Conditions: Alzheimer Disease Parkinson Disease
Keywords: Safety Tolerability Pharmacokinetics Pharmacodynamics Neurotoxic proteins Neurotransmitters Amyloid Precursor Proteins Inflammatory factors Synaptic factors CSF
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 75
Healthy volunteers: no

Brief summary:
Annovis is conducting a clinical study to investigate Posiphen in patients with Early Alzheimer's Disease (AD) and Early Parkinson's Disease (PD). Investigators are looking to recruit 68 patients in two parts of the study. In Part one of the study Investigators will recruit 14 AD and 14 PD patients who will either receive placebo (an inert pill which looks like the study drug) or the study drug Posiphen, both taken daily. In Part two of the study Investigators will recruit 40 PD patients who will receive different strengths of the study drug Posiphen taken daily. Patients will be required to come to the site for 3 face to face visits and have 4 phone calls, tests include but are not limited to, blood and CSF (spinal fluid) sampling, cognitive assessments, clinical examinations and laboratory safety tests. Primarily the Investigators are looking for the safety and tolerability of Posiphen, although Investigators will also evaluate the activity of Posiphen by a number of different biomarkers measuring pathway and target engagements.

Detailed description:
Part 1 is a study with 14 Early AD and 14 Early PD patients who are randomized to 80 mg of Posiphen or placebo. Participants will undergo a Screening Visit, provide informed consent and be evaluated for eligibility per the inclusion and exclusion criteria. If enrolled, participants will proceed to the randomized treatment portions of the study. Period 1 consists of first-time dosing in clinic with administration of 80 mg of Posiphen or Placebo. Period 2 consists of an at home dosing period of 25±2 days, with daily administration of 80 mg of Posiphen or Placebo. Period 3 will be comprised of a stay at the clinical research unit where the subject will undergo study procedures that include safety assessments (AE and concomitant medication monitoring, 12-lead ECGs, clinical laboratory testing, vital signs assessments, and physical examinations), the last dose of Posiphen or Placebo, and 6 hours of blood and CSF sampling. At the end of blood/CSF sampling, the subjects will need to stay for a minimum of 1 hour of observation but may stay if necessary for observation until the following day (e.g., if the subject has blood/CSF sampling on Day 25, he/she may stay for observation until Day 26). After all end-of-study procedures are complete, the subject will be discharged to home. A 24-hour follow-up call will occur to assess the participants current condition and if there are any additional adverse events to report.

After completion of Part 1 of the study, the plasma and CSF samples will be analyzed for the biomarkers to determine if changes are needed to the biomarkers to be measured in Part 2. Since the conduct of the study in Part 2 will be identical to the conduct of the study in Part 1, recruitment will continue uninterrupted. The only potential change between Part 1 and Part 2 are the biomarkers to be measured.

Part 2 is a study with 40 Early PD patients, 10 patients each who are randomized to one of 4 treatment conditions of Posiphen (5 mg, 10 mg, 20 mg, or 40 mg). Participants will undergo a Screening Visit, provide informed consent and be evaluated for eligibility per the inclusion and exclusion criteria. If enrolled, participants will proceed to the randomized treatment portions of the study. Period 1 consists of first-time dosing in clinic with administration of 5, 10, 20, or 40mg of Posiphen. Period 2 consists of an at home dosing period of 25±2 days, with daily administration of 5, 10, 20, or 40mg of Posiphen. Period 3 will be comprised of a stay at the clinical research unit where the subject will undergo study procedures that include safety assessments (AE and concomitant medication monitoring, 12-lead ECGs, clinical laboratory testing, vital signs assessments, and physical examinations), the last dose of Posiphen or Placebo, and 6 hours of blood and CSF sampling. At the end of blood/CSF sampling, the subject will need to stay for a minimum of 1 hour of observation but may stay if necessary for observation until the following day (e.g., if the subject has blood/CSF sampling on Day 25, he/she may stay for observation until Day 26). After all end-of-study procedures are complete, the subject will be discharged to home. A 24-hour follow-up call will occur to assess the participant's current condition and if there are any additional adverse events to report.

Arms:
arm 1: Posiphen Oral Capsule, 80mg, taken once per day for 25±2 days. arm 2: Posiphen Oral Capsule, 40mg, taken once per day for 25±2 days. arm 3: Posiphen Oral Capsule, 20mg, taken once per day for 25±2 days. arm 4: Posiphen Oral Capsule, 10mg, taken once per day for 25±2 days. arm 5: Posiphen Oral Capsule, 5mg, taken once per day for 25±2 days. arm 6: Placebo Oral Capsule, taken once per day for 25±2 days. arm 7: Placebo Oral Capsule, taken once per day for 25±2 days. arm 8: Posiphen Oral Capsule, 80mg, taken once per day for 25±2 days.

Interventions: intervention 1: Posiphen intervention 2: Placebo
Intervention descriptions: intervention 1: Solid oral dosage form, capsule intervention 2: Solid oral dosage form, capsule

Protocol document:
SAP_002.pdf: 
Statistical Analysis Plan 
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Sponsor: Annovis Bio Inc. 
Protocol:  ANVS-12003 
Statistical Analysis Plan 
A Multicenter, Randomized, Double-Blind, Placebo-Controlled, Dose-
Finding Study to Evaluate the Safety, Tolerability, Pharmacokinetic and 
Pharmacodynamic Effects of Posiphen® in Subjects with Early Alzheimer’s 
Disease (AD) or Early Parkinson’s Disease (PD) 
Part 
1-The 
evaluation 
of 
the 
safety, 
tolerability 
and 
pharmacokinetics of Placebo and 80 mg QD Posiphen in 14 
Early AD and 14 Early PD patients 
Part 
2-The 
evaluation 
of 
the 
safety, 
tolerability 
and 
pharmacokinetics of Placebo and Posiphen 5 mg QD, 10 mg QD, 
20 mg QD, 40 mg QD and 80 mg QD in 54 Early PD patients. 
Current Protocol 
ANVS-12003_AD-PD-CSFProtocol_v3_8-21-2020 
Sponsor: 
ANNOVIS BIO, Inc 
1055 Westlakes Drive, Suite 300 
Berwyn, PA 19312 
Authors & 
Qualifications 
David Fleet M.Sc. C.Stat 
Neil Parkinson, M.Sc. 
Data Magik Ltd 
Laburnum House 
Salisbury  
SP5 3RT 
Document Date: 
26th February 2021 
Version/Status: 
Final 1 
NCT 04524351




Statistical Analysis Plan 
 
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CONTENTS. 
1. INTRODUCTION. 
4 
2. OBJECTIVES. 
4 
3. STUDY DESIGN. 
6 
4. POPULATION 
8 
5. STUDY METHODS 
8 
6. STATISTICAL METHODS. 
17 
7. PRESENTATION. 
20 
8. REFERENCES. 
28 
9. FINALISATION OF STATISTICAL ANALYSIS PLAN. 
28 
 
Appendix 1 
Table Shells (Parts 1 & 2) 
Appendix 2     Plot Templates (Parts 1 & 2) 
Appendix 3  Listings shells (Parts 1 & 2) 
 


Statistical Analysis Plan 
 
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1. Introduction. 
 
Progressive neurodegenerative diseases such as Alzheimer’s (AD) and Parkinson’s disease 
(PD) share many common characteristics, including the central role of neurotoxic 
aggregating proteins in their pathogenesis.  
 
Posiphen has been shown to reduce the levels of several neurotoxic aggregating proteins 
that contribute to the progression of AD and PD. The aim of this study is to assess the 
potential effects of posiphen in patients with AD and PD in the assessment of target and 
pathway engagement in CSF and plasma.  Collectively, these endpoints will help assess 
efficacy and add valuable information to the Posiphen clinical development programme.  
 
The purpose of this Statistical Analysis Plan (SAP) is to describe the planned analyses and 
presentations of data collected from the three ‘patient cohorts’. The findings will then be 
interpreted in four statistical reports which will include tables, figures and lists (TFLs) based 
on this SAP and to support the subsequent production of the Clinical Study Report (CSR).   
 
This statistical analysis plan is based on protocol: ANVS-12003_AD-PD-CSFProtocol_v3_8-
21-2020 and will be finalised before the first cohort complete Part 1 and are unblinded for 
analysis.  
 
From a scheduling perspective, four separate analyses will be performed. The first once the 
first cohort of 14 PD patients have completed Part 1 and the second once the second cohort of 
14 AD patients complete Part 1.  The third analysis will be carried out when the third cohort 
(4 treatment groups) in Part 2 has completed all assessments but this will also include the 
Placebo and 80mg posiphen patients from the Part 1 PD cohort. The fourth analysis will  
include the pharmacokinetic data from both Part 1 (AD and PD patients) and Part 2 (PD 
patients), and, if required, a presentation of the results from the evaluation of infective 
particles in CSF and exosomes in plasma.  
 
Given the study design (patient type and treatment dose) all safety evaluations will be 
reported separately in each of the first three analyses.   
 
The following study documents were used for the preparation of this SAP: 
 
• ANVS-12003_AD-PD-CSFProtocol_v3_8-21-2020 (protocol) 
• ANVS 12003 (Annov 251151) SDTM 3.2 Annotated CRF v0.4 20200902 
 
The example layouts of the data presentations are shown in Appendices 1-3 for tables, figures 
and listings, respectively. 
 
 
 
2. Objectives. 
 
A broad range of objectives are specified in this early stage exploratory investigation. They 
include safety and tolerability in both patient groups (AD and PD). Pharmacodynamics and 
early Efficacy indicators with biomarker target engagement, pathway engagement and 
functional/cognitive measures respectively across both Part 1 and 2 of the study.  
 


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Drug pharmacokinetics in CSF and blood will also be characterised at the dose selected in AD 
and PD patients in Part 1 and across the range of doses selected in PD patients in Part 2.  
Part 1 - Safety, tolerability, pharmacodynamics, efficacy and pharmacokinetics of posiphen 
(80mg QD after 25 days) in early AD and PD patients. 
Part 2 - Safety, tolerability, pharmacodynamics, efficacy and pharmacokinetics of posiphen 
(5, 10, 20, 40mg QD after 25 days) in early PD patients. 
The primary comparisons of interest performed in Part 1 and Part 2 are the changes from 
baseline (Within treatment group). The null hypothesis throughout is that there is no 
difference between the Day 25 end of treatment and Pre-treatment baseline (H0. Xend=Xbase , 
H1. Xend^=Xbase).  
 
Between treatment group comparisons will also be done (but are of secondary interest due to 
the small sample size). Here, the null hypothesis is that there is no difference between any of 
the treatment groups at the end of treatment (H0. Placebo=X5mg=X10mg =X20mg=X40mg=X80mg , 
H1. Placebo^=X5mg^=X10mg^=X20mg^=X40mg^=X80mg).   
  
2.1 Endpoints 
 
2.1.1 Pharmacodynamics 
Pharmacodynamic effects are based on target engagement outcome measures with the levels 
of neurotoxic proteins measured in plasma and CSF. Potential drug efficacy is assessed 
through pathway engagement measurements of the levels of proteins obtained in blood, 
plasma and CSF.  
 
2.1.1.1 Target Engagement  
Levels of neurotoxic proteins measured in plasma and CSF are: 
 
• aSYN monomers  
• Aβ monomers:  Aβ40, Aβ42 
• soluble Amyloid Precursor Proteins:  sAPPα and sAPPβ 
• T-tau and P-tau protein  
 
2.1.1.2 Pathway Engagement  
Levels of proteins involved in the toxic cascade and measured in plasma and CSF are: 
 
• Axonal transport 
• Inflammatory factors 
• Synaptic factors 
• Control proteins  
 
Plus: 
Infective particles (oligomers) in CSF 
and 
Exosomes from platelet-poor plasma 
 
2.1.2 Efficacy (Functional/Cognitive)  
Although clinical benefit is not expected within this early phase, exploratory investigation, the 
AD and PD specific tests performed will provide valuable information on likely estimates of 


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variability for use in future power calculations to support sample sizes for posiphen efficacy 
clinical trials.  
 
For both AD and PD patients, the Mini-Mental State Examination (MMSE) will be used as a 
global measure of cognition and the Coding subtest from the Weschler Adult Intelligence 
Scales, 4th edition (WAIS-IV) will be used as a sensitive measure of CNS dysfunction.   
 
AD patients will undertake the Alzheimer’s disease Assessment Scale-cognitive subscale 
(ADAS-Cog 14) and functional impairment will be evaluated using the Clinical Dementia 
Rating (CDR) scale.  
 
PD patients will undertake the MDS-Unified Parkinson’s disease rating scale (MDS-UPDRS) 
to assess the non-motor and motor experiences of daily living. 
 
2.1.3 Safety and Tolerability  
Safety assessments for both parts of the study are based on physical examinations, clinical 
laboratory data, vital signs (blood pressure and heart rate), adverse events (AEs) and serious 
AEs [SAEs], ECGs (12 lead) and the Columbia suicide rating scale (C-SSRS).  Adverse 
events are assessed throughout but all other assessments are to be made at entry (Baseline) 
and on the Day 25 final assessment day. 
 
2.1.4 Pharmacokinetics  
PK parameters of the parent drug, posiphen, will be determined for both AD and PD patients 
at 80mg QD dose group after 25 days of repeated dosing (Part 1) and for PD patients at 5, 10, 
20, 40mg QD after 25 days of repeated dosing for PD patients (Part 2). 
 
Specific parameters of interest are Area under the curve (AUC 0-t, 0-inf), maximum 
concentration (Cmax), time of maximum concentration (Tmax), half-life (t½), and clearance 
(CL). 
 
 
 
3. Study Design. 
 
This study will be conducted in two parts: Part 1 will evaluate the safety, tolerability, and 
pharmacokinetics of Posiphen in 14 Early AD and 14 Early PD patients at 80 mg QD.  
 
Each group will include 14 Early AD patients, randomly assigned, 10 to posiphen and 4 to 
placebo and 14 Early PD patients randomly assigned, 10 to Posiphen and 4 to placebo.  
 
Part 1 will also include exploratory measures of pharmacodynamics and efficacy of Posiphen 
with target engagement, pathway engagement and functional/cognitive measures, including 
the degree of memory loss and cognitive function in AD patients as well as the degree of 
motor impairment and non-motor symptoms and quality of life in PD patients.  
 
Part 2 will evaluate the safety, tolerability, and pharmacokinetics of posiphen in 40 patients 
with early PD, who will be randomly assigned to each of 4 dose levels; 5 mg QD, 10 mg QD,  
20 mg QD and  40 mg QD. 
 


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Part 2 will also include exploratory measures of pharmacodynamics and efficacy of posiphen 
in Early PD patients with target engagement, pathway engagement and functional/cognitive 
measures, including the degree of motor impairment as well as other non-motor symptoms 
and quality of life in PD patients.  
 
Full clinical details are given in the protocol. 
 
3.1 Power and Sample Size Determination 
This 
is 
an 
exploratory 
investigation 
of 
safety, 
tolerability, 
pharmacokinetics, 
pharmacodynamics and efficacy focusing on target engagement, pathway engagement and 
functional/cognitive measures.  Ten patients per dose group will provide sufficient data to 
demonstrate within group changes for sAPPα and sAPPβ to be of the order of 20-40% (see 
Table 1 below) and to characterise the PK of posiphen and support potential dose 
proportionality analyses.   
 
