Conditions: Constipation Parkinson Disease
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 151
Healthy volunteers: no

Brief summary:
This study will be conducted as a multi-center, randomized, double-blind, placebo-controlled study. Approximately 72 patients will be randomized 3:1 to treatment or placebo, with approximately 54 patients allocated to receive the active investigational product and approximately 18 patients allocated to receive placebo.

\- Study Update-

Amendment 3 - In this amendment, an additional 80 patients (approximately) will be randomized 1:1 to treatment or placebo (double-blind) with approximately 40 subjects allocated to each group.

Detailed description:
The study will be conducted on an out-patient basis. Each patient will have 6 visits to the clinic: a screening visit, a randomization visit, 3 follow up visits, and 1 end of study visit.

Patient randomization will be stratified based upon the baseline weekly complete spontaneous bowel movement rate (CSBM) established during the screening period. Patients will be allowed to adjust their dosing, based upon protocol specifications. Rescue medications will be provided to all patients to ensure they move their bowels on a regular basis.

Patients will also be asked to participate in up to 2 sub-studies: a pk study and/or a stool microbiome study. The first 20 patients to consent to the pk study will have additional blood samples taken at randomization and at 2 follow up visits. The first 20 patients to consent to the stool microbiome study will provide stool samples at randomization and at 2 follow up visits.

Arms:
arm 1: ENT-01 tablet will be taken once daily by mouth. arm 2: Placebo tablet will be taken once daily by mouth.

Interventions: intervention 1: Active Investigational Treatment ENT-01 intervention 2: Placebo Treatment
Intervention descriptions: intervention 1: ENT-01 will be administered in tablet form, once daily. intervention 2: Placebo will be administered in tablet form, once daily.

Protocol document:
Prot_000.pdf: 
CONFIDENTIAL 
19April21_7.0 
Page 1 of 135 
 
 
 
 
 
 
Protocol Title: 
A Multicenter, Randomized, Double-Blind, Placebo-Controlled, Multiple 
Dose Study to Evaluate Safety, Tolerability and Efficacy of Orally 
Administered ENT-01 for the Treatment of Parkinson’s Disease-Related 
Constipation (KARMET) 
Sponsor 
Enterin, Inc. 
2005 Market Street, Suite 3125 
Philadelphia, PA 19103 
 
Investigational Product 
ENT-01 
 
Study Phase 
2b 
 
Protocol Number 
ENT-01-030 
 
Clinical Trial Registry Number 
NCT Number: 03781791 
 
IND NUMBER 
130770 
 
Protocol Version Number 
Version 7.0 
 
Protocol Version Date 
27April2021 
 
Original Protocol: 21August2018  
Amendment 1: 31December2018  
Amendment: 15March2019  
Amendment version #5:  8October2019 
Amendment version #6: 11Dec2019 
 
 


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STUDY NUMBER ENT-01-030 
AMENDMENT NUMBER 7 
 
SUMMARY OF CHANGES 
 
Section Number 
Summary of Changes 
Rationale for Change 
Section 4.0, 
Study Synopsis 
 
“80 subjects” changed to “up to 80 subjects” 
To correspond with “60 evaluable 
subjects”  
Section 4.0 
Study Synopsis 
 
“20 centers” changed to “25 centers” 
Slow recruitment during Covid 
Section 5.3 
Study Synopsis 
 
Requirement to take OTC laxatives for 6 
weeks prior to inclusion removed; duration of 
constipation required changed to 
approximately 6 months 
Unnecessary; subjects with more than 
3 CSBMs per week do not proceed to 
randomization anyway.  
Study Synopsis 
 
Discontinuation of clonazepam removed 
Sleep no longer being assessed  
Section 5.4, 5.7 
Study Synopsis 
 
Discontinuation of pimavanserin removed 
Subjects reluctant to participate and 
PIs feel it is an unnecessary exclusion 
Section 6.2 
Packaging and 
Labeling 
Picture of actual label inserted to replace the 
previous picture 
Provides a picture of the actual label 
and removable panel. 
Section 6.3 
Storage, 
Dispensing and 
Reconciliation of 
Study Drug and 
Identity of 
Investigational 
Products 
 
Inserted “Controlled room temperature” as the 
storage temperature requirement. A definition 
of Controlled room temperature is also 
provided. 
Allows for the minor temperature 
excursions outside the 20 - 25⁰C 
temperature range often experienced in 
the storage of IP at hospitals, 
pharmacies and clinical sites.  
Section 8.2 – NCI-
CTCAE Table 
Definitions of Dose Limiting Adverse Events 
- Vomiting 
Language was confusing to 
coordinators. More precise wording 
Section 8.10 
Reporting of Adverse Events (AEs) or Serious 
Adverse Events (SAEs) – AE’s and SAE’s 
begins after obtaining of informed consent 
To clarify the inconsistency within the 
Protocol  
Section 9.2.5 
Study Synopsis 
 
Sample size assumptions reworded to include 
standard deviation and 10% drop out rate 
assumption; sample size unchanged 
More precise wording 
Section 9.2.6 
Study Synopsis 
 
Data cut will be performed at 40 subjects 
changed to data cut may be performed at 40 
subjects 
To correspond to SAP 
Study Synopsis 
Section 9.2.6 
 
Mentioned that we performed an interim 
analysis earlier 
Also mentioned in earlier amendment 


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Section 4.0, 
4.1,4.2,4.3,4.5,4.6 
 
Visit window broadened 
Was too narrow; was leading to many, 
unnecessary minor protocol deviations 
Section  4.1 
 
Specifying that SAPS-PD and hallucination 
interview only need to be remeasured if 
hallucinations were present at baseline  
Eliminating unnecessary 
questionnaires 
Section 4.2 
 
Subjects allowed to continue stool diary 
longer to enable 11 days of data collection 
prior to randomization 
Several subjects needed a few extra 
days to collect the required data, 
especially those with cognitive 
impairment 
Section 4.2, 4.3,4.4, 
4.5, 4.6,  
 
Combined SAPS-PD and hallucination 
questionnaire 
To keep hallucination related questions 
together 
Section 4.1, 4.2, 4.3, 
4.4, 4.5 
 
L-Dopa sub-study related language clarified 
Was confusing to coordinators 
Section 4.1, 
Table 1 footnote, 
Appendix 2  
Language relating to orthostatic blood 
pressure measurements clarified. 
 
Section 3.3, 4.0, 4.1 
 
UPDRS certification requirement removed 
Covid is making it difficult to recertify 
coordinators every 6 months 
 
 
 


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CONFIDENTIALITY STATEMENT 
 
This study will be performed in accordance with ethical principles that 
have their origin in the Declaration of Helsinki and conducted in adherence 
to the study Protocol, Good Clinical Practices as defined in Title 21 of the 
US Code of Federal Regulations Parts 11, 50, 54, 56, and 312, as well as 
ICH GCP consolidated guidelines (E6) and applicable regulatory 
requirements. 
This document is confidential and proprietary to Enterin, Inc. Acceptance 
of this document constitutes agreement by the recipient that no unpublished 
information contained herein will be reproduced, published, or otherwise 
disseminated or disclosed without prior written approval of Enterin, Inc., 
except that this document may be disclosed in any medium to appropriate 
clinical investigators, Institutional Review Boards, and others directly 
involved in the clinical investigation that is the subject of this information 
under the condition that they keep the information strictly confidential. 


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PROTOCOL ENT-01-030 AMENDMENT 
 
Investigator Agreement 
 
I have read this protocol amendment “A Phase 2b Multicenter, Randomized, 
Double-blind, Placebo-controlled, Multiple Dose Study to Evaluate Safety, 
Tolerability, and Efficacy of Orally Administered ENT-01 for the Treatment of 
Parkinson’s Disease (PD)-related Constipation (KARMET)” and any auxiliary 
materials related to Study ENT-01-030 and agree to the following: 
1. 
To conduct the study as described in the protocol, protocol amendment, and 
any auxiliary materials 
2. 
To protect the rights, safety, and welfare of the participants in the study 
 
3. 
To provide oversight to all personnel to whom study activities have been delegated 
 
4. 
To maintain control all investigational product (IP) provided by the 
Sponsor and to maintain records of the disposition of those products 
5. 
To conduct the study in accordance with the protocol amendment and all 
applicable local and national regulations, including, but not limited to, the 
International Conference on Harmonisation (ICH) Guideline for Good 
Clinical Practice (GCP), and the standards set forth by the ethical principles 
that have their origin in the Declaration of Helsinki, GCPs as indicated in 
Food and Drug Administration (FDA) 
ICH E6 (R2) and ICH E3. 
 
6. 
To obtain approval for the protocol, protocol amendment, and all 
written materials provided to participants prior to initiating the study at 
my site 
7. 
To obtain informed consent from all participants enrolled at my study site prior 
to initiating any study-specific procedures or administrating IP to participants 
8. 
To maintain accurate records of each subject’s participation 
 
 
Investigator’s Signature 
Date 


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STATEMENT OF COMPLIANCE 
 
The study will be conducted in compliance with this clinical study protocol. Good 
Clinical Practices (GCP) pursuant to ICH E6 (R2), and all applicable local and national 
regulatory requirements. 
Enrollment at any clinical study site may not begin prior to that site receiving approval 
from the ethics committee/IRB of record for the protocol and all materials provided to 
potential participants. 
 
