depending upon your group assignment.   
You will be asked to bring all study drug blister cards (used and unused) with you to Visits 4 and 5 so 
that we can determine how many capsules you have taken.  
If a dose is missed, do not take the missed dose later in the day.  Just skip the missed dose and take 
the next scheduled doses.  
The study drug must be used only by you and should be stored at room temperature, protected from 
moisture and kept out of the reach of children. 
Day 14 Telephone Contact (Visit 3) 
You will be contacted 14 (+/- 4) days from the baseline visit for the following:  


 
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• You will be asked about any adverse events (changes in health or any side effects) that you 
may have experienced since your last visit. 
• A list of vitamins and medications you are currently taking will be collected. 
• Your study medication compliance will be discussed. 
Pre-Confinement Visit (Visit 4) 
Admission for the confinement visit should occur between 21 and 23 days following the first dose of 
study medication.  The pre-confinement visit will be conducted 1-3 days prior to admission. 
During the Pre-Confinement Visit (which includes your Study Partner): 
• You will be asked to bring all your used and unused study medication blister cards to this visit 
so your study medication compliance can be reviewed. 
• A list of vitamins and medications you are currently taking will be collected. 
• A physical and neurological exam will be performed. 
• Blood will be drawn from a vein in your arm for routine laboratory tests to make sure you do 
not have any medical conditions that would interfere with your continued participation in the 
study. 
• You will be given tests of your memory and thinking and you will be asked questions about 
your daily functioning and your behavior.   
• You will be asked about any adverse events (changes in health or any side effects) that you 
may have experienced since your last visit. 
Confinement Visit (Visit 5): 
The confinement visit will last approximately 3 days and will consist of four phases: Admission, 36-
Hour Sampling, 12-Hour Observation and Discharge.  You will be provided with meals and a place to 
sleep during this visit. 
Phase 1- Admission: 
• You should arrive early in the morning after fasting overnight (a minimum of 8 hours). 
• Blood will be drawn from a vein in your arm for routine laboratory tests to make sure you do 
not have any medical conditions that would interfere with your participation in the study. 
• You will be expected to bring all your used and unused study medication blister cards to this 
visit so your study medication compliance can be reviewed. 
• Your weight and vital signs will be recorded. 
• A brief physical and neurological exam will be performed. 
• Adverse events and your current vitamins and medications will be reviewed and recorded.  
Phase 2 – 36-Hour Sampling: 
During this phase, the following will take place over a 36 hour period: 


 
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• A brief physical and neurological exam will be performed. 
• Vital signs will be collected and any adverse events and your current vitamins and medications 
will be reviewed and recorded.  
• An intravenous (IV) line will be inserted into your vein to use for drawing blood intermittently 
for laboratory tests. 
• A second IV line will be inserted into your vein to administer a form of the amino acid leucine 
called “HEAVY LEUCINE”.  HEAVY LEUCINE is nearly identical to the leucine found in food 
except for a small difference in weight, and is harmless to humans and the environment. 
Amino acids are used by the body to make proteins.  You will be given the HEAVY LEUCINE in 
order to measure certain proteins associated with AD that are made by the brain.  
• You will take your study medication as instructed. 
• You will undergo a Lumbar Puncture (LP) with placement of a catheter (a long, thin flexible 
plastic tube) for Cerebral Spinal Fluid (CSF) collection every 2 hours for a total of 36 hours over 
the course of your stay.   
A local anesthetic will be administered to temporarily numb a small area of skin on the lower 
back.  A catheter (flexible tube) will be inserted through a lumbar spinal needle into the spinal 
fluid sac. 
After the catheter is in place, the lumbar spinal needle is removed and the catheter remains 
until the final CSF sample is taken.  After the CSF collection process is complete, the spinal 
catheter will be removed by the physician/nurse. 
You will remain at bedrest throughout the CSF collection (sampling) period of 36 hours.  You 
are allowed to use a bedside commode for toileting. 
To reduce the chance of a blood clot in your legs, you will wear anti-embolism stockings and 
also will be encouraged to do leg exercises periodically while lying flat in bed.   
You may be required to stay in the clinical study unit longer if the study doctor believes it is in 
your best interest.   
Additional physical exams and vital signs may be performed during your stay in the clinical 
study unit as required by the clinical study unit’s policy or at the discretion of the study doctor.  
The above tests may need to be repeated or additional tests may need to be performed for 
your safety. 
• You will be provided with a special low-leucine meal plan designed for this study so that your 
body uses the HEAVY LEUCINE to make proteins. Many foods that you are familiar with are 
included in this diet, including a variety of fruits (for example, bananas, pears and peaches), 
vegetables (for example, tomatoes, carrots and lettuce) and certain wheat products. 
• Over the course of your stay, you will be continually monitored by the study staff.  You will be 
asked questions about any symptoms or discomfort you may be experiencing as a result of the 
lumbar puncture and infusions. 
 


 
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Phase 3 – 12-Hour Observation:    
• Blood will be drawn from a vein in your arm for laboratory tests.  
• You will be observed for at least 2 hours following the removal of the LP catheter. 
• You will be asked if you are experiencing any pain or discomfort from the spinal tap, and given  
mild analgesics (Ibuprofen, Naproxen or Tylenol) if necessary. 
• All of your used and unused study drug blister cards should be returned to the study staff for 
final drug accountability. 
• You will be asked questions about how you feel about being in this study.  
Phase 4 – Discharge: 
• Your weight and vital signs will be collected and any adverse events will be reviewed and 
recorded.  
• Blood will be drawn from a vein in your arm for laboratory tests. 
• You will be discharged after the study staff determine you are stable to leave. 
Post-Confinement 24-Hour Phone Follow-up: 
You will receive a phone call from the study staff about 24 hours following discharge from the 
Confinement Visit.  You will be asked how you are feeling and if you experienced any new adverse 
events since your discharge. 
 Early Treatment Discontinuation  
If you stop participating in the study for any reason (for example, the study team feels it is in your 
best interest, you decide not to continue or the study is stopped), you will be asked to return to the 
clinic for a final evaluation visit.  
This final evaluation will include all of the procedures normally performed at the Pre-confinement 
Visit.  It is important for your health and safety to have these final procedures completed. 
Your participation in the study may also be stopped by the study doctor or Sponsor without your 
permission.  If this happens, it might be because: 
• You had a bad reaction to the study drug. 
• You were started on a medication that might cause you to have a bad reaction. 
• The sponsor decides to stop doing the study for any reason. 
• The study doctor stops doing the study for other reasons. 
Summary of Study Visit Activities 
The following chart summarizes the activities that will occur over the course of this study. 


 
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Visit Number 
1 
2 
3 
4 
5 
Visit Name Screening 
Baseline 
Phone 
Contact 
Pre-
Stay 
3-Day Stay in Clinic 
Length of Visit 
(hours) 
3-4 
5-6 
15 
Minutes 
1 
72 
Admit 
to 
Clinic 
36-Hour 
Sampling 
12-hour 
Observatio
n 
Discharge  
Informed Consent 
 
 
 
 
 
 
 
 
Personal Info,  
Medical & 
Psychiatric History 
 
 
 
 
 
 
 
 
Physical & 
Neurological Exam 
 
 
 
 
 
 
 
 
Current Vitamins & 
Medications 
Collected 
 
 
 
 
 
 
 
 
Vital Signs Recorded 
 
 
 
 
 
 
 
 
Weight Recorded 
 
 
 
 
 
 
 
 
Height Recorded 
 
 
 
 
 
 
 
 
Blood Sampling *** 
 
 
 
 
 
 
 
 
Urine Collected 
   
 
 
 
 
 
 
 
12-Lead ECG 
 
 
 
 
 
 
 
 
Memory & Thinking 
Skills Tests 
 
 
 
 
 
 
 
 
Functional & 
Behavioral 
Questionnaires 
 
 
 
 
 
 
 
 
Research 
Satisfaction Survey  
 
 
 
 
 
 
 
 
Adverse Events 
Collected 
 
 
 
 
 
 
 
 
Lumbar Puncture 
(LP) **** 
 
 
 
 
 
 
 
 
MRI of Brain * 
 
 
 
 
 
 
 
 
LP Catheter 
Placement &  CSF 
Sampling 
 
 
 
 
 
 
 
 
Study Drug 
Dispensed 
 
 
 
 
 
 
 
 
Study Drug 
Compliance 
 
 
 
 
 
 
 
 
Telephone Contact** 
 
 
 
 
 
 
 
 
Study Partner Attend 
Visit 
 
 
 
 
 
 
 
 
    *  A scan will be performed only if one had not been performed within 12 months of Visit 1 


 
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  **  The study team will call you within 24 hours after Visit 1 and after Discharge to see how you are 
feeling 
***  PK sampling also obtained at 36-Hour Confinement Visit 
****If an LP has been done within 90 days, the LP and samples may be used in place of Screening LP 
Genetic & Biomarker Research  
During this study, blood and CSF samples will be collected from you for genetic and biomarker 
research.  The samples will be sent to laboratories contracted for the study and to the ADCS 
Biomarker Core at the University of California, San Diego (UCSD).   
Genetic Study 
Previous studies have shown that a gene called apolipoprotein E (“APOE”) may influence the rate of 
disease progression or a subject’s response to treatment.  We will test your blood to see what form of 
the APOE gene you have.  This genetic testing is not optional.  
The results of these tests will be maintained in scientific databases for this research study.  These 
results are important only for research - not for helping to care for you.  For this reason, the results 
will not be released to you or your family.  
No information regarding your genetic or biomarker results will be entered into your regular medical 
record.  If you are concerned about a potential genetic disorder, you should discuss this with your 
primary care doctor.  You and your doctor may choose to test specifically for it, but this would require 
additional blood samples and would not be part of this research project. 
Data from your tests will not be revealed to other sites that are participating in the clinical study, 
family members, insurance companies, employers, or other individuals or organizations.  
Although the study researchers and the study Sponsor will have access to coded individual data, any 
information gained from this research will be reported in publications in an anonymous summary 
form.  Data will be stored in a locked file, and in a computer with restricted access.  
Any information that could be used to potentially identify you in the computer will be stored in a 
separate file and encrypted.  Only the researchers and their research assistants will have access to the 
original research data. 
Your samples will be stored for up to 20 years.  More research may be performed on your sample at a 
future date.  You will not be notified at the time this additional research is conducted, nor will any 
additional informed consent be obtained.  This research will be limited to the health information 
recorded by your study doctor or generated from any blood samples collected from this study.  You 
will not be contacted in the future to provide any further information. 
By signing this consent form, you give the Sponsor and other commercial or academic third parties 
that collaborate with the Sponsor, permission to use the material obtained from your sample for 
pharmacogenetics research.  At any time for any reason, you may withdraw consent for this genetic 
testing and request that your sample be destroyed with no further testing being performed. 
Sample Storage & Future Use  


 
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With your consent, we would like to store your DNA sample for future research studies related to 
Alzheimer’s disease and other neurodegenerative disorders.  If you decide you do not want to have 
your DNA sample stored for future research, you may still participate in this study.  
Storage of your remaining blood and CSF samples is required for this study and it is not optional.  If 
you decide that you do not want your blood and CSF samples stored for future research, you may not 
participate in this study. 
Your samples will be stored by code number and no identifying information will be included with 
them.  Samples will be stored indefinitely at the ADCS Biomarker Core and may be shared with other 
researchers studying AD or aging.  
If your samples are sent to other researchers, they will be identified only by a code number and 
descriptive data (such as your age and gender).  No other personal identifying information will be 
attached to your samples, so it will not be possible to identify you from any of the samples. 
MRI Image Storage & Future Use 
Your MRI images will be sent to the ADCS and will be analyzed by researchers at other institutions.  
ADCS investigators will maintain your imaging data and be responsible for deciding how it will be 
used for future research.    
Study investigators may make some of this data available to investigators at other scientific 
institutions for research purposes. 
Your name and all links to your identity will be removed from the data before it is shared.  Your 
imaging data will be labeled with a coded research identifier to protect your identity. 
Will this research data be shared? 
Data from this research will be shared with other researchers and the FDA.  Data sharing is important 
for further translation of research results into knowledge, products and procedures to improve 
human health.  All links with your identity will be removed from the data before it is shared. 
What are the risks and possible discomforts from being in this 
research study? 
Taking Posiphen daily may involve risks that are not known at this time due to the investigational 
nature of this study.  Some of these unknown risks may be life-threatening or result in death.  
However, you will be told of any new risks or significant findings that develop during the course of 
this study. 
You will be assigned to a study group at random (by chance).  Your assignment is based on chance 
rather than a medical decision made by the researchers.  The study group you are assigned to might 
not be the group you would prefer to be in.  Your assigned study group might also prove to be less 
effective or have more side effects than the other study group(s), or other treatments available for 
your condition. 


 
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Participation in this study may involve some added risks or discomforts, which are outlined below.  
Risks of Study Drug (Posiphen) 
To-date, a total of 101 individuals have taken Posiphen. The most common (occurring in 2 or more 
subjects) side effects reported after receiving Posiphen have included the following: 
• Dizziness 
• Headaches 
• Abnormal dreams 
• Nausea 
• Vomiting  
• Feeling warm 
• Heart rate increased 
• Constipation 
• Fainting 
• Orthostatic hypotension 
It is important to remember that Posiphen has been studied in a limited number of people.  
Therefore, Posiphen may involve other risks, including life threatening risks that are not known at 
present.  Because Posiphen is a drug that works on the nervous system, you should notify your study 
doctor immediately if you experience any changes in your thinking, behavior or mood.  
Risks of Drugs that Reduce β-Amyloid in the Brain 
Posiphen is designed to reduce β-amyloid protein in the brain. With some other drugs that reduce β-
amyloid protein cerebral vascular edema (swelling on the brain) was observed.   In particular, in a 
clinical study examining the ability of an experimental vaccine to reduce the amount of β-amyloid 
protein in the brain swelling was observed in 12 individuals.  The swelling in these patients was not 
permanent.  Brain swelling increases pressure inside the skull. This pressure can prevent blood from 
flowing to your brain, which deprives it of the oxygen it needs to function. Swelling can also block 
other fluids from leaving your brain, making the swelling even worse. Damage or death of brain cells 
may result.  To date brain swelling has not been observed after taking Posiphen. 
Symptoms of brain swelling vary, depending on the severity and the cause. Usually they begin 
suddenly. You may notice any of these symptoms: 
• Headache  
• Neck pain or stiffness 
• Nausea or vomiting 
• Dizziness  
• Irregular breathing 
• Vision loss or changes 
• Memory loss  
• Inability to walk 


 
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• Difficulty speaking 
• Stupor 
• Seizures  
• Loss of consciousness 
In the event you experience any of these symptoms, please contact your doctor and [SITE 
INVESTIGATOR NAME] immediately.  
Risks of Lumbar Punctures  
In total, up to 142 mL - approximately 9 tablespoons of CSF may be taken during the entire study.  
Your body will make up for this loss.  
During the lumbar puncture procedure, you may have temporary pain and discomfort in your back.  
Headache may occur in about 5% of people who undergo a lumbar puncture.  Less commonly, in 
about 1 - 4% of participants, is a persistent low-pressure headache that may develop, probably due to 
leakage of CSF.  If the headache persists it may require additional treatment.   
Uncommonly, a blood patch (injection of some of your blood into the lumbar puncture site to patch 
the CSF leak) may be required and should relieve the headache immediately.  
Although very rare, it is possible that you may have an allergic reaction to the local anesthetic 
(lidocaine 1%) used for the lumbar puncture.  An allergic reaction could cause swelling and a rash on 
your skin where the anesthetic was injected.  Please tell us if you have ever had a reaction to local 
anesthetic before (such as when you were visiting the dentist).  
Potential but rare risks of lumbar puncture include infection, damage to nerves in your back and 
bleeding into the CSF space.  The risk of these is much less than 1%.  To minimize these risks, the 
lumbar puncture will be performed by Dr. [INSERT NAME OF DOCTOR WHO WILL BE PERFORMING LP 
HERE] or by a neurologist specifically trained in the procedure.  
Risks of CSF Sampling 
The risks associated with continuous CSF sampling are similar to those associated with a single 
lumbar spinal tap. 
The most common side effects of this procedure are headache and backache.  If a headache occurs, it 
is generally relieved by drinking fluids and lying down.  If necessary, you may receive a pain reliever 
such as Acetaminophen or one or more doses of Caffeine through your IV line.  
Headaches can recur for several days, although on rare occasion headaches can last longer - up to a 
couple of weeks. 
If you experience a persistent headache, it is possible that a blood patch procedure will be 
performed.  The rate of post–lumbar puncture headache requiring a blood patch is about 10% in 
older individuals. This involves the injection of a small amount of your own blood into the region of 
the original tap but not into the CSF fluid space.  The blood-patch is performed by an anesthesiologist 
and has been effective in relieving headache in up to about 95% of cases.   


 
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In rare cases, this procedure may need to be repeated.  This may be done under X-ray guidance at the 
discretion of the anesthesiologist to guide the injection of blood.  Should you experience a headache 
during the period after the lumbar spinal tap, you must notify the study team. 
Most people experience only the minor discomfort of a needle prick when local anesthetic is 
administered.  Others experience a few moments of mild to moderate pain similar to that 
experienced when an injection is received.  A hypersensitivity or allergic reaction to the local 
anesthetic may occur.  The collection of CSF is usually painless.   
Another discomfort that can occur following a lumbar spinal tap is a brief pain or tingling sensation in 
either leg, hip, buttock or groin area.  This is caused by a brief stimulation of a nerve and ends quickly 
with no further complications. 
This discomfort may also occur once the spinal catheter is in place and is reversible with change in 
position, adjusting the catheter or removing the catheter. 
On rare occasions, a temporary weakness of the eye muscle that moves the eye from side to side may 
develop, producing double vision.  This complication has been temporary, and normal vision has been 
restored. 
Other complications are rare and include nerve root damage, epidural (the area outside the covering 
of the spine) or subdural (spinal) bleeding, infection, paralysis or death. 
All precautions will be taken to minimize these risks.  Subjects will be monitored for associated 
symptoms of infection: neck stiffness, headache, increased irritability, nausea, light sensitivity and 
fever following the procedure.  
Risks of Blood Draws & IV Line Insertions 
Drawing blood from a vein in your arm and inserting an intravenous (IV) line into your arm poses a 
small risk of infection, temporary pain, blood clot or bruising at the site of the needle stick.  Some 
people may feel light-headed, or experience upset stomach or fainting when their blood is drawn. 
To minimize these risks, experienced medical personnel will handle all the blood drawing and IV line 
insertion procedures and sterile conditions will be maintained.  
In total, 215 mL – approximately 14 tablespoons of blood - may be taken during this study.  Your body 
will make up for this loss. 
Risks of MRI Scans 
An MRI may cause possible discomfort for people due to the loud banging made by the machine and 
the confined space of the testing area.  There is also a risk of injury if metal is brought into the 
imaging room, which might be pulled into the MRI magnet.  
Risks of ECGs 
There is no pain or discomfort during an ECG; however, removing the pads may cause some irritation 
to your skin. 


 
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Risks of Reproduction 
It is not known if the study drug may affect an unborn or nursing baby.  If you are pregnant or 
breastfeeding, you cannot be in this study. 
Women who are capable of having children are excluded from participation in this study.  If you are a 
woman, you must either 1) be two years post-menopausal or 2) have had a documented surgical 
sterilization procedure.   
The effects of Posiphen on the developing fetus are unknown and there is a possibility it may cause 
physical or mental damage.  There may be risks to an unborn baby you father during or after the 
study.  
Avoiding sexual activity is the only certain method to prevent pregnancy.  However, if you choose to 
be sexually active and are not surgically sterile, you must agree to use a barrier method of birth 
control (such as a condom with spermicide) from the time you start taking the study drug until 30 
days after you receive the last dose of Posiphen. 
If you (or your partner) becomes pregnant during this time period, you must notify the study doctor 
or study staff immediately.  The study doctor may ask for information about the pregnancy and the 
birth of the baby.  The study doctor may share this information with the Sponsor or other parties 
listed in the section about confidentiality. 
If you are not willing to use acceptable birth control, you should not participate in this study or sign 
this consent form. 
Risks of Genetic Testing 
Variation in some genes is known to be directly related to risk of certain illnesses.  Under some 
circumstances, it can be a risk for genetic information to be known by others.  In some cases, 
knowledge of genetic information could have negative psychological consequences or could affect 
access to or retention of certain benefits or entitlements.  For example, the information could 
potentially be used against subjects if it were revealed to insurance companies or potential 
employers.   
A U.S. Federal law called the “Genetic Information Nondiscrimination Act” (GINA) generally makes it 
illegal for health insurance companies, group health plans and most employers to discriminate 
against you based on your genetic information.  This law generally will protect you in the following 
ways:  
• Health insurance companies and group health plans may not request your genetic information 
that we get from this research.  
• Health insurance companies and group health plans may not use your genetic information 
when making decisions regarding your eligibility or premiums.  
• Employers with 15 or more employees may not use your genetic information that we get from 
this research when making a decision to hire, promote or fire you or when setting the terms 
of your employment.  


 
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Be aware that this new Federal law does not protect you against genetic discrimination by companies 
that sell life insurance, disability insurance or long-term care insurance. 
You will not get the results of the genetic portion of the study nor will the results be made available in 
your medical record.   
Furthermore, the researchers have adopted strict privacy and confidentiality procedures for 
maintaining your genetic information as described in this consent form. 
Risks of Loss of Confidentiality 
Study personnel will not release your study information to your doctor without your permission, and 
will code your study information to protect your confidentiality. 
However, it is possible that information about your participation in this study could enter your 
medical record, particularly if you experience an adverse event requiring treatment.  
It is possible that information about your amyloid status or participation in this study could influence 
your ability to obtain life insurance, health insurance, or long-term care insurance or affect your 
future employability.   
To help us protect your privacy, a Certificate of Confidentiality (Certificate) for this study has been 
obtained from the National Institutes of Health. The researchers can use this Certificate to legally 
refuse to disclose information that may identify you in any federal, state, or local civil, criminal, 
administrative, legislative, or other proceedings, for example, if there is a court subpoena. The 
researchers will use the Certificate to resist any demands for information that would identify you, 
except as explained below. 
The Certificate cannot be used to resist a demand for information from personnel of the United 
States Government that is used for auditing or evaluation of Federally-funded projects or for 
information that must be disclosed in order to meet the requirements of the federal Food and Drug 
Administration (FDA).   
You should understand that a Certificate of Confidentiality does not prevent you or a member of your 
family from voluntarily releasing information about yourself or your involvement in this research. If 
an insurer, employer, or other person obtains your written consent to receive research information, 
then the researchers may not use the Certificate to withhold that information. 
The Certificate of Confidentiality will not be used to prevent disclosure to state or local authorities of 
[INSERT STATE]. 
Risks of Placebo  
 
If you receive placebo as your study drug, you may experience memory problems consistent with AD 
and aging.   
Risks of Testing & Questionnaires 
Repeated evaluations of mood and mental status may be slightly frustrating or produce fatigue and 
boredom. 