Table 1: Sample Size Estimation 
Biomarker 
SD 
Delta effect size 
Within treatment 
group change 
N/Group* 
sAPPα 
 
0.1695 
0.3 to 2 SD 
36% 
9 
 
 
 
 
 
sAPPβ 
0.1695 
0.3 to 2 SD 
24% 
9 
 
 
 
 
 
*With 80% power and 0.05 two tailed significance level 
NB. Alpha-synuclein (aSYN) and in neurotransmitters provide valuable supporting 
information but changes are anticipated to be smaller and would require larger group sizes.  
 
3.2 Interim analysis  
No interim analysis is planned.   
 
However, on completion of Part 1, the biomarkers in the plasma and CSF samples will be 
analysed unblinded to determine what changes are needed to the biomarkers to be measured 
in Part 2.  Since the conduct of the study in Part 2 will be identical to the conduct of the study 
in Part 1, recruitment will continue uninterrupted.  The only potential change between Part 1 
and Part 2 are the actual biomarkers to be measured. Due to COVID-19 causing recruitment 
challenges, at the time of intended analysis, Part 1 will now be analysed according to cohort. 
Therefore, the first analysis will occur after 14 PD patients have completed Part 1 and a 
second analysis will occur when 14 AD patients have completed Part 1.  
 
3.3 Randomisation 
For Part 1:  For each cohort (AD, PD) up to 14 subjects will be randomised to posiphen or 
placebo in a ratio of 10 Active: 4 Placebo.  
 
For Part 2:  A sample size of up to 40 Early PD patients will be randomized to one of the 4 
posiphen doses, 10 patients each dosed at 5 mg, 10 mg, 20 mg, or 40 mg.    
 
 
The study is conducted in a blind manner and the randomization schedule for each cohort will 
be available once all patients in each cohort have completed the required schedule and the 
data have been locked for that cohort.  


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4. Population 
Three analysis populations are defined for each of the three cohorts (Part 1 AD, Part 1 PD, 
Part 2 PD):  
 
Intent to treat population  
The Intention to treat population (ITT) includes all patients who received at least one dose of 
study medication and provide post baseline data. Analyses will be based on the treatment 
actually received and not necessarily that scheduled by the randomisation. 
 
Safety population 
The Safety population (Safety) includes all patients who received at least one dose of the 
study medication. Summaries of safety and tolerability will be based on the actual treatment 
received. 
 
Pharmacokinetics population 
The Pharmacokinetic population (PK) includes all patients who receive at least one dose of 
study medication and who provide sufficient data to enable parameter estimation. 
 
 
 
5. Study Methods  
Assessments will be made during three visits (Screening, Baseline and Confinement); in 
addition there are two telephone calls to assess for adverse events. Specific clinic procedures 
and details of all assessments performed are given in Section 9 of the protocol.   
 
5.1 Schedule. 
Details of all assessments performed with timings are given in Table 2. 


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Table 2: Clinical Assessments Schedule 
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 6-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 Baseline and 
after discharge from the Confinement Visit. 
e The Early Discontinuation Visit will contain the same assessments as the Pre-confinement Visit. 
f Sampling Times:  0,1,2,3,4,5,6 hrs (±10 mins for collection)  
g Hoehn & Yahr Stage only for inclusion determination 
h X-Ray is only required if LP insertion will not be performed with fluoroscopy 
Visit Timing 
Day -42 to 0 
Day 14 
(± 4) 
1-3 days prior 
to confinement 
Days 25 (± 2) 
Procedures 
Screening Baseline Phone 
Pre- 
Confinement 
Phone 
Contact 
Confinement 
Admission 
6-Hour 
Sampling 
Dischargee 
Informed Consent 
X 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
Medical and Psychiatric History 
X 
 
 
 
 
 
 
Height and BMI 
X 
 
 
 
 
 
 
Inclusion and Exclusion Criteria 
X 
X 
 
 
 
 
 
Urine Dipstick Pregnancy Test  
X 
 
 
 
 
 
 
Physical and Neurological 
Examination 
X 
X 
 
 
X 
 
X 
AD & PD:  MMSE 
X 
X 
 
 
X 
 
 
AD:  CDR 
X 
X 
 
 
X 
 
 
AD:  ADAS-Cog 14  
 
X 
 
 
X 
 
 
AD & PD:  WAIS-IV 
 
X 
 
 
X 
 
 
PD:  MDS-UPDRS  
Xg 
X 
 
 
X 
 
 
C-SSRS 
X 
X 
 
 
X 
 
 
Vital Signs b 
X 
X 
 
 
X 
X 
X 
Clinical Lab Safety Tests 
X 
Xi 
 
 
Xi 
 
 
12-Lead ECG 
X 
X 
 
 
X 
 
X 
MRI of Brainc 
X 
 
 
 
 
 
 
Adverse Events  
X 
X 
X 
X 
X 
X 
X 
Prior/Concomitant Medication  
X 
X 
X 
X 
X 
X 
X 
Randomization 
 
X 
 
 
 
 
 
Dispense Study Drug 
 
X 
 
 
 
 
 
Dosing Study Drug in Clinic 
 
X 
 
 
 
X 
 
Study Drug Compliance Check 
 
 
X 
X 
X 
 
 
APOE Genotype and DNA 
Markers 
 
X 
 
 
 
 
 
Blood Sampling for PK 
 
X 
 
 
 
X 
 
Lumbar Puncture for baseline 
Pharmacodynamics 
 
X 
 
 
 
 
 
LP Catheter Placementh 
 
 
 
 
X 
 
 
Collecting blood and CSF  for 
Biomarkers  in Plasma and CSF 
 
X 
 
 
 
Xf 
 
24-Hour Phone Follow-Upd 
 
X 
 
 
 
 
X 
Lumbar Xrayh (Optional) 
X 
 
 
 
 
 
 


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i Safety labs at baseline and confinement (for sample 0 hour) should be taken fasted  
 
5.2 Treatment Measures. 
Treatment measures are evaluated by pharmacodynamics and by efficacy endpoints.  
 
5.2.1 Pharmacodynamic Endpoints 
Pharmacodynamic endpoints are based on the assessments from protein levels in blood, 
plasma and CSF at the Baseline (0h Pre-dosing) and Day 25 (Confinement) visit (0h Pre-
dosing, 1, 2, 3, 4, 5, 6h). Pre-dosing values at Baseline will be compared with Day 25 
(Confinement) and derived as a percentage change from Baseline. The Percentage change 
from Day 25 0h pre-dose at 1, 2, 3, 4, 5 and 6h on Day 25 (Confinement) will also be derived 
and presented. The maximum percentage change determined over the 6h period may also be 
incorporated. 
  
Table 3: Pharmacodynamic Endpoints  
 
Pharmacodynamic measure  
 
CSF 
 
Plasma 
Target Engagement  
aSYN monomers  
X 
 
Aβ monomers:  
 
 
 
 Aβ40 
X 
X 
Aβ42 
X 
X 
Soluble Amyloid Precursor Proteins:  
 
 
sAPPα  
 
X 
 
 sAPPβ 
 
X 
 
Tau Proteins: 
T-tau  
 
X 
X 
P-tau 
 
X 
X 
Pathway Engagement  
Axonal Transport: 
                                            
 
 
Neurofilament light (NfL) 
X 
X 
Inflammatory factors:  
 
 
                              Complement 3  
X 
 


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YKL40  
X 
 
 sTREM2  
X 
 
                                                                           GFAp 
X 
X 
Synaptic factors: 
 
 
SNAP25 
X 
 
Synaptotagmin 
X 
 
Neurogranin 
X 
 
Control proteins:  
 
 
 
 
Beta-trace protein 
X 
 
Infective particles: 
 
 
 
 
aSYN oligomers 
X 
 
Aβ oligomers 
X 
 
Exosomes 
 
 
 
X 
 
 
5.2.2 Efficacy - Functional/Cognitive Measures 
Patients will complete the appropriate questionnaires at Baseline and Day 25 (Confinement).  
For each, the Total score and, where appropriate the item/domain scores, will be calculated, 
and the Change from Baseline derived. Actual scores and Effect sizes ((XBase-XDay25)/SD) will 
be presented.   
  
MMSE is a 30 point test that is widely used for cognitive function that includes tests of 
orientation, attention, memory, language and visual-spatial skills.    
 
WAIS-IV  is a test to assess cognitive ability in adults which helps to examine the relationship 
between intellectual function and memory in seven subtests: Spatial Addition, Symbol Span, 
Design Memory, General Cognitive Screener, Logical Memory (I & II), Verbal Paired 
Associates (I & II), and Visual Reproduction (I & II). Performance is reported as five Index 
Scores: Auditory Memory, Visual Memory, Visual Working Memory, Immediate Memory, 
and Delayed Memory.  Patients will be scored on the number of correct items from images 
and words shown in the space of two minutes. 
 
MDS-UPDRS has four parts: Part 1 – non-motor experiences of daily living (13 items), Part 2 
- motor experiences of daily living (13 items), and Part 3 – motor examination (20 scores 
based on 18 items plus Hoehn and Yahr Stage) and Part 4 – motor complications (6 


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questions).  Each item is scored on a scale of 0 (normal) to 4 (severe).  Scores from each part 
of the scale are combined to provide a total score.  
 
ADAS-Cog14 is a test to evaluate cognition and differentiates between normal and impaired 
cognitive functioning by using a number of tests to assess memory, language and 
comprehension. Different Word lists were used in different countries to allow for language 
differences but validated versions were always used and always presented in the correct and 
standardised manner. 
 
CDR is a global summary to identify the overall severity of dementia from domains; Memory, 
Orientation, Judgement and Problem solving, Community Affairs, Home and Hobbies and 
Personal care, each of which is rated on a 5 point score.     
 
 
 
Table 4: Clinical Function and Cognitive Endpoint Scoring  
Efficacy 
 
Score Range 
(units) 
Scale Direction 
Cognitive Function Tests 
WAIS-IV 
Total Score 
Scaled Score 
 0 - 100 
High score best  
ADAS-cog14 
(14 items) 
Total Score  
(Count incorrect items) 
0 - 90 
 
Low score best  
(0=All correct) 
ADAS-cog11 
(11 items) 
Total Score  
(Count incorrect items) 
Includes: Word Recall,  Naming 
Objects and Fingers,  Commands, 
Constructional Praxis, Ideational 
Praxis, Orientation,  Word 
Recognition, Language,  
Comprehension of Spoken 
Language,  Word Finding 
Difficulty and  Remembering Test 
Instructions 
0 - 70 
 
Low score best  
(0=All correct) 
ADAS-cog 6 
(6 items) 
Total Score  
(Count incorrect items) 
Includes Word Recall, Naming 
Objects and Fingers, Commands, 
Constructional Praxis, Orientation, 
and Word Recognition Score 
0 - 45 
 
Low score best  
(0=All correct) 
ADAS-cog 3 
(Memory) 
(3 items) 
Total Score  
(Count incorrect items) 
Includes Word Recall, Orientation, 
and Word Recognition 
0 - 30 
 
Low score best  
(0=All correct) 
CDR 
Individual item scores: 
Memory, orientation, 
judgement and problem 
solving, community affairs, 
home and hobbies, and 
personal care 
0 = Normal 
0.5 = Very Mild 
Dementia 
1 = Mild Dementia 
2 = Moderate 
Dementia 
3 = Severe 
Dementia 
Global Summary 
Score range = 0 – 3 
Low score best 


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Sum of Boxes (Total Score)  
0 - 18 
Low score best 
Non-Motor Experiences and Motor Function 
MDS-UPDRS 
 
Individual scores  
0 =Normal 
1 = Slight 
2 = Mild 
3 = Moderate 
4 = Severe 
Low score best 
Total Score  
0 - 208 
Low score best 
Part 1 – non-motor 
experiences of daily living 
 0 -52 
Low score best 
Part 2  - motor experiences of 
daily living 
0 -52  
Low score best 
Part 3 – motor examination 
0 -72 
Low score best 
Part 4 – motor complications 
0 -24 
Low score best 
Hoehn and Yahr 
0 = Asymptomatic 
1 = Unilateral 
involvement only 
2 = Bilateral 
involvement 
without any 
impaired balance 
3 = Severe 
disability 
4 – Wheelchair 
bound or bedridden 
unless aided 
Low score best 
Disease Staging 
MMSE 
Total Score 
(Count correct items) 
 
0 - 30 
 
High score best 
(0=Severe impairment 
/ 30=No impairment) 
Categorical scores 
 
24– 30 
18 - 23 
0 - 17 
No cognitive 
impairment 
Mild cognitive 
impairment 
Severe cognitive 
impairment 
 
 
5.3 Safety Measures. 
Drug safety is assessed by monitoring adverse events, vital signs, physical examinations, 
12-lead ECG, and concomitant medication data. These are supported by laboratory data 
assessments of blood and urine providing serum chemistry, haematology and urinalysis 


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evaluations.  Patients will also use the Columbia suicide rating scale (C-SSRS) as an 
additional safety assessment on entry and at the confinement visit.   
  
All reported adverse events will be coded into system organ class (SOC) and preferred term 
(PT) using the MedDRA dictionary version 23. The number of patients reporting adverse 
events during either the dosing or follow up periods is summarised according to the coded 
system organ class (SOC) and preferred term (PT). All adverse events are coded individually, 
even where combination events are reported simultaneously.  
 
Adverse events starting or pre-existing events with an increase in severity on or after the first 
dose of study IMP are defined as treatment emergent adverse events (TEAE).  An event 
starting before the first dose of study IMP will be identified as a pre-treatment sign or 
symptom (PTSS).  
 
All laboratory data will be merged with the subject Case Report Form (CRF) database and 
summarised. Laboratory values will be defined as clinically significant according to the pre-
specified abnormal limits, and will be summarised and listed by subject.  
 
The Columbia suicide rating scale is a series of six questions in which the patient is asked to 
indicate whether they have experienced several thoughts or feelings relating to suicide over 
the past month.  The responses to each question by patients in each cohort will be summarised 
by treatment group and visit.  
 
5.4 Pharmacokinetic Assessments 
Plasma samples are collected at Baseline (0h Pre-dosing) and Day 25 (Confinement) visit (0h 
Pre-dosing, 1, 2, 3, 4, 5, 6h). The actual values at 0h pre-dose and at 1, 2, 3, 4, 5 and 6h on 
Day 25 (Confinement) will also be derived and from these the parameters of the parent drug 
concentration vs time course profile will be determined for each individual patient.  
 
Drug concentrations will be summarised using the designated sampling times (not the actual 
sampling times) using descriptive statistics (mean, standard deviation, coefficient of variation, 
median, minimum, maximum, and number of observations) for each treatment group and 
cohort (Part 1 AD, Part 1 PD, Part 2 PD). However, all PK statistical analyses will be carried 
out on completion of Part 2. Therefore, the 80mg PD patient data from Part 1 will be included 
with the PK patient data from the four Part 2 PD patient groups for the PD PK analysis.   
 