Any amendments to the protocol or changes to the consent document will be approved 
before implementation of that amendment. Reconsent of previously enrolled 
participants may be necessary depending on the nature of the amendment. 
 
The Principal Investigator (PI) will ensure that changes to the study plan as defined by this 
Protocol will not be made without prior agreement from Enterin, Inc and documented 
approval from the ethics committee/IRB or record, unless such a change is necessary to 
eliminate immediate hazard to the study participants. 
 
All personnel involved in the conduct of this study have completed Human Subjects 
Protection and GCP Training as outlined by their governing situation. 


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STUDY CONTACT INFORMATION 
 
 
 
Contact Name 
Telephone 
E-mail address 
Denise Barbut, MD, FRCP 
Chief Medical Officer 
(917) 975 - 1377 
d.barbut@enterininc.com 
William Wetzler 
Executive Director,  
Clinical   Operations 
 (267) 317-6156 
 w.wetzler@enterinc.com 


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SPONSOR SIGNATURE PAGE 
 
 
PROTOCOL APPROVAL PAGES(S) 
 
 
Authored by: 
 
Denise Barbut, MD FRCP Printed Name 
 
 
 
 
 
(917) 975-1377 
Telephone Number 
 
 
d.b arbut@enterininc.com 
 
E-mail Address 
 
 
 
 
 
Date 


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Contents 
TABLE OF CONTENTS 
 
Protocol Title: ............................................................................................................................................... 1 
Sponsor.......................................................................................................................................................... 1 
Investigational Product ............................................................................................................................... 1 
Study Phase ................................................................................................................................................... 1 
Protocol Number .......................................................................................................................................... 1 
IND NUMBER ............................................................................................................................................. 1 
Protocol Version Number............................................................................................................................ 1 
Protocol Version Date .................................................................................................................................. 1 
CONFIDENTIALITY STATEMENT ....................................................................................................... 2 
STUDY CONTACT INFORMATION ...................................................................................................... 6 
SPONSOR SIGNATURE PAGE ................................................................................................................ 7 
LIST OF TABLES ..................................................................................................................................... 13 
LIST OF FIGURES ................................................................................................................................... 13 
LIST OF ABBREVIATIONS ................................................................................................................... 14 
1 
STUDY SYNOPSIS ........................................................................................................................ 16 
2 
INTRODUCTION .......................................................................................................................... 27 
2.1 
Background .................................................................................................................................... 27 
2.2 
Rationale for the Study .................................................................................................................. 29 
3 
OBJECTIVES AND ENDPOINTS .............................................................................................. 33 
3.1 
Primary Objective and Endpoints ................................................................................................ 33 


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4 
STUDY DESCRIPTION ............................................................................................................... 39 
4.1 
Visit 1: Screening Visit (Study days -14 to -1) ............................................................................. 42 
4.2 
Visit 2: Randomization (Study Day 1+/2 days) ............................................................................ 45 
4.3 
Visit 3: Study Day 11 (+/- 1 day) ................................................................................................... 47 
4.4 
Visit 4: End of treatment period/ Beginning of Placebo Period (Study Day 26) (- 2 days) ...... 49 
4.5 
Visit 5: End of placebo period/ beginning of Wash-Out (Study Day 39) (+/- 2 days) .............. 50 
4.6 
Visit 6: End of Study Visit (Study Day 69) (+/- 3 days) .............................................................. 52 
4.7 
Termination Visit (if appropriate) ................................................................................................ 53 
5 
STUDY POPULATION ................................................................................................................. 54 
5.1 
Number of Subjects in this amendment ....................................................................................... 54 
5.2 
Selection Criteria ............................................................................................................................ 54 
5.3 
Inclusion Criteria ........................................................................................................................... 54 
5.4 
Exclusion Criteria .......................................................................................................................... 55 
5.5 
Screen Failures ............................................................................................................................... 57 
5.6 
Discontinuation Criteria and Early Termination Procedures ................................................... 57 
5.7 
Concomitant and Prohibited Medication ..................................................................................... 58 
6.0 
MATERIALS ................................................................................................................................. 59 
6.1 
Study Drug ...................................................................................................................................... 59 
6.2 
Packaging and Labeling ................................................................................................................ 59 
6.3 
Storage, Dispensing and Reconciliation of Study Drug and Identity of Investigational 
Products .......................................................................................................................................... 60 
7 
WARNINGS AND PRECAUTIONS ........................................................................................... 61 
8 
STUDY PROCEDURES ................................................................................................................ 61 
8.0 
Observations and Measurements.................................................................................................. 61 


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8.1 
Instructions to Subjects ................................................................................................................. 62 
8.2 
NCI CTCAEv4.03 Definitions of Dose Limiting Adverse Events .............................................. 62 
8.3 
Criteria for Dose Limiting Toxicity (DLT) Endpoints ................................................................ 63 
8.4 
Emergency Unblinding .................................................................................................................. 63 
8.5 
Pre-Existing Medical Conditions .................................................................................................. 64 
8.6 
Treatment Emergent Adverse events ........................................................................................... 64 
8.7 
Laboratory Abnormalities ............................................................................................................ 64 
8.8 
Adverse Event Assessment and Recording .................................................................................. 65 
8.9 
Reporting Requirements ............................................................................................................... 66 
8.10 
Serious Adverse Events .................................................................................................................. 68 
8.11 
Notification of Serious Adverse Events ........................................................................................ 68 
8.12 
Reporting a Serious Adverse Event .............................................................................................. 69 
8.13 
Departure from Protocol for Emergency or Adverse Event ...................................................... 70 
8.14 
Safety and Quality Monitoring ..................................................................................................... 70 
8.15 
Stopping Rules ................................................................................................................................ 70 
8.16 
Follow-Up and Final Reports ........................................................................................................ 71 
8.17 
Regulatory Aspects ........................................................................................................................ 71 
9 
DATA MANAGEMENT AND STATISTICS ............................................................................. 71 
9.0 
Populations for Analyses ............................................................................................................... 71 
9.1 
Statistical Analyses ......................................................................................................................... 72 
9.2.1 
Analysis of Subject Disposition, History, and Baseline Characteristics .................................... 72 
9.2.2 
Safety Analyses ............................................................................................................................... 72 
9.2.3 
Tolerability Analyses ..................................................................................................................... 73 
9.2.4 
Efficacy Analyses ............................................................................................................................ 74 


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9.2.4.1 Primary Efficacy Endpoint Analyses ........................................................................................... 74 
9.2.4.2 Secondary Efficacy Endpoint Analyses ........................................................................................ 74 
9.2.4.3 Exploratory Efficacy Endpoint Analyses ..................................................................................... 75 
9.2.5 
Sample Size/Power Considerations .............................................................................................. 75 
9.2.6 
Data Cut Analyses .......................................................................................................................... 76 
10 
ESTIMATED DURATION OF THE STUDY ............................................................................. 76 
11 
ADMINISTRATIVE PROCEDURES AND ETHICAL CONSIDERATIONS ....................... 76 
11.0 
Subject Information and Informed Consent ............................................................................... 76 
11.1 
Study Monitoring ........................................................................................................................... 77 
11.2 
Audits and Inspections ................................................................................................................... 77 
11.3 
Ethics Committee Review .............................................................................................................. 78 
11.4 
Standards ........................................................................................................................................ 78 
11.5 
Confidentiality ................................................................................................................................ 78 
11.6 
Protocol Adherence ........................................................................................................................ 79 
11.7 
Amendments to the Protocol ......................................................................................................... 80 
11.8 
Protocol Deviations ........................................................................................................................ 80 
11.9 
Study Termination ......................................................................................................................... 80 
12 
DATA HANDLING AND RECORD KEEPING ........................................................................ 81 
12.0 
Inspection of Records..................................................................................................................... 81 
12.1 
Data Management .......................................................................................................................... 81 
12.2 
Data Capture and Management ................................................................................................... 81 
12.3 
Liability and Insurance ................................................................................................................. 82 
12.4 
Retention of Records ...................................................................................................................... 82 
12.5 
Data Quality Assurance ................................................................................................................. 82 


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13 
USE OF INFORMATION ............................................................................................................. 83 
14 
REFERENCES ............................................................................................................................... 84 
APPENDIX 1 LABORATORY TESTS ................................................................................................ 88 
APPENDIX 2 SCHEDULE OF EVENTS ............................................................................................. 92 
APPENDIX 3 DOSING SCHEDULE .................................................................................................... 96 
APPENDIX 4 ROME-IV CRITERIA FOR CONSTIPATION ......................................................... 97 
APPENDIX 5 EASE OF PASSAGE ...................................................................................................... 98 
APPENDIX 6 BRISTOL STOOL CHART .......................................................................................... 99 
APPENDIX 7 SUBJECT DIARIES..................................................................................................... 100 
APPENDIX 8 PD HALLUCINATION QUESTIONAIRE .............................................................. 133 