 
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Unknown Risks & New Information 
In addition to the side effects already described, the study drug and the study procedures may have 
other unknown risks.  As with any new medicine, there is a risk of rare or previously unknown side 
effects, and/or a chance that Posiphen might interact with other medicines.  You must tell your study 
doctor if you experience any side effects.   
As with any drug, you may experience an unexpected allergic reaction that could be life-threatening.  
Symptoms of an allergic reaction can include, but are not limited to: 
• Rash  
• Having a hard time breathing 
• Wheezing when you breathe 
• Sudden drop in blood pressure 
• Swelling around the mouth, throat or eyes 
• Fast pulse 
• Sweating 
If you suffer a serious or lasting injury because of your participation in this study, it may affect your 
ability to obtain private health insurance, your employability and/or your quality of life. 
If you should experience a skin reaction during your participation in the study, whether or not the 
reaction causes you discomfort or you believe the reaction is serious, a photograph of the area where 
the reaction appears may need to be promptly taken for evaluation and assessment purposes, as 
determined by the study doctor.  These photographs, if taken, will remain in confidential files and 
your identity will not be revealed. 
New information about Posiphen may become available during the study.  If this happens, your study 
doctor will tell you about the new information, and [he/she] will talk to you about whether you want 
to remain in the study.     
What are the benefits of taking part in this study?  
Participation in this study may help to improve your condition, but it is also possible that your 
condition may not improve or could worsen.  There is no guarantee that you will personally benefit by 
participating in this research study. 
Your participation in this study may help the investigators learn more about the safety and benefits of 
Posiphen for Alzheimer’s disease to help other people who have a similar medical problem in the 
future. 
What other choices do I have if I do not take part in this study?  
If you are currently experiencing memory problems, there are 5 drugs approved for the treatment of 
Alzheimer's disease: 
• Donepezil (Aricept®) 


 
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• Tacrine (Cognex®) 
• Galantamine (Razadyne®) 
• Rivastigmine (Exelon®) 
• Memantine (Namenda™) 
• Aducanumab (Aduhelm™) 
Thus, alternatives to participation in this study include the possible use of one of these approved 
medications.  
Because study results might be affected, participation in any other clinical studies will not be 
permitted until your participation in this study has ended. 
Another alternative is not to participate in this study and to continue under standard medical care, 
which may include receiving some of the above listed medications for memory problems and AD. 
Will my medical information be kept private/confidential?  
Research records will be kept as confidential as possible within the limitations of state and federal 
law.  The study staff and Sponsors will handle your personal health information in a confidential 
manner. 
Federal Privacy Regulations require that you authorize the release of any health information that 
may reveal your identity.    
For this research study, this includes information in your existing medical records needed for this 
study and new information created or collected during the study. 
The persons and entities that you are authorizing to use or disclose your personal health 
information may include the: 
• Study doctor 
• Study staff 
• Institution 
• Institution’s IRB 
• Alzheimer’s Disease Cooperative Study (ADCS)  
• Other ADCS research sites participating in this study and 
• Laboratories used for this study  
In order to analyze the data collected during this research study, all of the health information 
generated or collected about you during this study may be inspected by the study Sponsor and its 
authorized agents, the Food and Drug Administration (FDA), the Department of Health and Human 
Services (DHHS) agencies and the Institutional Review Board (IRB).   
Because of the need to release information to these parties, absolute confidentiality cannot be 
guaranteed. 


 
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Once your personal health information is released it may be re-disclosed, at which point your 
health information will no longer be protected by federal privacy regulations.  
The findings of this research will be presented at meetings or in publications; however, neither your 
name nor identity will be disclosed in those presentations.   
You have the right to review and copy the health information collected about you, however, you will 
not be allowed to look at your study-related information until after the research is completed.  
This authorization will have no expiration.  
What are the costs of taking part in this research study?  
There will be no costs to you for participation in this study. 
Procedures related to the study and study drug will be provided at no cost to you.   
Will I be paid for taking part in this research study?  
You may receive up to [INSERT AMOUNT] for participation in this the study.  This money covers the 
costs for time spent at the clinic and travel expenses to and from the clinic. 
If you choose to leave or are withdrawn by the study staff before finishing all study procedures, you 
will be paid a lesser amount that is based on the completed visits or procedures. You will also receive 
a hand-crafted quilt donated by the Quilt Guild for study participation.  
No other payment will be offered to you.  You will receive your payment within 2 weeks of your final 
visit. 
 
What happens if I am injured as a result of taking part in this 
research study?   
All forms of medical findings and treatments – whether routine or experimental – involve some risk of 
injury.  In spite of all safety measures, you might develop medical problems from participating in this 
study. 
You must report any suspected illness or injury to the study doctor immediately.  If such problems 
take place, the [SITE INVESTIGATOR’S INSTITUTION] will provide emergency medical treatment and 
will assist you in getting proper follow-up medical treatment. 
If you require medical treatment for an untoward event, the company that manufactures the drug 
( Annovis Bio, Inc.) in consultation with the study doctor, will pay the reasonable and customary costs 
of such treatment only if it is determined that the untoward event occurred as a result of 
manufacture of the study drug and the costs of treatment are not covered by any other health 
insurance, government health program or other third party providing coverage for health care. 


 
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Neither financial compensation nor reimbursement for such things as pre-existing conditions, illness 
or disease unrelated to study drug, lost wages, property damage, disability, or discomfort is available.   
The National Institute on Aging and the Alzheimer’s Disease Cooperative Study do not provide 
compensation for any other type of research-related injury related to participation in this study.  IN 
ADDITION TO THIS STATEMENT, ADD YOUR INSTITUTION’S SUBJECT INJURY CLAUSE HERE (you will / 
will not pay for subject injury, etc). By signing this consent form you do not give up any of your legal 
rights. 
If I have questions or concerns about this research, whom can I 
contact?   
If you have any questions regarding this research or if you believe that you may have experienced a 
research related injury or a reaction to the study drug, you should contact Dr. [Insert Site PI Name] 
(study doctor) at [TELEPHONE].  
                             OFFICE], at [TELEPHONE] for more information about this or to report research-
related problems. 
Is participation in this study voluntary? 
Your participation in this research study is entirely voluntary.  You have the right to refuse to 
participate, or may discontinue participation in this study at any time without jeopardy to the medical 
care you receive at this institution. 
You have the right to not sign this form that allows us to use and share your health information for 
research; however, if you don’t sign it, you cannot take part in this research study.  
You have the right to withdraw your permission to use or disclose personal information about your 
health.  If you choose to withdraw your permission, you must notify Dr. [Insert Site PI Name] in 
writing.  Dr. [Insert Site PI Name]’s mailing address is: 
(Insert Address) 
 
Dr. [Insert Site PI Name] will still be able to use the information collected about you prior to your 
withdrawal from the study.  Information that has already been sent to the Sponsor cannot be 
withdrawn. 
 
 
 
 
 


[INSERT LOCAL SITE NAME] 
 
 
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STATEMENT OF CONSENT 
You have read (or have had read to you) the above description of this research study.  You have been 
informed of the risks and benefits involved, and all of your questions have been answered to your 
satisfaction.  By signing this form, you voluntarily consent to participate in the research study and you 
authorize the use of your bodily fluid samples for the research described above. 
Unless you authorize the use and disclosure of your personal health information, you cannot 
participate in this research study.  If you refuse to give your authorization, your medical care will not 
be affected. 
You will receive a copy of this consent form. 
  
Yes  No  
Subject 
Initials 
 
 You voluntarily agree to participate. 
☐ 
 
☐ 
 
 
 
  
 
 
 
 
 
 
  
 
 
 
 
 
 
 
1BSample Storage 
 
 
 
 
 
 
 You agree that your DNA samples may be stored and used for future research. 
☐ 
 
☐ 
 
 
 
 (Optional) 
 
 
 
 
 
 
 You agree that your Blood samples may be stored and used for future research. 
☐ 
 
☐ 
 
 
 
 (Not Optional) 
 
 
 
 
 
 
 You agree that your CSF samples may be stored and used for future research. 
☐ 
 
☐ 
 
 
 
 (Not Optional) 
 
 
 
 
 
 
 
____________________________ 
Study Participant Name (print) 
 
 
_____________________________ 
Signature 
 
 
__________________ 
Date 
 
 
 
 
 
 
 
 
 
 
Person Obtaining Consent (print) 
 
Signature 
 
Date 
 
 
 
 
 
 
 
 
 
 
Legal Representative / Next of Kin  
If applicable (i.e. spouse, child, 
sister) (print) 
 
Signature 
 
Date 
 
 
 
 
 
 
 
 
 
 
Witness Name 
(print) 
 
Signature 
 
Date 


[INSERT LOCAL SITE NAME] 
 
 
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STUDY PARTNER INFORMATION & CONSENT 
As the participant’s study partner, you have important tasks that need to be carried out in order for the 
study to be conducted in the safest and best manner possible.  These responsibilities include: 
• You must have regular contact with the participant a minimum of 10 hours per week and 
provide supervision of drug administration as needed. 
• You must attend the entire screening and baseline visits. You should also stay for the first three 
hours of the confinement visit and return to drive the participant home.   
• You are an important source of information about the participant.  You must agree to be asked 
questions about your relationship with the participant, as well as their health, memory, daily 
functioning and behavior in order to find out whether there are any changes in the participant.  
• You must be able to make sure that the study participant is taking the study drug as directed.  
If for some reason you become unable to carry out your responsibilities, please tell the study team 
immediately.  You may be asked, if possible, to select a substitute who can take over your duties. 
You have read all the preceding information which describes both the participant’s participation in the 
study and your involvement as the participant’s study partner.  The study has been explained to you in 
detail.  All your questions have been answered to your satisfaction.   
 
  
Yes 
 
No 
 
Study 
Partner’s 
Initials 
 
 You voluntarily agree to participate as a Study Partner. 
☐ 
 
☐ 
 
 
 
  
 
 
 
 
 
 
 
 
 
 
 
 
Study Partner’s Name 
(print) 
 
Signature 
 
Date 
 
 
 
 
 
 
 
 
 
 
 
Person Obtaining Consent (print) 
 
Signature 
 
Date 
 
 
 


[INSERT LOCAL SITE NAME] 
 
 
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California Site Only: Insert the Experimental Bill of Rights 
 

Prot_000.pdf: 
 
 
 
 
 
 
 
 
 
HOWARD FELDMAN, MDCM, FRCP(C)  
PROFESSOR, DEPARTMENT OF NEUROSCIENCES 
DIRECTOR, ALZHEIMER’S DISEASE COOPERATIVE STUDY (ADCS) 
DEAN, ALZHEIMER’S DISEASE AND NEURODEGENERATIVE RESEARCH 
9500 Gilman Drive     La Jolla, California 92037-0949     TEL: (858) 246-1347   FAX: (858) 246-1414 
 
 
 
Protocol Title: 
 
 A Multicenter, Randomized, Double-Blind, Placebo-Controlled, 
Ascending Dose Study to Evaluate the Safety, Tolerability, 
Pharmacokinetics (PK) and Pharmacodynamic (PD) Effects of Posiphen® 
in Subjects with Early Alzheimer’s Disease (AD)  
 
Protocol Short Title: A Safety and PK/PD Study of Posiphen in Subjects 
with Early Alzheimer’s Disease 
 
  
Protocol Number: ADC-043-DISC (QR15001)  
 
 
NCT02925650 
 
 
Protocol Version 8.0 
 (23NOV2021) 
 
 
 
 
 


Protocol Number: ADC-043 DISC (QR15001) 
 
          Page 1 of 58 
 
                                                                                                                                                                                                                                                                                      
                                                                                                                                                                                                                                                                                      
Protocol Title: A Multicenter, Randomized, Double-Blind, Placebo-Controlled, Ascending 
Dose Study to Evaluate the Safety, Tolerability, Pharmacokinetics (PK) and 
Pharmacodynamic (PD) Effects of Posiphen® in Subjects with Early Alzheimer’s Disease 
(AD) 
Protocol Short Title: A Safety and PK/PD Study of Posiphen in Subjects with Early 
Alzheimer’s Disease 
 
Protocol Number: ADC-043-DISC (QR15001) 
 
US IND Number: 72,654 
 
November 23, 2021, V 8 
 
 
 
 
 
 
 
Confidentiality Statement 
The information in this document is privileged or confidential and may not be disclosed unless 
such disclosure is required by applicable laws and regulations. It is the property of Annovis Bio, 
Inc. and ADCS, and should not be copied by or distributed to persons not involved in the study. 
Persons to whom the information is disclosed must be informed that the information is privileged 
or confidential and may not be further disclosed by them. These restrictions on disclosure will 
apply equally to all future information supplied to you, which is indicated as privileged or 
confidential. Any unauthorized review, use, disclosure, copying or distribution is strictly 
prohibited.  
 


Protocol Number: ADC-043 DISC (QR15001) 
 
          Page 2 of 58 
 
 
PROJECT DIRECTORS 
DRUG SUPPLIER 
 
Martin Farlow, M.D. 
Project Director 
Indiana University 
Department of Neurology 
355 W. 16th Street, Suite 4700 
Indianapolis, IN 46202 
 
mfarlow@iupui.edu  
Tel: (317) 963-7402 
 
Douglas R. Galasko, MD 
Project Director 
UC San Diego 
9500 Gilman Drive MC 0948 
La Jolla, CA 92093-0948  
 
dgalasko@ucsd.edu 
Tel: (858) 552-8585  
 
 
Maria L. Maccecchini, Ph.D. 
President & CEO 
Annovis Bio, Inc. 
1055 Westlakes Drive, Suite 300 
Berwyn, PA 19312 
 
maccecchini@annovisbio.com  
Tel: (610) 727-3710 
 
COORDINATION & DATA 
MANAGEMENT CENTER 
MEDICAL MONITOR 
 
Howard Feldman, MDCM, FRCP(C)  
Director 
UC San Diego 
Alzheimer’s Disease Cooperative Study 
9500 Gilman Drive MC 0949 
La Jolla, CA 92093-0949 
 
hhfeldman@ucsd.edu  
Tel: (858) 246-1347 
 
 
Gabriel Leger, M.D. 
Medical Monitor  
UC San Diego 
Alzheimer’s Disease Cooperative Study 
9500 Gilman Drive, MC 0949 
La Jolla, CA 92093-0949  
 
gleger@ucsd.edu  
Tel: (702) 833-0916 
PROTOCOL VERSION 
VERSION DATE 
Final Protocol 
  November 23, 2021 V8 
 
 
 


Protocol Number: ADC-043 DISC (QR15001) 
 
          Page 3 of 58 
 
LIST OF ABBREVIATIONS  
 
5’UTR  
5’ untranslated region 
αSYN 
Alpha-synuclein 
Aβ 
Beta-amyloid 
Aβ38 
Beta Amyloid 38 
Aβ40 
Beta Amyloid 40 
Aβ42 
Beta Amyloid 42 
AChE 
Acetylcholinesterase 
AD 
Alzheimer’s Disease 
ADAS-Cog12 
Alzheimer’s Disease Assessment Scale – Cognitive 
ADCS 
Alzheimer’s Disease Cooperative Study 
AE 
Adverse Event 
AIDS 
Acquired Immunodeficiency Syndrome 
APOE/APOE4 
Apolipoprotein E /Apolipoprotein epsilon 4  
APP  
Amyloid Precursor Protein 
ANOVA 
Analysis of variance 
AUC 
Area under the curve 
BACE 
β-secretase 
BChE  
Butyrylcholinesterase 
BDNF 
Brain-derived neurotrophic factor 
BID 
Twice a Day 
BP 
Blood pressure 
CDR 
Clinical Dementia Rating  
CFR 
Code of Federal Regulations 
CTCAE 
Common Terminology Criteria for Adverse Events 
CLBP 
Chronic low back pain 
Cmax 
Maximum plasma and CSF concentration  
ChE 
Cholinesterase 
CJD 
Creutzfeldt-Jakob Disease 
C-SSRS 
Columbia Suicide Severity Rating Scale 
CNS 
Central Nervous System 
CSF 
Cerebrospinal Fluid 
CV 
Coefficients of variation 
CVD 
Cerebrovascular Dementia 
DNA 
Deoxyribonucleic Acid 
DSMB 
Data Safety Monitoring Board 
DSM 
Diagnostic and Statistical Manual of Mental Disorders 
ECG 
Electrocardiogram 
eCRF 
Electronic Case Report Form 
FCR  
Fractional clearance rate 
FDA 
Food and Drug Administration 
FSR 
Fractional Synthesis Rate 
FWA 
Federalwide Assurance 
GCP 
Good Clinical Practice 
GI 
Gastrointestinal 
GRAS  
Generally recognized as safe  
GWAS 
Genome Wide Association Studies 
HEK 293 
Human embryonic kidney cells 


Protocol Number: ADC-043 DISC (QR15001) 
 
          Page 4 of 58 
 
HEENT 
Head/ears/eyes/nose/throat 
hERG 
Human ether-a-go-go related gene  
HIPAA 
Health Insurance Portability and Accountability Act 
ICH 
International Conference on Harmonisation 
IRB 
Institutional Review Board 
IRE 
Iron-responsible element 
IRP1 
Iron Regulatory Protein 1 
LBD 
Lewy Bodies Dementia 
LP 
Lumbar Puncture 
MAD 
Multiple Ascending Dose 
MCI 
Mild Cognitive Impairment 
MMSE 
Mini-Mental State Examination 
MRI 
Magnetic Resonance Imaging 
mRNA 
Messenger Ribonucleic Acid 
NIA 
National Institute on Aging, under the NIH 
NIA-AA 
NIA and Alzheimer’s Association 
NIH 
National Institutes of Health 
NOAEL 
No observed adverse effect level 
NPH 
Normal Pressure Hydrocephalus 
NPI 
Neuropsychiatric Inventory 
OHRP 
Office for Human Research Protections 
p-tau  
Phosphorylated tau 
PD 
Pharmacodynamic 
PHI 
Protected Health Information 
PI 
Principal Investigator 
PK 
Pharmacokinetic 
PSP 
Progressive Supranuclear Palsy 
QD 
Once a Day 
QID 
4 Times a Day 
QT 
Interval seen in electrocardiogram (ECG) test 
QTc  
Interval seen in QT (ECG) test 
sAPP 
Soluble Amyloid Precursor Protein 
sAPPα 
Soluble Amyloid Precursor Protein alpha 
sAPPβ 
Soluble Amyloid Precursor Protein beta 
SAD 
Single Ascending Dose 
SAE 
Serious Adverse Event 
SH-SY-5Y 
Human Derived Cell Line 
SILK™ 
Stable Isotope Labeling and Kinetics 
SD 
Standard Deviation 
t1/2 
Half-life 
tau 
Tau protein 
TID 
Three Times a Day 
 
 
 
 


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SIGNATURE PAGE 
(SIGNATURES ON FILE AT THE COORDINATING CENTER) 
 
 
_______________________________________ 
 
___________________ 
Maria L. Maccecchini, Ph.D.  
 
 
 
Date (DDMMMYYYY) 
President & CEO 
Annovis Bio, Inc.  
 