PK parameters will be derived using non-compartmental methods (Table 5) using SAS 
programming. All derivations will be performed using the actual times of blood sampling. All 
derived parameters (except tmax) will also be presented as descriptive summary statistics 
(mean, SD, minimum and maximum).  The time to maximum concentration (tmax) will be 
displayed as median, minimum and maximum.  
 
 
 
 
 
 
 
 
 
 


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Table 5: Estimated Pharmacokinetic Parameters - Plasma 
Parameter 
Description 
Calculation 
Cmax 
Maximum drug 
concentration from 
individual concentration-
time curve 
Directly obtained from plot 
tmax 
Time to reach maximum 
concentration 
Directly obtained from plot 
Clast 
Last measurable drug 
concentration  
Directly obtained from plot* 
Tlast 
Time of the last measurable 
concentration 
Directly obtained from plot* 
λz (Kel) 
Terminal-phase rate 
constant (λz) calculated- 
The value of λz from the 
slope of the regression line 
of lnC vs time 
Assuming - Data points 
should be randomly 
distributed around a single 
straight line; 
At least 3 data points used 
in the regression; 
Correlation coefficient of 
regression> 0.9000; 
Period of data points at 
least 2-fold greater than the 
calculated half-life 
 
t1/2 
Observed terminal 
elimination half-life  
z
T
λ
)
2
λn(
2
/
1
=
 
AUC0-t 
Area under the plasma 
concentration-time curve 
from time-zero to the time 
of the last measurable 
concentration.  
Calculated using the linear trapezoidal rule ((t2-
t1)*(C1+C2)/2)) during the ascending portion of 
the curve and the log-trapezoidal rule ((t2-
t1)*(C2-C1)/ln(C2/C1)) during the descending 
portion of the curve. 
 
AUC0-∞ 
Area under the 
concentration time curve 
from time-zero 
extrapolated to infinity 
 
z
last
last
C
AUC
AUC
l
+
=
inf
 
CL/F 
Apparent systemic 
clearance  
inf
/
AUC
Dose
F
CL
=
 
*NB: Parameters used in derivations but not presented in data summaries. 
 
For the purpose of calculating AUCs when two consecutive plasma concentrations are below 
the lower limit of quantification (LLOQ) after tmax, all subsequent values will be excluded 
from derivations.  However, single ‘embedded’ missing values will be included in the 
derivations with extrapolation between the items.  


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5.5 Missing Data. 
All possible efforts will be made to ensure complete data collection and no imputation of data 
will be performed.  
 
5.6 Other Definitions  
Baseline, Study Day and Event Duration are specifically defined as: 
 
Baseline  
Two baselines are defined: 
1 Last assessment prior to first dose of study IMP on Day 0. 
2 At 0 hours on Day 25 of the Confinement visit  
Study Day 
Study Day = (Date of event - first dose of study IMP) + 1. 
Event Duration 
Event Duration= (Date of resolution – Date of onset) + 1 
 
 
 
 


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6. Statistical Methods.  
 
This statistical analysis plan specifies the requirements for the analysis and reporting of all 
data collected during the study.  
 
All analyses and data reporting procedures will be undertaken using SAS® v9.4 in a Windows 
version 10 operating system environment.  
 
The descriptive statistics produced will account for the nature and distribution of the data 
collected. For continuous variables (actual and percentage change from baseline), data will be 
presented as number (n), mean, median, standard deviation, and range. For discrete variables 
(Incidence, Response outcome) data will be presented as frequencies and proportions and the 
denominator will be the number of patients available.  
 
Data collected from patients in each of the three cohorts (Part 1 AD, Part 1 PD, Part 2 PD) 
will be derived and summarised as described in section 5 and analysed according to the 
defined analysis population in section 4. 
 
All comparisons of primary interest performed in Part 1 and Part 2 involve changes from 
baseline (Within treatment group). The null hypothesis throughout is that there is no 
difference between the Day 25 end of treatment and Pre-treatment baseline (H0. Xend=Xbase , 
H1. Xend^=Xbase).  
 
Between treatment groups comparisons are also possible (but of secondary interest due to the 
small sample size). Here the null hypothesis is that there is no difference between any of the 
treatment groups at the end of treatment (H0. Placebo=X5mg=X10mg =X20mg=X40mg=X80mg , H1. 
Placebo^=X5mg^=X10mg^=X20mg^=X40mg^=X80mg).   
 
 
6.1 Pharmacodynamic Endpoints 
For the Part 1 AD and Part 1 PD cohorts, each variable measured in plasma and CSF will be 
analysed using an analysis of variance (ANOVA) model of the percentage changes from 
Baseline (between Baseline Day 0 Pre-dose 0h and Day 25 Pre-dose 0h). This will include 
Treatment group as a fixed effect and LSmeans (with 95% CI) for this effect will be presented 
(Within group).   
 
Between group differences for 80mg vs Placebo will be assessed and presented (LSmeans +/-
95% CI) although results will be viewed cautiously given small control group samples size. 
 
Then the percentage change from Baseline (Day 25 Pre-dose 0h) for each of the 6 hourly 
assessments will be analysed using a Repeated measured mixed model (RMMM) including 
Treatment group as a fixed effect, Time as the repeated effect and Subject as random term. 
Compound symmetry will be assumed for the covariance/correlation matrix. An overall 
average LSmean (with 95% CI) for the Treatment effect will be presented together with 
individual LSmean (with 95% CI) estimates for each hour. Treatment group differences will 
also be presented for the comparison between 80mg vs Placebo but the same caveat used 
given the control group subject sample size.  
 


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The maximum percentage change from Baseline (Day 25 Pre-dose 0h) over the 6 hours will 
also be analysed with an ANOVA model for both the Within treatment group and Between 
group (80mg vs Placebo) comparisons. 
 
In the event of either model failing assumptions of normality or constant variance, a log 
transformed analysis of the actual values (not percentage change) will be substituted and the 
output estimates (means and 95% CIs) then back transformed by exponentiation to enable 
presentation of the ratio of values (%). 
 
For the Part 2 PD cohort with 4 treatment dose groups (5, 10, 20, 40mg) the same analyses 
models will be employed. However, the data from the Part 1 PD patients will also be included 
(for those biomarker variables retained from Part 1 after evaluation at that stage) from both 
the Placebo and 80mg patient groups and combined with the Part 2 data. Within and Between 
group ANOVAs and RMMM model analyses will then be employed in a similar manner to 
Part 1. The Part 2 report of treatment group differences will include Placebo (from Part 1) 
versus each of the five active treatment groups, namely 5mg, 10mg, 20mg and 40mg from 
Part 2, and 80mg group from Part 1.  
 
6.2 Functional Cognitive Endpoints 
Prior to any formal statistical analyses of the cognitive and functional endpoints, each 
question, sub-domain and total score will be descriptively summarised and evaluated.  
Subsequently, formal statistical analysis of individual questions will only be performed and 
reported if any notable change from baseline is observed in the descriptive summaries. This 
process will be repeated at each analysis stage, i.e. Part 1 PD, Part 1 AD and Part 2 PD 
database locks. 
 
For the Part 1 AD and Part 1 PD cohorts, the variables will be analysed using an ANOVA 
model of the Change from Baseline (Day 25 - Day 0). This will include Treatment group as a 
fixed effect, and LSmeans (with 95% CI) for this effect will be presented (Within group) 
together with Effect sizes ((XDay 25-XBase)/SD). Between treatment group differences will also be 
presented for the comparison between Placebo and 80mg but interpreted cautiously. 
 
For the Part 2 PD cohort with 4 treatment dose groups (5, 10, 20, 40mg) a similar ANOVA 
will be employed.  However, the data from the Part 1 PD patients will also be included from 
both the Placebo and 80mg patient groups and combined with the Part 2 Parkinson patient 
data. Within and Between group ANOVA model analyses will then be employed in a similar 
manner to Part 1. However, in the Part 2 report, treatment group differences will include 
Placebo (from Part 1) versus each of the five active treatment groups, namely 5mg, 10mg, 
20mg and 40mg from Part 2, and 80mg group from Part 1. 
 
6.3 Safety Endpoints 
All safety data will be summarised according to cohort and treatment dose group. Presentation 
formats of the data described in section 5 are given in section 7. For consistency with the 
pharmacodynamic and functional cognitive data analyses, the data from the Part 1 Parkinson 
patients will also be included from both the Placebo and 80mg patients and combined with the 
Part 2 Parkinson patient data for the third analysis. 
 
 
 
 


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6.4 Pharmacokinetic Endpoints 
All PK analyses will be done on completion of Part 2, including the data from the Part 1 AD 
and PD patients. 
 
6.4.1 Part 1 AD, Part 1 PD Cohort  
All derived parameters for these patient groups will be presented descriptively as outlined in 
section 5.4. No formal analyses will be performed with the AD patient data. However, the 
80mg PD patient data will be combined and included with the analysis of the four Part 2 
Posiphen treatment groups. 
 
For actual drug concentrations descriptive statistics (arithmetic mean, minimum, median, 
maximum, coefficient of variation (CV), standard deviation (SD) and number of observations) 
will be used and displayed graphically as arithmetic mean (±SD) plasma drug concentration 
value versus time (nominal) for each treatment/dose group using a log-linear format.  
 
The derived pharmacokinetic parameters will also be displayed using descriptive summaries 
(arithmetic mean, standard deviation, coefficient of variation, geometric mean, median, 
minimum, maximum and number of observations) for each cohort and treatment/dose group. 
If appropriate, geometric means will be displayed in addition to the arithmetic mean.  
 
6.4.2 Part 2 PD Cohort  
Although the PK parameters will be presented as for Part 1 (according to Treatment dose 
group) the Cmax and AUCs will also be formally analysed to evaluate the relationship between 
the ascending dose groups (Dose-proportionality as in Figure 1). In addition to the patients 
from the four treatment groups in Part 2 (5mg, 10mg, 20mg and 40mg), the 80mg group of 
PD patients from Part 1 will be included. 
 
This will involve an ANOVA power model using the log transformed individual patient 
derived parameters as an explanatory variable and with Treatment dose group as the 
dependent fixed effect. In the event of demonstrating an overall statistically significant 
difference, comparisons between all pairs of means will be performed using the Scheffe 
method. LSmeans point estimates with 95% CIs derived will be presented after back 
transformation (exponentiation) of all pairwise differences between doses as ratios (%). 
Significant differences (p<0.05) in the ratio of any two doses will be declared if the 95% 
confidence interval excludes 100%. 
 
The regression slope (Figure 1) of the log transformed PK parameter against log Dose 
represents the power model (log(AUC)=a + b*(Dose)) where ‘a’ represents the intercept and 
‘b’ the coefficient for linear slope). Therefore, a 90% confidence interval for the estimate of 
the slope excluding b=1 will indicate significant departure from linearity.  
 
 
 
 
 
 
 
 
 
 
 


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Figure 1: Dose Proportionality (Log PK Parameter vs Log Dose) 
 
 
 
For the dose difference analysis, all dose group estimates will be presented as LS means with 
95% CIs after back transformation of the log values and all pairwise comparisons with the 
lowest dose will be presented as ratios of LS means (%) with 95% confidence intervals. These 
will be presented graphically.  
 
For both the Differences between doses and Dose proportionality analyses of the Cmax and 
AUC parameters summaries of the model applied with Mean square error, Degrees of 
freedom, F statistic and resulting p-value for each model parameter will be displayed.  
 
For Dose proportionality the coefficient of the linear slope (b) will be presented as a point 
estimate with 90% CI together with plots of log Cmax vs Dose and log AUC vs Dose. 
 
 
 
7. Presentation. 
 
As each cohort completes (Part 1 AD, Part 1 PD, and Part 2 PD) these data will be unblinded 
and the pharmacodynamics (Biomarkers in CSF and plasma), efficacy, and safety will be 
evaluated and presented accordingly.    
 
Once all patients in Part 2 have completed, the PK data and infective particle and exosomes 
data from all three cohorts will be made available and analysed separately.  
  
Summaries will be presented by cohort and treatment, visit and time recorded as appropriate.   
 
The table below summarises the schedule for presentation of each cohort’s statistical report 
output:  
  
 
 
 
 
 
 
 
0
10
20
30
40
50
60
70
80
90
100
0
10
20
30
40
50
Single Dose
Linear (Single Dose)


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Analysis 
Cohort 
Deliverables 
1 
Part 1 – PD Patients (14) 
Disposition Tables and Listings 
 
Biomarker* Tables, Figures and Listings 
 
Functional/cognitive Tables, Figures and 
Listings 
 
Safety Tables and Listings 
 
 
 
2 
Part 1 – AD Patients (14) 
Disposition Tables and Listings 
 
Biomarker* Tables, Figures and Listings 
 
Functional/cognitive Tables, Figures and 
Listings 
 
Safety Tables and Listings 
 
 
 
On completion of Part 2: 
3 
PD Patients (54) – including 14 
PD patients from Part 1 and 40 
PD patients from Part 2  
Disposition Tables and Listings 
 
Biomarker* Tables, Figures and Listings 
 
Functional/cognitive Tables, Figures and 
Listings 
 
Safety Tables and Listings 
 
 
 
4 
PD Patients (50) – including ten 
80mg PD patients from Part 1 
and forty PD patients (5mg, 
10mg, 20mg and 40mg) from 
Part 2 
PK analyses Tables, Figures and Listings 
 
 
 
 
AD Patients (10) – 80mg 
patients from Part 1 
PK analyses Tables, Figures and Listings 
 
 
 
 
All patients (68) from Part1 and 
Part 2 
Infective Particles in CSF: Tables, Figures and 
Listings  
 
Exosomes in plasma: Tables, Figures and 
Listings 
 
 
 
*Biomarker results (excluding infective particles and exosomes) from assays conducted in Part 1 will be 
evaluated by the Sponsor and will inform the choice of assays and subsequent output from Part 2 
NB: Infective particles and exosomes biomarker data will not be presented until after the completion of Part 2 
 
Detailed ‘table shells’ for Part 1 and Part 2 will be produced to accompany this SAP. These 
not only describe table layout but table numbering formats and also the level of accuracy 
(number of decimal places) to be described in each table. 
 
The following described tables and figures will be included in the statistical report.  
  
7.1 Study Status. 
 
7.1.1 Disposition 
For each cohort enrolled into the study, patient disposition details will be presented by 
treatment group and overall. Disposition details include number Screened, number and 
percentage Randomised (ITT population), Treated (Safety population), Completed, 
Discontinued, and the Primary Reason for Discontinuation (Tables 14.1.1 and 14.1.2).   For 


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each cohort, discontinuations from the study will be listed by treatment group and patient with 
the date of discontinuation, primary reason and the corresponding AE number if applicable 
(Listing 16.2.1.5).   
 
7.1.2 Demographics. 
For each cohort in the Safety population, patient demographic details (age, sex, race and 
ethnicity) will be summarised according to treatment group and overall (Table 14.1.3). 
 
7.1.3 Protocol Deviations 
For each cohort, protocol deviations will be listed by treatment and patient, showing the 
deviation details (Listing 16.2.1.4). 
 
7.1.3 Concomitant Medication 
For each cohort, prior and concomitant medications will be summarised by treatment group 
and overall according to the WHO drug dictionary ATC code levels 2 and 4 (Tables 14.1.4 
and 14.1.5. 
 