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LIST OF TABLES 
Table 1. Study Schedule of Events ........................................................................ 23 
 
 
LIST OF FIGURES 
Figure 1: Study Design ............................................................................................ 42 


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LIST OF ABBREVIATIONS 
 
ADLs 
Activities of Daily Living 
AE 
Adverse Event 
BDI-II 
Beck Depression Inventory II 
CNS 
Central nervous system 
CRO 
Contract Research Organization 
CSBM 
Complete spontaneous bowel movement, defined as a bowel movement 
(BM) that did not occur within 24 hours following use of rescue 
medication, where the subject felt that he/she had no more stool to pass 
and was completely emptied out. 
C-SSRS 
Columbia-Suicide Severity Rating Scale 
CTCAE 
Common Terminology Criteria for Adverse Events 
DLT 
Dose Limiting Tolerability 
DSMB 
Data Safety and Monitoring Board 
eCRF 
Electronic Case Report Form 
ECG 
Electrocardiogram 
eCRF 
Electronic Case Report Form 
ENS 
Enteric nervous system 
EoS 
End of study 
EoT 
End of Treatment 
FDA 
Food and Drug Administration 
FSH 
Follicle-stimulating hormone 
GCP 
Good Clinical Practice 
GI 
Gastrointestinal 
ICF 
Informed Consent Form 
IRB 
Institutional Review Board 
IRT 
Interactive Response Technology. IRT includes Interactive Voice Response 
system and Interactive Web Response system 
IWRS  
Interactive Web Response System 
L-Dopa 
Levodopa 
LFT 
Liver function test 


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MDS-UPDRS 
Movement Disorder Society’s Unified Parkinson’s Disease 
Rating Scale 
mITT 
Modified Intent-to-treat 
 
MMSE 
Mini Mental State Examination 
MTD 
Maximum Tolerable Dose 
PAC-QOL 
Patient Assessment of Constipation - Quality of Life 
PAC-SYM 
Patient Assessment of Constipation Symptoms 
PD 
Parkinson’s Disease 
PDSS 
Parkinson’s Disease Sleep Scale 
PFS 
Parkinson's Disease Fatigue Scale 
RBDQ 
REM Sleep Behavior Disorder Questionnaire 
REM 
Rapid eye movement 
SAE 
Serious Adverse Event 
SAPS-PD 
Scale for Assessment of Positive Symptoms – Parkinson’s Disease 
SBM 
Spontaneous bowel movement, defined as any bowel movement 
(BM) that did not occur within 24 hours following use of rescue 
medication 
TEAE 
Treatment Emergent Adverse Event 
ULN 
Upper Limit of Normal 
UM-PDHQ 
University of Miami Parkinson’s Disease Hallucination Questionnaire 
UPDRS 
Unified Parkinson’s Disease Rating Scale 
WHO 
World Health Organization 


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1 
STUDY SYNOPSIS 
Study Title 
A Multicenter, Randomized, Double-Blind, Placebo-Controlled, Multiple 
Dose Study to Evaluate Safety, Tolerability and Efficacy of Orally 
Administered ENT-01 for the Treatment of Parkinson’s Disease-Related 
Constipation (KARMET) 
Protocol Number 
ENT-01-030 
Study Phase 
2b 
Methodology 
This first part of this study was conducted as a multicenter, randomized, 
double-blind, placebo-controlled study. A subject’s total study duration was 
no more than 10 weeks and total active dosing days was a maximum of 25 
days. Approximately 72 subjects were randomized 3:1 to treatment or 
placebo (double-blind), with approximately 54 subjects allocated to the 
treatment groups and approximately 18 to placebo. 
In this amendment, up to an additional 80 subjects (approximately) will 
be  randomized 1:1 to treatment or placebo (double-blind) with 
approximately 40 subjects allocated to each group. 
Randomization will be stratified on baseline weekly CSBM rate. Subjects 
with a baseline weekly CSBM rate of 1-3.0 will start with 3 tablets daily (3 
placebo tablets or equivalent to 75 mg of ENT-01 if randomized to active 
treatment), while subjects with a baseline weekly CSBM rate of 0 - 
0.9 will start with 6 tablets daily (6 placebo tablets, or equivalent to 150 
mg of ENT- 01 if randomized to active treatment). 
Dose will then be increased by 25 mg (or 1 placebo tablet) every three days, 
until an effective dose is reached, up to a maximum dose of 10 tablets (i.e., 
250 mg of ENT-01 if randomized to active treatment) or maximum tolerated 
dose (MTD) is reached, whichever comes first. 
After completing up to 25 days of dosing, all subjects will be given placebo 
for 2 weeks (single-blind) followed by 4 weeks of wash-out. 
The study will be conducted on an out-patient basis. Each subject will have 6 
visits to the clinic: Visit 1/Screening Visit; Visit 2/Randomization Visit; Visit 
3/Follow up Visit (during the dosing period); Visit 4/Single- 
Blind Placebo Withdrawal Period Visit; Visit 5/Washout Period; and 
Visit 6/ End of Study Visit. 
Study Duration 
Each subject will participate in the study for approximately 10 weeks. 
The duration of the amended study, from first subject enrolled to last 
subject last visit, is anticipated to be 6 to 9 months. 
Study Centers 
This is a multicenter trial at up to 25 U.S. sites. 


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Objectives 
Primary Objectives: 
To evaluate the safety, tolerability, and efficacy of repeated oral 
doses of ENT-01 for up to 25 days in subjects with Parkinson’s 
Disease- related constipation in a randomized, double-blind, placebo- 
controlled study. 
• 
Primary Safety Objective: The primary safety objective of 
this study will be evaluation of the safety of repeated daily 
oral doses of ENT-01 for up to 25 days in subjects with 
PD- related constipation. 
• 
Primary Tolerability Objective: The primary tolerability 
objective will be evaluation of recurrent vomiting, recurrent 
diarrhea, abdominal pain and dizziness. 
• 
Primary Efficacy Objective: The primary efficacy objective 
will be evaluation of the efficacy of repeated daily oral doses 
of ENT-01 for up to 25 days in improving CSBM rate. 
Secondary Objectives: 
• To determine if the correct starting dose (75 mg or 150 mg) can 
be  predicted from the baseline weekly CSBM rate. 
• To collect information on the effect of ENT-01 on CSBM, SBM, ease 
of passage, consistency, suppository/enema use and subjective bowel 
function and quality of life (QOL). 
Exploratory Objectives: 
• 
To determine any change in efficacy in the blinded placebo 
withdrawal and washout periods. 
• 
To collect information on the effect of ENT-01 on motor and non- 
motor symptoms of Parkinson’s disease including mood, memory, 
and hallucinations/delusions. 
• 
To collect information on the effect of ENT-01 on levodopa 
absorption. 
• To collect information on the effect of ENT-01 on the stool microbiome 
Number of Subjects 
The study will have approximately 152 randomized subjects, with 
up to  approximately 80 subjects being randomized in this 
extension and captured in this amendment. 
Study Description 
Multicenter, randomized, double-blind, placebo-controlled, study of daily 
oral doses of ENT-01 for up to 25 days 
Study Population 
Parkinson’s Disease with constipation 
Study Drug 
Administration 
Oral 
Inclusion/Exclusion 
Criteria 
The study population is defined as subjects who meet the following 
criteria: Inclusion Criteria: 


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1. Subjects aged 30-90 years, both genders. 
2. Subjects must provide written informed consent and be willing and 
able to comply with study procedures. 
3. Subjects must be diagnosed with Parkinson's Disease defined as the 
presence of at least three of the following cardinal features, in the 
absence of alternative explanations or atypical features: rest tremor, 
rigidity, bradykinesia and/or akinesia, postural and gait abnormalities. 
4. There are insufficient criteria for Irritable Bowel Syndrome (IBS). 
5. Constipation which has been present for approximately 6 months. 
6. Body mass index (BMI) of 18-40 kg/m2. 
7. Subjects must fulfill Rome IV criteria for functional constipation which 
includes 2 or more of the following: 
• 
Straining during at least 25% of defecations 
• 
Lumpy or hard stools in at least 25% of defecations 
• 
Sensation of incomplete evacuation for at least 25% 
of defecations 
• 
Sensation of anorectal obstruction/blockage for at least 25% 
of defecations 
• 
Manual maneuvers to facilitate at least 25% of defecations (e.g., 
digital evacuation, support of the pelvic floor)  
8. Self-report of fewer than 3 complete spontaneous bowel movements 
(CSBMs) per week 
9. Loose stools are rarely present without the use of laxatives. 
10. Subjects must be able to read, understand, and accurately record data into 
the diary to guarantee full participation in the study. 
11. Female subjects must have negative serum or urine pregnancy tests and 
must not be lactating. For females able to bear children, hormonal (i.e., 
oral, implantable, or injectable), abstinence, same sex relationships, 
single-barrier method, or a double-barrier method of birth control must 
be used throughout the study. A vasectomized partner will be allowed as 
one in conjunction with another single-barrier method. 
12. Female subjects unable to bear children must have this documented in the 
CRF (i.e., tubal ligation, hysterectomy, or postmenopausal [defined as a 
minimum of one year since the last menstrual period]). Post- menopausal 
status will be confirmed by follicle stimulating hormone (FSH) in women 
less than 60 years of age. 
Exclusion Criteria: 
1. Unable or unwilling to provide informed consent or to comply with study 