_______________________________________ 
 
___________________ 
Gabriel Leger, M.D. 
 
 
 
                        Date (DDMMMYYYY) 
Medical Monitor 
UC San Diego 
Alzheimer's Disease Cooperative Study (ADCS) 
 
_______________________________________ 
 
___________________  
Martin Farlow, M.D. 
 
 
 
 
 
Date (DDMMMYYYY) 
Project Director 
Indiana University 
 
_______________________________________ 
 
___________________ 
Douglas Galasko, M.D. 
 
 
                       Date (DDMMMYYYY) 
Project Director 
UC San Diego 
 
_______________________________________ 
 
___________________ 
Howard Feldman, MDCM, FRCP(C)                                      Date (DDMMMYYYY) 
Director, ADCS 
Coordination & Data Management Center 
UC San Diego 
ADCS 
 
 
 


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1 
PROTOCOL SYNOPSIS .................................................................................... 10 
2 
INTRODUCTION .............................................................................................. 122 
2.1 
Primary Aims ................................................................................................... 12 
2.2 
Secondary Aims ............................................................................................ 133 
3 
PRELIMINARY STUDIES .................................................................................. 13 
3.1 
Summary of Non-Clinical Findings .................................................................. 13 
3.2 
Summary of Clinical Findings .......................................................................... 14 
4 
BACKGROUND AND SIGNIFICANCE .............................................................. 15 
5 
POTENTIAL RISKS AND BENEFITS OF INVESTIGATIONAL PRODUCT AND 
STUDY PROCEDURES ..................................................................................... 18 
5.1 
Risks and Benefits Associated with Posiphen or Placebo............................... 18 
5.2 
Risk/Benefit Associated with Blood Collections .............................................. 19 
5.3 
Risk/Benefit Associated with Stable Isotope Labeling Kinetics (SILK™) 
Procedure and Cerebrospinal Fluid (CSF) Sampling ...................................... 20 
    5.4 
Risk/Benefit Associated with Investigational Drugs Intended to Reduce β-
amyloid in the Brain ........................................................................................ 20 
6 
SAMPLE SIZE AND STATISTICAL PLAN ........................................................ 20 
6.1 
Randomization ................................................................................................ 20 
6.2 
Power and Sample Size Determination .......................................................... 21 
6.3 
Safety and Tolerability Analysis ...................................................................... 21 
6.4 
PK Analysis ..................................................................................................... 22 
6.5 
SILK™ Analysis .............................................................................................. 22 
6.6 
Criteria for the termination of the trial .............................................................. 23 
7 
STUDY DRUG AND CONCOMITANT MEDICATIONS ..................................... 23 
7.1 
Name and Description of IP and Comparator ................................................. 23 
7.2 
Dosage............................................................................................................ 23 
7.3 
Packaging/Dispensing/Labeling ...................................................................... 23 
7.4 
Storage ........................................................................................................... 24 


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7.5 
Drug Accountability ......................................................................................... 24 
7.6 
Compliance ..................................................................................................... 24 
7.7 
Breaking the Blind ........................................................................................... 25 
7.8 
Overdose ........................................................................................................ 25 
7.9 
Concomitant Medications ................................................................................ 25 
7.9.1 
Prohibited Concomitant Medications........................................................ 25 
7.9.2 
Permitted Concomitant Medications ........................................................ 26 
8 
STUDY POPULATION ....................................................................................... 26 
8.1 
Inclusion Criteria ............................................................................................. 26 
8.2 
Exclusion Criteria ............................................................................................ 28 
9 
DESCRIPTION OF STUDY VISITS .................................................................... 29 
9.1 
Pre-screening .................................................................................................. 29 
9.2 
Screening Visit ................................................................................................ 29 
9.2.1 
Re-screens .............................................................................................. 30 
9.3 
Baseline Visit .................................................................................................. 30 
9.4 
Day 14 Telephone Contact ............................................................................. 30 
9.5 
Confinement Visits .......................................................................................... 30 
9.5.1 
Pre-Confinement Visit .............................................................................. 30 
9.5.2 
Confinement Visit ..................................................................................... 30 
9.5.3 
Post-Confinement 24-Hour Phone Follow-up .......................................... 33 
10 
EARLY TREATMENT/STUDY DISCONTINUATION ......................................... 33 
10.1 
Reasons for Early Discontinuation .................................................................. 33 
11 
STUDY-SPECIFIC INSTRUMENTS ................................................................... 34 
11.1 Cognitive and Clinical Evaluations ..................................................................... 34 
11.1.1 
Alzheimer’s Disease Assessment Scale-Cognitive (ADAS-Cog12) ......... 34 
11.1.2 
Columbia Suicide Severity Rating Scale (C-SSRS) ................................. 34 


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11.1.3 
Logical Memory Test (Wechsler Memory Scale-Revised) ....................... 35 
11.1.4 
Mini-Mental State Examinations (MMSE) ................................................ 35 
11.1.5 
Neuropsychiatric Inventory (NPI) ............................................................. 35 
11.1.6 
Clinical Dementia Rating (CDR) .............................................................. 35 
12 
STUDY-SPECIFIC PROCEDURES ................................................................... 36 
12.1 
Safety Assessments ....................................................................................... 36 
12.1.1 
Physical and Neurological Examination ................................................... 36 
12.1.2 
Electrocardiogram (ECG) ......................................................................... 36 
12.1.3 
Clinical Laboratory Evaluations ................................................................ 36 
12.2 
Biofluids .......................................................................................................... 37 
12.2.1 
CSF for Biomarkers ................................................................................. 37 
12.2.2 
Apolipoprotein (APOE) Genotype and other Deoxyribonucleic Acid (DNA) 
Markers .................................................................................................... 38 
12.2.3 
Plasma for Biomarkers and Pharmacokinetics ........................................ 39 
12.2.4 
DNA/Genome Wide Association Studies (GWAS)/APOE Sample ........... 39 
13 
PERSONNEL REQUIREMENTS ....................................................................... 39 
14 
ADVERSE EVENTS (AES) ................................................................................ 40 
14.1 
Definition ......................................................................................................... 40 
14.2 
Following Up on Adverse Events .................................................................... 40 
15 
SERIOUS ADVERSE EVENTS (SAE) ............................................................... 40 
15.1 
Definition ......................................................................................................... 40 
15.2 
Reporting SAEs .............................................................................................. 40 
16 
DATA AND SAFETY MONITORING BOARD (DSMB)...................................... 40 
17 
RECORDING AND COLLECTION OF DATA .................................................... 41 
17.1 
Case Report Form .......................................................................................... 41 
17.2 
Study Files and Source Documents ................................................................ 41 
18 
ETHICS AND REGULATORY CONSIDERATIONS .......................................... 42 


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18.1 
Good Clinical Practice ..................................................................................... 42 
18.2 
Institutional Review Board (IRB) ..................................................................... 42 
18.3 
Informed Consent and Health Insurance Portability and Accountability Act 
(HIPAA) Compliance ....................................................................................... 43 
18.4 
Genetic Research and Storage of Genetic Material ........................................ 44 
18.5 
Storage of Biospecimen Samples ................................................................... 44 
19 
STUDY MONITORING ....................................................................................... 44 
20 
AUDIT ................................................................................................................ 45 
21 
RECORD RETENTION ...................................................................................... 45 
22 
PUBLICATION POLICY ..................................................................................... 45 
23 
SHARING OF FINAL RESEARCH DATA.......................................................... 45 
24 
LITERATURE CITED ......................................................................................... 47 
25 
APPENDICES .................................................................................................... 49 
 
APPENDIX 1: SCHEDULE OF EVENTS ........................................................... 50 
 
APPENDIX 2: STUDY SCHEMATIC DIAGRAM ................................................ 50 
 
APPENDIX 3: CONFINEMENT VISIT DIAGRAM .............................................. 51 
 
Appendix 3.1 Confinement Visit Phase 1: Admission Diagram ....................... 51 
Appendix 3.2a Confinement Visit Phase 2: Sampling – 36-Hour Observation   
Diagram .......................................................................................................... 52 
Appendix 3.2b Confinement Visit Phase 2: Sampling – 36-Hour Observation 
Schedule of Events ......................................................................................... 53 
 
Appendix 3.3 Confinement Visit Phase 3: 12-Hour Observation Diagram ...... 54 
 
Appendix 3.4 Confinement Visit Phase 4: Discharge Diagram ....................... 55 
 
APPENDIX 4: SAMPLE LOW LEUCINE MENU ................................................ 57 
APPENDIX 5: SEVERITY EVALUATION OF CHOLINERGIC AES .................. 57 
 
 
 


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1 
PROTOCOL SYNOPSIS 
PROTOCOL TITLE 
A Multicenter, Randomized, Double-Blind, Placebo-Controlled 
Ascending Dose Study to Evaluate the Safety, Tolerability, PK and 
PD Effects of Posiphen® in Subjects with Early AD 
PROJECT DIRECTORS 
Martin Farlow, M.D. and Douglas Galasko, M.D. 
COORDINATING 
CENTER 
Alzheimer’s Disease Cooperative Study (ADCS) 
STUDY DESIGN 
A Phase 1b multicenter, randomized, double-blind, placebo-
controlled study evaluating safety, tolerability, PK and PD of 23-25 
days exposure to multiple ascending doses of Posiphen in up to 
24 participants with a clinical profile consistent with early AD 
including Mild Cognitive Impairment due to AD (MCI-AD) or Mild 
AD with the following planned doses: 
• 
60 mg Once a Day (QD) (1x60mg) 
• 
60 mg Twice a Day (BID) (2x60mg) 
• 
60 mg Three Times a Day (TID) (3x60mg) 
Each dose group will include 8 subjects, 5 randomly assigned to 
Posiphen and 3 randomly assigned to placebo. 
Each dose group will be completed and evaluated for safety and 
tolerability by the standing ADCS Data Safety Monitoring Board 
(DSMB).  The ADCS DSMB will determine whether to approve 
dose escalation.   
Subjects who do not tolerate study drug during the initial 3-week 
at-home treatment period will be discontinued and replaced by 
new subjects. If two subjects at a given dose are discontinued due 
to adverse events, no additional subjects will be added at this dose 
level until a safety review has been conducted. The safety 
committee will determine the appropriate dose level for additional 
subjects. The safety committee has the option to change the 
allocation of future patients to active drug versus placebo based 
on statistical advice to maintain an appropriate balance between 
placebo subjects and subjects on active drug. 
DURATION OF STUDY 
PARTICIPATION 
Each subject will have up to a 42-day screening period, followed 
by approximately 21-23 days of treatment at home, and an in-clinic 
visit that includes treatment and a 36-hour lumbar puncture (LP) 
Stable Isotope Labeling and Kinetics (SILK™) procedure. 
SUMMARY OF 
INVESTIGATIONAL 
PRODUCT 
Posiphen 60 mg QD, 60 mg BID or 60 mg TID, or placebo 
capsules taken orally for up to 25 days. 


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SUMMARY OF KEY 
ELIGIBILITY CRITERIA  
• 
Clinical profile consistent with early AD including MCI-AD or 
mild AD according to the core clinical criteria outlined in the 
National Institute Aging (NIA) and Alzheimer’s Association 
(NIA-AA) Guidelines 
• 
Cerebrospinal fluid (CSF) AD biomarkers consistent with AD 
(Beta Amyloid 42(Aβ42)/Beta Amyloid 40(Aβ40) ratio less than 
0.131 as measured by mass spectrometry) 
• 
Age 55-89  (inclusive), in good health with no frailty 
• 
Mini-Mental State Examination (MMSE) score between 17 – 30 
(inclusive) - 
• 
Clinical Dementia Rating (CDR) global score of 0.5 with a 
memory score of 0.5 or greater, or global score of 1.0  
 
PRIMARY OUTCOME 
MEASURE 
Aim 1. Safety and Tolerability 
Aim 2. PK of Posiphen/metabolites in Plasma and CSF 
Aim 3. PD on fractional synthesis rate (FSR) of Beta Amyloid 40 
(Aβ40) in CSF 
OTHER KEY OUTCOME 
MEASURES / STUDY 
PROCEDURES 
Aim 4. Feasibility of SILK™ in a multicenter trial 
Aim 5. PD on other biomarkers in CSF including Beta Amyloid 38 
(Aβ38), Aβ40, Aβ42, Soluble Amyloid Precursor Protein alpha 
(sAPPα), Soluble Amyloid Precursor Protein alpha (sAPPβ), and 
T-Tau     
Aim 6. Cognitive and/or neuropsychiatric effects of Posiphen 
 
 
 


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2 
INTRODUCTION 
The medications that are currently approved for AD target symptoms such as memory loss, but 
the magnitude of the effect is small, and the clinical benefits are limited. Aducanumab 
(commercially known as Aduhelm™), an Aβ-directed monoclonal antibody, received accelerated 
approval by the FDA for the treatment of early AD in June 2021.  
 
Posiphen® is a selective inhibitor of amyloid precursor protein (APP) production and has 
potential utility as a disease modifying treatment for AD (Cullen 2006; Utsuki 2006; Lahiri 2007). 
Posiphen was discovered at the National Institute on Aging and was selected from a series of 
structurally related compounds designed for APP specificity with no minimal or minimal 
acetylcholinesterase inhibitory activity. Posiphen was shown to reduce APP and consequently 
beta-amyloid (Aβ) production in relevant preclinical in vitro and in vivo studies.  
 
Posiphen, developed by Annovis Bio, Inc., is a small molecule that is thought to act by lowering 
soluble APP protein levels through a post-transcriptional mechanism. Iron increases the 
translation of APP by binding to an iron-responsive element (IRE) that forms a stem loop 
structure in the 5’ untranslated region (5’UTR) of APP Messenger Ribonucleic Acid (mRNA). 
Iron regulatory proteins bind to these stem loop structures and suppress the iron activation of 
mRNA translation. Iron Regulatory Protein 1 (IRP1) selectively binds the APP IRE with an 
affinity of 35 pM. When IRP1 is bound to the stem loop, it prevents the complex from binding to 
the ribosome and prevents translation of the mRNA. Addition of Posiphen to human derived cell 
line (SH-SY-5Y) neuroblastoma cells increases the affinity of IRP1 to the stem loop structure to 
15 pM and lowers the rate of translation of the mRNA by the ribosome (Venti 2004; Cahill 2009; 
Cho 2010; Rogers 2011). A study conducted to analyze the effect of orally administered 
Posiphen on Αβ levels in the mouse brain, revealed that Posiphen lowered Αβ levels at plasma 
concentrations that are attainable with oral dosing in humans (IB  2015). Three Phase 1 studies 
have been completed with Posiphen. The first study was a single ascending dose (SAD) study 
in healthy volunteers; the second study was a multiple ascending dose (MAD) study in healthy 
volunteers, and the third study was one with a PK/PD study of biomarkers in CSF obtained from 
mild cognitively impaired participants.  
 
The present study will confirm the pharmacokinetics of Posiphen and its metabolites in plasma 
and CSF. More importantly, it will expand on the biomarker reduction seen in the mechanism of 
action study (Maccecchini 2012) by measuring the effects of a 23-25 day treatment period with 
Posiphen on the CSF and plasma levels of a number of biomarkers, inflammatory factors and 
control proteins. It will also expand the safety data in humans by extending the treatment period 
from 10 days to a treatment period from 23-25 days. In addition, this study will measure 
concentrations of various soluble biomarkers in CSF and use the SILK™ assay methods to 
directly measure the effect of Posiphen on the fractional synthesis rate of Aβ40 in CSF, which 
will help guide the further development of Posiphen and determine the feasibility of SILK™ in a 
multicenter trial. 
2.1 
Primary Aims 
Aim 1: To determine the safety and tolerability of multiple ascending doses of Posiphen on 
subjects with early AD with up to 23-25 days of daily usage. 
Aim 2: To determine the PK of Posiphen and its metabolites in plasma and CSF of subjects with 
early AD treated with QD, BID and TID dosing regimens.  


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Aim 3: To assess the PD effects on the FSR of Aβ 40 using the SILK™ technique with multiple 
doses of Posiphen. 
2.2 
Secondary Aims 
Aim 4: To implement a multicenter lumbar CSF catheter study employing standardized 
methodology and SILK™ technology. The following criteria will be evaluated as outcomes to 
determine feasibility:  
• 
Enrollment comparisons of research subjects at each of the 3 sites 
• 
Adequacy of sample collections to permit analyses of PK and PD effects of Posiphen  
• 
Subject satisfaction questionnaires 
• 
Comparison of protocol deviations and variances in data between the sites 
Aim 5: To assess the PD effects of treatment with Posiphen versus placebo on the levels of the 
following biomarkers in CSF of subjects with early AD: 
• 
Aβ 38 and 40 
• 
sAPPα 
• 
sAPPβ 
• 
T-tau protein (T) 
Aim 6: To assess whether there are short-term cognitive and/or neuropsychiatric effects of 
Posiphen in subjects with early AD, either positive or negative.  
• 
ADAS-Cog12 
• 
MMSE 
• 
Neuropsychiatric Inventory (NPI) 
3 
PRELIMINARY STUDIES 
The following is a brief summary of the available information on Posiphen. Detailed information 
can be found in the Posiphen Clinical Investigational Brochure (2015). 
3.1 
Summary of Non-Clinical Findings 
In wild type mice, Posiphen lowered brain APP and Aβ levels by up to 60% (Lahiri 2007, 
Cullen 2006). In APP transgenic AD mice, Posiphen led to a decrease in APP levels, improved 
neuronal stem cell survival and increased levels of brain-derived neurotrophic factor (BDNF) 
(Marutle 2007; Kadir 2008, Lilja 2013). Chronic administration of Posiphen to APP transgenic 
mice totally prevented decline in memory and learning at plasma concentrations that are easily 
attainable with oral dosing in humans (Maccecchini, Sambamurti & Arancio in preparation). In 
alpha-synuclein (αSYN) transgenic PD mice, Posiphen restored gut motility to normal 
(Maccecchini, Kuo & Nussbaum in preparation). 
Posiphen was not mutagenic or clastogenic as assessed by in vitro assays.  


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The cardiac electrophysiological properties of Posiphen tartrate were negative in vitro using 
human ether-a-go-go related gene (hERG) transfected human embryonic kidney cells (HEK 
293). Posiphen did not adversely affect the interval seen in an electrocardiogram (ECG) test 
(QT (or QTc)) interval.  
Toxicity studies in dogs showed brain toxicity (ataxia and tremors/twitching) and gastrointestinal 
(GI) toxicity at 30 mg/kg/day, which was dose-dependent and reversible. The no observed 
adverse effect level (NOAEL) was 20 mg/kg/day in dogs. Posiphen readily crosses the blood-
brain barrier. The signs/symptoms noted at high doses of Posiphen may be related to 
cholinergic manifestations. In in vitro assays, Posiphen showed minimal inhibition of AChE or 
BChE (Butyrylcholinesterase) activity, however, a metabolic product of Posiphen, N1-
norposiphen demonstrated acetyl cholinesterase inhibitory activity (Yu 2013). There were no 
effects on the reproductive organs associated with 4-week exposure to Posiphen in male or 
female rats or dogs.  
3.2 
Summary of Clinical Findings  
Three Phase 1 studies have been conducted with Posiphen (Annovis Bio, Inc. 2019). The first 
was an SAD study in healthy volunteers; the second was an MAD study in healthy volunteers, 
and the third one was a PK/PD study of CSF obtained from mild cognitively impaired 
participants.  
In the multiple ascending dose trial, Posiphen was administered orally in doses of 20, 40, and 
60 mg 4 times a day (QID). The first two treatments were administered for 7 days, and the third, 
for 10 days. Single doses were given on the first and last day to determine the pharmacokinetics 
of the drug. In general, the drug was well tolerated, resulting in no serious or severe adverse 
events and only one premature discontinuation, a subject in the 60 mg group discharged 
because of nausea, vomiting, dizziness and “feeling warm.” The incidence of adverse events, all 
of either mild or moderate severity also occurred with similar frequency in the placebo group. 
The most common AEs were dizziness, headache, and nausea/vomiting.  
Posiphen was absorbed rapidly after oral administration, achieving maximum plasma and CSF 
concentration (Cmax) within 1.2 - 1.5 hours. For the 40 and 60 mg doses, with fully defined 
plasma profiles, the mean terminal t½’s were 3.80  ± 0.88 and 5.23 ± 1.24 hours, respectively 
after a single dose and 3.53 ± 1.03 and 4.104  ± 0.91 hours, respectively, after repeat dosing. 
The half-life (t½) of the plasma concentrations at the lower doses could not be calculated 
accurately. The Cmax increased disproportionally with dose (24, 144, and 2310 ng.h/mL after a 
single dose of 20, 40, and 60 mg, respectively and 110, 134, and 2101 ng.h/mL after multiple 
doses of 20, 40, and 60 mg, respectively). 
In the proof of mechanism of action study, the PK of Posiphen was measured after 10 days of 
administration (4x60 mg) over 12 hours in CSF and plasma of the AD participants. The PD of a 
number of biomarkers was compared for 12 hours before the first dose at day 0 and after the 
last dose at day 11 of Posiphen administration. We found that the plasma concentrations of 
Posiphen overlapped with the plasma concentrations found in the MAD study. In this study, the 
N1 metabolite reached about 10 to 15%, while the level of the N8 metabolite reached about 20 
to 25% of the Posiphen levels measured in plasma.  
Because a substantial proportion of the adverse events observed in AD participants treated with 
cholinesterase inhibitors appear to reflect the cholinomimetic properties of molecules in this 


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class, Posiphen’s highest tolerated dose is determined by the levels of the N1-norposiphen in 
blood and brain. 
While the t1/2 of Posiphen in plasma was 5 hours as seen in the SAD and MAD studies, the t1/2 
in CSF/brain was longer than 12 hours. Concentrations of Posiphen in the brain, extrapolated 
from blood and CSF, were 8 times higher than in plasma. 10 days of treatment with Posiphen 
normalized CSF levels of Soluble Amyloid Precursor Protein (sAPP) and tau, reduced αSYN 
and a series of inflammatory markers. The concentration and persistence of Posiphen in the 
brain suggest that much lower doses of drug administered once daily could achieve the desired 
pharmacological effect.  
4 
BACKGROUND AND SIGNIFICANCE 
A major pathological hallmark of AD is the appearance in the brain of senile plaques composed 
of aggregated forms of Aβ, which is a proteolytic product of APP. Both accumulation of Aβ and 
other pathological changes, such as development of neurofibrillary tangles, synaptic loss and 
brain atrophy, are thought to occur one or two decades prior to overt dementia. Soluble Aβ 
oligomers have been demonstrated to show toxicity in vitro and in vivo and represent a target 
for drug development in AD. Recent research has shown that soluble forms of phosphorylated 
tau (p-tau) and αSYN may also contribute to neuronal loss and that AD as well as Parkinson’s 
Disease carry a high load of inflammation and microglia activating factors that contribute to 
neurodegeneration. 
For small molecules that target Aβ protein metabolism or other key biochemical pathways in AD, 
studies using CSF and plasma sampling in humans can provide data to demonstrate that a drug 
crosses the blood-brain barrier and engages the relevant target, and to describe the relationship 
between blood levels and central nervous system (CNS) effects. Repeated sampling through a 
lumbar CSF catheter for up to 36 hours, and the additional technique of SILK™ to measure 
parameters such as fractional synthesis rate of Aβ, can help guide the development of drugs 
targeting Aβ.  
Posiphen is the (+) enantiomer of phenserine, an AChE inhibitor which had been tested in 
several AD studies without significant evidence of efficacy. Preliminary data shows that 
Posiphen is affected by food; however, when the drug is taken in a series there is no effect on 
the PK analyses results. However, the failure of those studies with phenserine may have been 
related to the formulation, which is significantly affected by food. While phenserine inhibits 
AChE, Posiphen has no AChE activity itself and develops some activity in vivo with the 
metabolism to N1-bisnorposiphen. In vivo phenserine is about 200 times more potent in regards 
to AChE activity than Posiphen, when metabolized to N1-bisnorposiphen. 
Posiphen has been found to significantly reduce soluble APP and Aβ as well as tau, p-tau and 
αSYN in the rodent brain and in human CSF. In preliminary studies in animals and humans, 
inhibition of APP, Aβ, tau, p-tau and αSYN occurs at levels 6 to 10 times lower than the levels 
causing cholinomimetic effects. 
Posiphen is thought to act at the 5’UTR of the APP mRNA and lowers APP and Aβ protein 
levels in animal models, and decreased sAPP levels in human CSF. Preliminary data suggests 
that these effects are achieved via the same mechanism of binding to the 5’UTR of mRNAs and 
inhibiting their translation. 