7.1.4 Medical History. 
For each cohort, medical history details will be summarised according to the MedDRA 
system organ class and preferred term by treatment group and overall (Table 14.1.6). Height, 
weight and BMI will also be summarised (Table 14.1.7). 
 
7.1.5 Drug Administration. 
For each cohort, compliance with study drug administration will be summarised by treatment 
group as both percentage compliance and classified according to whether patients were less 
than or equal to/greater than 80% compliant (Table 14.1.8).  
 
7.2 Pharmacodynamics 
For each cohort, the levels of neurotoxic proteins in plasma and CSF collected at hourly 
intervals during the 6 hour post dose confinement, the actual values and change from baseline 
and percentage change from baseline will be summarized by treatment group.  
 
7.2.1.1 Target Engagement - CSF 
 
The target engagement CSF variables include: Aβ-40, Aβ-42, sAPPα, sAPP β, t-tau, p-tau-
181 and a-SYN and are summarised in Tables 14.2.1.1.1 – 14.2.1.1.7.  
 
Subsequently, the results from the Within group ANOVA model of the percentage changes 
from Baseline (between Baseline Day 0 Pre-dose 0h and Day 25 Pre-dose 0h) will be 
presented, together with Maximum Percentage Change over 6h (Day 25) from Baseline (Day 
25 Pre dose 0h) (Model terms – Table 14.2.1.2; Model estimates – Table 14.2.1.3) and plotted 
as treatment estimates (Figures 14.2.1.1, Template 1). 
 
Additionally, the percentage change from baseline (Day 25 Pre-dose 0h - Day 0 Pre-dose 0h) 
and the Maximum Percentage Change over 6h (Day 25) from Baseline (Day 25 Pre-dose 0h) 
will be analysed for Between group differences. This will initially involve analysing the 
change from baseline differences between the Placebo treatment groups in Part 1 and the 
80mg patient groups from Part 1 in both the AD and PD cohorts, and subsequently the 
differences between the PD Placebo patients from Part 1 and the 5mg, 10mg, 20mg and 40mg 
Parkinson patient groups from Part 2; and repeating the PD Placebo versus PD 80mg 


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comparison (Model terms – Table 14.2.1.2; Model estimates – Table 14.2.1.3) and plotted as 
treatment estimate differences (Figures 14.2.1.2, Template 2). 
 
Furthermore, the percentage change from Baseline (Day 25 Pre-dose 0h) for each of the 6 
hourly assessments will be analysed using a Repeated measured mixed model (RMMM) and 
will be presented in Tables 14.2.1.4 and 14.2.1.5 for Within and Between treatment group 
model terms and estimates, respectively, together with corresponding plots (Figures 14.2.1.3, 
Templates 3 and 4, respectively).  
 
On completion of the two Part 1 cohorts, the target engagement CSF biomarkers will be 
evaluated and this will inform a decision about which should be assayed and statistically 
analysed after Part 2 has been completed. 
 
7.2.1.2 Pathway engagement – CSF 
The pathway engagement CSF variables include: NfL, GFAp, Complement C3, YKL40, 
sTREM2, SNAP25, Synaptotagmin, Neurogranin and Beta-trace protein and are summarised 
in Tables 14.2.2.1.1 – 14.2.2.1.9. 
 
Subsequently, the results from the Within group ANOVA model of the percentage changes 
from Baseline (between Baseline Day 0 Pre-dose 0h and Day 25 Pre-dose 0h) will be 
presented, together with Maximum Percentage Change over 6h (Day 25) from Baseline (Day 
25 Pre dose 0h) (Model terms – Table 14.2.2.2; Model estimates – Table 14.2.2.3) and plotted 
as treatment estimates (Figures 14.2.2.1, Template 1). 
 
Additionally, the percentage change from baseline (Day 25 Pre-dose 0h - Day 0 Pre-dose 0h) 
and the  Maximum Percentage Change over 6h (Day 25) from Baseline (Day 25 Pre dose 0h) 
will be analysed using a Between group ANOVA model. This will initially involve analysing 
the change from baseline differences between the Placebo treatment group from Part 1 and the 
80mg Parkinson patient group from Part 1, and subsequently the differences between the 
Placebo from Part 1 and the 5mg, 10mg, 20mg and 40mg Parkinson patient groups from Part 
2 (Model terms – Table 14.2.2.2; Model estimates – Table 14.2.2.3) and plotted as treatment 
estimate differences (Figure 14.2.2.2, Template 2). 
 
Furthermore, the percentage change from Baseline (Day 25 Pre-dose 0h) for each of the 6 
hourly assessments will be analysed using a Repeated measured mixed model (RMMM) and 
will be presented in Tables 14.2.2.4 and 14.2.2.5 for Within and Between treatment group 
model terms and estimates, respectively, together with corresponding plots (Figures 14.2.2.3, 
Templates 3 and 4, respectively).  
 
On completion of the two Part 1 cohorts, the pathway engagement CSF biomarkers will be 
evaluated and this will inform a decision about which should be assayed and statistically 
analysed after Part 2 has been completed. 
 
 
7.2.2.1 Target Engagement - Plasma 
The target engagement plasma variables include: Aβ-40, Aβ-42, t-tau and p-tau-181 and are 
summarised in Tables 14.2.3.1.1 – 14.2.3.1.4. 
 
These variables are analysed as for the CSF variables: 
Within group ANOVA model: Model terms – Table 14.2.3.2; Model estimates – Table 
14.2.3.3) and plotted as treatment estimates (Figures 14.2.3.1, Template 1). 


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Between group ANOVA model: Model terms – Table 14.2.3.2; Model estimates – Table 
14.2.3.3) and plotted as treatment estimate differences (Figure 14.2.3.2, Template 2). 
 
Repeated measured mixed model (RMMM) – Within Group: Model terms – Table 14.2.3.4 
and Model Estimates Table 14.2.3.5 and Between group also in Tables 14.2.3.4 and 14.2.3.5, 
respectively, together with corresponding plots (Figures 14.2.3.3, Templates 3 and 4, 
respectively). 
 
On completion of the two Part 1 cohorts, the target engagement plasma biomarkers will be 
evaluated and this will inform a decision about which should be assayed and statistically 
analysed after Part 2 has been completed. 
 
7.2.2.2 Pathway Engagement - Plasma 
The pathway engagement plasma variables include: NfL and GFAp and are summarised in 
Tables 14.2.4.1.1 – 14.2.4.1.2. 
 
These variables are analysed as for the CSF variables: 
Within group ANOVA model: Model terms – Table 14.2.4.2; Model estimates – Table 
14.2.4.3) and plotted as treatment estimates (Figures 14.2.4.1, Template 1). 
 
Between group ANOVA model: Model terms – Table 14.2.4.2; Model estimates – Table 
14.2.4.3) and plotted as treatment estimate differences (Figure 14.2.4.2, Template 2). 
 
Repeated measured mixed model (RMMM) – Within Group: Model terms – Table 14.2.4.4 
and Model Estimates Table 14.2.4.5 and Between group also in Tables 14.2.4.4 and 14.2.4.5, 
respectively, together with corresponding plots (Figures 14.2.4.3, Templates 3 and 4, 
respectively). 
 
On completion of the two Part 1 cohorts, the pathway engagement plasma biomarkers will be 
evaluated and this will inform a decision about which should be assayed and statistically 
analysed after Part 2 has been completed. 
 
7.2.3 Efficacy - Functional/Cognition 
Prior to any formal statistical analyses of the cognitive and functional endpoints, each 
question, sub-domain and total score will be descriptively summarised and evaluated.  
Subsequently, formal statistical analysis will only be performed and reported if any notable 
change from baseline is observed in the descriptive summaries. This process will be repeated 
at each analysis stage, i.e. Part 1 PD, Part 1 AD and Part 2 PD database locks. 
 
Depending on the outcome of these original evaluations:  
 
All Cohorts: every patient is evaluated using the MMSE and WAIS-IV: 
MMSE 
The total score and absolute and standardised change from baseline for the MMSE functional 
cognitive measure or any sub-score (after initial evaluation) will be summarised by treatment 
group (Table 14.3.1.1). Scores from the individual questions will also be listed. The results 
from the Within treatment group ANOVA on the change from baseline will also be presented 
in Table 14.3.1.2 (model terms) and Table 14.3.1.3 (model estimates, LS means ±95% CIs) 
and plotted as treatment estimates (Figure 14.3.1.1, Template 1).  
 


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Additionally, the change from baseline differences between the Placebo treatment groups in 
Part 1 and the 80mg patient groups from Part 1 in both the AD and PD cohorts, and 
subsequently the differences between the PD Placebo patients from Part 1 and the 5mg, 10mg, 
20mg and 40mg Parkinson patient groups from Part 2; repeating the PD Placebo versus PD 
80mg comparison will be analysed. The results from these Between treatment group 
ANOVAs on the change from baseline will also be presented in Table 14.3.1.2 (model terms) 
and Table 14.3.1.3 (model estimates, LS means ±95% CIs differences) and plotted as 
treatment estimate differences (Figure 14.3.1.2, Template 2).  
 
WAIS-IV 
This will be summarised (Table 14.3.2.1) and analysed in a similar manner to the MMSE with 
results from the Within treatment group ANOVA on the change from baseline being 
presented in Table 14.3.2.2 (model terms) and Table 14.3.2.3 (model estimates, LS means 
±95% CIs) and plotted as treatment estimates (Figure 14.3.2.1, Template 1). 
 
Additionally, the change from baseline differences between the Placebo treatment groups in 
Part 1 and the 80mg patient groups from Part 1 in both the AD and PD cohorts, and 
subsequently the differences between the PD Placebo patients from Part 1 and the 5mg, 10mg, 
20mg and 40mg Parkinson patient groups from Part 2; repeating the PD Placebo versus PD 
80mg comparison will be analysed. The results from these Between treatment group 
ANOVAs on the change from baseline will also be presented in Table 14.3.2.2 (model terms) 
and Table 14.3.2.3 (model estimates, LS means ±95% CIs differences) and plotted as 
treatment estimate differences (Figure 14.3.2.2, Template 2).  
 
Parkinson Patients – Parts 1 and 2 
MDS-UPDRS 
The MDS-UPDRS total score and four individual domain scores (Non Motor Aspects of 
Experiences of Daily Living; Motor Aspects of Experiences of Daily Living; Motor 
Examination; Motor Complications) or any sub-score (after initial evaluation) will be 
summarised (Tables 14.3.3.1) and analysed in a similar manner to the MMSE. The results 
from the Within treatment group ANOVA on the change from baseline and Between 
treatment group ANOVA differences (Part 1: Placebo vs 80mg; Part 2: Placebo vs 5mg, 
10mg, 20mg, 40mg and 80mg) will be presented in Tables 14.3.3.2 (model terms) and Tables 
14.3.3.3 (model estimates, LS means ±95% CIs) and plotted as treatment estimates (Figures 
14.3.3.1 (Template 1) and 14.3.3.2 (Template 2), for Within and Between estimates, 
respectively). 
 
Furthermore, the Hoehn and Yahr scores will be categorically summarised by treatment group 
and visit in both Parts 1 and 2 (Table 14.3.3.4). 
 
Alzheimer Patients – Part 1 only 
ADAS-cog 
The ADAS-cog 14, 11, 6, 3 total score or any sub-score (after initial evaluation) will be 
summarised (Table 14.3.4.1) and analysed in a similar manner to the MMSE with results from 
the Within treatment group ANOVA on the change from baseline and change from baseline 
Between treatment group ANOVA difference between Placebo and 80mg AD patients being 
presented in Table 14.3.4.2 (model terms) and Table 14.3.4.3 (model estimates, LS means 
±95% CIs) and plotted as treatment estimates (Figures 14.3.4.1 (Template 1) and 14.3.4.2 
(Template 2), respectively). 
 
 


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CDR 
The CDR scale will be categorically summarised by item (Memory, orientation, judgement 
and problem solving, community affairs, home and hobbies, and personal care) and overall 
Global summary score by visit (Table 14.3.5). Additionally, the sum of boxes score will also 
be summarised and analysed following initial evaluation as described for the ADAS-cog 
(Tables 14.3.5.1, 14.3.5.2 and 14.3.5.3, for descriptive summaries, model and treatment 
estimates, respectively, together with corresponding Figures 14.3.5.1 (Template 1) and 
14.3.5.2 (Template 2)).   
 
7.3 Safety. 
For consistency with the pharmacodynamic and cognitive/functional summaries and analyses 
and for comparative purposes, safety data from the Placebo and 80mg Part 1 PD patients will 
be combined with the data collected from the PD patients in Part 2. Consequently, all safety 
summaries will include: the Placebo and 80mg PD patients for the Part 1 PD cohort analysis 
report; Placebo and 80mg AD patients for the Part 1 AD cohort analysis report; and, for the 
Part 2 analysis report, Placebo and 80mg PD patients from Part 1 together with the 5mg, 
10mg, 20mg and 40mg PD patients from Part 2. 
 
7.3.1.1 Adverse Event Overview. 
For each cohort, all adverse events and serious adverse events will be listed by treatment 
group and patient (Listing 16.2.1.1). 
 
For each cohort and treatment group, an overview summary will display the frequency, 
number and percentage of patients, with any adverse event, pre-treatment sign and symptom 
(PTSS) and treatment emergent adverse events (TEAEs). Additionally, the table will include 
the number of TEAEs classed by CTCAE Grade (mild, moderate, severe, life threatening, 
death), relationship to drug (unrelated, possibly related, related), by severity evaluation (mild, 
moderate, severe), and overall numbers related to a study procedure, AEs leading to 
discontinuation of study IMP, SAEs, and death (Table 14.4.1.1). 
  
7.3.1.2 Adverse Events (AEs) and Serious Adverse Events (SAEs). 
For each cohort and treatment group, the number and percentage of patients with PTSS will 
be summarised by SOC and PT (Table 14.4.1.2). Similar tables will be presented for TEAEs 
(Table 14.4.1.3), Related TEAEs (14.4.1.4), TEAE and CTCAE grade (14.4.1.5.1 – 
14.4.1.5.5), SAEs (14.4.1.6, TEAEs leading to discontinuation of study IMP(14.4.1.7) and 
TEAEs leading to death (14.4.1.8).  
 
7.3.2 Laboratory Data. 
For each cohort and treatment, laboratory data will be presented as normal summaries, actual 
values and change from baseline according to profile (Biochemistry: Tables 14.4.2.1.1 – 
14.4.2.1.14; Haematology: Tables 14.4.2.2.1 – 14.4.2.2.10; Coagulation: Table 14.4.2.3; 
Urinalysis: Tables 14.4.2.4). The data will also be presented as Categorical (‘Not Done’, 
‘Abnormal NCS’, ‘Abnormal CS’ and ‘Normal’) summaries by Treatment group according to 
profile (biochemistry, haematology and urinalysis) and visit as Tables 14.4.2.5. Urinalysis 
variables will also be presented as Categorical summaries (according to type) by Treatment 
group and visit as Tables 14.4.2.6.1 – 14.4.2.6.20. Scatter plots of pre-treatment vs. post-
treatment values for all haematology and clinical chemistry variables will be displayed 
(Figures 14.4.2, Template 5). 
 