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procedures. 
2. Diagnosis of secondary constipation beyond that of Parkinson’s Disease. 
3. Review of Screening Period diaries indicates either of the following: 
• 
Fewer than 11 days of diary completion 
• 
More than 3 complete spontaneous bowel movements per week 
based upon the average CSBM rate reported during the Screening 
Period. 
4. A compromised gastrointestinal system which includes: 
• 
Structural, metabolic, or functional GI diseases or disorders 
• 
Acute GI illness within 2 weeks of the screening visit 
• 
History of major GI surgery within 30 days of the screening visit (a 
history of cholecystectomy, polypectomy, hernia repair or 
appendicectomy are not exclusionary as long as they were performed 
more than 30 days before the screening visit). 
5. Unable or unwilling to withdraw from laxatives, opiates,or any 
medications which may cause constipation 2 weeks prior to the dose 
adjustment period and throughout the rest of the study. 
6. Unable or unwilling to withdraw from proton pump inhibitors 
and antacids at the end of the screening period. 
7. Any clinically significant abnormalities on screening laboratories 
or physical examination requiring further evaluation or treatment. 
8. Neurological disorder other than Parkinson’s Disease that in the 
opinion of the investigator might interfere with the conduct of the 
study. 
9. On treatment with intra-jejunal dopamine or carbidopa/levodopa 
(i.e. Duopa). 
10. Subjects starting a new Parkinson’s Disease medication or modifying 
an existing medication within 2 weeks prior to enrollment. 
11. Unable to maintain a stable diet regimen. 
12. Subjects with a cognitive impairment that preclude them 
from understanding the informed consent. 
13. Subjects placed under legal guardianship. 
14. Females who are pregnant or breastfeeding. 
15. History of excessive alcohol use or substance abuse. 
16. Participation in an investigational drug trial within the month prior 
to dosing in the present study. 
17. Any other reason, which, in the opinion of the investigator, would 
confound proper interpretation of the study. 
Study Product, Dose, 
Route, Regimen 
ENT-01 and placebo will be administered in tablet form, once daily, on 
an empty stomach, first thing in the morning, with 8 oz of water. No food 


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intake will be allowed for 60 minutes after taking medication or placebo. 
For subjects randomized to active treatment, their tablets will be 25 mg 
of ENT-01 each. Subjects randomized to active treatment with a baseline 
weekly CSBM rate of 0-0.9 will begin at 150 mg of ENT-01 and subjects 
randomized to active treatment with a baseline weekly CSBM rate of 1.0- 
3.0 will begin at 75 mg of ENT-01. Subjects randomized to placebo 
with a baseline weekly CSBM rate of 0-0.9 will begin at 6 placebo 
tablets and subjects randomized to placebo with a baseline weekly 
CSBM rate of 1.0-3.0 will begin at 3 placebo tablets. 
Study Endpoints 
Primary Endpoints: 
Primary Safety Endpoints: 
Adverse events as evaluated with subject self-reporting, vital signs, 
clinical chemistry and hematology analyses on blood samples, urinalysis, 
and electrocardiograms (ECG). 
Primary Tolerability Endpoints: 
• 
Recurrent vomiting. 
• 
Recurrent diarrhea. 
• 
Abdominal pain. 
• 
Dizziness. 
Primary Efficacy Endpoint: 
Change from baseline in the weekly CSBM rate during the 
Fixed Dose period. 
Secondary Efficacy Endpoints: 
• 
Frequency of dose adjustments in the Dose Adjustment period. 
• 
Prokinetic bowel response during the Fixed Dose period. 
• 
Change from baseline in the weekly SBM rate during the 
Fixed Dose period. 
• 
SBM bowel response to ENT-01 during the Fixed Dose period. 
• 
Change from baseline in stool ease of passage during the 
Fixed Dose period. 
• 
Change from baseline in stool consistency during the Fixed 
Dose  period. 
• 
Change from baseline in suppository/enema use during the 
Fixed Dose period. 
• 
Time to first CSBM in the Fixed Dose period. 
• 
Time to first SBM in the Fixed Dose period. 
• 
Time to first rescue medication use in the Fixed Dose period. 
• 
Stool-related subjective assessments using the PAC-SYM 
and PAC-QOL. 
Exploratory Efficacy Endpoints: 


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• 
Change from baseline in the weekly CSBM rate during the 
Placebo Withdrawal and Washout periods. 
• 
Change from baseline in the weekly SBM rate during the 
Placebo Withdrawal and Washout periods. 
• 
Change from baseline in stool ease of passage during the 
Placebo Withdrawal and Washout periods. 
• 
Change from baseline in stool consistency during the 
Placebo Withdrawal and Washout periods. 
• 
Change from baseline in suppository/enema use during 
Placebo Withdrawal and Washout periods. 
• 
Change from baseline in suppository/enema use during 
Placebo Withdrawal and Washout periods. 
Improvement in frequency and/or severity of 
hallucinations/delusions during the Fixed Dose period over 
baseline.: Improvement is defined as an improvement of 2.33 points 
or greater reduction in score from baseline on the SAPS-PD. Up to 
40% of subjects are expected to have minor or major hallucinations 
and/or delusions. The SAPS-PD will be administered at Screening, 
and subsequently will only be administered to those subjects with 
hallucinations and/or delusions (as determined by a SAPS-PD score 
>0 at the Screening and Randomization visits). 
• 
Improvement in symptoms of Parkinson’s Disease as assessed 
by the Movement Disorder Society UPDRS (MDS-UPDRS). 
• 
Improvement in mood as assessed by the Beck Depression 
Index (BDI-II). 
• 
Improvement in cognition as assessed by the MMSE. 
• 
Change in plasma L-Dopa levels 20, 40, and 60 minutes after 
L- Dopa and ENT-01 dose. 
• 
Change in stool microbiome during treatment. 
Statistical Analysis 
Sample Size Consideration 
Sample size for the first portion of this study was determined based on the 
results from the phase 2a study. In the phase 2a study, efficacy was 
demonstrated in 34 subjects treated with ENT-01, with a mean change from 
baseline in CSBMs of 2.4. 
In this protocol amendment, enrolling up to an additional 80 subjects in a 
1:1 randomization will yield a conditional power of approximately 93% and 
an additional 60 subjects will yield a conditional power of 88%. This is 
based on the additional subjects having a least squares mean change from 
baseline for the Fixed Dose Period of 3.55 and 1.28 for ENT-01 and 
placebo, respectively, and a common standard deviation of 3.88. The power 
calculations incorporated a 10% drop-out rate.  These parameter estimates 


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and conditional power calculations are based on the results of the initial 
cohort of subjects in this Phase 2b study. 
 
Analysis Populations 
• 
Safety Population: The Safety Population will consist of all 
subjects who receive at least one dose of study medication 
during the study. 
• 
Modified ITT Population: The modified Intent-to-Treat (mITT) 
Population will include subjects who are randomized and have 
the stool diary assessments both at baseline and at least one post 
baseline time-point. 
• 
Fixed Dose Population: The Fixed Dose Population will include 
all subjects who are in mITT Population who enter the Fixed Dose 
period of the study. 
• 
Efficacy Evaluable Population: The Efficacy Evaluable 
Population will include subjects who are in the Fixed Dose 
Population, complete the Fixed Dose period, are compliant with 
study medication in the Fixed Dose period and have no major 
protocol deviations. 
Analysis Methods 
Descriptive summary statistics will be presented for the safety and 
tolerability endpoints by treatment group and overall. Continuous variables 
will be summarized using the number of observations, mean, standard 
deviation, median, minimum, and maximum. Categorical variables will be 
summarized using frequency counts and percentages. 
The primary efficacy endpoint is the change from baseline in weekly 
CSBM rate during the Fixed Dose period. The primary analysis will be 
based on the Efficacy Evaluable Population. The primary efficacy analysis 
will be performed via an analysis of covariance (ANCOVA) model with 
change from baseline in weekly CSBM rate as the dependent variable, 
treatment (active or placebo) as a factor, and a subject’s baseline weekly 
CSBM rate as a covariate. The comparison of the active versus the placebo 
treatment group will be at the two-sided 0.05 significance level. 
Interim Analysis 
An unblinded interim analysis was performed after the initial cohort of 
the study. A blinded second interim analysis may be performed after 
the 40th subject (in the 1:1 randomized cohort) completes the Fixed  
Dose period (i.e. Visit 4). 
Initial Date 
December 2018 
Completion Date 
November 2021 


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Table 1. Study Schedule of Events 
 
 
 
Visit 1 
Screening 
Visit 2 
Randomization 
 
Visit 3 
Follow-Up 
Visit 4 
End of Treatment 
 
Visit 5  
End of 
Placebo 
Visit 6  
End of Study 
Early 
Termination 
 
Day -21 
Day 1  
(+/-8 days) 
 