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As Posiphen is thought to inhibit the synthesis of APP, as well as other neurotoxic aggregating 
proteins, it might have a broader spectrum of activity and protect neurons and have a disease-
modifying effect in AD as well as other neurodegenerative disorders.  
CSF Sampling and SILK™ 
Sampling CSF repeatedly for as long as 36 hours through a lumbar catheter had been initially 
used in a few studies related to AD treatment during the 1990s (Cutler 1998). The technique 
has gained greater utility in relation to measuring pharmacological effects on Aβ with the 
addition of SILK™, a technique developed by Dr. R. Bateman and colleagues at Washington 
University (Bateman 2006). This technique involves placement of a lumbar CSF catheter, 
followed by intravenous infusion of 13C6-labeled “heavy” leucine for 9 hours. Leucine is an 
essential amino acid that undergoes active transport into the brain. 13C6 leucine is non-
radioactive, harmless to humans and the environment, and is physically identical to unlabeled 
(12C6) leucine except for an additional proton difference in the mass of the amino acids. The 
leucine is incorporated into a fraction of newly synthesized proteins throughout the body, 
including the brain, which are secreted into the CSF. Samples of CSF are collected and Aβ is 
immune-precipitated from the CSF, and the ratio of Aβ containing 13C6-leucine to that containing 
unlabeled leucine is measured using mass spectrometry. During hours 4-12 after start of the 
leucine infusion, the fraction of labeled Aβ in the lumbar CSF increases, reaching a maximum of 
about 10%. The slope (change in % labeled Aβ/time) is called the fractional synthesis rate 
(FSR) and is a measure of the rate of Aβ synthesis in the brain. During late stages of the 
catheter procedure (hours 24-36), the fractional % of Aβ that is labeled decreases linearly, and 
the slope is measured to calculate the fractional clearance rate (FCR). Using this method, 
Bateman and colleagues estimated that Aβ has a rapid FSR and FCR in CSF. 
As part of their CSF sampling studies, Bateman and colleagues measured CSF Aβ hourly for 36 
hours in healthy adults and participants with AD. They found that CSF Aβ in normal subjects 
fluctuate from hour to hour, in a circadian pattern, and to a much greater extent than in 
participants with AD (Huang 2012). This variability complicates the design of studies in which 
CSF is sampled in a serial manner through a single lumbar puncture before and after treatment 
from the same person.  
In this study 13C6-leucine will be infused during the CSF catheterization procedure. CSF 
samples will be collected for mass spectrometry analysis of Aβ40 and Aβ42 to see if treatment 
with Posiphen decreases CSF levels of Aβ40 and of sAPPβ measured initially by mass 
spectrometry.  
During the leucine infusion, participants must comply with a low leucine diet – approximately 
1500 mg or less of leucine per day (refer to Appendix 4, Sample Low Leucine Menu). The 
kitchen on site will be responsible for preparing low leucine meals and snacks while maintaining 
a food journal for each subject. Subjects who break the low leucine diet will not be excluded 
from the study, since this will not be evident until the leucine analyses are completed.  
At the end of the study, all unused leucine should be returned to C2N.  
Study population: The study will be conducted in participants with a clinical profile consistent 
with early AD including MCI due to AD and mild Alzheimer’s disease, consistent with the core 
clinical criteria outlined in the NIA-AA guidelines (2011), MMSE inclusion score between 17 – 30 
(inclusive), Clinical Dementia Rating (CDR) global score of 0.5 with a memory score of 0.5 or 


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greater, or global score of 1.0, and by the presence of a CSF Aβ42/ Aβ40 ratio less than 0.131 
consistent with AD (as measured by C2N at screening). All subjects will have a screening 
lumbar puncture to measure the CSF Aβ42/40 ratio. These biomarkers can support a diagnosis 
of AD. CSF will be stored to allow analysis of baseline (pre-treatment) levels of larger 
biomarkers. 
Dose selection: Posiphen will be tested at doses of 60 mg QID, 60 mg BID (120 mg total daily), 
and at 60 mg TID (180 mg total daily). Whereas the plasma t½ of Posiphen has been  
determined to be 5 hours, the CNS t½ is estimated to be over 12 hours, allowing that  once daily 
dosing may be feasible.  However, previous studies indicated that likely cholinergic adverse 
events are mostly driven by Cmax. In particular, in the SAD study significant nausea, vomiting 
and dizziness occurred at a single dose of 160mg. In the MAD study 40mg QD (=160 mg/day) 
was well tolerated. Thus, the same total daily dose is better tolerated if divided into several 
doses. The use of multiple doses compared to once a day in this study will help evaluate the 
PK/PD relationship in the context of tolerability of different administration schemes.  
Study Design: This study is designed as a Phase 1b multicenter, randomized, double-blind, 
placebo-controlled study evaluating safety, tolerability, PK and PD of 23-25 days exposure to 
multiple ascending doses of Posiphen in up to 24 participants with a clinical profile consistent 
with early AD including MCI-AD or Mild AD with the following planned doses: 
• 
60 mg QD (1x60mg) 
• 
60 mg BID (2x60mg) 
• 
60 mg TID (3x60mg) 
Each dose group will include 8 subjects, 5 randomly assigned to Posiphen and 3 randomly 
assigned to placebo. 
There is some risk of excess cholinergic adverse events given that some subjects will be 
included who are on cholinesterase inhibitor treatment for Alzheimer’s disease which may 
become additive with the first metabolite of Posiphen (N1-norPosiphen) that has 
anticholinesterase properties. 
Therefore, each dose group will be completed and evaluated for safety and tolerability by the 
standing ADCS DSMB before the next dose group is initiated.  
Subjects who do not tolerate study drug during the initial 3-week at home treatment period will 
be discontinued and replaced by new subjects. If two subjects at a given dose are discontinued 
due to adverse events, no additional subjects will be added at this dose level until a safety 
review has been conducted. The safety committee will determine the appropriate dose level for 
additional subjects. The safety committee has the option to change the allocation of future 
patients to active drug versus placebo based on statistical advice to maintain an appropriate 
balance between placebo subjects and subjects on active drug. 
Outcome measures to assess pharmacodynamics: Transcriptional inhibition of the APP 
gene will result in decreased production of APP, which may be reflected by decreased secretion 
of the processed parts fragments of APP, namely sAPP and Aβ, in CSF. We have selected 
Aβ40 as a primary outcome measure because it has a relatively high concentration in CSF and 
is unaffected by the deposition of Aβ as plaques in the brain, which will be present in the study 
population. We also will measure secreted forms of APP (sAPPα and sAPPβ) and levels of tau 
in CSF as secondary outcome measures. sAPP reflects processing of APP by α-secretase to 


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produce sAPPα and β-secretase (BACE) to produce sAPPβ. Tau levels are being measured 
since tau transcription may be under the regulation of a similar IRE that may also be affected by 
Posiphen treatment. 
Safety outcome measures: Reports of adverse events (AEs) and serious adverse events 
(SAEs) during exposure to Posiphen will be collected to evaluate if there are any significant 
clinical safety issues for the study population. All SAEs occurring up to 30 days after last study 
drug administration that are considered drug-related must also be reported. Extensive clinical 
and laboratory safety data already exist for Posiphen, therefore, this approach will be sufficient 
in the proposed study. 
Clinical and cognitive assessment measures: Although clinical benefit is unlikely due to the 
short duration of this study, standard cognitive measures (MMSE, Logical Memory subtest of 
the Wechsler memory Scale (Story 1 only), and ADAS-cog) will be collected to describe the 
study cohort, which allows comparison to subjects in other clinical trials. It is predicted that 
cognition will remain stable over the course of drug exposure; however, this will be assessed 
formally in clinic.   
5 
POTENTIAL RISKS AND BENEFITS OF INVESTIGATIONAL PRODUCT AND STUDY 
PROCEDURES  
5.1 
Risks and Benefits Associated with Posiphen or Placebo 
There are no benefits to the subjects other than receiving medical and selective cognitive 
evaluations. 
The clinical investigator should advise all potential subjects of the possibility of unexpected side 
effects and carefully evaluate each person exposed to Investigational Product for possible AEs. 
Side effects to placebo are not uncommon but are obviously not due to a pharmacological agent 
as an industry standard placebo (non-lactose compound) will be provided. The placebo used for 
the study consists of microcrystalline cellulose which is generally recognized as safe (GRAS), 
with no known side effects anticipated. 
In clinical studies to date, Posiphen has been well tolerated with single doses of 80 mg or less 
and QID doses of 60 mg. A higher single dose of 160 mg was associated with an increased 
incidence of nausea and vomiting, potentially cholinergic based side effects, which resulted in 
the decision not to test higher single doses. Please refer to table below from the single, 
ascending dose study, which included healthy men and women: 


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Aside from nausea and vomiting, which are well-known responses to treatment with ChE 
inhibitors, the only consistent pattern of AEs entailed dizziness/fainting, headache, and 
reduction in total serum protein. These effects were seen to varying degrees at all doses of 
Posiphen and also in the placebo group. There was a tendency, but no definitive pattern of 
increased incidence of AEs with increasing dose of Posiphen. There have been no SAEs in 
prior clinical studies with Posiphen. 
Definitive reproductive and developmental toxicity studies have not been conducted with 
Posiphen. As a result, women of childbearing potential will be excluded from participating in this 
study. 
5.2 
Risk/Benefit Associated with Blood Collections 
Phlebotomy is associated with mild to moderate discomfort due to piercing of the skin. This can 
be minimized with the use of a well-trained phlebotomist/nurse. Sometimes the blood draw site 
may become discolored with a “bruised” appearance that is transient and not painful. Rarely, the 
blood draw site may become infected and require antibiotic treatment. 


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5.3 
Risk/Benefit Associated with SILK™ Procedure and CSF Sampling 
The most frequent adverse event associated with these studies was post-lumbar puncture 
headache. This was successfully managed by blood patching if a participant is unresponsive to 
orally administered fluids and/or caffeine.  
Other potential risks include meningitis, pain or tingling sensation in the lower extremities, 
temporary eye weakness or double vision, infection or bleeding at the site of catheter insertion, 
pain at the site of insertion, backache, epidural or subdural bleeding, infection, paralysis and 
potential injury to the nerve root.  
In studies conducted at C2N Diagnostics and at Washington University, the rate of post-lumbar 
puncture headache requiring a blood patch is about 10% in older individuals (Randall Bateman, 
MD, personal communication). 
5.4 
Risk/Benefit Associated with Investigational Drugs Intended to Reduce β-amyloid 
in the Brain 
In 2010, Food and Drug Administration (FDA) sent a notice to all sponsors of investigational 
compounds that are intended to treat Alzheimer's disease by reducing Aβ in the brain of the 
occurrence of imaging abnormalities believed to represent cerebral vasogenic edema. These 
imaging abnormalities were noted in a study of a monoclonal antibody (Salloway, 2013) and 
were, in the majority of instances, asymptomatic, and their presence was detected by routine 
brain magnetic resonance imaging (MRI). Symptoms, when present in association with such 
imaging abnormalities, were reported to include headache, worsening cognitive function, 
alteration of consciousness, seizures, unsteadiness, and vomiting. In most instances, the 
occurrence of such imaging abnormalities, even when symptomatic, did not require treatment 
beyond discontinuation of the investigational compound, with imaging abnormalities then 
resolving. Infrequently, high-dose steroid therapy was administered in the presence of 
prominent symptoms.  
More recently (2014), FDA has determined that, following a comprehensive review of existing 
data, development programs investigating use of a monoclonal antibody that targets β-amyloid 
(regardless of mechanism) must incorporate serial clinical and MRI monitoring. However, 
development programs for small molecules that target Aβ are only required to inform 
investigators of the potential risk of cerebral vasogenic edema including its imaging 
manifestations, accompanying clinical signs and symptoms, and appropriate management 
should symptomatic cases occur. Informed consent documents for these programs should also 
include a description of the manifestations of cerebral vasogenic edema, including its possible 
symptoms (i.e., headache, worsening cognitive function, alteration of consciousness, seizures, 
unsteadiness, and vomiting). 
6 
SAMPLE SIZE AND STATISTICAL PLAN 
A sample size of up to 24 subjects (5 randomized to Posiphen and 3 randomized to placebo per 
treatment group) is planned across 3-6 clinical research centers.  


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6.1 
Randomization 
Subjects who have signed an informed consent and meet screening eligibility requirements will 
be randomly assigned to receive 60 mg Posiphen daily or placebo (with an allocation of 5 on 
Posiphen, 3 on placebo) by a stratified, random permuted blocked treatment assignment 
method, stratified by site. Following dosing of the initial 8 subjects and review of safety and 
tolerability, the next dose group will be 60 mg BID Posiphen or placebo (5 on Posiphen, 3 on 
placebo). Following the dosing of this group of 8 subjects and review of safety and tolerability, 
the next dose group will be 60 mg TID Posiphen or placebo (5 on Posiphen, 3 on placebo).  
6.2 
Power and Sample Size Determination 
 
SILK™ Analyses 
Power estimates for SILK™ studies involving measurements of FSR of Aβ40 were determined 
by analysis of data provided by C2N, Inc., and modeled to evaluate a primary analytic goal of 
demonstrating a monotonic dose response relationship to Posiphen treatment. 
Based on a two-sided t-test for the difference between two independent means at a significance 
level (alpha) of 0.05 and a statistical power of 80% (1-beta), a sample size of 5 subjects in each 
treatment group (n.active = 15) and 9 controls will provide at least 80% statistical power to 
detect a 27% change in the mean Aβ40 FSR.  The following table provides estimated power 
analysis to detect different effect size reductions of FSR Aβ40 of 27% with 2-sided alpha and 
significance set at 0.05, assuming no dropouts during the SILK™ study: 
Table 2. Estimated Power Analysis  
 
n.active= 12 
n.active= 15 
n.active= 18 
n.cont= 8 
0.783 
0.822 
0.848 
n.cont= 9 
0.812 
0.851 
0.876 
n.cont= 10 
0.836 
0.874 
0.898 
This serves as justification for our sample size estimate of 24. We expect that if dropouts are to 
occur, they will occur before the SILK™ studies at day 23-25 and can be replaced to allow the 
full sample size of 24 for SILK™ being achieved.  
6.3 
Safety and Tolerability Analysis 
Safety and tolerability will be assessed with physical examinations, vital signs, clinical laboratory 
values, use of concomitant medications, and AE reports. The frequencies of adverse events, 
serious adverse events and laboratory abnormalities between the participants across the three 
groups will be compared. The severity of cholinergic adverse events will be carefully evaluated 
taking the criteria in Appendix 5 into consideration (modified from Common Terminology Criteria 
for Adverse Events (CTCAE)” version 4.03) as there is potential for an interaction between 
Posiphen and Donepezil through the inhibition of AChE, thereby resulting in increased 
cholinergic activity.   
 


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Each dose group will be completed and evaluated for safety and tolerability by the standing 
ADCS DSMB before the next dose group is initiated. Drug administration at the next higher 
dose level may not proceed until written confirmation is provided from the ADCS DSMB 
indicating that the results of the previous dose level were evaluated and that it is permissible to 
proceed to a higher dose level. 
Subjects who do not tolerate study drug during the initial 3-week at-home treatment period will 
be discontinued and replaced by new subjects. If two subjects at a given dose are discontinued 
due to adverse events, no additional subjects will be added at this dose level until a safety 
review has been conducted. The ADCS DSMB will determine the appropriate dose level for 
additional subjects. The ADCS DSMB has the option to change the allocation of future patients 
to active drug versus placebo based on statistical advice to maintain an appropriate balance 
between placebo subjects and subjects on active drug.  
 
6.4 
PK Analysis 
Plasma concentration-time data will be analyzed by non-compartmental methods using 
WinNonlin version 6.2.1 or greater. Calculations will be based on the actual sampling times 
recorded during the study. Only the subjects receiving Posiphen will be included in the PK 
analysis. From the plasma concentration-time data, the following PK parameters will be 
determined, as data permit: Area under the curve (AUC)τ, AUC0-∞, Cmax, tmax, t½, eff, Vss, Cτ, 
AR, and CL. 
Dose-proportionality may be assessed on Cmax and AUCτ as appropriate. Serial pre-dose 
trough concentrations Cτ will be assessed for the attainment of steady-state. 
The focus of the PK analysis will be to characterize the preliminary PK of Posiphen. Plasma 
concentration-time data for Posiphen will be listed and summarized descriptively (number of 
subjects (N), mean, median, standard deviation (SD), minimum (min), and maximum (max)) and 
graphically presented. PK parameters will be listed and summarized descriptively (N, mean, 
median, SD, min, max, and 95% CI) by each dose regimen. Also, a plasma/CSF comparison of 
the Posiphen PK will be conducted.  
In addition, geometric means and between-subject coefficients of variation (CV) will be 
calculated for AUCτ, AUC0-∞, Cmax, tmax, t½, eff, Vss, Cτ, AR, and CL. All PK analyses will be 
performed by the ADCS Biomarker Core. 
6.5 
SILK™ Analysis 
In SILK™ studies, the mass spectrometry data will be used to calculate the fraction of labeled 
Aβ for Aβ38, 40, and 42 during 36 hours after administration of study drug for each subject, as 
described. The FSRs will be calculated by fitting a line to this data. FSRs for treated and 
untreated subjects will be compared by analysis of variance (ANOVA). The percentage of newly 
synthesized and degraded Aβ will be estimated over 36 hours by multiplying the mass 
spectrometry concentration of Aβ by the % of Aβ that contains 13C6-leucine at each time point. 
This will be used to estimate the AUC of the drug for decreasing the amount of newly produced 
Aβ (Bateman 2009). 


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6.6 
Criteria for the termination of the trial  
The trial may be terminated by the Project Directors and/or Annovis Bio, Inc., based on issues 
of safety, feasibility, and Data Safety Monitoring Board (DSMB) recommendations. 
7 
STUDY DRUG AND CONCOMITANT MEDICATIONS 
7.1 
Name and Description of IP and Comparator 
Posiphen will be prepared in 60 mg gelatin capsules, without excipients or fillers. Matching 
placebo capsules (non-lactose compound) will be prepared with an inert inactive excipient 
generally recognized as safe for human pharmaceutical use. Annovis Bio, Inc.’s vendor, 
Frontage Laboratories, will provide both study drug and placebo. 
7.2 
Dosage 
The study drug is to be taken orally; one capsule, up to three times per day depending on 
treatment group (one in the morning, one  in the middle of the day, and one in evening/bed 
time), with or without food, for 21-23 days at home (depending on when the confinement visit is 
scheduled). An additional 2 days of drug will be administered during the confinement visit for a 
total 23-25 days of Posiphen exposure.  The study drug will be size 5, hard gelatin capsules.  
The following table describes the number and type of capsules to be taken each day based on 
treatment group. The quantity of 60 mg Posiphen capsules and placebo capsules to be taken 
each day will depend on the assigned treatment group: 
Table 3. Quantity of Capsules per Day 
 
Quantity of Capsules per day 
Treatment Group 
60 mg Posiphen 
 
Matching Placebo 
Posiphen 60 mg Treatment Group 
1 
2 
Posiphen 120 mg Treatment Group 
2 
1 
Posiphen 180 mg Treatment Group 
3 
0 
Placebo 
0 
3 
Based on when the confinement visit is scheduled, participants will take 21-23 days of the drug 
at home. Participants must take 80% of all doses (across 21-23 days), and should take at least 
80% of all doses in the last week before the confinement visit. Blister packs will be examined in 
clinic at the pre-confinement visit to assess compliance overall and in the last week.  
7.3 
Packaging/Dispensing/Labeling 
Frontage Laboratories will prepare the study drug (Posiphen capsules and placebo capsules) to 
preserve the blind, i.e., identical color and shape opaque hard gelatin capsule shells. The study 
drug (Posiphen) and placebo capsules will be packaged in identical blister packs with labels 
including the requisite cautionary statement “CAUTION: Limited by Federal (United States) Law 
to Investigational Use Only”.  
In order to preserve the blind, Frontage Laboratories will provide the initial drug supply and 
subsequent resupply to the sites to dispense to participants.  


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The dosing schedule and storage requirements will be clearly explained to the participants and 
caregiver before dispensing the study drug, and it will be printed on the labels.  
All clinical trial material dispensed under this protocol will bear the requisite cautionary 
statement “CAUTION: Limited by Federal (United States) Law to Investigational Use Only”. 
7.4 
Storage 
Both Posiphen capsules and the matching placebo capsules (non-lactose compound) must be 
stored at room temperature (15-30 °C; 59-86 °F), protected from moisture, in a locked area with 
limited staff access. 
7.5 
Drug Accountability  
The investigator is responsible for investigational product reconciliation and records 
maintenance. In accordance with all applicable regulatory requirements, the investigator or 
designated site staff must maintain investigational product accountability records throughout the 
course of the study. The responsible person(s) will document the amount of investigational 
product dispensed to and returned by subjects. All used and unused study drug containers must 
be returned to the site for accountability. Once authorized by a clinical monitor, unused study 
drug (including partially used blister packs) may be destroyed on site per appropriate site SOPS 
or should be shipped back to Annovis Bio, Inc. for destruction.  
7.6 
Compliance 
Site personnel will assess compliance based on the amount of study drug dispensed to and 
returned by the participants, together with any related information, including the administration 
of study drug by study staff during the confinement visit. Participants must take 80% of all doses 
at home (21-23 days) depending on the confinement visit date and should take at least 80% of 
all doses in the last week prior to the confinement visit. Site personnel will count capsules and 
evaluate compliance from the blister packs at the pre-confinement visit.  
Prior to the pre-confinement visit, site personnel will also contact the participant and study 
partner to remind them of the dose regimen, in particular to ensure that the necessary level of 
compliance has been achieved to continue with the confinement visit procedures. 
Site personnel will assess compliance at the pre-confinement visit and again at the time of 
admission for the confinement visit. If participants are found to be < 80% compliant for all doses 
at home and in the last week there is an insufficient level of compliance, the site must consult 
with the Project Directors (and/or Medical Monitor) regarding timing and conduct of the 
confinement visit procedures. It is possible that subjects will be discontinued early from the 
study due to insufficient drug compliance. 
During the leucine infusion, participants must comply with a low leucine diet – 1500 mg or less 
of leucine per day. The kitchen on-site will have the responsibility of preparing low leucine meals 
meals/snacks and maintaining a food journal for each individual. (Refer to Appendix 4 for a 
sample low leucine menu). 