 


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7.3.3 Vital Signs  
For each cohort, vital sign data and the change from baseline will be presented as numerical 
summaries by treatment group overall and visit, separated into individual test (Weight, BMI, 
Temperature, Systolic BP, Diastolic BP, Heart Rate (Pulse Rate), and Respiratory Rate Tables 
14.4.3.1 – 14.4.3.7). 
  
7.3.4 ECG  
For each cohort, ECG data will be presented as numeric summaries (PR Interval, RR Interval, 
QRS Complex, QT Interval, and QTC Interval Measurement by Treatment group and overall 
according to visit (Tables 14.4.4.1 – 14.4.4.5). The ECG data will also be presented 
categorically overall (‘Not Done’, ‘Abnormal NCS’, ‘Abnormal CS’ and ‘Normal’) summary 
separately by cohort by Treatment group and overall according to visit (Tables 14.4.4.6).  
 
7.3.5 Columbia suicide rating scale (C-SSRS) 
For each cohort, the scores from each category of the C-SSRS will be summarised by 
treatment group and visit if any response is recorded as ‘Yes’ (Table 14.4.5). 
 
7.3.6 Physical Examination and Neurological Examination. 
For each cohort, the results of the physical and neurological examinations will be summarised 
by treatment group, overall and by visit (Table 14.4.6 and 14.4.7, respectively).  
 
7.4 Pharmacokinetics 
All PK statistical analysis will be carried out after the completion of Part 2. 
 
For each cohort, the plasma concentration data and derived pharmacokinetic parameters of 
Posiphen will be listed by treatment group, patient and time point. 
 
For each cohort, the actual values and derived pharmacokinetic parameters will be 
summarised using descriptive summaries (arithmetic mean, standard deviation, coefficient of 
variation, mean, median, minimum, maximum and number of observations) for each 
treatment dose group. If appropriate, geometric means will be displayed in addition to the 
arithmetic mean (Actual drug concentrations: Table 14.5.1; and, Parameter summaries: Table 
14.5.2). Supporting plots will display drug concentration over the 6-hour confinement period 
by treatment group (Figures 14.5.1, Template 6), together with parameter summary Geometric 
LS Means +/- 90%CI (Figures 14.5.2, Template 7). 
 
For Part 2, the statistical analysis of the Differences between doses for all Parkinson patients 
(including 5mg, 10mg, 20mg and 40mg from Part 2, and 80mg from Part 1) and Dose 
proportionality of both Cmax and AUC parameters will include details of the model applied 
with Mean square error, Degrees of freedom, F statistic and resulting p-value for each factor 
or model parameter. For the dose difference analysis, all dose group estimates will be 
presented LS means with 95% confidence intervals after back transformation of the log values 
and all pairwise comparisons between doses will be presented as ratios of LS means (%) with 
95% confidence intervals (Table 14.5.3). These will also be plotted as shown in Section 6.4.2 
(Figure 14.5.3). 
 
 
 
 


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8. References. 
 
ICHE9 “Statistical Principles For Clinical Trials” February 1998. 
ICHE8 “General Considerations For Clinical Trials” July 1997. 
 
 
 
9. Finalisation of Statistical Analysis Plan. 
 
The statistical analysis plan, and all relevant table shells will be finalised before the database 
lock of the Part 1, PD patient cohort. 

Prot_000.pdf: 
NCT04524351 


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CAUTION: Investigational New Drug 
Limited by Federal (or United States) Law to Investigational Use 
----------------------------------------------- 
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 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.  
 


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PRINCIPAL INVESTIGATOR SIGNATURE PAGE 
 
I, the undersigned, have reviewed this protocol and I agree to conduct this protocol in accordance 
with the ethical principles set forth in the Declaration of Helsinki, the Guideline for Good 
Clinical Practice (ICH E6), the U.S. Code of Federal Regulations (CFR) governing the protection 
of human subjects (21CFR§50), Institutional Review Boards (21CFR§56) and the requirements 
for conducting clinical investigations (21CFR§312), and all applicable local, state and federal 
government regulations and laws. 
 
SIGNATURE 
 
_______________________________________  
___________________ 
Name:  
 
 
 
 
 
Date  
 
Institution:_________________________________________________________ 
 
 


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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-Cog 
Alzheimer’s Disease Assessment Scale – Cognitive 
ADL 
Activities of Daily Living 
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 
DS  
Down Syndrome 
DSMB 
Data Safety Monitoring Board 
DSM 
Diagnostic and Statistical Manual of Mental Disorders 
ECG 
Electrocardiogram 
eCRF 
Electronic Case Report Form 


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FCR  
Fractional clearance rate 
FDA 
Food and Drug Administration 
FSR 
Fractional Synthesis Rate 
FWA 
Federal wide Assurance 
GCP 
Good Clinical Practice 
GI 
Gastrointestinal 
GRAS  
Generally recognized as safe  
GWAS 
Genome Wide Association Studies 
HEK 293 
Human embryonic kidney cells 
HEENT 
Head/ears/eyes/nose/throat 
hERG 
Human ether-a-go-go related gene  
HIPAA 
Health Insurance Portability and Accountability Act 
HPMC 
Hydroxypropyl methylcellulose  
H&Y 
Hoehn and Yahr 
ICH 
International Conference on Harmonization 
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 
MDS-UPDRS 
MDS-United Parkinson’s Disease Rating Scale 
MMSE 
Mini-Mental State Examination 
MRI 
Magnetic Resonance Imaging 
mRNA 
Messenger Ribonucleic Acid 
NIH 
National Institutes of Health 
NOAEL 
No observed adverse effect level 
NPH 
Normal Pressure Hydrocephalus 
OHRP 
Office for Human Research Protections 
p-tau  
Phosphorylated tau 
PD 
Parkinson’s disease 
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 


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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 
TPN 
Total Parenteral Nutrition 
WAIS-IV 
Weschler Adult Intelligence Scales, 4th edition 
 
 


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Table of Contents 
1 
PROTOCOL SYNOPSIS ............................................................................................... 11 
2 
INTRODUCTION........................................................................................................... 14 
2.1 
Objectives ..................................................................................................................... 15 
2.2 
Endpoints ...................................................................................................................... 16 
3 
PRELIMINARY STUDIES ........................................................................................... 18 
3.1 
Summary of Non-Clinical Findings .............................................................................. 18 
3.2 
Summary of Clinical Findings ...................................................................................... 18 
4 
BACKGROUND AND STUDY OVERVIEW ............................................................. 20 
5 
POTENTIAL RISKS AND BENEFITS OF INVESTIGATIONAL PRODUCT AND 
STUDY PROCEDURES ............................................................................................................ 23 
5.1 
Risks and Benefits Associated with Posiphen or Placebo ............................................ 23 
5.2 
Risk/Benefit Associated with Blood Collections .......................................................... 23 
5.3 
Risk/Benefit Associated with CSF Sampling ............................................................... 23 
6 
SAMPLE SIZE AND STATISTICAL PLAN .............................................................. 24 
6.1 
Randomization .............................................................................................................. 24 
6.2 
Power and Sample Size Determination......................................................................... 24 
6.3 
Safety and Tolerability Analysis ................................................................................... 27 
6.4 
PK Analysis .................................................................................................................. 28 
6.5 
Exploratory Pharmacodynamic and Efficacy Analysis ................................................ 28 
6.6 
Criteria for the termination of the trial .......................................................................... 28 
6.7 
Interim Analysis ............................................................................................................ 28 
7 
STUDY DRUG AND CONCOMITANT MEDICATIONS ........................................ 30 
7.1 
Name and Description of IP and Comparator ............................................................... 30 
7.2 
Dosage........................................................................................................................... 30 


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7.3 
Packaging/Dispensing/Labeling ................................................................................... 30 
7.4 
Storage .......................................................................................................................... 30 
7.5 
Drug Accountability...................................................................................................... 30 
7.6 
Compliance ................................................................................................................... 32 
7.7 
Breaking the Blind ........................................................................................................ 32 
7.8 
Overdose ....................................................................................................................... 32 
7.9 
Concomitant Medications ............................................................................................. 33 
7.9.1 
Prohibited Concomitant Medications ....................................................... 33 
7.9.2 
Permitted Concomitant Medications ........................................................ 33 
8 
STUDY POPULATION ................................................................................................. 34 
8.1 
Inclusion Criteria .......................................................................................................... 35 
8.2 
Exclusion Criteria ......................................................................................................... 36 
9 
DESCRIPTION OF STUDY VISITS ........................................................................... 38 
9.1 
Screening Visit .............................................................................................................. 38 
9.2 
Baseline Visit ................................................................................................................ 38 
9.3 
Day 14 Telephone Contact............................................................................................ 39 
9.4 
Confinement Visit ......................................................................................................... 39 
9.4.1 
Pre-Confinement Telephone Contact ...................................................... 39 
9.4.2 
Confinement Visit ..................................................................................... 39 
9.4.3 
Post-Confinement 24-Hour Phone Follow-up .......................................... 42 
10 
EARLY TREATMENT/STUDY DISCONTINUATION ........................................... 43 
10.1 
Reasons for Early Discontinuation ............................................................................... 43 
11 
STUDY-SPECIFIC INSTRUMENTS ........................................................................... 44 
12 
STUDY-SPECIFIC PROCEDURES ............................................................................ 44 
12.1 
Safety Assessments ....................................................................................................... 44 


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12.1.1 
Physical and Neurological Examination ................................................... 44 
12.1.2 
Electrocardiogram (ECG) ......................................................................... 44 
12.1.3 
Clinical Laboratory Evaluations ................................................................ 44 
12.2 
Biofluids ........................................................................................................................ 45 
12.2.1 
CSF for Biomarkers ................................................................................. 45 
12.2.2 
APOE Genotype and other DNA Markers ................................................ 46 
12.2.3 
Plasma for Biomarkers and Pharmacokinetics ........................................ 46 
12.2.4 
Blood Samples for Exosomes .................................................................. 47 
13 
PERSONNEL REQUIREMENTS ................................................................................ 47 
14 
ADVERSE EVENTS (AES) ........................................................................................... 47 
14.1 
Definition ...................................................................................................................... 47 
14.2 
Following Up on AEs ................................................................................................... 48 
15 
SERIOUS ADVERSE EVENTS (SAE) ........................................................................ 48 
15.1 
Definition ...................................................................................................................... 48 
15.2 
Reporting SAEs ............................................................................................................ 48 
16 
DATA AND SAFETY MONITORING BOARD (DSMB) ......................................... 49 
17 
RECORDING AND COLLECTION OF DATA ......................................................... 49 
17.1 
Electronic Case Report Form (eCRF) ........................................................................... 49 
17.2 
Study Files and Source Documents .............................................................................. 50 
18 
ETHICS AND REGULATORY CONSIDERATIONS ............................................... 50 
18.1 
Good Clinical Practice .................................................................................................. 50 
18.2 
Institutional Review Board (IRB) ................................................................................. 51 
18.3 
Informed Consent and Health Insurance and Portability and Accountability Act 
(HIPAA) Compliance ............................................................................................................... 51 
19 
STUDY MONITORING ................................................................................................ 52 
20 
AUDIT .............................................................................................................................. 52 


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21 
RECORD RETENTION ................................................................................................ 52 
22 
LITERATURE CITED .................................................................................................. 54 
23 
APPENDIX 1: SCHEDULE OF EVENTS ................................................................... 56 
24 
APPENDIX 2: SEVERITY EVALUATION OF CHOLINERGIC ADVERSE 
EVENTS (ADVERSE EVENTS OF SPECIAL INTEREST) ................................................. 58 
 
 


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1 
PROTOCOL SYNOPSIS 
PROTOCOL TITLE 
A Multicenter, Randomized, Double-Blind, Placebo-Controlled, Dose-
Finding Study to Evaluate the Safety, Tolerability, Pharmacokinetic and 
Pharmacodynamic Effects of Posiphen® in Subjects with Early 
Alzheimer’s Disease (AD) or Early Parkinson’s Disease (PD) 
STUDY DESIGN 
This study will be conducted in two parts.   
Part 1 will evaluate the safety, tolerability, and pharmacokinetics of 
Posiphen in 14 Early AD and 14 Early PD patients at the following dose: 
• 
80 mg QD 
Each group will include 14 Early AD patients, randomly assigned, 10 to 
Posiphen and 4 to placebo and 14 Early PD patients randomly assigned, 
10 to Posiphen and 4 to placebo. 
Part 1 of the study will also include exploratory measures of 
pharmacodynamics and efficacy of Posiphen with target engagement, 
pathway engagement and functional/cognitive measures, including the 
degree of memory loss and cognitive function in AD patients as well as 
the degree of motor impairment and non-motor symptoms and quality of 
life in PD patients. 
Subjects who drop out during the initial 3-week at-home treatment period 
will be discontinued and replaced by new subjects. If 4 subjects are 
discontinued due to adverse events, no additional subjects will be 
recruited until a safety review has been conducted. 
After Part 1 has been completed study data will be locked to allow a full 
analysis of biomarker data to sensitize analysis plans in Part 2.  
Part 2 will evaluate the safety, tolerability, and pharmacokinetics of 
Posiphen in 40 patients with early PD, randomly assigned to the 
following dose levels, 10 patients per dose level:  
• 
5 mg QD 
• 
10 mg QD 
• 
20 mg QD 
• 
40 mg QD 
Part 2 of the study will also include exploratory measures of 
pharmacodynamics and efficacy of Posiphen in Early PD patients with 
target engagement, pathway engagement and functional/cognitive 
measures, including the degree of motor impairment as well as other non-
motor symptoms and quality of life in PD patients. 


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Subjects who drop out during the initial 3-week at-home treatment period 
will be discontinued and replaced by new subjects. If 4 subjects are 
discontinued due to adverse events, no additional subjects will be 
recruited until a safety review has been conducted. 
DURATION OF STUDY 
PARTICIPATION 
Each subject will have up to a 42-day screening period, a baseline visit 
including a lumbar puncture (LP), followed by 25±2 days of treatment at 
home, followed by an in clinic visit that includes treatment and a 6-hour 
LP procedure. 
SUMMARY OF 
INVESTIGATIONAL 
PRODUCT 
Part 1: Posiphen/placebo 80 mg QD, taken orally for 25±2 days by Early 
AD and PD patients. 
 