Day 11 
(+/- 3 day) 
Day 26 
(- 2 days) 
 
Day 39 
(+/- 8 days) 
Day 69  
(+/- 8 days) 
Informed Consent 
X 
 
 
Inclusion/Exclusion Criteria a 
X 
X 
 
 
Demographics/Medical History 
X 
 
 
Complete Physical Exam 
X 
 
 
X 
Brief Physical Exam 
 
X 
X 
X 
X 
Height 
X 
 
 
Weight 
X 
X 
X 
X 
X 
X 
X 
Orthostatic Vital Signsb 
X 
X 
X 
X 
X 
X 
X 
12-lead ECG 
X 
X 
X 
X 
X 
X 
X 
Labs and Urinalysis 
X 
X 
X 
X 
X 
X 
Pregnancy Testingc 
X 
X 
X 
X 
X 
X 
X 
In-clinic IP/Placebo Dosing d 
 
X 
X 
X 
In-clinic Levodopa Dosing 
 
X 
X 
X 
X 
X 
X 
Levodopa PK Drawse 
 
Xe 
Xe 
Xe 
Xe 
Xe 
Xe 
IP/Placebo Dispense 
 
X 
X 
X 
 
IP/Placebo Collection 
and Accountability 
 
 
X 
X 
 
X 
X 
X 
Commented [A1]: Confirm whether +/- 8 days or only +8 
days 


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Visit 1 
Screening 
Visit 2 
Randomization 
Visit 3 
Follow-Up 
Visit 4 
End of 
Treatment 
 
Visit 5  
End of 
Placebo 
Visit 6  
End of  Study 
Early 
Termination 
 
Day -21 
Day 1  
(+/-8 days) 
Day 11 
(+/- 3 day) 
Day 26 
(- 2 days) 
 
Day 39 
(+/- 8 days) 
Day 69  
(+/- 8 days) 
Instructions to 
Discontinue Laxatives and 
other BM Medications 
 
X 
 
Dispense Rescue Medicationsf 
X 
X 
X 
X 
X 
Stool Diary Instruction/Reviewg 
 
X 
X 
X 
X 
 
X 
X 
X 
AE Review 
X 
X 
X 
X 
X 
X 
X 
Prior and Con Meds 
X 
X 
X 
X 
X 
X 
X 
Rome IV questionnaire 
X 
 
MDS-UPDRSh 
X 
X 
X 
X 
X 
X 
C-SSRS 
X 
X 
X 
X 
X 
X 
X 
BDI-II 
X 
X 
X 
X 
X 
X 
MMSE 
X 
X 
X 
X 
X 
X 
SAPS-PD, 
Hallucination/Delusion 
Questionnairei 
 
X 
X 
X 
X 
 
X 
X 
X 
PAC-QOL and PAC-SYM 
 
X 
X 
X 
X 
X 
X 
Dispense Stool Collection Kits 
X 
X 
           
 
Collect Stool Samplej 
 
X 
X 
          
          X 


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a. Confirmation of eligibility: To be randomized to receive double-blind study medication in this study, subjects must meet the following criteria: 
• 
Have had at least 11 “Available Data Days” (days with electronic diary compliance sufficient for determining whether a BM occurred) 
• 
Have had <3 CSBMs/week based upon the average CSBM rate during the first 2 screening weeks (baseline period). 
• 
Be approved for randomization by the enrollment committee 
 
b. Three sets will be taken. Time points to be defined. Orthostatic vital signs will be taken in the supine position and will include BP and HR, sitting or standing BP 
and HR, RR, and body temperature. . Sitting or standing measurements will be taken within approximately 5 minutes of sitting or standing. 
c. For women of child-bearing potential, on-site with urine pregnancy test kit. If the on-site UPT is positive, a serum pregnancy test will be done. For post- 
menopausal women <60 years or of age, a serum FSH will be done at the screening visit only. 
 
d. Sites will use a centralized computer-based system (IWRS) to obtain the randomization bottle(s) assigned to each subject. 
 
• 
Subjects with baseline CSBMs from 0 to 0.9 will be randomized to begin treatment with ENT-01 150 mg or 6 tablets of placebo. 
• 
Subjects with baseline CSBMs from 1.0 to 3.0 will be randomized to begin treatment with ENT-01 75 mg or 3 tablets of placebo. 
• 
Subjects will self-administer study medication at home. The dose will be taken upon awakening, on an empty stomach with 8 oz. of water and no solid food 
will be taken for another 60 minutes. Subjects will remain in a sitting upright or standing position for at least 60 minutes after taking medication. 
• 
Subjects will be advised to stay at home for 4 to 6 hours after taking the first dose of medication in case there is a need to evacuate urgently. After the 
first dose, subjects will know if and when to leave the house. 
 
e. A subset of 20 subjects from this study population will participate in an L-Dopa PK Study, as follows: 
• 
Visits 2 and 6: Blood draws for L-dopa levels (PK) will be drawn at Time zero. 
• 
The first daily dose of L-Dopa will be administered on site after the first blood draw. There will be another blood draw at 20, 40, and 60 minutes post- L- 
Dopa dose. 
• 
Visits 3. 4, and 5: Immediately upon arrival at the study site, the subject will take study medication (ENT-01 or placebo) on an empty stomach. Blood will 
be drawn at Time zero. 30 to 60 minutes after taking ENT-01 or placebo, the subject will take their first daily dose of L-Dopa. Blood will be drawn again 
20, 40, and 60 minutes post-L-Dopa dose. The subject may have food 60 minutes after taking ENT-01. 
 
f. 
Rescue medications will be dispensed as needed and instructions given concerning laxatives and other medications to treat constipation. 
 
g. Subjects will be educated concerning use of the electronic diary to report stools immediately after a bowel movement. 


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h. The first 3 parts of the MDS-UPDRS should be administered while the subject is in an ON state, approximately one hour after taking L-Dopa (if applicable). In 
some cases, subjects will reach the “on” state earlier or later than at 60 minutes. In those cases, MDS-UPDRS should be performed when the Subject is “on”. 
 
i. 
Those subjects who have a SAPS-PD score >0 on Visits 1 and 2 will have the SAPS-PD and hallucination/delusion questionnaire at all subsequent visits. 
• 
If only Visit 1 or Visit 2 has a score >0, then SAPS-PD will not be repeated in subsequent visits. 
 
j. 
The stool sample should be collected during the EoT visit.  If subjects discontinue from the study prior to the EoT visit, a stool sample should be collected at the Early 
Termination visit (as applicable).  
 
AE=adverse event; BDI-II= Beck Depression Inventory II; BM=bowel movement; BP=blood pressure; C-SSRS= Columbia-Suicide 
Severity Rating Scale; ECG=electrocardiogram; EoT=end of treatment; FSH=follicle-stimulating hormone; HR=heart rate; L- 
Dopa=levodopa; MDS-UPDRS=Movement Disorder Society’s Unified Parkinson’s Disease Rating Scale; MMSE=Mini-mental State 
Examination; PAC-QoL=Patient Assessment of Constipation - Quality of Life; 
PAC-SYM=Patient Assessment of Constipation Symptoms; PK=pharmacokinetic(s); RR=respiratory rate; SAPS-PD=Scale for 
Assessment of Positive Symptoms – Parkinson’s Disease; UPT=urine pregnancy test. 


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2 
INTRODUCTION 
 
Constipation is a common problem worldwide, affecting 2% to 27% of the population, with 
most estimates varying from 12% to 20% (Bouras and Tangalos, 2009). The prevalence of 
constipation increases to 30%-40% among people aged >65 years and women are 
disproportionately affected. In North America, 63M people meet the Rome IV criteria for 
constipation and in the US alone, constipation is responsible for over 2M physician visits 
annually. Laxatives are prescribed to 2- 3M patients every year and furthermore, in most 
patients, the condition is chronic requiring life- long treatment (Bouras and Tangalos, 2009). 
Constipation is much more common among patients with Parkinson’s Disease (PD) than in the 
general population. There are 1M people suffering from Parkinson’s Disease in the US, of 
which roughly 60%, or 600,000 suffer from chronic constipation and in most, the condition is 
chronic, severe and unresponsive to standard therapy (Ondo et al., 2012; Zangaglia et al., 2007). 
This represents an economic burden to the individual with Parkinson’s Disease and to the 
healthcare system. According to the most recent Federal Supply Schedule (FSS; April 2016), 
the average 30- day reimbursed price for a basket of orally administered drugs for constipation is 
approximately $260 or $3120 per year.   This represents about $1.8B of prescription laxatives 
just for patients with Parkinson’s Disease. Constipation not only constitutes a major economic 
burden, but it also significantly affects the quality of life of the individual with Parkinson’s 
Disease, contributing to social isolation and depression. Furthermore, the severity of the 
symptoms correlates negatively with patient reported quality of life. 
An effective prokinetic medication for individuals with Parkinson’s Disease would be a useful 
addition to the currently available treatments for this disabling condition. 
2.1 
Background 
 