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7.7 
Breaking the Blind  
Only in the case of an emergency, when knowledge of whether the participant has received the 
investigational product is essential for the clinical management or welfare of the participant, may 
the investigator request to unblind a participant’s treatment assignment. If the investigator needs 
the blind to be unmasked for a subject for any reason, the investigator must contact the Medical 
Monitor (and/or Project Directors) to obtain an approval. The blind will need to be broken 
centrally at the ADCS  Coordinating Center.  If the blind is broken, whether it be by accident or 
for the welfare of the participant, the investigator MUST contact the Medical Monitor. Refer to 
the study procedures manual for detailed procedures related to breaking the blind and reporting. 
7.8 
Overdose  
No specific antidote for the overdose of Posiphen is known; however, signs or symptoms of 
possible overdose should be noted and treated. The Medical Monitor and Project Directors 
should be notified as well. Because strategies for the management of overdose are continually 
evolving, it is advisable to contact a poison control center to determine the latest 
recommendations for the management of an overdose of any drug. Although Posiphen is a 
weak AChE inhibitor, based upon overdose information published in prescribing information for 
related products, overdose with AChE inhibitors can result in cholinergic crisis characterized by 
severe nausea, vomiting, salivation, sweating, bradycardia, hypotension, respiratory depression, 
collapse and convulsions. Increasing muscle weakness is a possibility and may result in death if 
respiratory muscles are involved. Tertiary anti-cholinergics such as atropine may be used as an 
antidote for overdosage. Intravenous atropine sulfate titrated to effect is recommended. Atypical 
responses in blood pressure (BP) and HR have been reported with other cholinomimetics when 
co-administered with quaternary anticholinergics such as glycopyrrolate. It is not known whether 
dialysis (hemodialysis, peritoneal dialysis, or hemofiltration) is an effective treatment for 
overdose. 
7.9 
Concomitant Medications 
7.9.1 Prohibited Concomitant Medications 
Current use of anti-coagulants (e.g., warfarin (Coumadin), dabigatran (Pradaxa) or anti-platelet 
drugs (e.g., aspirin, Plavix)), other than aspirin up to a dose of 325 mg per day, is prohibited. 
Prospective subjects who can safely discontinue these medications for 4 weeks before 
screening and remain off these medications throughout the study, can be considered for 
inclusion in the study.    
Use of the following medications is prohibited within 4 weeks of screening and throughout the 
study except as noted in Section 7.9.2: 
• 
specific psychoactive medications (tricyclic antidepressants, antipsychotics (except ≤50 
mg quetiapine daily, risperidone ≤1.5 mg/day, olanzapine ≤5 mg/day, and aripiprazole 
≤10 mg/day) 
• 
mood-stabilizing psychotropic agents (e.g., lithium salts) 
• 
psychostimulants 
• 
opiate analgesics 
• 
antiparkinsonian medications 


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• 
anticonvulsant medications 
• 
systemic corticosteroids 
• 
CNS active antihistamines for allergy or sleep (daily use)  
Investigational agents are prohibited 4 weeks prior to entry and for the duration of the trial. Also, 
exclusionary, is previous treatment with an investigational small molecule with anti-amyloid 
properties or passive immunization against Aβ within 1 year of entry or previous treatment with 
an active immunization against amyloid. This will also include Aduhelm™, an Aβ-directed 
monoclonal antibody that received accelerated FDA approval for treatment of early AD but 
which is prohibited in this study. 
Initiation of prohibited medications during the course of the study is discouraged, however, if an 
excluded medication is initiated after screening, the site should consult with the Project 
Directors and Medical Monitor for further guidance.  
7.9.2 Permitted Concomitant Medications 
This protocol allows concomitant treatment with cholinesterase inhibitors and/or memantine if on 
a stable dose (FDA approved drug ranges) for 12 weeks prior to screening. However, 
participants are only permitted to be on a maximum of 10 mg/day of Aricept (donezepil).  
Initiation of or modifications to concomitant AD medications during the course of the study is 
discouraged; however, if a change occurs or a new medication is initiated, the site should 
consult with the Project Directors and Medical Monitor for further guidance. The addition of 
Aduhelm is not permitted during the trial.  
Gabapentin and pregabalin for non-seizure indications, stable doses of bladder medication, and 
non-opiate analgesics for pain management are permitted.  
Use of benzodiazepines for treatment on an as-needed basis for insomnia or daily dosing as 
anxiolytics is permitted. If they are being given chronically, use of sedatives or hypnotics should 
be avoided for 8 hours before administration of cognitive tests. If sedating medication is given 
for any short-term use, then all cognitive assessments must be administered at least 24 hours 
after administration of the sedative. Sedating medication should not be taken the night before 
the baseline visit, the day of the baseline visit, the night before the pre-confinement visit, or the 
day of the pre-confinement visit.  
8 
STUDY POPULATION 
Up to 24 participants with early AD including those with a clinical profile consistent with MCI or 
mild AD will be enrolled in this study, in accordance with the criteria specified below. Subjects 
who do not meet all inclusion criteria, disease diagnostic criteria, or who meet any exclusion 
criteria may not be randomized into the study without prior approval from the Project Directors 
and Medical Monitor. 
8.1 
Inclusion Criteria 
Subjects must meet the following criteria:  
1. Male or female aged 55 to 89 years (inclusive), in good health, no frailty. 


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2. Female participants must be post-menopausal for at least 2 consecutive years or 
surgically sterile (bilateral tubal ligation, hysterectomy or bilateral oophorectomy) for at 
least 6 months prior to screening.  
3. Female participants will be given a urine pregnancy test at the screening visit for which 
they should test negative.  
4. Clinical profile consistent with MCI or mild AD, consistent with the core clinical criteria 
outlined in the NIA-AA Guidelines (2011).  
5. MMSE score between 17 and 30 (inclusive). 
6. CDR global score of 0.5 with a memory score of 0.5 or greater, or CDR global score of 
1.0. 
7. Participant likely to tolerate all study procedures per PI judgment. 
8. To qualify for entry, subjects will have a CSF Aβ42/ Aβ40 ratio below 0.131 that is 
consistent with Alzheimer’s disease as measured via mass spectrometry by C2N.  
9. General cognition and functional performance sufficiently preserved that the subject can 
provide written informed consent. 
10. A minimum of 6 years of education or good work history. 
11. Study partner is available who has frequent contact with the subject (e.g., average of 10 
hours per week or more), and can accompany the subject to most visits to answer 
questions about the subject. The study partner is required to attend the entire screening 
visit and the baseline visit. The study drug is dispensed to the participant at the baseline 
visit and the study partner (or other individual) should also oversee study drug 
administration if needed to ensure compliance with dose regimen. At a minimum, the 
study partner should stay for the first 3 hours of the confinement visit and return at the 
discharge to drive the subject home.   
12. No evidence of current suicidal ideation or previous suicide attempt in the past month as 
evaluated in the Columbia Suicide Severity Rating Scale. 
13. MRI scan within the 12 months prior to screening without evidence of infection, 
infarction, or other focal lesions and without clinical symptoms suggestive of intervening 
neurological disease. Lacunes that are not believed to contribute to the subject's 
cognitive impairment are permissible. If there is no MRI available within a 12-month 
timeframe, then an MRI must be performed as part of the screening procedures for 
eligibility.  
14. Stability of permitted medications for 4 weeks prior to baseline. NOTE: Cholinesterase 
inhibitors and or memantine are allowable only if stable for 12 weeks prior to screen. If 
taking Arciept (donezepil), no more than 10 mg/day is permitted during the course of the 
study.  
15. Adequate visual and hearing ability (physical ability to perform all the study 
assessments). 
16. Good general health with no disease expected to interfere with the study. Subjects may 
have common age-related disorders (i.e., hypertension, type II diabetes, dyslipidemia, 
and hypothyroidism) as long as these disorders are being controlled by diet or 
medication. 
17. Must speak English, Spanish, or Korean fluently. 


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8.2 
Exclusion Criteria 
Subjects meeting any of the following criteria must not be included in the study: 
1. Has a history of a psychiatric disorder such as schizophrenia, bipolar disorder or major 
depression according to the criteria of the most current version of the Diagnostic and 
Statistical Manual of Mental Disorders (DSM). Mild depression or history of depression 
that is stable on treatment with a tricyclic antidepressant SSRI or SNRI medication at a 
stable dose is acceptable. 
2. Other neurodegenerative diseases, including Parkinson’s disease and Huntington’s 
disease, or cerebral tumor. 
3. Dementia other than AD, such as Acquired Immunodeficiency Syndrome (AIDS), 
Creutzfeldt-Jakob disease (CJD), Lewy Bodies dementia (LBD), Cerebrovascular 
dementia (CVD), Progressive Supranuclear Palsy (PSP), or normal pressure 
hydrocephalus (NPH). 
4. History of a seizure disorder. 
5. Clinically significant abnormalities in screening laboratory or ECG results. 
6. Has current serious or unstable illness including cardiovascular, hepatic, renal, 
gastroenterologic, respiratory, endocrinologic, neurologic, psychiatric, immunologic, or 
hematologic disease or other conditions that, in the investigator’s opinion, makes them 
ineligible for participation in this study. 
7. Has four or more microinfarcts as noted in the MRI scan. 
8. Has cancer or has had a malignant tumor within the past 3 years, except patients who 
underwent potentially curative therapy with no evidence of recurrence. (Patients with 
stable untreated prostate cancer or skin cancers are not excluded). 
9. According to the criteria of the most current version of the DSM, alcohol abuse, alcohol 
dependence, or drug abuse in the past 5 years. 
10. Participation in another clinical trial with an investigational agent and have taken at least 
one dose of study medication, unless unblinded on placebo, within 16 weeks prior to 
screening. (The end of a previous investigational trial is the date the last dose of an 
investigational agent was taken). 
11. Resides in a skilled nursing facility. 
12. Subjects with infection or inflammation of the skin or skin disease at or in proximity to the 
lumbar puncture site. 
13. History of lumbar spine surgery or chronic low back pain (CLBP). 
14. Subjects whom the site PI deems to be otherwise ineligible. 
15. Has a deep brain stimulator (DBS).  
 


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9 
DESCRIPTION OF STUDY VISITS 
Each subject will have a 42 day screening period followed by approximately 21-23 days of 
treatment at home. The study visits are described below and outlined in the Schedule of Events 
in Appendix 1 as well as in the Study Schematic Diagram in Appendix 2.  
9.1 
Pre-screening 
During the prescreen phase, sites will identify potential participants through a variety of 
mechanisms (i.e., reviewing participants enrolled in ADRC centers, de novo recruitment, and 
referrals) and assess if they may potentially qualify for the study following the site’s standard 
practice.  
9.2 
Screening Visit 
The screening visit should occur up to 42 days before the baseline visit. The screening visit 
procedures may be completed over multiple days and will typically require at least two in-clinic 
visits, one of which will be to perform the lumbar puncture. 
Potential participants, or legally authorized representatives, and their study partners must sign 
an informed consent form prior to administration of any study-related procedures. Information 
regarding the subject’s demographics, concurrent medications, and medical history will be 
collected along with cognitive assessments, physical examination, and neurological 
examination. 
Safety assessments will include an ECG and MRI scan that will need to be read locally and 
reviewed by the site investigator to confirm eligibility. The clinical safety laboratory blood and 
urine tests will be sent to a central lab for analysis. The screening safety laboratory tests will be 
comprised of a metabolic panel, complete blood count, coagulation panel, and urinalysis as 
listed in Section 12.1.3. Coagulation panel results must be obtained prior to performing the 
lumbar puncture. The urinalysis will include a pregnancy test for female participants. 
A lumbar puncture will be performed as part of the screening visit, in the morning (before 10 am 
is recommended), after an overnight fast. 16-20 mL CSF will be typically obtained. 2 mL CSF 
will be sent for analysis by the site (total protein, glucose, cell count) to their local lab and the 
remainder will be shipped to the ADCS Biomarker Core. The CSF sent to the ADCS Biomarker 
Core should be placed into a 14 ml conical polypropylene tube, inverted gently 3-4 times, then 
centrifuged at 1500 x g for 10 minutes at room temperature. Next, the supernatant should be 
aliquoted into 2.0 ml tubes, frozen at -80oC, and shipped overnight to the ADCS Biomarker core.  
If CSF cannot be shipped on the day of draw, it will be frozen and stored at -80 oCby the site 
until it can be shipped on dry ice. The ADCS Biomarker Core will send CSF samples to C2N 
and C2N will analyze the screening CSF for Aβ42/ Aβ40 levels. All LP supplies will be provided 
to sites by ADCS. 
Blood will be drawn at the time of the LP for research procedures. This will consist of 3 mL of 
plasma, and 10 mL of blood for APOE genotyping and storage of DNA. Blood samples should 
be shipped overnight at ambient temperature to the ADCS Biomarker Core. For a complete list 
of procedures, refer to the Schedule of Events (Appendix 1). 
Cognitive assessments must be done either prior to the lumbar puncture or within 48 hours after 
the lumbar puncture. Cognitive assessments should not be administered when the subject is in 


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a fasted state. Given the timing of the MRI and the LP, during the screening visit the cognitive 
testing can be scheduled up to 7 weekdays prior to the LP visit date. 
Once all screening visit procedures are completed, all information related to eligibility, including 
screening lab results, must be reviewed by the site investigator to assess the subject’s eligibility 
before proceeding to the baseline visit. 
9.2.1 
Screening LP Exception 
If a participant has undergone an LP for another study at your site within 90 days prior to the 
baseline visit, this LP and collected CSF samples may be used in place of the screening LP with 
prior approval from ADCS Clinical Operations. ADCS approval may be requested by email 
(csf@ucsd.edu) 
9.2.2 Re-screens 
Unless otherwise approved by the Project Directors, only one re-screen is allowed per 
participant, should the original screen be a failure. The re-screen should typically occur at least 
3 months after the original screen failure. Individuals who fail screening due to ineligible results 
from the CSF testing may not be re-screened.  
9.3 
Baseline Visit 
Baseline procedures include cognitive assessments, behavioral assessments, Research 
Satisfaction Survey, safety assessments, and review of concurrent medications and adverse 
events that occur in clinic.  
For a complete list of all visit procedures, refer to the Schedule of Events (Appendix 1). 
Following completion of all baseline procedures, subjects who continue to meet all protocol 
inclusion criteria and no exclusion criteria, should be scheduled for the confinement visit. The 
person should then be randomized, dispensed study drug and instructed on proper storage and 
dosing. The subject should be instructed on 1) when to take the first dose of the study drug and 
2) the duration of study drug administration (21-23 days at home) based on the scheduled date 
of the confinement visit.   
9.4 
Day 14 Telephone Contact  
A telephone call will be conducted at 14 ± 4 days from the baseline visit to assess for any 
changes in status, adverse events, or concomitant medications. Study medication compliance 
will be discussed and reminders given about the upcoming confinement visit. If there is an 
insufficient level of drug compliance (less than 80% overall and or less than 80% in the last 
week), the site must consult with the Project Directors and Medical Monitor regarding timing and 
conduct of the remaining confinement visit procedures. It is possible that subjects will be 
discontinued early from the study due to insufficient drug compliance. 
9.5 
Confinement Visits 
Admission for the confinement visit should occur 21-23 days (+2 days) following the first dose of 
study medication. A pre-confinement visit will be conducted 1-3 days prior to admission and a 


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post-confinement phone follow-up will be conducted approximately 24 hours following 
discharge. The confinement visit is outlined in Appendix 3, Confinement Visit Diagram. 
9.5.1 Pre-Confinement Visit  
The Pre-Confinement Visit procedures include a physical and neurological examination, 
cognitive and behavioral assessments, certain safety assessments (including clinical safety 
labs), and a review of compliance, concurrent medications and adverse events. The clinical 
safety labs will be sent to a central lab for analysis. For a complete list of all visit procedures, 
refer to the Schedule of Events (Appendix 1). 
If there is an insufficient level of drug compliance, the site must consult with the Project 
Directors and Medical Monitor regarding timing and conduct of the Confinement Visit 
procedures. It is possible that subjects will be discontinued early from the study due to 
insufficient compliance. 
9.5.2 Confinement Visit 
The confinement visit will last just over 2 days and will consist of four phases: admission, 36-
hour sampling, 12-hour observation, and discharge. Subjects will be provided with meals (note: 
some restrictions are specified below) and sleeping accommodations during this visit. For a 
complete list of all visit procedures, refer to the Schedule of Events (Appendix 1) and to the 
Confinement Visit Diagram (Appendix 3). 
Phase 1: Admission 
Subjects should arrive fasted in the early morning after an overnight fast. Clinical safety labs will 
be drawn and sent to the local lab for analysis.  
Study drug compliance over the preceding days will be assessed. If there is an insufficient level 
of drug compliance (less than 80% overall and or less than 80% in the last week), the site must 
consult with the Project Directors and Medical Monitor regarding timing and conduct of the 
remaining confinement visit procedures. It is possible that subjects will be discontinued early 
from the study due to insufficient study drug compliance. 
Admission visit procedures include conducting a brief Physical and Neurological Exam, 
collecting vital signs, reviewing and recording adverse events and concurrent medications. 
Please refer to Appendix 3.1, Confinement Visit Phase 1: Admission Diagram. 
Phase 2: 36-Hour Sampling 
For an overview of the 36-Hour Sampling time points, please refer to the Confinement Visit 
Schedule of Events (Appendix 3.2b). 
If no medical or compliance concerns are identified during the admission procedures, the 
subject will move on to the 36-Hour Sampling Phase, which entails the following procedures: 
1. Place an intravenous line (for infusion) as well as a second venous line for blood 
sampling 
2. Place the CSF catheter 
3. Collect initial samples of blood and CSF 


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4. Administer assigned study drug (will vary based on blinded treatment assignments) at 0 
hours 
5. Begin the 13C6-labeled “heavy” leucine infusion. Subjects will receive an initial bolus of 3 
mg/kg over 10 minutes, followed by 8 hours and 50 minutes of continuous intravenous 
infusion at a rate of 2 mg/kg/hour. The total leucine administration with the bolus is 9 
hours 
6. Provide a low leucine meal (NOTE: all food provided during the heavy leucine infusion 
must conform to low leucine dietary guidelines; refer to Appendix 4, Sample Low 
Leucine Menu and to the Procedures Manual for more details) 
7. Continue with the following for 36 hours: 
a. Collect CSF (6 mL) and blood (10mL) every 2 hours at the following time points: 
0, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34 and 36 hours; all 
collection times should be calculated from the start of the heavy leucine infusion. 
Samples should be in separate tubes and sent to the ADCS Biomarker Core  
NOTE: If unable to obtain a total of 6 mL CSF at any given collection time (e.g., 
due to participant headache or participant’s CSF production level) please obtain 
the maximum amount possible and notify ADCS Clinical Operations  
b. At 0 hours, an additional 1-2 ml CSF (or volume per Local laboratory 
requirements) should be sent ambient to the local laboratory for protein, glucose, 
and cell counts   
c. Perform safety monitoring every 4 hours while the subject is awake. This 
includes questions about any symptoms in relation to the CSF catheter (including 
potential adverse events such as leg or back pain, dizziness or headaches) and 
vital sign monitoring. Participants also should be asked about pain, redness or 
swelling at the IV site in the arm  
d. Administer assigned study drug (will vary based on blinded treatment 
assignments) every 8 hours – at 8, 16, 24, and 32 hours  
e. Provide low leucine meals and snacks as needed during the duration of the 
heavy leucine infusion; return to normal diet after completion of the infusion 
(Refer to Appendix 4, Sample Low Leucine Menu) 
f. 
Subjects remain at bedrest throughout the CSF sampling period of 36 hours. 
They are allowed to use a bedside commode for toileting 
8. 36 hours after the start of the heavy leucine infusion, collect the final blood and CSF 
samples. An extra 9mL of whole blood should be sent to the central lab for the metabolic 
panel and a CBC/Diff.  An extra 2mL of CSF should be sent to the local lab for protein, 
glucose, and cell counts 
9. Remove the blood and LP catheters  
10. Perform final safety monitoring, a brief physical and a brief Neurological Exam 
If the subject experiences intolerable AE(s) that require early discontinuation of the sampling 
procedures and removal of the LP catheter, contact the Medical Monitor and Project Directors 
as soon as possible to discuss the situation. If feasible, the standard 12-hour observation and 
discharge procedures should be followed to allow for management and follow-up on the adverse 
events. 


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Phase 3: 12-Hour Observation 
The subject should be observed for at least 2 hours following removal of the LP catheter. 
Subjects are allowed to be up and out of bed during this period. The Research Satisfaction 
Survey should also be administered.  
Any clinical safety labs from the pre-confinement visit that were clinically high should be 
repeated and sent to the local laboratory. A blood patch will be administered as required if a 
subject exhibits signs of a post-spinal tap headache that is not managed with bed rest, oral and 
intravenous fluids, caffeine, and/or mild analgesics (e.g., Ibuprofen, Naproxen or Tylenol). 
Do not continue administration of study drug after completion of the 36-Hour Sampling Phase. 
All used and unused study drug should be returned to the clinical site for final accountability. 
Please refer to Appendix 3.3, Confinement Visit Phase 3:12-Hour Observation Diagram. 
Phase 4: Discharge 
Prior to discharge, vital signs should be recorded and a review of AE(s) and Concurrent 
Medications should be conducted. Upon determination by the investigator (or qualified 
designee) that the subject is stable, the participant will be discharged. If a participant 
experiences an unstable AE, please contact the Medical Monitor and Project Directors for 
guidance on the appropriate course of action to be taken. Please refer to Appendix 3.4, 
Confinement Visit Phase 4: Discharge Diagram. 
9.5.3 Post-Confinement 24-Hour Phone Follow-up 
Approximately 24 hours following discharge from the Confinement Visit the subject or a person 
designated to speak for them will be contacted by phone to confirm the subject’s well-being and 
to query about any new AEs. 
10 
EARLY TREATMENT/STUDY DISCONTINUATION 
The investigators at each site will make every reasonable effort to maximize subject retention. 
However, if an investigator removes a subject from treatment or study, or if a subject declines 
further treatment or study participation, an early discontinuation visit will be completed as soon 
as possible following discontinuation. The early discontinuation visit will contain the same 
assessments as the pre-confinement visit. If an in-person visit is not possible, site personnel will 
complete as much of the early discontinuation visit as possible by telephone. 
Subjects who discontinue from the study prior to completing all protocol procedures may be 
replaced at the discretion of the Coordinating Center, in consultation with the Sponsor. 
10.1 
Reasons for Early Discontinuation 
Participants may withdraw from the study at any time as stated in the informed consent 
document given to the participant at the screening visit. Participants may also be discontinued 
from treatment/study for reasons such as the following: 
• 
Adverse experience: The participant has experienced an adverse event that, in the 
opinion of the investigator, requires early termination. This may include abnormal 
laboratory values. 