Part 2: Posiphen at 5, 10, 20, or 40 mg QD, taken orally for 25±2 days by 
Early PD patients.  
SUMMARY OF KEY 
ELIGIBILITY CRITERIA:  
FOR AD 
• 
Diagnosis of early AD  
• 
Age 45 or older 
• 
MMSE 18-28 
• 
Clinical Dementia Rating (CDR) of 0.5 or 1.0  
SUMMARY OF KEY 
ELIGIBILITY CRITERIA:  
FOR PD 
• 
Diagnosis of early PD  
• 
Age 45 or older 
• 
MMSE 18-30 
• 
Hoehn & Yahr ≤ 4 
PRIMARY OUTCOME 
MEASURES 
Aim 1.  Safety and Tolerability 
Aim 2.  PK of Posiphen in Plasma  
EXPLORATORY  
MEASURES 
TARGET ENGAGEMENT 
Levels of neurotoxic proteins in blood, plasma and CSF: 
• 
aSYN and Aβ monomers and dimers, and infective oligomers  
• 
Aβ38, Aβ40, Aβ42  
• 
soluble Amyloid Precursor Proteins:  sAPPα and sAPPβ 
• 
T-Tau and P-Tau proteins 
PATHWAY ENGAGEMENT 
Levels of proteins involved in the toxic cascade leading to nerve cell 
death in blood, plasma and CSF 


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• 
Neurotransmitters 
• 
Neurotrophic factors 
• 
Inflammatory factors 
• 
Synaptic factors 
• 
Control proteins that should not change 
FUNCTIONAL/COGNITIVE MEASURES 
PD: MMSE, WAIS-IV, MDS-UPDRS 
AD: MMSE, WAIS-IV, ADAS-Cog 14, CDR  
 
 


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2 
INTRODUCTION 
Currently, there is no treatment available to stop or reverse the progression of Alzheimer’s 
(AD) and Parkinson’s disease (PD). Neurodegenerative diseases such as AD and PD share 
many common characteristics, including the central role of neurotoxic aggregating proteins 
in their pathogenesis. Amyloid β (Aβ) and tau aggregates (senile plaques and neurofibrillary 
tangles, respectively) have been traditionally associated with AD, while α-synuclein (αSYN) 
aggregates (Lewy bodies) have been associated with PD.  However, it is becoming 
increasingly clear that all these proteins are involved in both diseases and that aggregation 
of one can lead to accumulation of another. Furthermore, in several studies of brains from 
older patients, a high percentage of all AD brains present mixed pathologies, such as Lewy 
body disease. Several clinical trials targeting just one (often Aβ) of the neurotoxic 
aggregating proteins have failed. Finally, other fragments of Amyloid β Precursor Protein 
(APP) have been implicated in AD pathology. Collectively, these facts point to the need for 
development of combination therapies that target multiple neurotoxic aggregating proteins 
simultaneously, if we are to have a good chance of at least halting disease progression. 
Posiphen has a unique mechanism of action, in that it inhibits the translation and, therefore, 
reduces the levels of several neurotoxic aggregating proteins both in vitro and in vivo, including 
α-Synuclein (αSYN), Amyloid Precursor Protein (APP), its fragments, and tau. All three proteins 
have been implicated in the pathogenesis of PD.  Furthermore, at older ages, there is a high 
incidence of patients with mixed pathologies, such as PD dementia.  Therefore, it is reasonable to 
hypothesize that inhibiting expression of all three proteins should lead to a better efficacy 
outcome in PD patients than inhibiting just one. 
Posiphen’s safety has been established in three Phase I clinical studies by Maccecchini et al. 
Importantly, Posiphen normalized levels of αSYN, APP, and tau in the cerebrospinal fluid (CSF) 
of mildly cognitively impaired (MCI) patients at a dose of 4x60 mg/day (Protocol QR12001). 
Preclinical data proves Posiphen’s efficacy in restoring colonic motility in a human SNCAA53T tg 
mouse model of early PD, in restoring memory and learning in an APP/PS1 transgenic (tg) 
mouse model of AD, restoring axonal transport in DS trisomic mice, preserving memory and 
learning in traumatic brain injury rats and preserving the retina in acute glaucoma. 
αSYN, APP, and tau contribute to the progression of AD and PD in similar ways: they impair 
axonal transport and lower neurotransmitter and neurotrophic factor release, they cause 
inflammation, they form aggregates, and, finally, they lead to nerve cell death. In our in vitro and 
in vivo preclinical studies, Posiphen has normalized all those actions. 
We have conducted a proof-of-concept study in mildly cognitive impaired patients at 4x60 
mg/day (Protocol QR12001) and found that Posiphen lowers levels of neurotoxic proteins and of 
inflammatory markers.  Presently, we are testing Posiphen in the DISCOVER study at 1x60, 
2x60 and 3x60 mg/day for up to 25 days to see if it changes the synthesis and degradation 


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kinetics of Aβ in a stable isotope labelling kinetic (SILK) study.  The DISCOVER study is on-
going through the ADCS and is planned for 24 patients to be enrolled; currently, approximately 
half of the study has been enrolled, and there have been no severe AEs or SAEs attributed to 
Posiphen reported.  
From our animal data, we found that Posiphen works in more than Alzheimer animals, because it 
inhibits more than one neurotoxic protein.  We found that in transgenic AD and PD animals it 
lowers levels of neurotoxic proteins and restores the affected function.  Therefore, we want to 
test Posiphen in a comparison study of AD and PD patients and measure not just target 
engagement, but also pathway engagement.  
From our animal data, we also found that the doses used to show an effect on neurotoxic proteins 
are very high and that we can see an effect of Posiphen at much lower doses. Therefore, we are 
testing Posiphen in a randomized dose de-escalation study to see what the lowest effective dose 
is in humans. 
In this study, we want to find out if the animal Posiphen data can be replicated in humans and 
show target and pathway engagement in CSF.  Target engagement means that Posiphen would 
lower and normalize the levels of neurotoxic proteins in CSF.  Pathway engagement means that 
Posiphen would increase levels of neurotransmitters, neurotrophic factors, and lower levels of 
inflammatory and neurodegeneration markers in both AD and PD patients.  By examining all 
these factors in early AD and PD patients, we will specify the biomarkers that change with 
Posiphen treatment, which will be used in later phase studies.  Additionally, we will assess 
function and cognition, so we can make sure that Posiphen has no deleterious effect and 
potentially identify any improvement.  Collectively, these endpoints will help assess the efficacy 
of Posiphen and drive it to late phase clinical studies. 
2.1 
Objectives 
Study objectives include assessing safety, tolerability, and pharmacokinetics of Posiphen as well 
as exploring pharmacodynamics and efficacy in Early AD and Early PD patients with target 
engagement, pathway engagement and functional/cognitive measures being the focus of the 
exploratory analyses.  This study will be conducted in two parts as follows.   
Part 1  
Primary objectives are to assess the safety, tolerability, and pharmacokinetics of Posiphen in 14 
Early AD and 14 Early PD patients at a dose of 80 mg QD for 25±2 days. 
Each group will include 14 Early AD patients, randomly assigned, 10 to Posiphen and 4 to 
placebo and 14 Early PD patients randomly assigned, 10 to Posiphen and 4 to placebo. 
Exploratory objectives will include investigating the pharmacodynamics and efficacy of 
Posiphen as well as the relationship between them.  Efficacy assessments will focus on the 
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• degree of memory loss and cognitive function in AD patients 
• degree of motor impairment and non-motor symptoms and quality of life in PD patients. 
Part 2  
Primary objectives are to assess the safety tolerability, and pharmacokinetics of Posiphen in 40 
patients with early PD, 10 patients randomly assigned to each of the following dose levels, given 
for 25±2 days:  
• 5 mg QD 
• 10 mg QD 
• 20 mg QD 
• 40 mg QD 
Exploratory objectives will include investigating the pharmacodynamics and efficacy of 
Posiphen as well as the relationship between them.  Efficacy assessments will focus on the 
degree of motor impairment in PD as well as other non-motor symptoms and quality of life. 
2.2 
Endpoints 
 
SAFETY AND TOLERABILITY 
• adverse events 
• physical examinations  
• vital signs  
• 12-lead ECGs 
• clinical laboratory values 
 
PHARMACOKINETICS 
PK parameters will minimally be determined, as data permit: Area under the curve (AUC), 
Cmax, Tmax, t½, and CL. 
 
Additionally, the following Exploratory Objectives will be evaluated: 


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TARGET ENGAGEMENT 
Levels of neurotoxic proteins in blood, plasma and CSF 
• aSYN and Aβ monomers and dimers, and infective oligomers 
• Aβ38,40,42 
• soluble Amyloid Precursor Proteins:  sAPPα and sAPPβ 
• T-tau and P-tau protein  
 
PATHWAY ENGAGEMENT 
Levels of proteins involved in the toxic cascade leading to nerve cell death in blood, plasma and CSF 
• neurotransmitters 
• neurotrophic factors 
• Inflammatory factors 
• Synaptic factors 
• Control proteins that should not change 
 
FUNCTIONAL/COGNITIVE MEASURES 
PD: MMSE, WAIS-IV, MDS-UPDRS 
AD: MMSE, WAIS-IV, ADAS-Cog 14, CDR 
 
 


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3 
PRELIMINARY STUDIES 
 
The following is a brief summary of the available information on Posiphen. Detailed information 
can be found in the current Posiphen Clinical Investigational Brochure. 
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 as well as in long-term potentiation at 
brain and plasma concentrations that are 10 times lower than originally published. These plasma 
levels are attainable with low oral dosing in humans (Teich 2018). In alpha-synuclein (αSYN) 
transgenic PD mice, Posiphen restored gut motility to normal and lowered aSYN in the brain and 
gut of the tg PD mice. Again, the efficacious levels were 10 times lower than originally 
published (Kuo 2019). In DS trisomic mice Posiphen fully restored axonal transport in vitro and 
in vivo (Mobley Lab, UCSD, 2019; manuscript submitted for publication). In summary, Posiphen 
is a translational inhibitor of APP and aSYN and fully restores function at doses that are very 
low and very safe. 
Posiphen was not mutagenic or clastogenic as assessed by in vitro assays.  
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 (IB 2019). The first was a single 
ascending dose (SAD) study in healthy volunteers; the second was a multiple ascending dose 
(MAD) study in healthy volunteers, and the third one was a pharmacokinetic 


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(PK)/pharmacodynamic study of CSF obtained from mild cognitively impaired participants 
(Protocol QR12001).  
In the MAD 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 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 (Protocol QR12001), the PK of Posiphen was 
measured after 10 days of administration (4x60 mg) over 6 hours in CSF and plasma of the AD 
participants. The pharmacodynamics of a number of biomarkers was compared for 6 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 patients treated with 
cholinesterase inhibitors appear to reflect the cholinomimetic properties of molecules in this 
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 sAPP, tau, 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. 
 
 


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4 
BACKGROUND AND STUDY OVERVIEW 
4.1 
Background 
A major pathological hallmark of AD is the appearance of plaques and tangles in the brain and of 
PD the appearance in the gut and in the brain of aSYN aggregates that form Lewy bodies. In 
both diseases these aggregates are thought to occur one or two decades prior to overt symptom 
development. Recent research has shown that soluble forms of phosphorylated tau (p-tau) and 
Aβ may also contribute to neuronal loss and that PD as well as AD carry a high load of 
inflammation and microglia activating factors that contribute to neurodegeneration. 
For small molecules that target biomarker levels, 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 CNS effects. 
Repeated sampling through a lumbar CSF catheter for 6 hours can mirror image, what happens in 
the brain as to the levels of aSYN monomers and dimers, Aβ monomers and dimers as well as 
other biomarkers and help guide the development of drugs targeting neurotoxic aggregating 
proteins.  
Posiphen is the (+) enantiomer of phenserine, an AChE inhibitor which had been tested in 
several AD studies without significant evidence of efficacy. 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 has about 200 times more AChEI activity than 
Posiphen’s metabolite N1-bisnorposiphen. Posiphen absorption is affected by food; however, at 
steady state there is no effect on the PK analyses results. 
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 acts at the 5’UTR of the aSYN and APP mRNA and lowers their protein expression 
levels in animal models; it also decreased aSYN and sAPP levels in human CSF. Our data 
suggests that these effects are achieved via the same mechanism: the 5’UTRs of these mRNAs 
form a complex with iron regulatory protein 1 and Posiphen stabilizes the complex, thereby 
inhibiting the translation of these mRNAs. 
As Posiphen inhibits the synthesis of aSYN and APP, as well as other neurotoxic aggregating 
proteins, it might have a broader spectrum of activity than PD and AD. By protecting neurons 
from dying it has a disease-modifying effect in PD, AD as well as other neurodegenerative 
disorders.  
4.2 
Study Overview 


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Study population: The study will be conducted in patients with early AD or early PD. 
Dose selection: In Part 1 Posiphen will be tested at 80 mg QD and in Part 2, Posiphen will be 
tested at 5, 10, 20, 40 mg QD, for 25±2 days.   
Whereas, the plasma t½ of Posiphen has been determined to be 5 hours, the CNS t½ has been 
measured to be over 12 hours, allowing that once daily dosing is feasible.  Previous studies 
indicated that likely cholinergic adverse events are mostly driven by Cmax and occur at 160mg 
QD.  Since our maximum dose is 80 mg QD, we do not expect any cholinomimetic effects. 
Study Design:  Part 1 is a study with 14 Early AD and 14 Early PD patients who are randomized 
to 80 mg of Posiphen or placebo.  They will undergo a Screening Visit, and if they provide 
informed consent and are considered eligible per the inclusion and exclusion criteria, will 
proceed to participate in the treatment period.  Period 1 consists of first-time dosing in clinic 
with administration of 80 mg of Posiphen or Placebo.  Period 2 consists of an at home dosing 
period of 25±2 days, with daily administration of 80 mg of Posiphen or Placebo.  Period 3 will 
be comprised of a stay at the clinical research unit where the subject will undergo study 
procedures that include safety assessments (AE and concomitant medication monitoring, 12-lead 
ECGs, clinical laboratory testing, vital signs assessments, and physical examinations), the last 
dose of Posiphen or Placebo, and 6 hours of PK blood and CSF sampling.  At the end of 
blood/CSF sampling, the subjects will need to stay for a minimum of 1 hour of observation but 
may stay if necessary for observation until the following day (e.g., if the subject has blood/CSF 
sampling on Day 25, he/she may stay for observation until Day 26).   After all end-of-study 
procedures are complete, the subject will be discharged to home.  A 24-hour follow-up call will 
occur to assess the participants current condition and if there are any additional adverse events to 
report.   
After completion of Part 1 of the study, the plasma and CSF samples will be analyzed for the 
biomarkers to determine if changes are needed to the biomarkers to be measured in Part 2.  Since 
the conduct of the study in Part 2 will be identical to the conduct of the study in Part 1, 
recruitment will continue uninterrupted.  The only potential change between Part 1 and Part 2 are 
the biomarkers to be measured. 
Part 2 is a study with 40 Early PD patients, 10 patients each who are randomized to one of 4 
treatment conditions of Posiphen (5 mg, 10 mg, 20 mg, or 40 mg). They will undergo a 
Screening Visit, and if they provide informed consent and are considered eligible per the 
inclusion and exclusion criteria, will proceed to participate in the treatment period.  Period 1 
consists of first-time dosing in clinic with administration of 5, 10, 20, or 40mg of Posiphen.  
Period 2 consists of an at home dosing period of 25±2 days, with daily administration of 5, 10, 
20, or 40mg of Posiphen.  Period 3 will be comprised of a stay at the clinical research unit where 
the subject will undergo study procedures that include safety assessments (AE and concomitant 
medication monitoring, 12-lead ECGs, clinical laboratory testing, vital signs assessments, and 
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sampling.  At the end of blood/CSF sampling, the subject  will need to stay for a minimum of 1 
hour of observation but may stay if necessary for observation until the following day (e.g., if the 
subject has blood/CSF sampling on Day 25, he/she may stay for observation until Day 26).  After 
all end-of-study procedures are complete, the subject will be discharged to home.  A 24-hour 
follow-up call will occur to assess the participant’s current condition and if there are any 
additional adverse events to report.   
For both parts, subjects who drop out during the initial 3-week at home treatment period will be 
discontinued and replaced by new subjects.  If 4 subjects are discontinued due to adverse events 
a safety review will be conducted.  
Outcome measures to assess pharmacodynamics: Translational inhibition of the aSYN mRNA 
will result in decreased production of aSYN, which may be reflected by decreased secretion of 
the aSYN monomers and oligomers into the spinal fluid. Similarly, translational inhibition of the 
APP mRNA will result in decreased production of APP and its fragments, which may be 
reflected by decreased secretion of the Aβ monomers and oligomers into the spinal fluid. We 
also will measure secreted sAPPα and sAPPβ and levels of tau and p-tau in CSF. sAPP reflects 
processing of APP by α-secretase to produce sAPPα and β-secretase (BACE) to produce sAPPβ. 
Tau levels are being measured since we found that tau translation may be under the regulation of 
a similar iron-responsible element (IRE) that may also be affected by Posiphen treatment. In fact, 
we have measured in human CSF and in mouse brain that tau and p-tau levels are lowered 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, functional and cognitive assessment measures: Although clinical benefit is unlikely 
due to the short duration of this study, AD and PD specific tests will be performed. It is predicted 
that cognition and function will remain stable over the course of drug exposure; however, this 
will be assessed formally in the clinic.  For both populations, the MMSE (Folstein 1975) will be 
administered as a global measure of cognition and the Coding subtest from the Weschler Adult 
Intelligence Scales, 4th edition (WAIS-IV) will serve as a sensitive measure of CNS dysfunction.  
The subjects with AD will also be administered the ADAS-Cog 14 (Schafer 2012).  The subjects 
with PD will be administered the Hoehn & Yahr (1967) for determination of inclusion into the 
study.  Functional impairment will be evaluated using the Clinical Dementia Rating (CDR) scale 
(Berg1988) for AD and the MDS-UPDRS (Goetz 2008) for those with PD.    