Constipation is common in Parkinson’s Disease and often becomes symptomatic years before 
the onset of the motor dysfunction and the subsequent diagnosis of Parkinson’s Disease (Lin et 
al., 2014; Savica et al., 2009). There is substantial evidence that the neurodegenerative process 


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associated with Parkinson’s Disease, namely the accumulation of toxic aggregates of alpha- 
synuclein, occurs within the enteric nervous system years before they appear within the brain. 
(Braak et al., 2006). It is believed that the enteric nervous system (ENS), with its vast surface 
area, is subject to continuous insults from infectious agents and toxic substances. Although the 
function of alpha- synuclein is not known, inflammation within the nervous system leads to an 
increase in its intracellular levels (Allen Reish and Standaert, 2015). In individuals with 
Parkinson’s Disease the increase in alpha- synuclein leads to the formation of neurotoxic 
aggregates, perhaps because of a failure by the neuron (due to genetic factors) to effectively 
dispose of them (Sahay et al., 2016). The aggregates of alpha- synuclein then traffic along the 
vagal nerve to the dorsal motor nucleus within the brainstem, and from there to more rostral 
structures (Greene et al, 2014). 
The individual with Parkinson’s Disease suffers from a form of constipation that is believed to 
be caused principally by delayed transit through the colon. In addition, defecation is often 
impaired by dysfunction of the Parkinson’s Disease subject’s anorectal reflex. For many 
individuals, bowel issues represent a significant detriment to quality of life. Failure to effectively 
manage this problem can also lead to bowel obstruction, especially as the terminal phase of 
Parkinson’s Disease approaches. A limited number of therapies have been subjected to clinical 
trials and they include agents that increase the fluid content of the stool, either by blocking fluid 
resorption or increasing the osmolar load within the intestine (Ondo et al., 2012; Zangaglia et 
al., 2007). 
Squalamine is a small molecule, originally discovered in the liver of the dogfish shark, where 
it is delivered following a meal via the biliary tract into the intestine (Moore et al., 1993). When 
squalamine is applied to the isolated colon and/or jejunum of a mouse, the compound stimulates 
organized peristaltic waves (Kunze et al, manuscript in preparation). This effect occurs through 
direct stimulation by squalamine of the intrinsic primary afferent neurons (IPANS) which are 
the most abundant type of neuron within the ENS. The IPANs, in turn, excite the ganglia of the 
myenteric plexus, and promote propulsive peristalsis. In addition, intense electrical signals are 
directed to the brainstem via the vagus following exposure of the ENS to squalamine which 
persist for some time. Of relevance to this clinical study, squalamine reverses the impaired 
motility of  the colon from aged animals engineered to express a Parkinson’s Disease phenotype. 


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Furthermore, in a recent study, squalamine has been shown to prevent the formation of toxic 
alpha- synuclein aggregates (Perni et al., 2017). The molecule accomplishes this as a 
consequence of its mechanism of action and the mechanism by which alpha-synuclein 
aggregates to form toxic polymers. Squalamine enters a neuron and binds to the negatively 
charged phospholipids that are present on the cytoplasmic face of the neuron’s plasma 
membrane, effectively neutralizing the negative charge of the membrane surface. Alpha- 
synuclein normally binds to the cytoplasmic face of the membrane via the positive charge on its 
N-terminus. Thus, when squalamine binds to the cytoplasmic face of the plasma membrane it 
displaces alpha-synuclein into the cytoplasm. As the concentration of alpha-synuclein on the 
membrane decreases, the likelihood that monomers will aggregate into higher order polymers 
decreases, thus reducing the rate of formation of aggregates and thereby rescuing neurons from 
certain death. At the same time, squalamine excites the neuron by displacing certain ion 
channels and transporters from the membrane. Thus, squalamine both excites the neurons of 
the ENS and protects them from alpha-synuclein induced cytotoxicity. It is for these reasons 
that we will evaluate squalamine as a therapeutic to treat the constipation associated with 
Parkinson’s Disease and why we will explore its effect on sleep architecture and 
hallucinations/delusions. 
2.2 
Rationale for the Study 
 
Constipation is a major clinical component of Parkinson’s Disease and is reported to occur in 
greater than 60% of affected individuals. The pathophysiological basis of constipation in 
Parkinson’s Disease is generally believed to be due to delayed transit through the colon 
(Edwards et al., 1991; Klingelhoefer and Reichmann, 2015). Several studies have demonstrated 
that transit of stool through the colon of an individual with Parkinson’s Disease is about 50% 
that measured in age matched controls. Consequently, both stool frequency and stool 
consistency are abnormal in Parkinson’s Disease. For many patients, as well as those caring for 
these individuals, constipation remains a significant morbidity associated with the condition 
(Salat-Foix and Suchowersky, 2012; Sung et al., 2014). 
Few placebo-controlled clinical trials have been conducted in the Parkinson’s Disease 
population to assess the efficacy of therapeutics that could be of value. Addition of fiber to the 
diet, although increasing stool volume, is reported to have no effect on colon transit time (Ondo 


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et al., 2012; Zangaglia et al., 2007). An osmotic laxative, polyethylene glycol (Magrogol) has 
been studied in a small placebo controlled clinical trial of individuals with mild constipation, and 
shown to provide benefit with respect to stool frequency and consistency (Zangaglia et al., 
2007). A short term placebo controlled trial of Lubiprostone, a chloride channel activator which 
increases intestinal fluid secretion, was only effective in about 50% of those treated, and 
resulted in passage of loose stools/diarrhea in place of constipation (Ondo et al., 2012). 
Furthermore, Lubiprostone delays gastric emptying (Camilleri et al., 2006), a function already 
compromised in Parkinson’s Disease. 
The pathophysiology of the gastrointestinal (GI) dysfunction in Parkinson’s Disease involves 
deposition of alpha-synuclein within both the ENS as well as within the brainstem. For reasons 
that remain unknown alpha-synuclein, which is a protein normally produced in neurons, forms 
neurotoxic intracellular aggregates in Parkinson’s Disease. Numerous studies suggest that the 
alpha- synuclein aggregate formation begins in the ENS of the Parkinson’s Disease individual 
many years before the onset of the motor symptoms (Braak et al., 2006). As a consequence of 
the normal retrograde neuronal trafficking that occurs within the vagus nerve, toxic aggregates 
are transported from the neurons of the ENS to the dorsal motor nucleus of the vagus, and then, 
gradually to sites within the brain that are involved in physical movement and balance (Greene, 
2014; Visanji et al., 2014). Because the constipation is fundamentally of an acquired 
neurodegenerative nature, it differs, so far as we now understand, from other forms of this 
condition (Visanji et al., 2014). 
The mechanism of action of squalamine has been elucidated over the past decade. The molecule 
is structurally related to a bile salt, but bears a polyamine, giving it a net positive charge. It is 
produced naturally in the liver of the dogfish shark and delivered along with bile salts to small 
intestine via the bile duct. Based on studies conducted ex vivo with isolated segments of mouse 
jejunum and colon, and our knowledge of squalamine’s mechanism of action at a cellular level, 
we can understand the prokinetic effects of squalamine on intestinal motility. Once within the 
lumen of the intestine, squalamine is transported from the epithelium into the lamina propria 
where it enters neurons of the ENS, including the intrinsic primary afferent neurons, the most 
abundant neuron of the ENS (Furness et al., 1999). Once within the neuron, the positively 
charged squalamine molecule is attracted to the inner face of the plasma membrane, which has 


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a negative surface charge, due to the presence of anionic phospholipids, neutralizing the 
negative charge (Zasloff et al., 2011). As a consequence, many proteins that are bound to the 
inner face of the cytoplasmic face of the plasma membrane are displaced or functionally altered, 
including ion channels and transporters (Sumioka et al., 2009; Yeung et al., 2008). By this 
mechanism, squalamine activates the firing of the IPANs, which in turn, activate the myenteric 
plexus, resulting in propulsive migrating motor complexes (MMCs), and direct electrical 
activity rostrally to the brainstem via the vagus (Kunze et al, manuscript in preparation). 
Alpha-synuclein normally binds to membranes composed of a high proportion of anionic 
phospholipids via its positively charged N-terminus (Shi et al., 2015). When the intracellular 
concentration of alpha-synuclein increases, the concentration of alpha-synuclein molecules 
bound to the inner face of the cytoplasmic membrane increases proportionally, and as it does 
the probability that they will physically contact one another and aggregate increases. In 
individuals with Parkinson’s Disease, these aggregates grow in size and eventually overwhelm 
the cell’s capacity to digest the inclusions, whereas in those without Parkinson’s Disease these 
aggregates are cleared by autophagy (Recchia et al., 2004). In a recent study we show that 
squalamine effectively displaces alpha-synuclein from the surface of a negatively charged 
membrane surface, and completely prevents aggregate formation. These studies have been 
conducted in specialized in vitro systems as well as in a validated in vivo model of Parkinson’s 
Disease, a genetically engineered strain of C. elegans expressing human alpha-synuclein. Oral 
treatment of C. elegans with squalamine prevents the aggregation of alpha-synuclein within the 
muscles of the animal and reverses the paralysis caused by these aggregates (Perni et al., 2017). 
Thus, squalamine has the remarkable property of stimulating peristaltic activity by directly 
activating the neurons of the ENS and via the same mechanism, inhibiting the formation of 
neurotoxic alpha-synuclein aggregates within the same neurons. In principle, squalamine could 
reverse the neurodegenerative process within the ENS of the individual with Parkinson’s 
Disease. If indeed the pathophysiology of the central nervous system (CNS) component of 
Parkinson’s Disease occurs as a consequence of the trafficking of alpha-synuclein aggregates 
from the ENS to the brain via the connecting nerves, then squalamine could have an impact on 
the disease process occurring within the CNS both therapeutically and prophylactically. 
A Phase 2a study, Evaluation of Safety and Tolerability of ENT-01 for the Treatment of 