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• 
Death. 
• 
Safety risk: Any participant who becomes a safety risk to themselves during the trial will 
be withdrawn. 
• 
Protocol violation: The participant fails to meet protocol entry criteria or does not adhere 
to protocol requirements. 
• 
Non-compliance: The participant is non-compliant with completion of study-related 
evaluations and/ or intake of study drugs. 
• 
In the investigator’s judgment, it is in the participant’s best interest to discontinue 
participation in the study. 
• 
Development of suicidal or homicidal ideation requiring hospitalization or confinement. 
• 
Participant has need for a medication prohibited by the protocol. 
• 
Consent is withdrawn. The participant wishes to withdraw from the study, or the legally 
authorized representative wishes that the participant be withdrawn. 
• 
The study is terminated by the Sponsor/Coordinating Center, alone or at the 
recommendation of the Data Safety Monitoring Board. 
• 
Lost to follow up. Participant could not be recalled back to conduct follow up visits. 
• 
Loss of informed study partner. The participant no longer has a responsible study 
partner to oversee participant visits and administration of study drug. 
• 
Coordinating Center (ADCS) request. The Coordinating Center determines it is in the 
participant’s best interest to discontinue participation from the treatment and or study. 
11 
STUDY-SPECIFIC INSTRUMENTS 
11.1 
Cognitive and Clinical Evaluations  
11.1.1 Alzheimer’s Disease Assessment Scale-Cognitive (ADAS-Cog12) 
The ADAS-Cog (Rosen 1984) is a structured scale that evaluates memory (word recall, word 
recognition), reasoning (following commands), language (naming, comprehension), orientation, 
ideational praxis (placing letter in envelope) and constructional praxis (copying geometric 
designs). Ratings of spoken language, language comprehension, word finding difficulty, and 
ability to remember test instructions are also obtained. The test is scored in terms of errors, with 
higher scores reflecting poorer performance and greater impairment. An ADAS-Cog12 version 
will be used in the current study that includes Delayed Word Recall – a measure of episodic 
memory (Mohs 1997). Total ADAS scores can range from 0 (best) to 70 (worse) on the standard 
11 items, and then the delayed recall item is added with a score ranging from 0-10, for a 
maximum score of 80. The ADAS-Cog has been the primary cognitive instrument in previous 
and ongoing ADCS trials. Psychometrists at each site are required to complete an ADAS-Cog 
training and certification process prior to administering the evaluation. 
11.1.2 Columbia Suicide Severity Rating Scale (C-SSRS)  
Consistent with FDA regulatory guidance (FDA 2012), occurrence of suicide-related thoughts 
and behaviors will be assessed. The C-SSRS is a scale that captures the occurrence, severity, 


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and frequency of suicide-related thoughts and behaviors during the corresponding assessment 
period (Posner 2011). The scale includes suggested questions to elicit the type of information 
needed to determine if a suicide-related thought or behavior occurred. 
If a suicide-related thought or behavior is identified at any time during the study, a thorough 
evaluation will be performed by a study physician, and appropriate medical care will be 
provided.  
11.1.3 Logical Memory Test (Wechsler Memory Scale-Revised) 
The Logical Memory Test I and II (Delayed Paragraph Recall) is a modification of the episodic 
memory measure from the Wechsler Memory Scale-Revised (WMS-R) (Wechsler 1987). In this 
modified version, free recall of one short story that consists of 25 bits of information will be 
elicited immediately after it is read aloud to the participant and again after an approximate thirty-
minute delay. The total bits of information from the story that are recalled immediately 
(maximum score = 25) and after the delay interval (maximum score = 25) are recorded. A 
retention or “savings” score can be computed by dividing the score achieved during delayed 
recall by the score achieved during immediate recall. 
11.1.4 Mini-Mental State Examinations (MMSE) 
The MMSE (Folstein 1975) is a brief, frequently used screening instrument in Alzheimer’s 
disease drug studies. The MMSE scale evaluates orientation, memory, attention, concentration, 
naming, repetition, comprehension, and ability to create a sentence and to copy two overlapping 
pentagons. The MMSE is scored as the number of correctly completed items with a lower score 
indicative of poorer performance and greater cognitive impairment. The total score ranges from 
0 (worse) to 30 (perfect performance). 
11.1.5 Neuropsychiatric Inventory (NPI) 
The NPI is a well-validated, reliable, multi-item instrument to assess psychopathology in AD 
based on interview with the study partner (Cummings 1997). The NPI evaluates both the 
frequency and severity of 12 neuropsychiatric features, including delusions, hallucinations, 
agitation/aggression, dysphoria, anxiety, euphoria, apathy, disinhibition irritability/lability, and 
aberrant motor behavior, as well as sleep and appetite/eating disorders. Frequency ratings 
range from 1 (occasionally, less than once per week) to 4 (very frequently, once or more per 
day or continuously). Severity ratings range from 1 (mild) to 3 (severe). The score for each 
subscale is the product of severity and frequency and the total score is the sum of all subscales. 
11.1.6 Clinical Dementia Rating (CDR) 
The CDR (Berg 1998) describes five degrees of impairment in performance on each of six 
categories of cognitive functioning including memory, orientation, judgment and problem 
solving, community affairs, home and hobbies, and personal care. The ratings of degree of 
impairment obtained on each of the six categories of function are synthesized into one global 
rating of dementia (ranging from 0 to 3), with more refined measure of change available by use 
of the Sum of Boxes. Reliability and validity has been established, as has high inter-rater 
reliability. This will be used as a global measure of severity of dementia. Where a full CDR 
interview is not possible, the abbreviated CDR (Davis 1991) can be utilized. 


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11.1.7 
Research Satisfaction Survey (RSS) 
The Research Satisfaction Survey (RSS) was adapted from the work of Larsen (Larsen 1979) to 
measures the satisfaction of elderly volunteers in clinical trials to treat or prevent cognitive loss 
or dementia.  The RSS builds on the Client Satisfaction Questionnaire (CSQ-8) which 
demonstrates a high degree of validity and internal consistency and measured consumer 
satisfaction with health and human service programs. Higher scores on the CSQ indicate a 
greater level of satisfaction in service programs102; conversely, lower CSQ-8 scores have been 
associated with higher dropout rates (Lunnen 1998) and missed appointments in treatment 
settings (Larsen 1983) and in urban Community Mental Health programs (Larsen 1979). Open-
ended questions about satisfaction are part of the CSQ, and allow participants to provide 
feedback about specific operational procedures of a particular service program (Larsen 1979). 
The RSS is similarly designed with a 7- item survey and may include additional items to inquire 
about the specific aspects of a given trial. Four items are designed to be answered with a 
modified Likert scale and can be used to create a summary score with a range of 4-16, which 
can be analyzed with parametric statistics; three items provide for open ended responses about 
the intervention, the assessments and the frequency of visits and may be used to understand 
retention rates. 
12 
STUDY-SPECIFIC PROCEDURES  
12.1 
Safety Assessments 
Safety will be evaluated by monitoring for changes in the parameters summarized below, 
including any AEs/SAEs as reported by subjects or observed by the clinical staff, or by the use 
of concomitant medication during the study. 
12.1.1 Physical and Neurological Examination 
A medically qualified professional will perform a brief physical examination that consists of a 
review of the major body systems (i.e., skin, head/ears/eyes/nose/throat (HEENT), 
cardiovascular, pulmonary, abdomen, musculoskeletal, and extremities) and a brief neurological 
examination which will include an assessment of cranial nerves, strength, coordination, reflexes, 
sensation, tremor, gait and mental status. Assessments of height, weight, and vital signs 
(systolic and diastolic blood pressure, pulse, temperature, and respiration) are included. 
12.1.2 Electrocardiogram (ECG) 
An appropriately qualified individual will conduct a standard 12-lead resting ECG. The ECG 
report must be reviewed, signed, and dated by the site PI (or a medically-qualified individual 
delegated by the site PI). Those with clinically significant ECG findings will be referred for follow-
up as deemed appropriate by the investigator and may be excluded from the study. 
12.1.3 Clinical Laboratory Evaluations 
Blood and urine samples will be obtained for clinical safety lab assessments as described in the 
Schedule of Events (Appendix 1). The following table lists the clinical safety lab tests that will be 
assessed by the central lab at three time points: the screening visit, the pre-confinement visit, 
and the end of the 36-hour sampling period.  


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Table 4. Clinical Safety Lab Tests 
CLINICAL SAFETY LAB TESTS 
METABOLIC PANEL 
COMPLETE BLOOD COUNT 
URINALYSIS 
Sodium (Na) 
Potassium (K) 
Chloride (Cl) 
Carbon Dioxide (CO2) 
Blood Urea Nitrogen (BUN) 
Glucose  
Calcium (Ca)  
Creatinine (Crn) 
Bilirubin Total 
Albumin 
Protein (NOS) Total 
Glutamic-
OxaloaceticTransferase (AST, 
SGOT) 
Glutamic-PyruvateTransferase 
(ALT, SGPT) 
Alkaline Phosphatase NOS 
White Blood Cell Count (WBC) 
Red Blood Cell Count (RBC) 
Hemoglobin (Hb) 
Hematocrit (HCT) 
Mean Corpuscular Volume (MCV) 
Mean Corpuscular Hemoglobin (MCH) 
Mean Corpuscular Hemoglobin 
Concentration (MCHC) 
Red Blood Cell Distribution Width 
(RDW) 
Mean Platelet Volume (MPV) 
Platelet Count (PLT) 
 
 
Color 
Appearance 
Specific Gravity 
pH 
Blood 
Glucose 
Protein 
Ketones 
Leukocyte Esterase  
Nitrite 
Urobilinogen 
Bilirubin 
Pregnancy Test 
(only screening visit) 
COAGULATION PANEL  
PPT/PTT 
[Screening Visit ONLY] 
Routine laboratory samples (including CSF) that are collected at the admission before the LP, 
the post-confinement visit, and the 12-hour observation (if necessary) should be analyzed by a 
CLIA/CAP (or equivalent) certified local laboratory. Lab reports will be reviewed, signed and 
dated by the site PI (or a medically qualified individual delegated by the PI). If a value is outside 
of the laboratory’s normal range, the clinician will indicate if the value has clinical significance. 
Those results that are deemed clinically significant may need to be repeated and may require 
follow up with the subject’s primary care physician for further evaluation.  
12.2 
Biofluids 
12.2.1 CSF for Biomarkers 
Standard Lumbar Puncture: 
A lumbar puncture will be performed as part of the screening visit, in the morning (before 10am 
is recommended), after an overnight fast. 2ml CSF will be sent for analysis by the site (total 
protein, glucose, cell count) to a local lab. The remaining CSF will be placed into a 14 ml conical 
polypropylene tube, inverted gently 3-4 times, then centrifuged at 1500 x g for 10 minutes at 
room temperature. The supernatant should be aliquoted into 2.0 ml tubes, frozen at -80o, and 
shipped overnight to the ADCS Biomarker Core. The ADCS Biomarker Core will subsequently 
send these samples to C2N for processing and analysis according to Biomarker core SOPs. 
Analyses will include AD biomarkers (e.g., Aβ, sAPPs, tau). 
CSF samples will be banked in the biospecimen repository of the ADCS Biomarker Core for 
various future research studies; access to samples will be granted according to SOPs 
established by ADCS Biomarker Core. 


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Prior to the LP, a coagulation panel will be obtained to rule out a clotting disorder. Participants 
taking an anti-platelet agent (e.g., Plavix) must be discontinued from that agent for a minimum 
of 5 days prior to the lumbar puncture. These participants may continue with the agent after a 
minimum of 24 hours post LP. Participants who are taking anticoagulants (e.g., warfarin 
(Coumadin) and/or dabigatran (Pradaxa) may not undergo an LP and are not suitable to 
participate in this study. 
When done at the same visit, the LP should occur after the MRI scan. When done at the same 
visit, if the MRI scan can only be scheduled prior to the LP, the site must contact ADCS Clinical 
Operations for permission before proceeding. In the unplanned event that the MRI does not 
precede the LP, there should be a minimum window of 72 hours between the MRI and the LP. 
LP under fluoroscopy is permitted; however, the site must contact ADCS Clinical Operations for 
permission before proceeding. Site personnel should advise the subject that use of fluoroscopy 
(x-rays) involves exposure to radiation. 
Each study participant or a person designated to speak for them will be contacted by phone 
approximately 24 hours after the LP to confirm participant well-being and any adverse events. 
CSF Sampling:  
A total volume of 116 mL of CSF should be collected during the CSF catheter sampling during 
the confinement visit. At hours 0, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 
and 36, 6 mL of CSF is collected.  
NOTE: If unable to obtain a total of 6 mL CSF at any given collection time (e.g., due to 
participant headache or participant’s CSF production level) the maximum amount will be 
obtained and ADCS Clinical Operations must be notified.  
To clear blood associated with needle insertion, the first 1-2 mL (or more if needed) of CSF 
should will be discarded. In addition, at 0 hours, 1-2 ml CSF (or volume per Local laboratory 
requirements) will be sent ambient to the local laboratory for protein, glucose, and cell counts. 
At the 36 hours, an extra 2mL of CSF should be sent to the local lab for protein, glucose, and 
cell counts. The remaining CSF should be placed into a 14 ml conical polypropylene tube, 
inverted gently 3-4 times, and then centrifuged at 1500 x g for 10 minutes at room temperature. 
Then the supernatant should be aliquoted into 2.0 ml tubes and frozen at -80o and shipped 
overnight to the ADCS Biomarker Core.  
Each study participant or a person designated to speak for them will be contacted by phone one 
day after being discharged from the confinement visit. The participant will be asked about their 
well being and the occurrence of any adverse events. 
12.2.2 APOE Genotype and other DNA Markers 
DNA will be extracted from participant blood samples at the screening visit and will be analyzed 
for APOE genotype. This will allow secondary analyses of data on the impact of the APOE 
genotype on putative biomarkers of AD, clinical outcome measures, and adverse events. APOE 
polymorphisms are an important genetic risk factor for AD. Individuals with one or two 
Apolipoprotein epsilon 4 (APOE4) alleles have a higher risk of developing AD and an earlier age 
of onset. Furthermore, those with one or two APOE4 alleles exhibit a more rapid rate of clinical 
change in some, but not all studies.  


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Blood is collected in a uniform fashion using EDTA as anti-coagulant, as described in Section 
12.2.3.  Once blood is collected into a 10 mL EDTA plastic tube, it is shipped overnight at 
ambient temperature to the ADCS Biomarker Core for APOE genotyping. Participants will be 
asked to consent to optional DNA banking for future research studies.  
12.2.3 Plasma for Biomarkers and Pharmacokinetics 
Plasma samples will be collected at screening visit and during the confinement visit every 2 
hours for analysis of AD biomarkers (e.g., Aβ40 & 42 and exploratory biomarkers), 
cholinesterase inhibition, and drug level analyses. 
Biomarker and PK analyses at the screening visit will be performed on the 10 ml EDTA plastic 
tube whose collection is described in Section 12.2.2.    
In addition, 10 ml of blood will be collected in a uniform fashion into 10mL lavender top EDTA 
tubes at hours 0, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, and 36 during the 
confinement visit. Once blood is collected, it should be centrifuged at 1500 x g for 10 minutes at 
room temperature. 2 ml of plasma should be aliquoted into 2.0 ml freezer tubes, frozen at -80o, 
and shipped overnight on dry ice to the ADCS Biomarker Core. 
At the last collection (36 hours), an extra 9mL of whole blood should be sent to the central lab 
for metabolic panel and CBC/Diff.   
Plasma samples will be banked in the biospecimen repository of the ADCS Biomarker Core for 
future research studies; access to samples will be granted according to SOPs established by 
the ADCS Biomarker Core. 
12.2.4 DNA/Genome Wide Association Studies (GWAS)/APOE Sample 
A single 10 mL EDTA tube of whole blood will be collected for DNA/GWAS and APOE 
genotyping and epigenetic analysis at the screening visit. The EDTA tube will be shipped 
overnight at ambient temperature to the ADCS Biomarker Core and processed per SOP for 
APOE genotyping. If the sample condition is compromised or if there is poor sample yield, 
subjects will be asked to agree to re-sampling. 
In addition, subjects will be asked to consent to optional DNA banking for future research 
studies. The DNA will extracted and banked in biospecimen repository of the ADCS Biomarker 
Core. 
13 
PERSONNEL REQUIREMENTS 
The site PI is responsible for the overall conduct of the study at the site. The PI is to supervise 
project personnel and ensure that clinical raters maintain a high level of skill and accuracy in 
conducting assessments. Additionally, the PI will perform or supervise clinical evaluation of all 
subjects and ensure protocol adherence. Additional key personnel will be required, as outlined 
in the procedures manual.  


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14 
ADVERSE EVENTS (AEs)  
14.1 
Definition 
An AE is defined as per the US Code of Federal Regulation, Title 21, Part 312.32 
[http://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/cfrsearch.cfm?fr=312.32]. An AE is any 
untoward medical occurrence associated with the use of a drug in humans, whether or not 
considered drug related. 
Collection of adverse events will begin once informed consent is signed and will continue until 
the follow-up phone call performed on the day after discharge.  Adverse events include but are 
not limited to: (1) worsening or change in nature, severity, or frequency of conditions or 
symptoms present at the start of the study; (2) Subject deterioration due to primary illness; (3) 
intercurrent illness; and (4) drug interaction. An abnormal laboratory value will only be reported 
as an AE if the investigator considers it clinically significant, or if it leads to the subject being 
withdrawn from the study. 
The investigator should attempt to establish a diagnosis of the event based on signs, symptoms, 
and or other clinical information. In such cases, the diagnosis should be documented as the AE 
and not the individual signs or symptoms. Symptoms and conditions present at the beginning of 
the study will be characterized, so that AEs can be defined as any new symptom, or any 
increase in frequency or severity of an existing symptom. Adverse events should be described 
with medical terminology so that the event can be matched against a medical coding dictionary, 
such as MedDRA (Medical Dictionary for Regulatory Activities). 
Investigators should report their assessment of the potential relatedness of each AE to the 
protocol procedure, and to the investigational product, and/or drug delivery system used in the 
protocol.  
Following questioning and evaluation, all AEs, whether determined to be related or unrelated to 
the study drug by a medically qualified site PI or clinician must be documented in the subject’s 
records, in accordance with the investigator’s normal clinical practice, and on the AE electronic 
case report form (eCRF). 
14.2 
Following Up on AEs 
The investigator is obliged to follow subjects with AEs until the events have subsided, the 
conditions are considered medically stable, or the subjects are no longer available for follow up. 
Subjects who discontinue due to adverse experiences will be treated and followed according to 
established medical practice. All pertinent information will be entered into the eCRF. Adverse 
events will be reported to the independent DSMB, per ADCS SOPs and study DSMB Charter. 
Because of the potential of additive cholinergic adverse events caused by a combination of 
Posiphen and cholinesterase inhibitors in some subjects, the severity of such adverse events 
should be carefully evaluated taking the criteria in Appendix 5 into consideration (modified from 
CTCAE version 4.03.) 


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15 
SERIOUS ADVERSE EVENTS (SAE) 
15.1 
Definition  
A SAE is defined as per the US Code of Federal Regulation, Title 21, Part 312.32 
[http://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/cfrsearch.cfm?fr=312.32]. 
An adverse event 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 subject 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 in-patient hospitalization, or the 
development of drug dependency or drug abuse. 
15.2 
Reporting SAEs 
Collection of serious adverse events will begin once informed consent is signed (regardless of 
study drug exposure) and will continue until 30 days following the last dose of study drug.  All 
SAEs must be reported to the Medical Monitor, Project Directors and the Coordinating Center 
within 24 hours of learning of the event. This in turn will trigger an alert to the appropriate 
Coordinating Center personnel and Protocol Project Directors, which will lead to the initiation of 
the creation of the report. A notification will be sent to all participating sites and the DSMB once 
the report is available. Sites will inform their IRB of the event based on local IRB requirements. 
Annovis Bio, Inc. the IND holder is responsible for submitting any SAEs according to the FDA 
reporting requirements. 
16 
DATA AND SAFETY MONITORING BOARD (DSMB) 
The Coordinating Center currently has a DSMB that reviews the safety of all subjects enrolled in 
trials on an ongoing basis. The initial task of the DSMB will be to review the protocol to identify 
any necessary modifications. If modifications are necessary, revisions will be reviewed by the 
DSMB prior to its recommendation on initiation of the project. The DSMB, based on its review of 
the protocol, will work with the Medical and Safety Core personnel to identify the study-specific 
data parameters and format of the information to be regularly reported. The DSMB will initially 
be provided with data blinded to treatment status, but they may request unblinded data if there 
is a safety concern. The DSMB and NIA representative will meet in person or by conference call 
on a quarterly basis. 
Additionally, the DSMB will be informed of the occurrence of any serious adverse events within 
7 days of being reported to the Coordinating Center. The DSMB may at any time request 
additional information from the Coordinating Center. 
Based on the review of safety data, the DSMB will make recommendations regarding the 
conduct of the study. These may include amending safety monitoring procedures, modifying the 
protocol or consent, terminating the study or continuing the study as designed. Using the ADCS 


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Safety Review Process (review of lab data, vitals, and adverse events) and the DSMB, there is 
substantial oversight and case review to alert the investigators, in a timely manner, to any safety 
issues that may arise. For further details, please refer to the DSMB charter.  
Lastly, the DSMB will evaluate safety and tolerability for each dosing group.  
17 
RECORDING AND COLLECTION OF DATA  
17.1 
Case Report Form 
The PI or designee will record all data collected (either written or electronic record of data). 
Written or electronic data of record must be entered on the eCRF provided for that purpose. In 
some instances, no prior written or electronic record of data may exist and data recorded 
directly on the eCRF is considered source data. The site will be suitably trained on the use of 
the eCRF and appropriate site personnel will be authorized to provide electronic signatures. The 
PI is responsible to verify the integrity of the data and acknowledge as such by signature. 
All site entries will be made in a secured web site and the PI will review the record for 
completeness. If corrections are necessary to the eCRFs, the PI or designee will update the 
eCRF and provide documentation for the reason for change. 
Completed eCRFs will be submitted according to provided instructions, and reviewed by the 
Coordinating Center to determine their acceptability. If necessary, data correction requests will 
be generated for resolution by the study site.  
17.2 
Study Files and Source Documents 
Subject confidentiality is strictly held in trust by the participating investigators, research staff, 
and the Coordinating Center and/or sponsoring institution and their agents. This confidentiality 
is extended to cover testing of biological samples and genetic tests in addition to the clinical 
information relating to subjects. 
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 sponsoring institution. Authorized 
representatives of the sponsoring institution may inspect all documents and records required to 
be maintained by the investigator, including but not limited to, medical records (office, clinic or 
hospital) and pharmacy records for the subjects in this study. The clinical study site will permit 
access to such records. Any data, specimens, forms, reports, video/audio recordings, and other 
records that leave the site will be identified only by a subject identification number (Subject ID, 
PID) to maintain confidentiality. All records will be kept in a locked file cabinet. All computer 
entry and networking programs will be done using PIDs only. Information will not be released 
without written permission of the subject, except as necessary for monitoring by the Institutional 
Review Board (IRB), the FDA, the NIA, and the Office for Human Research Protections 
(OHRP). 
Information about study subjects will be kept confidential and managed according to the 
requirements of the Health Insurance Portability and Accountability Act (HIPAA) of 1996. Those 
regulations require a signed Subject HIPAA Authorization informing the subject of the following:  
• 
What protected health information (PHI) will be collected from subjects in this study 


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• 
Who will have access to that information and why 
• 
Who will use or disclose that information 
• 
The rights of a research subject to revoke their authorization for use of their PHI  
In the event that a subject revokes authorization to collect or use PHI, the investigator, by 
regulation, retains the ability to use all information collected prior to the revocation of subject 
authorization. Each site PI, under the guidance of his/her IRB, is responsible for ensuring that all 
applicable HIPAA regulations and State laws are met. 
18 
ETHICS AND REGULATORY CONSIDERATIONS 
18.1 
Good Clinical Practice 
This study will be conducted in compliance with the protocol and accordance with Good Clinical 
Practice (GCP) guidelines, as defined by the International Conference on Harmonisation (ICH) 
Guideline, Topic E6, the United States Code of Federal Regulations (CFR), Title 21, Part 50 – 
Protection of Human Subjects, and Part 56 – IRBs, HHS Regulations (45 CFR part 46, 160, 164 
HHS Regulations for the Protection of Human Subjects; HIPAA; 42 CFR part 50), Subpart F 
HHS Regulations for Responsibility of Applicants for Promoting Objectivity in Research for 
Which PHS Funding Is Sought HIPAA, State and Federal regulations and all other applicable 
local regulatory requirements and laws. 
Study personnel involved in conducting this study will be qualified by education, training and 
experience to perform their respective task(s) in accordance with GCP. 
No study document shall be destroyed without prior written agreement between the 
Coordinating Center and the investigator. Should the investigator wish to assign study records 
to another party or move them to another location, he/she may do so only with the prior written 
consent of the Coordinating Center. 
18.2 
Institutional Review Board (IRB)  
Institutional Review Boards must be constituted and their authority delegated through the 
institution’s normal process of governance according to applicable State and Federal 
requirements for each participating location. Each participating institution must provide for the 
review and approval of this protocol and the associated informed consent documents and 
recruitment material by an appropriate IRB registered with the OHRP. Any amendments to the 
protocol or consent materials must also be approved before they are placed into use. In the 
United States, only institutions holding a current US Federal wide Assurance (FWA) issued by 
OHRP may participate. Refer to: http://www.hhs.gov/ohrp/assurances/. 
The investigator must obtain approval from the IRB for all subsequent protocol amendments 
and, when warranted, changes to the informed consent document. Protocol and informed 
consent form amendments can be made only with the prior approval of the Coordinating Center. 
The investigator may not implement any protocol deviation without prior notification, review, and 
documented approval from the ADCS, except where necessary to eliminate an immediate 
hazard to study subjects, or when change(s) involve only logistical or administrative aspects of 
the trial. The investigator shall notify the IRB of deviations from the protocol or serious adverse 
events occurring at the site, in accordance with local procedures. 