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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 functional and 
cognitive evaluations. 
The clinical investigator must 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 the 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 up to 25 days. 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. 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. 
5.3 
Risk/Benefit Associated with 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, 


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pain at the site of insertion, backache, epidural or subdural bleeding, infection, paralysis and 
potential injury to the nerve root.  
6 
SAMPLE SIZE AND STATISTICAL PLAN 
For Part 1:  A sample size of up to 2x14 subjects (10 randomized to Posiphen and 4 to Placebo 
for AD and PD respectively) is planned.  
For Part 2:  A sample size of up to 40 Early PD patients is planned to be randomized to 
Posiphen, 10 patients each dosed at 5 mg, 10 mg, 20 mg, or 40 mg.   
Definitions of the analysis populations and a detailed description of the analysis rules will be 
presented in a separate Statistical Analysis Plan (SAP). 
6.1 
Randomization 
Subjects who have signed an informed consent and meet screening eligibility requirements will 
be randomly assigned to the active and placebo treatment groups (10 randomized to Posiphen 
and 4 to Placebo for AD and PD, respectively) in Part 1. In Part 2, 40 Early PD patients will be 
randomly assigned, 10 patients each to one of the 4 different dose levels of Posiphen (5, 10, 20 
or 40mg).  
6.2 
Power and Sample Size Determination 
This is an exploratory study to assess safety, tolerability and pharmacokinetics for Early AD and 
Early PD patients in addition to investigating pharmacodynamics and efficacy focusing on target 
engagement, pathway engagement and functional/cognitive measures.  10 patients per dose 
group provides enough data to characterize the PK of Posiphen as well as adequately supporting 
potential dose proportionality analyses.   
 
Due to the sample size computation below, we will state that sAPPα, sAPPβ and aSYN were 
given specific consideration in deciding the need to have 10 Posiphen patients in each treatment 
group given their specific importance in the emergence and progression of AD and PD. The 
statistical background and rational for having 10 Posiphen patients in each treatment group to 
have a meaningful interpretation of sAPPα, sAPPβ and aSYN data are outlined below. 
The statistical evaluation of our previous study in MCI patients with soluble APPα and β levels 
measured in spinal fluid before and after Posiphen administration (Protocol QR12001) can be 
used to power this study (Maccecchini 2012). All assay data collected were analyzed using a 
repeated measure mixed model analysis of variance. The model included Day (Day 11/Day0) as 
a fixed effect, Time (9 time points per person between 0 and 6 hours) as a repeat measure effect 


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and Patient (18 samples per person total) as a random effect. Data are presented as means ±SEs. 
The statistical evaluations were undertaken by Data Magik (Salisbury, UK). 
 
 
Analysis of sAPPα and β using type, day and id as variables shows: 
The results of the 10 days at 8 hours on sAPPα and β is SD = 0.1695.  
Assuming type one error, α=0.05 and type two error, β=0.2, or a power of 0.8, the calculated 
sample range of delta effect sizes was 0.3 to 2 SD. 


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sAPPα 
Using the above data, we can calculate that a power analysis of APPα shows the requirement of 
9 people to see a 36% change in sAPPα levels at a statistical significance level of >0.05
 
sAPPβ 
Using the above data, we can calculate that a power analysis of APPβ shows the requirement of 9 
people to see a 24% change in sAPPβ levels at a statistical significance level of >0.05. We are 
expecting changes in sAPPα and β levels to be between 20 and 50% and for the higher doses an 
n<5 would be adequate to detect those changes.  
aSYN 
However, we do not expect the changes in alpha-synuclein and in neurotransmitters to be that 
pronounced. Therefore, we chose an n=10. If dropouts are to occur, they will be replaced to 
allow the full sample size of 5 x 10 participants to be achieved. 
6.3 
Safety and Tolerability Analysis 
Safety and tolerability will be assessed with physical examinations, vital signs, clinical 
laboratory values, 12-lead ECG, use of concomitant medications, and AE reports. The 
frequencies of adverse events, serious adverse events and laboratory abnormalities between the 
participants across the treatment groups will be compared. The severity of cholinergic adverse 
events will be carefully evaluated taking the criteria in Appendix 2 into consideration (modified 
from the latest version of the Common Terminology Criteria for Adverse Events (CTCAE) ) 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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Subjects who dropout during the initial 3-week at-home treatment period will be discontinued 
and replaced by new subjects. If 4 subjects are discontinued due to adverse events a safety 
review will be conducted.  
6.4 
PK Analysis 
Plasma concentration-time data will be analyzed by non-compartmental methods using SAS 
version 9.4 or greater.  Calculations will be based on the actual sampling times recorded during 
the study.  Since the study is blinded, all subjects’ plasma will be included in the PK analysis.  
From the plasma concentration-time data, the following PK parameters will minimally be 
determined, as data permit: Area under the curve (AUC), Cmax, Tmax, t½, and CL.  Dose-
proportionality may be assessed on Cmax and AUC as appropriate. Serial pre-dose trough 
concentrations Cmax will be assessed for the attainment of steady state. 
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.  In addition, 
geometric means and between-subject coefficients of variation (CV) will be calculated. 
6.5 
Exploratory Pharmacodynamic and Efficacy Analysis 
Pharmacodynamics and efficacy analyses will focus on exploratory endpoints outlining target 
engagement, pathway engagement and functional/cognitive measures being the reference stones 
of the exploratory analyses both for Part 1 and Part 2. 
Analysis will include descriptive summary tables, listings and figures for Part 1 and Part 2 as 
well as combined analyses of dose/treatment groups from Part 1 and Part 2. Statistical 
comparisons will be descriptively performed for PD and efficacy parameters as well as the 
relationships between them using mixed models. 
6.6 
Criteria for the termination of the trial  
The trial may be terminated by the Project Directors and/or the Sponsor based on issues of 
safety, feasibility, and Data Safety Monitoring Board (DSMB) recommendations. 
6.7 
Interim Analysis 
No interim analysis is planned.  However, after completion of Part 1 of the study, the plasma and 
CSF samples will be analyzed unblinded for the biomarkers to determine if changes are needed 
to the biomarkers to be measured in Part 2.  Since the conduct of the study in Part 2 will be 
identical to the conduct of the study in Part 1, recruitment will continue uninterrupted.  The only 
potential change between Part 1 and Part 2 are the biomarkers to be measured. Due to COVID-
19 potentially causing recruitment challenges, at the time of intended analysis, Part 1 data may 
be analyzed should only 1 of the 2 cohorts in Part 1 be complete (e.g., 14 PD Patients, 5 AD 


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Patients). Only the full cohort would be unblinded, any patients not analyzed as part of the Part 1 
analysis will be analyzed when that cohort is fully completed. Full details of all analyses will be 
detailed in the SAP.  
 


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7 
STUDY DRUG AND CONCOMITANT MEDICATIONS 
7.1 
Name and Description of IP and Comparator 
Posiphen will be prepared in 5 mg, 10 mg, 20 mg, 40mg, and 80 mg HPMC hard capsule shells.  
Matching Placebo capsules (non-lactose compound) will be prepared with an inert inactive 
excipient generally recognized as safe for human pharmaceutical use.  
7.2 
Dosage 
The study drug is to be taken orally; one capsule, once a day in the morning, with or without 
food, for 25 days ± 2 days at home (depending on when the confinement visit is scheduled).  
Participants must take 80% of all doses (across 25±2 days).  Bottles will be examined in clinic at 
the pre-confinement visit to assess compliance overall and in the last week.  
7.3 
Packaging/Dispensing/Labeling 
The study drug (Posiphen capsules and placebo capsules) will be manufactured under cGMP, in 
a manner to preserve the blind, i.e., identical color and shape opaque HPMC hard capsule shells 
and packaged 30 capsules per bottle.  
The investigational drug supply will be shipped directly to the clinical sites to dispense to 
participants.  
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 must be stored at room temperature 
(15-30 °C; 59-86 °F), protected from moisture, in a locked, temperature-controlled area with 
restricted 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 


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drug (including partially used bottles) may be destroyed on site per appropriate site SOPS or 
should be shipped back to the Sponsor for destruction.  
 


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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 (25±2 days) depending on the confinement visit date. Site personnel will count capsules 
and evaluate compliance from the bottle 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, 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 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. 
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. 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 


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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., Plavix)see section 12.1.1, other than aspirin up to a dose of 325 mg per day, is 
prohibited. Prospective subjects who in consultation with the prescribing physician 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: 
• tricyclic antidepressants  
• antipsychotics prescribed for any reason, except ≤50 mg quetiapine daily, risperidone 
≤1.5 mg/day, olanzapine ≤5 mg/day, and aripiprazole ≤10 mg/day 
• psychostimulants 
Investigational agents are prohibited 4 weeks prior to entry and for the duration of the trial.  
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 anti-parkinsonian medications at stable doses 
for 4 weeks or greater prior to screening and with cholinesterase inhibitors and/or memantine if 
on a stable dose (FDA approved drug ranges) for at least 12 weeks prior to screening. 
Anticonvulsant medications used for epilepsy or mood stabilization; neuropathic pain 
indications. Dosing must be stable for at least 4 weeks prior to Screening. 
Mood-stabilizing psychotropic agents, including but not limited to lithium. Dosing must be 
stable for at least 4 weeks prior to Screening. 
Use of short acting benzodiazepines and hypnotics for treatment on an as-needed basis for 
insomnia or daily dosing as anxiolytics is permitted but 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.  


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8 
STUDY POPULATION 
Participants in this study must be 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. 
 


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8.1 
Inclusion Criteria 
Subjects must meet the following criteria:  
1. Male or female aged 45 years and over. 
2. Female participants must be of non-childbearing potential or 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. A) AD - CDR = 0.5 or 1.  
 B) PD - Hoehn & Yahr ≤ 4; PD criteria by MDS-UPDRS. 
5. A) AD MMSE score between the range of 18 to 28. 
B) PD MMSE score between the range of 18 to 30. 
6. General cognition and functional performance sufficiently preserved that the subject can 
provide written informed consent. 
7. No evidence of current suicidal ideation or previous suicide attempt in the past month as 
evaluated in the Columbia Suicide Severity Rating Scale. 
8. 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.  
9. Stability of permitted medications prior to screening.  
a. Stable for at least 12 weeks: Cholinesterase inhibitors and/or memantine 
medication  
b. Stable for at least 4 weeks:  
i. Anti-parkinsonian medication 
ii. Anticonvulsant medications used for epilepsy or mood stabilization; 
neuropathic pain indications 
iii. Mood-stabilizing psychotropic agents, including, but not limited to, 
lithium. 


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10. Adequate visual and hearing ability (physical ability to perform all the study 
assessments). 
11. Good general health with no disease expected to interfere with the study.  
12. Subjects previously exposed to Posiphen may be included in the study. 
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 SSRI or SNRI medication at a stable dose is acceptable. 
2. History of a seizure disorder. 
3. Has a history or current evidence of long QT syndrome, Fridericia’s formula corrected 
QT (QTcF) interval ≥ 450ms, or torsades de pointes. 
4. Has bradycardia (<50 bpm) or tachycardia (>100 bpm) on the ECG at screening. 
5. Has uncontrolled Type-1 or Type-2 diabetes . A Subject with HbA1c levels up to 7.5% 
can be enrolled if the investigator believes the subject’s diabetes is under control. 
6. Has clinically significant renal or hepatic impairment. 
7. Has any clinically significant abnormal laboratory values. Subjects with liver function 
tests (aspartate aminotransferase [AST] or alanine aminotransferase [ALT]) greater than 
twice the upper limit of normal will be excluded. 
8. Is at imminent risk of self-harm, based on clinical interview and responses on the C 
SSRS, or of harm to others in the opinion of the Investigators. Subjects must be excluded 
if they report suicidal ideation with intent, with or without a plan or method (e.g. positive 
response to Items 4 or 5 in assessment of suicidal ideation on the C SSRS) in the past 2 
months, or suicidal behavior in the past 6 months. 
9. Has four or more signal hypointensities on T2*-weighted gradient recalled echo magnetic 
resonance sequences that are thought to represent hemosiderin deposits including 
microhemorrhages and superficial siderosis or evidence of acute or sub-acute micro or 
microhemorrhage as noted on the MRI scan. 
10. Has cancer or has had a malignant tumor within the past year, except patients who 
underwent potentially curative therapy with no evidence of recurrence. (Patients with 
stable untreated prostate cancer or skin cancers are not excluded). 


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11. Alcohol / Substance use disorder, moderate to severe, in the last 5 years according to the 
most current version DSM. 
12. Participation in another clinical trial with an investigational agent and have taken at least 
one dose of study medication, unless unblinded on placebo, within 60 days prior to the 
start of screening. (The end of a previous investigational trial is the date the last dose of 
an investigational agent was taken), or five half-lives of the investigational drug, 
whichever is greater. 
13. Subjects with infection or inflammation of the skin or skin disease at or in proximity to 
the lumbar puncture site. 
14. History of lumbar spine surgery or chronic low back pain (CLBP). 
15. Subjects with learning disability or developmental delay. 
16. Subjects whom the site PI deems to be otherwise ineligible. 
 