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Parkinson’s Disease Related Constipation (RASMET) was conducted. The purpose of the study 
was to assess the safety, tolerability, pharmacokinetics, and pharmacodynamics of an orally- 
administered medication to relieve symptoms of constipation associated with Parkinson's 
Disease. The study consisted of an 8 to 12-week Stage 1 period, and an 8 to 10-week Stage 2 
period. All subjects received the study drug during one of the observational periods of the study. 
The study evaluated the safety of ENT-01, and its effect on bowel function and neurologic 
symptoms. 
In Stage 1, 10 subjects received escalating single doses from 25 mg to 200 mg/day or maximum 
tolerated dose (MTD). The goal was to determine safety and tolerability. In Stage 2, 34 subjects 
received daily doses escalating from 75 mg to a maximum of 250 mg/day, a dose that induced 
change in bowel function or MTD, followed by a “fixed” dose for 4 days, and a 2-week washout. 
Primary efficacy endpoint was defined as increase of 1 complete spontaneous bowel movement 
(CSBM)/week, or 3 CSBM/week over 14-day baseline period. Safety was also assessed. 
In Stage 2, 82.4% subjects achieved the primary efficacy endpoint. Mean CSBM/week 
increased from 1.2 at baseline to 3.6 during fixed dose (p=1.2x10-7). Common adverse events 
included nausea in 21/44 (47%) and diarrhea in 18/44 (40%) subjects. No study-drug related 
serious adverse events were reported. Significant and lasting improvements were observed in 
MDS-UPDRS scores (p=0.0008), cognition (MMSE) (p=0.0004), hallucinations, sleep, and 
circadian rhythm (p=0.016). Systemic absorption was < 0.3%. 
The results of the study indicated that orally administered ENT-01 was safe and significantly 
improved bowel function and neurologic symptoms. Minimal systemic absorption suggests that 
improvements result from local stimulation of the ENS. Data from this study informed the 
development of the 2b study. 
Phase 2b (ENT-01-030) Dosing Rationale 
 
Dosing will range from 75 mg to 250 mg. The dose will be taken upon awakening on an empty 
stomach along with 8 oz. of water simultaneously to levodopa (if applicable). The compound 
is highly charged and will absorb to foodstuffs, so subjects will not be allowed to ingest any food 
for at least 60 minutes after taking study medication. 


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Subjects are not allowed to miss a dose, however, in the event this does happen, subjects must 
wait a minimum of 5 hours post ingestion of a meal and at least 1 hour prior to the next meal 
to take the missed dose. 
In the recently completed Phase 2a study involving subjects with Parkinson’s Disease-related 
constipation, ENT-01 was administered orally both in single-dose (Stage 1: n=10) and multiple 
daily doses (Stage 2: n=34) and was well tolerated to doses up to 250 mg. Maximal treatment 
duration in Stage 2 was 25 days. In this 2b study, daily doses of ENT-01 will be administered 
orally to subjects with Parkinson’s Disease-related constipation and maximum treatment 
duration will also be 25 days. 
 
Humans have long been exposed to low doses of squalamine (milligram to microgram) in the 
various commercial dogfish shark liver extracts available as nutraceuticals (e.g., Squalamax). 
In addition, following systemic administration squalamine is cleared by the liver and excreted 
as the intact molecule (in mice) into the duodenum through the biliary tract. Drug related GI 
toxicology has not been reported in published clinical trials involving systemic administration 
of squalamine. ENT-01 has limited bioavailability in rats and dogs. Based on measurement of 
portal blood concentrations following oral dosing of radioactive ENT-01 to rat’s absorption of 
ENT-01 from the intestine is low. As a consequence, the principal focus of safety is on local 
effects on the gastrointestinal tract. However, ENT-01 appears to be well tolerated in both rats 
and dogs. 
Because local gastrointestinal intolerance is the primary adverse effect, safety margins based 
on mg/kg/day of ENT-01 are considered an appropriate safety metric based on allometric 
comparisons of gastrointestinal dimensions between species (Kararli, 1995; Sharma and 
McNeill, 2009). 
 
3 
OBJECTIVES AND ENDPOINTS 
 
3.1 
Primary Objective and Endpoints 
 
The purpose of this study is to evaluate the safety, tolerability, and efficacy of repeated oral doses of 
ENT- 01 for up to 25 days in subjects with Parkinson’s Disease-related constipation in a randomized, 
double- blind, placebo-controlled study. 


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Safety: 
The primary safety objective of this study will be the evaluation of the safety of repeated daily 
oral dose of ENT-01 for up to 25 days in subjects with PD-related constipation. 
Safety endpoints will include adverse events as evaluated with subject self-reporting, vital 
signs, clinical chemistry and hematology blood analyses, urinalysis, and electrocardiogram 
(ECG). 
It is anticipated that no more than 2 subjects out of 60 (3.3%) will have an adverse event of 
grade 4 or 5 that is at least possibly related to ENT-01 in this Study. Should there be more than 
2 subjects with an adverse event grade 4 or 5 that is at least possibly related to ENT-01 in the 
cohort of subjects randomized to ENT-01, the study will be put on an immediate clinical hold. 
In addition, individual safety stopping rules will include: 
 
• Reaching dose-limiting tolerability (DLT) before a prokinetic effect 
 
• Having a non-DLT gastrointestinal adverse event > grade 3 within 24 hours of taking 
study medication or placebo, that is at least possibly attributable to ENT-01 
• A fall in systolic blood pressure to < 80 mm Hg upon rising from lying to sitting or 
standing 
 
• A fall in diastolic blood pressure to < 40 mm Hg upon rising from lying to sitting or 
standing 
 
• Elevation of liver function tests (LFTs) > 3 times the upper limit of normal(ULN) 
Tolerability 
The primary tolerability objectives will be evaluation of recurrent vomiting, recurrent diarrhea, 
abdominal pain and dizziness. 
 
Tolerability endpoints are referred to as the DLT and will include: 
 
• Recurrent vomiting defined as 3-5 episodes of vomiting within 24 hours of taking 
study medication/placebo 
• Recurrent diarrhea defined as 7 or more episodes of diarrhea within 24 hours of taking 
study medication/placebo 
• Abdominal pain defined as moderate pain that limits instrumental activities of daily 
living (ADL) (e.g., preparing meals, shopping for groceries or clothes, using the 
telephone, managing money, etc.) within 24 hours of taking study medication / 


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placebo. 
• Dizziness defined as lightheadedness or fainting on rising from lying to sitting or 
standing that limits instrumental ADL and severe enough to indicate non-urgent medical 
intervention within 24 hours of taking study medication/placebo. 
It is anticipated that all subjects receiving active treatment will exhibit a prokinetic effect at a 
dose lower than DLT. 
Efficacy: 
The primary efficacy objective will be evaluation of the efficacy of repeated daily doses of ENT- 
01 for up to 25 days in improving the CSBM rate. 
A CSBM is a bowel movement where the subject indicated that he/she was “done for the day”, that 
he/she had “no more poop left to pass” and the bowel movement was not a1 or 2 on the Bristol stool 
scale and did not occur within 24 hours after taking rescue medication. For each subject, the Fixed Dose 
period begins on the first day of the subject’s prokinetic dose, (or the highest dose at which the 
subject did not have a DLT), following the incremental dosing algorithm defined in Section 4.2, Visit 
2: Randomization (Study Day1). 
 