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18.3 
Informed Consent and Health Insurance and Portability and Accountability Act 
(HIPAA) Compliance 
Informed consent will be obtained in accordance with 21CFR§50.25, and ICH Good Clinical 
Practice. Applicable HIPAA privacy notifications will be implemented and HIPAA authorizations 
signed before protocol procedures are carried out. Information should be given in both oral and 
written form as deemed appropriate by the Site’s IRB. 
Prior to the beginning of the trial, the investigator must obtain the IRB’s written approval of the 
informed consent form and any other written information to be provided to subjects and be 
acceptable by ADCS Regulatory Affairs. Consent forms must be in a language fully 
comprehensible to the prospective subjects and/or their authorized representatives and study 
partners. Subjects, their relatives, guardians, or authorized representatives and study partners will 
be given ample opportunity to inquire about the details of the study. Prior to a subject’s 
participation in the trial, the written informed consent form must be signed and personally dated by 
the subject or by the subject’s legally acceptable representative, and by the person who 
conducted the informed consent discussion. Subjects will be provided a copy of the signed ICF. 
The informed consent will not only cover consent for the trial itself, but for the genetic 
samples/data/storage and biomarker samples/data/storage as well. Consent forms will specify 
that DNA and biomarker samples are for research purposes only; the tests on the DNA and 
biomarker samples are not diagnostic in nature and subjects will never receive results. 
18.4 
Genetic Research and Storage of Genetic Material  
Only DNA from consenting subjects will be banked and used to facilitate future research on 
aging and dementia, particularly in the discovery of genetic polymorphisms that may influence 
risk of developing AD. Collection of DNA will permit investigators to probe candidate genetic 
polymorphisms as predictors of outcome in future studies. The samples will be stored by the 
ADCS as long as funding is available from the National Institutes of Health (NIH). If funding 
should lapse completely, the UCSD ADRC will provide responsible custodianship of the ADCS 
Biomarker Core. 
18.5 
Storage of Biospecimen Samples  
All biospecimens being banked for future AD biomarker research will be shipped to and stored 
by the ADCS Biomarker Core. Sample tubes will be barcoded and inventoried by subject ID 
number only and banked without personal identifiers, in accordance with ADCS Biomarker Core 
SOPs. The presence of the sample is recorded into a computerized inventory database that is 
encrypted and password-protected. 
Only specimens from consenting subjects will be banked and used to facilitate future research 
on aging and dementia. The samples will be stored by the ADCS as long as funding is available 
from the NIH. If funding should lapse completely, the UCSD ADRC will provide responsible 
custodianship of the ADCS Biomarker Core. 
19 
STUDY MONITORING  
The clinical monitor is responsible for inspecting the online case report forms and source 
documentation at specific time points throughout the study to verify adherence to the protocol, 


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completeness and accuracy of the data, and adherence to local regulations on the conduct of 
clinical research. The monitoring visits must be conducted according to the applicable ICH and 
FDA guidelines to ensure protocol adherence, quality of data, drug accountability, compliance 
with regulatory requirements and continued adequacy of the investigational site and its facilities. 
The Site Investigator will cooperate in the monitoring process by ensuring the availability of the 
eCRFs, source documents and other necessary documents at the time of the monitoring visits. 
The Site Investigator will promptly address any matters brought to his/her attention by the 
monitor. The Site Investigator may also be asked to meet in-person with the site monitor during 
certain visits. 
20 
AUDIT 
In accordance with ICH E6 (Good Clinical Practices) representatives of the Coordinating Center 
and/or Sponsor and/or regulatory agency may select this study for audit. The investigator and 
study staff are responsible for maintaining the site master file containing all study-related 
regulatory documentation as outlined by ADCS Regulatory Affairs that will be suitable for 
inspection at any time by ADCS, Sponsor, its designees, and/or regulatory agencies. Inspection 
of site facilities (e.g., pharmacy, laboratories) to evaluate the trial conduct and compliance with 
the protocol may also occur. 
21 
RECORD RETENTION 
Essential documents and study records must be retained for a minimum of seven years 
following primary publication of study results. The Coordinating Center will notify sites when 
retention of such documents is no longer required. Essential documents should be retained for 
at least 2 years after the last approval of a marketing application in an ICH region and until there 
are no pending or contemplated marketing applications in an ICH region or at least 2 years have 
elapsed since the formal discontinuation of clinical development of the investigational product. 
However, these documents should be retained for a longer period if required by the applicable 
regulatory requirements or by an agreement with the Sponsor. It is the responsibility of the 
Sponsor to inform the investigator/institution as to when these documents no longer need to be 
retained.  
22 
PUBLICATION POLICY 
The results of this study will be published. To coordinate dissemination of data from this study, a 
publication committee will be formed. The committee will consist of the The study PIs, interested 
site PIs, and appropriate ADCS and Coordinating Center personnel The committee will solicit 
input and assistance from the Sponsor and other investigators, as appropriate, and adhere to all 
ADCS Publications Policies. 
23 
SHARING OF FINAL RESEARCH DATA  
Data from this research will be shared with other researchers pursuant to the 02/26/2003 “NIH 
Final Statement on Sharing Research Data”. The ADCS grant contains a data sharing policy 
consistent with the goals of the NIH but which also respects the rights of commercial partners. 
The NIH policy can be found at:  


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http://grants.nih.gov/grants/guide/notice-files/NOT-OD-03-032.html 
NIH believes that data sharing is important for further translation of research results into 
knowledge, products, and procedures to improve human health. The NIH endorses the sharing 
of final research data to serve these and other important scientific goals. To protect subjects’ 
rights and confidentiality, identifiers will be removed from the data before they are shared. 
 
 


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24 
LITERATURE CITED 
Annovis Bio, Inc. (2019). Investigational Drug Brochure for Posiphen. 
Bateman R.J., Siemers E.R., Mawuenyega K.G., Wen G, Browning K.R., Sigurdson W.C., 
Yarasheski K.E., Friedrich S.W., Demattos R.B., May P.C., Paul S.M., Holtzman D.M. 
(2009). A gamma-secretase inhibitor decreases amyloid-beta production in the central 
nervous system. Ann Neurol, 66:48-54.  
Bateman R.J., Munsell L.Y., Morris J.C., Swarm R., Yarasheski K.E., Holtzman D.M. (2006). 
Human amyloid-beta synthesis and clearance rates as measured in cerebrospinal fluid 
in vivo. Nat Med. 12:856-61.  
Berg, L. (1988). Clinical Dementia Rating (CDR). Psychopharmacol Bull, 24(4), 637-639. 
Cahill C.M., Lahiri D.K., Huang X., Rogers J.T. (2009). Amyloid precursor protein and alpha 
synuclein translation, implications for iron and inflammation in neurodegenerative 
diseases. Biochim Biophys Acta 1790:615-28. 
Cho H.H., Cahill C.M., Vanderburg C.R., Scherzer C.R., Wang B., Huang X., Rogers J.T. 
(2010). Selective translational control of the Alzheimer amyloid precursor protein 
transcript by iron regulatory protein-1. J Biol Chem. 285 (No. 41):31217-32.  
Cullen E. et al, Geneva-Springfield Symposium poster (2006). Axonyx, Brain Aβ42 in mice 
treated orally with Posiphen is significantly lower than in vehicle control. 
Cummings J.L. (1997). The Neuropsychiatric Inventory: assessing psychopathology in dementia 
patients. Neurology Suppl 6:S10-6. 
Cutler N.R., Polinsky R.J., Sramek J.J., Enz A., Jhee S.S., Mancione L., Hourani J., Zolnouni P. 
(1998). Dose-dependent CSF acetylcholinesterase inhibition by SDZ ENA 713 in 
Alzheimer’s disease. Acta Neurol Scand. 97:244-50. 
Davis, P.B., White, H., Price, J.L., McKeel, D., Robins, L.N., (1991) Retrospective Postmortem 
Dementia Assessment: Validation of a New Clinical Interview to Assist Neuropathologic 
Study.  Arch Neurol 48, 613-617. 
[FDA] United States Food and Drug Administration. Guidance for Industry. Suicidal Ideation and 
Behavior: Prospective Assessment of Occurrence in Clinical Trials [draft guidance], 
August 2012. 
Folstein M.F., Folstein, S.E., McHugh, P.R. (1975). “Mini-mental state”. A practical method for 
grading the cognitive state of patients for the clinician. Journal of Psychiatric Research, 
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Huang Y., Potter R., Sigurdson W., Santacruz A., Shih S., Ju Y.E., Kasten T., Morris J.C., 
Mintun M., Duntley S., Bateman R.J. (2012). Effects of age and amyloid deposition on 
Aβ dynamics in the human central nervous system. Arch Neurol. 69:51-8. 
Kadir A., Andreasen N., Almkvist O., Wall A., Forsberg A., Engler H., HagmanG., Lärksäter 
M.,Winblad B., Zetterberg H., Blennow K., Längström B., Nordberg A. (2008). Effect of 
Phenserine Treatment on Brain Functional Activity and amyloid in Alzheimer ’s disease. 
Ann Neurol 621-631. 
Lahiri D.K., Chen D., Maloney B., Holloway H.W., Yu Q.S., Utsuki T., Giordano T., Sambamurti 
K., Grieg N. (2007). The experimental Alzheimer’s Disease drug Posiphen 


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[(+)-phenserine] lower amyloid-β peptide levels in cell cultures and in mice. J Pharmacol 
Exp Ther 320:386-396. 
Larsen, D.L., Attkisson, C.C., Hargreaves W.A., & Nguyen T.D. Assessment of client/patient 
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Larsen, D.L., Nguyen T.D., Green R.S., & Attkisson, C.C. Enhancing the utilization of outpatient 
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Lilja A.M., Röjdner J., Mustafiz T., Thome C.M., Storelli E., Gonzalez D., Lithner C.U., Greig 
N.H., Nordberg A., Marutle A. (2013) Age-Dependent Neuroplasticity Mechanisms in 
Alzheimer Tg2576 Mice Following Modulation of Brain Amyloid-β Levels 8(3). 
Lunnen, K.M., & Ogles B.M. A multiperspective, multivariable evaluation of reliable change. J 
Consult Clin Psychol., 66(2), 400-410 (1998). 
Maccecchini M.L., Chang M.Y., Pan C., John V., Zetterberg H., Greig N. (2012). Posiphen as a 
candidate drug to lower CSF amyloid precursor protein, amyloid-b peptide and s levels: 
target engagement, tolerability and pharmacokinetics in humans. J Neurol Neurosurg 
Psychiatry 83:894-902. 
Mohs R.C., Knopman D., Petersen R.C., Ferris S.H., Ernesto C., Grundman M., Sano M., 
Bieliauskas L., Geldmacher D., Clark C., Thal L.J. (1997). Development of cognitive 
instruments for use in clinical trials of antidementia drugs: additions to the Alzheimer’s 
Disease Assessment Scale that broaden its scope. The Alzheimer’s Disease 
Cooperative Study. Alzheimer Dis Assoc Disord, 11Suppl 2: S13-21. 
Marutle A., Ohmitsu M., Nilbratt M., Grieg N.H., Nordberg A., Sugaya K. (2007) Modulation of 
human neural stem cell differentiation in Alzheimer (APP23) transgenic mice by 
phenserine. PNAS 104(30) 
Posner K., Brown G.K., Stanley B., Brent D. A., Yershova K.V., Oquendo M.A., Currier G.W., 
Melvin G.A., Greenhill L., Shen S., Mann J.J. (2011) The Columbia – Suicide Severity 
Rating Scale: Initial Validity and Internal Consistency Findings From Three Multisite 
Studies With Adolescents and Adults. Am. J Psychiatry 168(12) 1266-1277. 
Rogers J.T., Mikkilineni S., Castelvetri I.C., Smith D.H., Huang X.H., Bandyopadhyay S., Cahill 
C.M., Maccecchini M.L., Lahiri D.K., Greig N.H. (2011). The alpha-synuclein 
5’untranslated region targeted translation blockers: anti-alpha synuclein efficacy of 
cardiac glycosides and Posiphen. J Neural Transm 118, 493–507. 
Rosen W.G., Mohs, R.C., Davis K. L. (1984). A new rating scale for Alzheimer’s disease. 
American Journal of Psychiatry, 141(11), 1356-1364. 
Rosen W.G., Mohs, R.C., Davis K. L. (1984). A new rating scale for Alzheimer’s disease. 
American Journal of Psychiatry, 141(11), 1356-1364. 
Salloway S., Sperling R., Gilman S., Fox N.C., Blennow K., Raskind M., Sabbagh M., Honig 
L.S., Doody R., van Dyck C.H., Mulnard R., Barakos J., Gregg K.M., Liu E., Lieberburg 
I., Schenk D., Black R., Grundman M., Baplneuzumab 201 Clinical Trial Investigators 
(2009). A Phase 2 multiple ascending dose trial of bapineuzumab in mild to moderate 
Alzheimer disease, Neurology, 73(24), 2061-2070. 
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Psychiatric Association (APA). (2000) Diagnostic and Statistical Manual of Mental 
Disorders. http://www.ncbi.nlm.nih.gov/books/NBK44358 


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Utsuki T., Yu Q.S., Davidson D., Chen D., Holloway H.W. (2006). Identification of novel small 
molecule inhibitors of amyloid precursor protein synthesis as a route to lower 
Alzheimer’s disease amyloid-beta peptide. J Pharmacol Exp Ther 318: 855-862. 
Venti A., Giordano T., Eder P., Bush A.L., Lahiri D.K., Greig N.H., Rogers J.T. (2004). The 
integrated role of desferrioxamine and phenserine targeted to aniron-responsive element 
in the APP-mRNA 5’-untranslated region. Ann N Y Acad Sci. 1035:34-48. 
Wechsler D. (1987). WMS-R Wechsler Memory Scale – Revised Manual. New York: The 
Psychological Corporation, Harcourt Brace Jovanovich, Inc. 
Yu QS1, Reale M, Kamal M.A., Holloway H.W., Luo W., Sambamurti K., Ray B., Lahiri D.K., 
Rogers JT, Greig NH.(2013). Syntheis of the Alzheimer drug Posiphen into its primary 
metabolic products (+)-N1-norPosiphen, (+)-N8-norPosiphen and (+)-N1,N8-
bisnorPosiphen, their inhibiton of amyloid precursor protein, a Synuclein synthesis, 
interleukin-1β release and cholinergic action. Antiinflamm Antiallergy Agents Med Chem. 
12:117-28. 
 
 


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25 
APPENDICES 
APPENDIX 1: SCHEDULE OF EVENTS 
Visit Timing a 
Day -42 to 0 
Day 14 
(+/- 4) 
1-3 days prior to 
confinement 
Days 21-23 (+/- 2) 
 
Screening 
Baseline 
Phone 
contact 
Pre- 
confinement e 
Confinement 
Admission 
36-Hour 
Sampling 
12-Hour 
Observation 
Discharge 
Informed Consent 
X 
 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
 
Medical and Psychiatric 
History 
X 
 
 
 
 
 
 
 
CDR 
X 
 
 
 
 
 
 
 
Logical Memory 
X 
 
 
 
 
 
 
 
MMSE 
X 
 
 
X 
 
 
 
 
ADAS-Cog12 
 
X 
 
X 
 
 
 
 
NPI 
 
X 
 
X 
 
 
 
 
Physical and Neurological 
Examination 
X 
X 
 
X 
X 
X 
 
 
C-SSRS 
X 
 
 
X 
 
 
 
 
Research Satisfaction Survey 
 
X 
 
 
 
 
X 
 
Vital Signs b 
X 
X 
 
X 
X 
X 
 
X 
Height 
X 
 
 
 
 
 
 
 
Clinical Safety Lab Testsg 
X 
 
 
X 
X 
X  
PRN f 
PRN f 
12-Lead ECG 
X 
 
 
 
 
 
 
 
MRI of Brainc 
X 
 
 
 
 
 
 
 
Lumbar Puncture 
X 
 
 
 
 
 
 
 
Blood Sampling for Research 
Labsh 
X 
 
 
 
 
X 
 
 
PK Sampling 
 
 
 
 
 
X 
 
 
Adverse Events  
X 
X 
X 
X 
X 
X 
 
X 
Concomitant Medication  
X 
X 
X 
X 
X 
X 
 
 
Dispense Study Drug 
 
X 
 
 
 
X 
 
 
Dosing 
 
X 
 
 
 
X 
 
 
Study Drug Compliance 
 
 
X 
X 
X 
 
X 
 
LP Catheter Placement 
 
 
 
 
 
X 
 
 
Heavy Leucine Infusion  
 
 
 
 
 
X 
 
 
CSF Catheter Sampling  
 
 
 
 
 
X 
 
 
24-Hour Phone Follow-Up d 
X 
 
 
 
 
 
 
X 
a Refer to Section 9 for more details on visit timing and windows. 
b Vital signs will include sitting blood pressure, pulse, temperature, respiration rate, and weight, except during the 36-hour Sampling 
Phase of the Confinement Visit when weight will not be included. 
c An MRI of the brain will only be conducted if there is no brain MRI available from within the window specified in Inclusion Criteria. 
d Phone Follow-up should occur approximately 24-hours after completion of the LP procedure at Screening and after discharge from 
the Confinement Visit. 
e The Early Discontinuation Visit will contain the same assessments as the Pre-confinement Visit. 
f If safety labs are clinically indicated during the Confinement Visit, they should be sent to a local laboratory rather than the central 
lab.  
g. Refer to Section 12.1.3 for the list of tests to be performed 
h Refer to Section 9.2 for details 
 


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APPENDIX 2: STUDY SCHEMATIC DIAGRAM 
 


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APPENDIX 3: CONFINEMENT VISIT DIAGRAM 
Appendix 3.1 Confinement Visit Phase 1: Admission Diagram 
 
Conditions 
Check 
Arriving early pre-lunch 
 
Overnight fasting 
 
Study Drug Compliance 
 
Physical exam 
 
Neurological exam 
 
Clinical safety labs 
 
Vital signs 
 
AEs reviewed and recorded 
 
Con Meds reviewed and recorded 
 
 
 


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Appendix 3.2a Confinement Visit Phase 2: Sampling – 36-Hour Observation Diagram 
             
 
   
Intravenous line for infusion
Venous line for blood sampling
CSF catheter
Initial sample of blood
Initial sample of CSF
Administer study drug
Begin 13C6-labeled "heavy" leucine 
infusion 3mg/kg over 10 minutes
Continue 8 hours and 50 minutes 
of 13C6-labeled "heavy" leucine 
infusion 2mg/kg/hour


Protocol Number: ADC-043 DISC (QR15001) 
 
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Appendix 3.2b Confinement Visit Phase 2: Sampling – 36-Hour Observation Schedule of Events  
   
All sample collection times should be calculated from the start of the heavy leucine infusion 
 
Hours 
0 
2 
4 
6 
8 
10 
12 
14 
16 
18 
20 
22 
24 
26 
28 
30 
32 
34 
36 
Blood draw(10mL) includes PK 
sampling 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Posiphen dosing 
X 
 
 
 
X 
 
 
 
X 
 
 
 
X 
 
 
 
X 
 
 
CSF catheter sampling   (6mL) 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Extra blood 9mL (central labs) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Extra CSF 2mL (local labs) 
x 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Leucine infusion (3mg/kg– 
10min) 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Leucine infusion (2mg/kg/hour – 
8 hours 50min) 
X 
X 
X 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Safety Monitoring including vital 
signs (when subject is awake) 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
Removing blood and LP 
catheters 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Low leucine meal 
Includes meals, snacks, and 
beverages 
Normal diet 
Physical and Neurological Exam 
 
X 
 


Protocol Number: ADC-043 DISC (QR15001) 
 
          Page 55 of 58 
 
Appendix 3.3 Confinement Visit Phase 3: 12-Hour Observation Diagram 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
12-Hour Observation 
• Observe subject for at least two hours following 
LP catheter removal 
• Safety labs collected for local lab if clinically 
indicated 
• Administer Research Satisfaction Survey 
• Blood patch to be administered as required per 
subject condition in case oral intravenous fluids 


Protocol Number: ADC-043 DISC (QR15001) 
 
          Page 56 of 58 
 
Appendix 3.4 Confinement Visit Phase 4: Discharge Diagram 
 
 
 
 


Protocol Number: ADC-043 DISC (QR15001) 
 
          Page 57 of 58 
 
APPENDIX 4: SAMPLE LOW LEUCINE MENU  
Low leucine meals are 1,500 mg of leucine or less a day. Individualized diet/menu is based on 
food preferences and coffee consumption since coffee contains leucine. Foods used for the 
menu are determined by availability in research kitchen and leucine values based on nutritional 
database values (i.e. Nutritionist Pro or Esha.com). Below is the sample of meals, beverages, 
and snacks menu. 
  