 


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9 
DESCRIPTION OF STUDY VISITS 
Each subject will have a 42-day screening period followed by 25±2 days of treatment at home. 
The study visits are described below and outlined in the Schedule of Events in Appendix 1.  
9.1 
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.  
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 MRI scan (up to 12 months prior) and ECG 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. Urine dipstick pregnancy test will be completed for female 
subjects.  
Cognitive and functional assessments should not be administered when the subject is in a fasted 
state. 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 Baseline. 
Sites should try to enroll patients who already have an MRI scan available to limit patient 
burden. Any patient who requires a new MRI must have all other screening assessments 
completed prior to performing the MRI.  
9.2 
Baseline Visit 
Baseline procedures include functional, cognitive, and behavioral assessments, safety 
assessments, and review of concurrent medications and adverse events that occur in clinic.  
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 
subject should then be randomized, dispensed study drug, and instructed on proper storage and 
at-home dosing based on the scheduled date of the confinement visit. The subject should be 
instructed on 1.) to take the first dose immediately after the baseline spinal tap and venipuncture  
2.) the Day 14 Contact and the Pre-Confinement Phone Contact and 3.) Study drug compliance 
and study drug return.  


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A lumbar puncture will be performed as part of the baseline visit, in the morning (before 10 am 
is recommended), after an overnight fast. After the collection of the baseline lumbar puncture 
and venipuncture, the first dose of Posiphen will be administered to the patient. 
The subject will stay at the unit for a 1 hour observation period and may be safely discharged to 
home per investigator discretion.  Subjects will have the option to stay overnight in the 
occurrence of adverse events associated with the LP procedure. 
Subject baseline visit can be split over two (2) consecutive days should the investigator feel that 
the neuropsychological tests are better completed without fatigue (i.e., after LP or unfasted).  
9.3 
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), 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. 
9.4 
Confinement Visit 
Admission for the confinement visit should occur 25 ± 2 days following the first dose of study 
medication. A pre-confinement phone call will be conducted 1-3 days prior to admission and a 
post-confinement phone follow-up will be conducted approximately 24 hours following 
discharge. Confinement procedures include functional, cognitive, and behavioral assessments, 
safety assessments, and review of concurrent medications and adverse events that occur in clinic, 
blood and CSF collection.  
9.4.1 
Pre-Confinement Telephone Contact  
A telephone call will be conducted at 1-3 days prior to confinement 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), 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 compliance. 
9.4.2 
Confinement Visit 
The confinement visit will consist of the following: admission procedures, Time 0 blood and 
CSF sampling, administration of last dose of study drug, 6-hours of blood and CSF sampling, at 
least a 1-hour observation period while subject remains on bed rest with the observation period 


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to be extended as clinically necessary as per the Investigator’s decision, and discharge 
procedures. Subjects will be provided with sleeping accommodations during this visit, if 
necessary, and maybe permitted to remain overnight per investigator discretion in the event there 
are adverse events associated with the study medications or study procedures.  
 
Subject confinement visit can be split over two (2) consecutive days should the investigator feel 
that the neuropsychological tests are better completed one day before the lumbar puncture.   
Period 1: Admission 
Subjects should arrive fasted in the early morning after an overnight fast. Clinical safety labs will 
be drawn and sent to the central 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), 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 Physical and Neurological Exam, administering 
functional and cognitive evaluations and C-SSRS, collecting vital signs, ECG, clinical labs, 
reviewing and recording adverse events and concurrent medications. 
Period 2: 6-Hour Sampling 
If no medical or compliance concerns are identified during the admission procedures, the subject 
will move on to the 6-Hour Sampling Phase, which entails the following procedures: 
1. Place an intravenous line for blood sampling.  Alternatively, blood may be sampled by 
direct venipuncture if deemed appropriate by the PI.  
2. Place the CSF catheter.  
3. Collect initial samples of blood and CSF (Hour 0). 
4. Administer last dose of assigned study drug after the 0 hour sampling. 
a. Collect CSF (6 mL) and blood (10mL) every hour at the following time points: 1, 
2, 3, 4, 5, 6 hours. 
i. CSF and blood should be taken within ± 10 mins of hourly times. 
ii.  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), reduce 


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the collection volume to a tolerated amount (minimum 2mL). If there is 
poor flow at any of the time points, then the next collection could be 
skipped to allow for better flow at the subsequent collection time. For 
example, if less than 2mL of CSF is collected at the 1-hour time point, 
then the 2-hour CSF collection could be skipped, and CSF collection 
would resume at the 3-hour time point. 
b. Perform periodic safety monitoring during the collection period. 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 blood collection site in the arm.  
c. Subjects remain on bed rest throughout the CSF sampling period of 6 hours. They 
are allowed to use a bedside commode for toileting. 
5. After the final 6-hour blood and CSF samples have been collected, remove the blood 
collection line and LP catheter. 
6. Perform final safety monitoring, a physical, and a neurological exam. 
The site should complete the LP catheter via their local practice/procedures; however, sites can 
refer to the LP catheter supporting documentation which will be provided.  
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 6-hour observation and 
discharge procedures should be followed to allow for management and follow-up on the adverse 
events.  
Period 3: Observation 
The subject should be observed for at least 1 hour following removal of the LP catheter.  The 
subject should remain on bed rest during this 1 hour follow up.  
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). All unused study drug should be returned to 
the clinical site for final accountability.  
Subjects may be permitted to stay overnight in the event there are ongoing adverse events that 
require monitoring per investigator discretion. 
 


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Period 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. 
9.4.3 
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. 
 
 


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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 confinement (admission) 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 time of enrollment. 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. 
• 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. 
• Development of suicidal or homicidal ideation requiring hospitalization or confinement. 
• 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. 


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11 
STUDY-SPECIFIC INSTRUMENTS  
We are planning a number of tests that measure states of function, depression, dementia, memory 
and attention.  Those measures include minimally the following: 
PD: MMSE, WAIS-IV, MDS-UPDRS 
AD: MMSE, WAIS-IV, ADAS-Cog 14, CDR 
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 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 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 supine 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 6-hour sampling period.  Refer to the Laboratory Manual for additional 
details. 
 
CLINICAL SAFETY LAB TESTS 


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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 (LP): 
A LP will be performed as part of the baseline visit, in the morning (before 10 am is 
recommended), after an overnight fast.   
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. 
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 Catheter Sampling:  
CSF should be collected at each timepoint during the CSF catheter sampling. At hours 0, 1, 2, 3, 
4, 5, 6 (± 10min), each 6 mL of CSF is collected. 
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 
 
COAGULATION PANEL  
PPT/PTT 
[Screening Visit ONLY] 


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NOTE: The final dose of study drug should be given after the 0 hour sample but before the 1 
hour sample  
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), reduce sampling size and obtain the 
maximum tolerated amount (minimum 2mL). 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), 
reduce the collection volume to a tolerated amount (minimum 2mL). If there is poor flow at any 
of the time points, then the next collection could be skipped to allow for better flow at the 
subsequent collection time. For example, if less than 2mL of CSF is collected at the 1-hour time 
point, then the 2-hour CSF collection could be skipped, and CSF collection would resume at the 
3-hour time point. 
To clear blood associated with needle insertion, the first 1-2 mL (or more if needed) of CSF will 
be discarded.  Refer to the CSF Manual for additional details.  
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 
wellbeing 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 baseline 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.  
Blood is collected in a uniform fashion using EDTA as anti-coagulant, as described in the 
Laboratory Manual.  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 the baseline visit and during the confinement visit every hour 
for analysis of biomarkers, inflammatory factors, dopamine, acetylcholine and drug level 
analyses. 
Biomarker and PK analyses sample collection will be performed on the 10 mL EDTA plastic 
tube whose collection is described in the Laboratory Manual.    


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12.2.4 
Blood Samples for Exosomes 
Blood will be collected in the morning, after an overnight fast, in EDTA tubes and will be 
subjected to two sequential centrifugations to generate platelet-poor plasma within one hour from 
the blood draw, as described in the Laboratory Manual. Note - for exosomes draws at the 
confinement visit after the 0-hour sample, strict fasting is no longer required. Low lipid foods 
(e.g., apple, banana, bread) are allowed.  
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 procedure manual.  
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 
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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 Medical Monitor, Sponsor, and DSMB, per Coordinating Center 
SOPs and the 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 2 into consideration (modified from 
CTCAE version 4.03.) 
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 inpatient 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, the Sponsor, Project Directors and the 
Coordinating Center within 24 hours of learning of the event. This in turn will trigger an alert to 


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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. The Sponsor is the IND holder and is responsible for submitting any SAEs 
according to the FDA reporting requirements. 
16 
DATA AND SAFETY MONITORING BOARD (DSMB) 
A DSMB will review the safety information from the study on an ongoing basis. The DSMB, 
will identify the study-specific data parameters and format of the information to be reported, as 
well as the timing of reports based on the enrollment status of the study. The DSMB will initially 
be provided with data blinded to treatment status, but they may request unblinded data if there is 
a safety concern. 
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 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. Further details will be provided in the DSMB charter. 
17 
RECORDING AND COLLECTION OF DATA  
17.1 
Electronic Case Report Form (eCRF) 
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 into the eCRF provided for that purpose. In 
some instances, no prior written or electronic record of data may exist, and data recorded directly 
into 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 eCRF 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.  


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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, 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 
• 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 Harmonization (ICH) 


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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 by the Study Director, 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. 
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. 


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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 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 should 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. 
19 
STUDY MONITORING  
The clinical monitor is responsible for inspecting the case report forms (CRFs) and source 
documentation at specific time points throughout the study to verify adherence to the protocol, 
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 
CRFs, source documents and other necessary documents at the time of the monitoring visits. 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 Regulatory Affairs that will be suitable for inspection at 
any time by the CRO, 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 


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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. These 
documents should be retained for a longer period however 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.  
 
 


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22 
LITERATURE CITED 
Berg, L. (1988). Clinical Dementia Rating (CDR). Psychopharmacol Bull, 24(4), 637-639. 
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. 
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, 12(3), 
189-198. 
Goetz CG, Tilley BC, Shaftman SR, Stebbin GT, et al. (2008). Movement Disorder Society-
Sponsored Revision of the Unified Parkinson’s Disease Rating Scale (MDS-UPDRS): Scale 
Presentation and Clinimetric Testing Results.  Movement Disorders. Vol. 23, No. 15, pp. 2129–
2170. 
Hoehn MM, Yahr MD (1967). Parkinsonism:  onset, progression, and mortality.  Neurology. 
Vol. 17. No. 5.   
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. 
Kuo YM, Nwankwo EI, Nussbaum RL, Rogers J, Maccecchini ML. Translational inhibition of 
α-synuclein by Posiphen normalizes distal colon motility in transgenic Parkinson mice. Am J 
Neurodegener Dis 2019;8(1):1-15 
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 
[(+)-phenserine] lower amyloid-β peptide levels in cell cultures and in mice. J Pharmacol Exp 
Ther 320:386-396. 
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). 
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. 
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) 


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Posiphen Investigational Drug Brochure (ANNOVIS BIO, Inc., 2019). 
Schafer K, Aisen P., et al. (2012). Administration and Scoring Manual  Alzheimer’s Disease 
Assessment Scale – Cognitive (ADAS-cog). A Publication of the Alzheimer’s Disease 
Cooperative Study.  
Teich AF, Sharma E, Barnwell E, Zhang H, Staniszewski A, Utsuki T, Padmaraju V, Mazell C, 
Tzekou A, Sambamurti K, Arancio O, Maccecchini ML. Translational inhibition of APP by 
Posiphen: Efficacy, pharmacodynamics, and pharmacokinetics in the APP/PS1 mouse. 
Alzheimers Dement Transl Res Clin Interv 2018;4:37–45. doi:10.1016/j.trci.2017.12.001. 
PMCID: PMC6021259 
Yu QS1, Reale M, Kamal M.A., Holloway H.W., Luo W., Sambamurti K., Ray B., Lahiri D.K., 
Rogers JT, Greig NH.(2013). Synthesis of the Alzheimer drug Posiphen into its primary 
metabolic products (+)-N1-norPosiphen, (+)-N8-norPosiphen and (+)-N1,N8-bisnorPosiphen, 
their inhibition 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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23 
APPENDIX 1: SCHEDULE OF EVENTS  
Visit Timinga 
Day -42 to 0 
Day 14 
(± 4) 
1-3 days prior 
to confinement 
Days 25 (± 2) 
Procedures 
Screening Baseline Phone 
Pre- 
Confinement 
Phone 
Contact 
Confinement 
Admission 
6-Hour 
Sampling 
Dischargee 
Informed Consent 
X 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
Medical and Psychiatric History 
X 
 
 
 
 
 
 
Height and BMI 
X 
 
 
 
 
 
 
Inclusion and Exclusion Criteria 
X 
X 
 
 
 
 
 
Urine Dipstick Pregnancy Test  
X 
 
 
 
 
 
 
Physical and Neurological 
Examination 
X 
X 
 
 
X 
 
X 
AD & PD:  MMSE 
X 
X 
 
 
X 
 
 
AD:  CDR 
X 
X 
 
 
X 
 
 
AD:  ADAS-Cog 14  
 
X 
 
 
X 
 
 
AD & PD:  WAIS-IV 
 
X 
 
 
X 
 
 
PD:  MDS-UPDRS  
Xg 
X 
 
 
X 
 
 
C-SSRS 
X 
X 
 
 
X 
 
 
Vital Signs b 
X 
X 
 
 
X 
X 
X 
Clinical Lab Safety Tests 
X 
Xi 
 
 
Xi 
 
 
12-Lead ECG 
X 
X 
 
 
X 
 
X 
MRI of Brainc 
X 
 
 
 
 
 
 
Adverse Events  
X 
X 
X 
X 
X 
X 
X 
Prior/Concomitant Medication  
X 
X 
X 
X 
X 
X 
X 
Randomization 
 
X 
 
 
 
 
 
Dispense Study Drug 
 
X 
 
 
 
 
 


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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 6-
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 Baseline and after 
discharge from the Confinement Visit. 
e The Early Discontinuation Visit will contain the same assessments as the Pre-confinement Visit. 
f Sampling Times:  0,1,2,3,4,5,6 hrs (±10 mins for collection)  
g Hoehn & Yahr Stage only for inclusion determination 
h X-Ray is only required if LP insertion will not be performed with fluoroscopy 
i Safety labs at baseline and confinement (for sample 0 hour) should be taken fasted  
 
 
Dosing Study Drug in Clinic 
 
X 
 
 
 
X 
 
Study Drug Compliance Check 
 
 
X 
X 
X 
 
 
APOE Genotype and DNA 
Markers 
 
X 
 
 
 
 
 
Blood Sampling for PK 
 
X 
 
 
 
X 
 
Lumbar Puncture for baseline 
Pharmacodynamics 
 
X 
 
 
 
 
 
LP Catheter Placementh 
 
 
 
 
X 
 
 
Sampling for Biomarkers in Blood, 
Plasma and CSF 
 
X 
 
 
 
Xf 
 
24-Hour Phone Follow-Upd 
 
X 
 
 
 
 
X 
Lumbar Xrayh (Optional) 
X 
 
 
 
 
 
 


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24 
APPENDIX 2: SEVERITY EVALUATION OF CHOLINERGIC ADVERSE 
EVENTS (ADVERSE EVENTS OF SPECIAL INTEREST)  
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