Amount 
Unit 
Kcal 
(kcal) 
Protein (g) 
Carb (g) 
Fat (g) 
Leuc (mg) 
  
  
2268.830 
17.847 
528.804 
20.605 
901.059 
Breakfast 
  
318.215 
5.040 
67.456 
4.304 
199.980 
Juice, Orange, Unsweetened, Prepared from 
Frozen Concentrate 
4.000 
fl. oz. 
56.025 
0.847 
13.421 
0.075 
16.185 
Banana No peel ( prep just before serving 
101.500 
g 
90.335 
1.106 
23.183 
0.335 
69.020 
Bread, Wheat 
1.000 
sl. 
67.500 
2.593 
12.365 
0.860 
97.000 
Jelly (includes Grape) with Leucine 
25.000 
g 
66.500 
0.038 
17.488 
0.005 
0.000 
Margarine, Spread, 60% Fat, Stick or Tub or Bottle 
w/ Leucine 
5.000 
g 
26.300 
0.030 
0.000 
2.959 
0.000 
Coffee, Brewed 
12.000 
fl. oz. 
3.555 
0.427 
0.000 
0.071 
17.775 
SWEET N LOW Sugar Substitute, Packet 
2.000 
item 
8.000 
0.000 
1.000 
0.000 
0.000 
Morning Snack 
  
275.270 
1.661 
65.782 
1.552 
100.007 
Crackers, Graham, Plain 
14.000 
g 
59.220 
0.966 
10.752 
1.414 
66.500 
Pear, Halves, Canned in Juice, Drained 
197.100 
g 
98.550 
0.670 
25.505 
0.138 
33.507 
Drink, Cranberry Juice Cocktail, Prepared from 
Frozen Concentrate w/ Leucine 
8.000 
fl. oz. 
117.500 
0.025 
29.525 
0.000 
0.000 
Lunch 
  
477.229 
3.660 
114.135 
3.677 
199.994 
Lettuce, Iceberg Leaves 
50.000 
g 
7.000 
0.450 
1.485 
0.070 
12.500 
Tomatoes, Red 
15.000 
g 
2.700 
0.132 
0.584 
0.030 
3.750 
Carrots 
15.000 
g 
6.150 
0.140 
1.437 
0.036 
15.300 
WISH BONE Salad Dressing, Italian, Light 
20.000 
g 
23.333 
0.000 
2.000 
1.667 
0.000 
Crackers, Saltine 
18.000 
g 
75.780 
1.710 
13.381 
1.593 
117.360 
Peach, Halves, Canned in Juice, Drained 
141.900 
g 
62.436 
0.894 
16.418 
0.043 
51.084 
Juice, Apple, Prepared from Frozen Concentrate w/ 
Leucine 
8.000 
fl. oz. 
112.330 
0.335 
27.581 
0.239 
0.000 
PEPSI Soda, Regular Cola 
15.000 
fl. oz. 
187.500 
0.000 
51.250 
0.000 
0.000 
Afternoon Snack 
  
357.310 
2.353 
78.447 
5.483 
100.780 
Cookie, Wafer, Vanilla 
26.000 
g 
122.980 
1.118 
18.486 
5.044 
78.780 
Mandarin Oranges, Canned in Light Syrup, Drained 
200.000 
g 
122.000 
0.900 
32.380 
0.200 
22.000 
Juice, Apple, Prepared from Frozen Concentrate w/ 
Leucine 
8.000 
fl. oz. 
112.330 
0.335 
27.581 
0.239 
0.000 
Dinner 
  
538.445 
3.264 
129.832 
4.007 
200.294 
Lettuce, Iceberg Leaves 
50.000 
g 
7.000 
0.450 
1.485 
0.070 
12.500 
Tomatoes, Red 
15.000 
g 
2.700 
0.132 
0.584 
0.030 
3.750 
Carrots 
15.000 
g 
6.150 
0.140 
1.437 
0.036 
15.300 
WISH BONE Salad Dressing, Italian, Light 
20.000 
g 
23.333 
0.000 
2.000 
1.667 
0.000 
Crackers, Saltine 
22.200 
g 
93.462 
2.109 
16.503 
1.965 
144.744 
Applesauce, Unsweetened, Canned 
240.000 
g 
100.800 
0.408 
27.048 
0.240 
24.000 
Drink, Cranberry Juice Cocktail, Prepared from 
Frozen Concentrate w/ Leucine 
8.000 
fl. oz. 
117.500 
0.025 
29.525 
0.000 
0.000 
PEPSI Soda, Regular Cola 
15.000 
fl. oz. 
187.500 
0.000 
51.250 
0.000 
0.000 
Evening Snack 
  
302.360 
1.870 
73.153 
1.582 
100.004 
Crackers, Graham, Plain 
14.000 
g 
59.220 
0.966 
10.752 
1.414 
66.500 
Pineapple, Canned in Juice, Drained 
209.400 
g 
125.640 
0.879 
32.876 
0.168 
33.504 
Drink, Cranberry Juice Cocktail, Prepared from 
Frozen Concentrate w/ Leucine 
8.000 
fl. oz. 
117.500 
0.025 
29.525 
0.000 
0.000 
 
 
 


Protocol Number: ADC-043 DISC (QR15001) 
 
          Page 58 of 58 
 
APPENDIX 5: SEVERITY EVALUATION OF CHOLINERGIC ADVERSE EVENTS   
Because of the potential of additive cholinergic adverse events caused by a combination of 
Posiphen and cholinesterase inhibitors in some subjects the severity of such adverse events 
should be carefully evaluated taking the criteria below into consideration (modified from 
“CTCAE” version 4.03.) 
 
Adverse 
Event 
Mild 
Moderate 
Severe 
Nausea 
Loss of appetite 
without 
alteration in 
eating habits 
Oral intake decreased 
without significant weight 
loss, dehydration or 
malnutrition 
Inadequate oral caloric or fluid 
intake; tube feeding, TPN, or 
hospitalization indicated 
Vomiting 
1 - 2 episodes 
(separated by 5 
minutes) in 24 
hrs 
3 - 5 episodes (separated 
by 5 minutes) in 24 hrs 
>=6 episodes (separated by 5 
minutes) in 24 hrs; tube feeding, 
TPN or hospitalization indicated 
Diarrhea 
Increase of <4 
stools per day 
over baseline 
Increase of 4 - 6 stools per 
day over baseline 
Increase of >=7 stools per day 
over baseline; incontinence; 
hospitalization indicated; limiting 
self care ADL 
Anorexia 
Loss of appetite 
without 
alteration in 
eating habits 
Oral intake altered without 
significant weight loss or 
malnutrition; oral 
nutritional supplements 
indicated 
Associated with significant 
weight loss or malnutrition (e.g., 
inadequate oral caloric and/or 
fluid intake); tube feeding or 
TPN indicated 
Fatigue 
Fatigue relieved 
by rest 
Fatigue not relieved by 
rest; limiting instrumental 
ADL 
Fatigue not relieved by rest, 
limiting self care ADL 
 

SAP_001.pdf: 
2 
ALZHEIMER'S DISEASE COOPERATIVE STUDY (ADCS) 
A Safety and PK/PD Study of Posiphen in Subjects with 
Early Alzheimer’s Disease (DISCOVER) 
Statistical Analysis Plan (SAP) 
February 7, 2022 
Project Principal Investigators: 
Douglas R. Galasko, MD and Martin Farlow, M.D. 
U-19 Principal Investigator:
Howard Feldman, MD 
Analysis Team:  
ADCS Biostatistics Core  
Contributing Investigators (alphabetical): 
Martin Farlow, M.D. 
Howard Feldman, MD 
Douglas R. Galasko, MD 
Shelia Jin, MD MPH 
Jeremiah Momper, PhD 
Robert Rissman, PhD 
Ron Thomas, PhD 
 
NCT02925650




3 
 H W 8S P H1 3 9U C 92 S D0 9 3
Discover Statistical Analysis Plan 
Prepared by: Dr. Ron Thomas, ADCS Biostatistics Core, February 7, 2022 
Table of Contents 
1 Study Eligibility .............................................................................................................................. 3 
2 Study Aims ..................................................................................................................................... 4 
3 Randomization ............................................................................................................................... 4 
4 Power and Sample Size Determination .......................................................................................... 4 
4.1 
Estimated Power Analysis ..................................................................................................... 4 
Table 1.  
Estimated Power Analysis ......................................................................................... 4 
4.2  
Study Populations ................................................................................................................ 5 
5 Statistical Analysis.......................................................................................................................... 5 
5.1 
Primary Aims ....................................................................................................................... 5 
5.1.1  
Aim 1 ....................................................................................................................... 5 
5.1.2  
Aim 2 ....................................................................................................................... 5 
5.1.3  
Aim 3 ....................................................................................................................... 6 
5.2  
Secondary Aims .................................................................................................................... 7 
5.2.1  
Aim 1 ....................................................................................................................... 7 
5.2.2  
Aim 2 ....................................................................................................................... 7 
5.2.3  
Aim 3 ....................................................................................................................... 8 
5.3  
Tertiary Analyses .................................................................................................................. 8 
 
 
Study Title: A Multicenter, Randomized, Double-Blind, Placebo-Controlled, Ascending Dose Study to 
Evaluate the Safety, Tolerability, Pharmacokinetics (PK) and Pharmaco- dynamic (PD) Effects of Posiphen® 
in Subjects with Early Alzheimer’s Disease (AD) 
Study Short Title: A Safety and PK/PD Study of Posiphen in Subjects with Early Alzheimer’s Disease 
Protocol Number: ADC-043-DISC (QR15001)  
1 
Study Eligibility 
Clinical profile consistent with early AD including MCI-AD or mild AD according to the core clinical criteria 
outlined in the National Institute Aging (NIA) and Alzheimer’s Association (NIA-AA) Guidelines 
Cerebrospinal fluid (CSF) AD biomarkers consist  with AD (Beta Amyloid 42(Ab42)/Beta Amyloid 40(Ab40) 
ratio less than 0.131 as measured by mass spectrometry) Age 55-89 (inclusive), in good health with no 
frailty Mini-Mental State Examination (MMSE) score between 17 – 30 (inclusive) Clinical Dementia 
Rating (CDR) global score of 0.5 with a memory score of 0.5 or greater, or global score of 1.0 
 


4 
 R G TL A B4 7
2 
Study Aims 
Aim 1. Safety and Tolerability 
Aim 2.  PK of Posiphen/metabolites in Plasma and CSF 
Aim 3.  PD on fractional synthesis rate (FSR) of Beta Amyloid 40 (Ab40) in CSF  
Aim 4.  Feasibility of SILK in a multicenter trial 
Aim 5.  PD on other biomarkers in CSF including Beta Amyloid 38 (Ab38), Ab40, Ab42, Soluble Amyloid 
 
Precursor Protein alpha (sAPPa), Soluble Amyloid Precursor Protein alpha (sAPPb), total Tau 
Aim 6.  Cognitive and/or neuropsychiatric effects of Posiphen 
 
3 
Randomization 
Subjects who have signed an informed consent and meet screening eligibility requirements will be randomly 
assigned to receive 60 mg Posiphen daily or placebo (with an allocation of 5 on Posiphen, 3 on placebo) by 
a stratified, random permuted blocked treatment assignment method, stratified by site. Following dosing 
of the initial 8 subjects and review of safety and tolerability, the next dose group will be 60 mg BID 
Posiphen or placebo (5 on Posiphen, 3 on placebo). Following the dosing of this group of 8 subjects and 
review of safety and tolerability, the next dose group will be 60 mg TID Posiphen or placebo (5 on 
Posiphen, 3 on placebo). 
 
4 
Power and Sample Size Determination 
Power estimates for SILK™ studies involving measurements of FSR of Ab40 were deter- mined by analysis 
of data provided by C2N, Inc., and modeled to evaluate a primary analytic goal of demonstrating a 
monotonic dose response relationship to Posiphen treatment. Based on a two-sided t-test for the 
difference between two independent means at a significance level (alpha) of 0.05 and a statistical power 
of 80% (1-beta), a sample size of 5 subjects in each treatment group (n active = 15) and 9 controls will 
provide at least 80% statistical power to detect a 27% change in the mean Ab40 FSR. The following table 
provides estimated power analysis to detect different effect size reductions of FSR Ab40 of 27% with 2-
sided alpha and significance set at 0.05, assuming no dropouts during the SILK™ study: 
 
4.1 
Estimated Power Analysis 
A sample size of up to 24 subjects (5 randomized to Posiphen and 3 randomized to placebo per 
treatment group) is planned across 3-6 clinical research centers. 
 
Table 1.  
Estimated Power Analysis 
 
n.active= 12 
n.active= 15 
n.active= 18 
n.cont= 8 
0.783 
0.822 
0.848 
n.cont= 9 
0.812 
0.851 
0.876 
n.cont= 10 
0.836 
0.874 
0.898 
 
This serves as justification for our sample size estimate of 24. We expect that if dropouts are to occur, 
they will occur before the SILKTM studies at day 23-25 and can be replaced to allow the full sample size of 
24 for SILKTM being achieved. 
 


5 
 
4.2  
Study Populations  
 
a. Enrolled Population: All participants who consented to screening  
b. Intention To Treat (ITT) Population: list of all randomized participants 
c. Modified Intention-to-Treat (mITT) Population: Randomized participants who took at least one 
dose of study medication confirmed by compliance assessment and had at least one post 
randomization study visit  
d. Per Protocol (completers): Randomized participants who were 
i. compliant through Confinement Visit,  
ii. compliant to with 20% of dose 1 week prior Pre-Confinement, and 
iii. completed the 36-hour sampling 
e. Safety Population: Randomized participants who:  
i. took at least one dose of drug confirmed by compliance assessment  
 
5 
Statistical Analysis 
5.1 
Primary Aims   
 
5.1.1  Aim 1 
To determine the safety and tolerability of multiple ascending doses of Posiphen on subjects with early 
AD with up to 23-25 days of daily usage. Safety and tolerability will be summarized using the following 
tables, listing and figures: 
 
• Vital signs 
• Adverse events onset before randomization 
• Adverse events onset after randomization 
• AE overall 
• AE by severity 
• MedDRA coded AEs 
• Percentage of AEs summarized by SOC 
• Number of participants with at least one AE 
• Number of AEs summarized by System Organ Class (SOC) and Preferred Terms (PT)  
• Serious AEs (SAE) onset after randomization 
• Death summary 
• Hospitalization summary 
• Safety Labs 
 
5.1.2  Aim 2 
To determine the PK of Posiphen and its metabolites in plasma and CSF of subjects with early AD treated 
with QD, BID and TID dosing regimens., samples will be analyzed at baseline time 0 pre first dose then 
at times 2H, 4H, 8H, 12 H, 16H. 20H and 24H post dose. Further doses are provided every 8 hours. 
Curves and area under the curve will be constructed for each of the three sequential dose cohorts of 60 
mg daily, 60 mg BID, and 60 mg TID. 
Standard PK parameters will be calculated for each subject, including AUC, Cmax, tmax, t½, Vss. Dose-
proportionality will be assessed on Cmax and AUC statistics. Plasma concentrations will be displayed with 
time series plots including individual and means by dose.  PK parameters summarized descriptively (N, 


6 
 
mean (SD) and 95% CI, median, min, max, and geometric mean (%CV) as appropriate) by each dose 
regimen. Accumulation ratio will be determined. CSF levels will be evaluated by dose levels and 
correlated where possible to blood levels.  
The Charles River Lab in Montreal will analyze PK of Posiphen and itsmetabolites in Plasma and CSF. This 
will include Posiphen, N1-Desmethyl Posiphen (Phenserine) and N8-Desmethyl Posiphen (Phenserine).  
Phenserine is an enantiomer of Posiphen. The chromatographic assay used does not distinguish 
between the two enantiomers, so these metabolites represent the total amount of both. 
 
5.1.3  Aim 3 
To assess the PD effects on the FSR of Ab 40 using the SILK™ technique with multiple doses of Posiphen. 
 
The schema for evaluating the Fractional Synthesis and Clearance Rate, the Absolute Synthesis and 
Clearance and Area under the concentration curve are presented in Figure 1.  
 
 
 
The analytic techniques to calculation these values are undertaken by C2N.  
 
The primary outcome measure of Fractional Synthesis Rate (FSR) of Abeta 40 will be compared across all 
Posiphen treated groups to placebo. A linear regression model will be used to evaluate treatment 
effects on the FSR Abeta 40, with age and gender as covariates. Quality control of samples for the FSR 40 
will be ascertained to determine sufficiency of the sample values prior to this regression analysis.  
 
A secondary analysis will test for possible dose group by treatment interaction.  
 
Exploratory analyses will include PK AUC correlation to Abeta 40 FSR and other SILK 
measured analytes (Abeta 38, 42, ratio 42/40, total Abeta peptides) including absolute 
synthesis and clearance.  


7 
 4 7H W 81 3 94 7U C 92 S D0 9 3
 
C2N will analyze Beta Amyloid 40 (Ab40).  
 
5.2  
Secondary Aims  
 
5.2.1  Aim 1 
To implement a multicenter lumbar CSF catheter study employing standardized methodology and SILK™ 
technology. The following criteria will be evaluated as outcomes to determine feasibility: 
 
Enrollment comparisons of research subjects at each of the 5 sites. Screen failure rates will be 
evaluated by cause, by number and by site. Demographics will be described across sites.  There will be 
comparison of protocol deviations and sampling results between the sites. Adequacy of sample 
collections to permit analyses of PK and PD effects of Posiphen during the SILK will be tabulated. 
Description of the adequacy of SILK sampling, Pk and PD during the confinement visits will be 
undertaken with comparisons across sites.  
 
Research Satisfaction Questionnaires:  Descriptive ratings of participants in response to the research 
study will be undertaken across visits including baseline and post-confinement when this questionnaire 
was administered. The responses on the survey will be grouped and compared to address the 
experience across the trial with characterization of the most frequent terms. Cross tabulations and shift 
tables will be used to address the response to the SILK procedure through its categorical variables and 
its influence on participant satisfaction. We will explore whether there is any indication of an effect of 
treatment on these participant profiles.  
 
5.2.2  Aim 2 
To assess the PD effects of treatment with Posiphen versus placebo on the levels of the following 
biomarkers in CSF of subjects with early AD: Ab 38 and 40, sAPPa, sAPPb, total tau protein. 
 
Mean changes in these biomarkers will be compared across treatment arms between baseline and day 
21 time 0 prior to the leucine infusion pooling all participants treated with Posiphen vs placebo across 
cohorts. Linear regression modelling will allow the comparison of mean change scores in the analytes 
between the active and placebo groups while adjusting for covariates including age, gender,  MMSE at 
baseline, and Apo-e status as well as armcode and dose cohort and cohort-by-armcode interaction.  
 
To evaluate the PK PD relationships, trough levels of Posiphen at day 21 (time 0) will be examined as a 
predictor of change in the CSF levels of each of these biomarkers between baseline and day 21. A linear 
regression model will be used to predict the change in CSF analyte with the covariates of trough PK, age, 
and gender. Other PK measures will be similarly evaluated including AUC, and Cmax. 
 
In a secondary analysis to evaluate the effect of dose, we will use a similar regression model with terms 
for placebo and each dose group as predictors, including age and gender as covariates.   
 
5.2.3  Aim 3 
To assess whether there are short-term cognitive and/or neuropsychiatric effects of Posiphen in 
subjects with early AD, either positive or negative on ADAS-Cog12, MMSE, Neuropsychiatric Inventory 
(NPI). 


8 
 
 
These cognitive and neuropsychiatric measures will be compared between Posiphen and placebo using 
multiple linear regression. This methodology will allow the comparison of mean change scores between the 
active and placebo groups while initially adjusting for covariates; age, gender, treatment arm and baseline 
MMSE. Dose effects will be assessed as part of this analysis to investigate trends. The effects of ApoE 
genotype on these clinical outcome measures will also be explored as a tertiary analysis.  
 
C2N will analyze Beta Amyloid 38 (Ab38), Ab40, and Ab42. The ADCS Biomarker Core will analyze Soluble 
Amyloid Precursor Protein alpha (sAPPa), Soluble Amyloid Precursor Protein alpha (sAPPb), and total 
Tau protein.   
 
5.3  
Tertiary Analyses 
 
The data of the normalized SILK curve sampling will provide for exploratory analyses of fractional 
synthesized rates of Abeta 38, 42, and total Abeta as well as the ratio curve of Abeta 42/40. The 
fractional clearance rate will be explored as well using the normalized SILK curve data for these same 
analytes also including Abeta 40. To compare all Posiphen treated groups to placebo a linear regression 
model will be used to evaluate treatment effects on the fractional and absolute synthesis rates, with age 
and gender as covariates. Quality control of samples for each of these analytes will be ascertained to 
determine sufficiency of the sample values prior to this regression analysis. 
 
The absolute synthesis and clearance rates of newly generated Abeta 38, 40, 42, total Abeta and ratio of 
Abeta 42/40 will also be analyzed compared across all Posiphen treated groups to placebo. A linear 
regression model will be used to evaluate treatment effects on the both the fractional and absolute 
synthesis rates, with age and gender as covariates. Quality control of samples for each of these analytes 
will be ascertained to determine sufficiency of the sample values prior to this regression analysis. The 
area under the curve will be calculated for newly generated Abeta. 
 
Software  
Statistical software R (version 4.1.2) will be used http:www.r-project.org. 
 
Concluding Comments 
This phase 1B trial of Posiphen in the treatment of early AD, investigates its pharmacokinetics, 
pharmacodynamics and target engagement, as well as safety and tolerability. The SAP addresses each of 
the study aims with plans for preplanned analyses