Primary Efficacy Endpoint: The primary efficacy endpoint is the change from baseline in the 
weekly CSBM rate during the Fixed Dose period. Baseline is defined as the weekly CSBM rate 
during the 2-week screening period. The fixed dose response will be obtained as the weekly 
CSBM rate during the Fixed Dose period. For each subject, the Fixed Dose period begins on 
the first day of the subject’s prokinetic dose, (or the highest dose at which the subject did not 
have a DLT), following the incremental dosing algorithm defined in section 4.2 Visit 2: 
Randomization (Study Day 0, treatment begins the following day at home). 
The frequency of CSBMs will be determined via a web-based electronic diary where the 
passage of a bowel movement or absence thereof will be recorded daily throughout the study by 
the subject. 
3.2 
Secondary Objectives and Endpoints 
The secondary objectives of the study are: 
1. 
To determine if the correct starting dose (75 mg or 150 mg) can be predicted from 
the baseline weekly CSBM rate 
2. 
To collect information on the effect of ENT-01 on CSBM, SBM, ease of passage, 


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consistency, suppository/enema use and subjective bowel function and quality of 
life (QOL) 
Secondary efficacy endpoints will include the following: 
 
• 
Frequency of dose adjustments in the Dose Adjustment period: The number of dose 
adjustments a subject has from the initial starting dose to the subject’s fixed dose. 
• 
Prokinetic bowel response during the Fixed Dose period: A prokinetic response is 
defined as a weekly CSBM rate increase of 1 or more compared to baseline or an 
absolute rate of 3 or more CSBMs per week. 
• 
Change from baseline in the weekly SBM rate during the Fixed Dose period: An 
SBM is a bowel movement which did not occur within 24 hours after taking rescue 
medication. 
• 
SBM bowel response to ENT-01 during the Fixed Dose period: An SBM bowel 
response is defined as a weekly SBM rate increase of 1 or more compared to 
baseline or an absolute rate of 3 or more SBMs per week. 
• 
Change from baseline in stool ease of passage during the Fixed Dose period: Stool 
ease of passage is rated on a 1-7 ordinal point scale (see APPENDIX 5). A subject’s 
ease of passage score for a given time period is the average of the ease of passage 
scores associated with SBMs during that time period. 
 
• 
Change from baseline in stool consistency during the Fixed Dose period: Stool 
consistency is measured using the 7-point Bristol Stool Chart (see APPENDIX 6). 
A subject’s stool consistency score for a given time period is the average of the stool 
consistency scores associated with SBMs during that time period. 
• 
Change from baseline in suppository/enema use during the Fixed Dose period: 
Suppository/enema use is measured as a weekly average during the Fixed Dose 
period. This is captured in the subject stool diary. 
• 
Time to first CSBM in the Fixed Dose period: Time to first CSBM in the Fixed Dose 
period is defined as the time from the subject taking the first dose of study medication 
in the Fixed Dose period until the subject has a CSBM. 


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• 
Time to first SBM in the Fixed Dose period: Time to first SBM in the Fixed Dose 
period is defined as the time from the subject taking the first dose of study medication 
in the Fixed Dose period until the subject has an SBM. 
• 
Time to first rescue medication use in the Fixed Dose period: Time to first rescue 
medication use in the Fixed Dose period is defined as the time from the subject 
taking the first dose of study medication in the Fixed Dose period until the subject 
first uses rescue medication. 
• 
Stool-related subjective assessments using the PAC-SYM: Subjective bowel 
function data is captured using the Patient Assessment of Constipation-Symptoms: 
The PAC-SYM is a 12- item questionnaire, and each question has a 0 (Absent) – 4 
(Very Severe) point response scale. The PAC-SYM overall score is the score 
obtained by averaging the response from the 12 items. The PAC-SYM scores and 
change from baseline will be calculated for each scheduled assessment time-point. 
• 
Stool-related subjective assessments using the PAC-QOL: Constipation-related 
quality of life is assessed using the Patient Assessment of Constipation-Quality of 
Life. The PAC-QOL is a 28-item questionnaire, and each question has a 5-point 
ordinal response scale. The PAC-QOL endpoints consist of an overall average score 
and the four subscale scores (i.e., physical discomfort, psychosocial discomfort, 
worries/concerns, and satisfaction). The PAC-QOL scores and change from 
baseline will be calculated for each scheduled assessment time-point. 
3.3 
Exploratory Objectives and Endpoints 
Exploratory objectives of the study are: 
• 
To determine the decrease in efficacy in the blinded placebo withdrawal and 
washout periods. 
 
• 
To collect information on the effect of ENT-01 on motor and non-motor 
symptoms of Parkinson’s disease including mood, memory, and 
hallucinations/delusions. 
• 
To collect information on the effect of ENT-01 on levodopa absorption. 
 
• 
To collect information on the effect of ENT-01 on the stool microbiome. 


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Exploratory endpoints include: 
 
• 
Change from baseline in the weekly CSBM rate during the Placebo Withdrawal 
and Washout periods. 
• 
Change from baseline in the weekly SBM rate during the Placebo Withdrawal and 
Washout periods. 
• 
Change from baseline in stool ease of passage during the Placebo Withdrawal and 
Washout periods. 
• 
Change from baseline in stool consistency during the Placebo Withdrawal and 
Washout periods. 
• 
Change from baseline in suppository/enema use during Placebo Withdrawal and 
Washout periods. 
• 
Improvement in frequency and/or severity of hallucinations/delusions during the 
Fixed Dose period over baseline: Improvement is defined as an improvement of 
2.33 points or greater reduction in score from baseline on the SAPS-PD. Up to 40% 
of subjects are expected to have minor or major hallucinations and/or delusions. The 
SAPS-PD will be administered at Screening and at Visit 2, and subsequently will 
only be administered to those subjects with hallucinations and/or delusions on both 
visits (as determined by a SAPS-PD score >0 at Visit 1 and 2). Subjects with SAPS- 
PD score >0 at baseline and randomization visits will be included in the analysis. 
• 
Improvement of the symptoms of Parkinson’s Disease as assessed by the MDS- 
MDS-UPDRS: The first 3 parts of the MDS-UPDRS will be administered to determine 
whether there is any deterioration in overall scores during the treatment period. Motor 
deterioration will be assessed from Part 3 of the MDS-UPDRS, but total score (Parts 
1+2+3) will also be reported. The assessment will be performed during the “ON” 
phase, approximately 60 minutes after taking levodopa. In individuals who are “on” 
earlier or later, the UPDRS will be performed when they are “ON”.  
• 
Improvement in mood as assessed by the Beck Depression Index (BDI-II): The 21- 
item BDI– II assesses the intensity of depression. Each item is a list of four 
statements arranged in increasing severity about a particular symptom of depression. 


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• 
Improvement in cognition as assessed by the MMSE: The MMSE is an 11-item 
examination with a score ranging from 0 – 30 with a higher score indicating better 
memory. The MMSE will be summarized over time. 
 
• 
Change in plasma L-Dopa levels 20, 40, and 60 minutes after L-Dopa and ENT-01 
dose: An increase of >30% in peak plasma level during treatment versus baseline 
would indicate clinically significant increase in absorption attributable to ENT-01 
(20 subjects). 
• 
Change in stool microbiome during treatment: Since ENT-01 has antimicrobial 
properties in vitro, it may alter the intestinal microbiome; the subject’s stool will be 
analyzed for common intestinal bacteria and viruses at Baseline and Fixed Dose, 
periods (20 subjects).  If subjects discontinue from the study prior to the end of the 
Fixed Dose period, stool will be collected and analyzed at the Early Termination 
visit, as applicable.  
 
4 
STUDY DESCRIPTION 
 
The first part of this Phase 2b study was conducted as a multicenter, randomized, double-blind, 
placebo-controlled study. Each subject’s total study duration was no more than 10 weeks and 
total active dosing days were 25 days. 72 subjects were randomized in a 3:1 ratio to active 
treatment or placebo, respectively. 54 subjects received active treatment and 18 received 
placebo. Randomization was stratified on baseline weekly CSBM rate. Subjects with baseline 
CSBM rates of 1.0-3.0/week were randomized to begin with 75 mg of ENT-01 (3 tablets) or 
3 placebo tablets: subjects with baseline CSBM rates of 0-0.9/week were randomized to begin 
with 150 mg of ENT-01 (6 tablets) or 6 placebo tablets. The dose was then escalated every 3 
days in increments of 25 mg (or 1 placebo tablet) until either an effective dose, a maximum 
tolerated dose or 250 mg was reached, whichever came first. The dose was then “fixed” at that 
dose for the remainder of the 25-day treatment period. 
In this amendment of the protocol, the study will be conducted as a multicenter, randomized 
double-blind, placebo-controlled study. Each subject’s total study duration will be no more than 
10 weeks and total active dosing days will be 25 days. Up to 80 subjects will be randomized 
across up to 25 U.S. sites to ensure approximately 60 subjects will complete all visits, assuming 
a 25% early termination rate. Randomization will be 1:1 to active treatment or  placebo. 


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Randomization will be stratified based on baseline weekly CSBM rate. Subjects with 
baseline CSBM rates of 1.0-3.0/week will be randomized to begin with 75 mg of ENT-01 (3 
tablets) or 3 placebo tablets: subjects with baseline CSBM rates of 0-0.9/week will be 
randomized to begin with 150 mg of ENT-01 (6 tablets) or 6 placebo tablets. The dose will then 
be escalated every 3 days in increments of 25 mg (or 1 placebo tablet) until either an effective 
dose, a maximum tolerated dose or 250 mg is reached, whichever comes first. The dose will then 
be “fixed” at that dose for the remainder of the 25-day treatment period. The dosing logic is 
illustrated in the diagram below. 
• 
The subject will be randomized to either placebo or 
ENT-01, with the starting number of tablets based on the 
baseline CSBM rate. 
• 
The dose will be escalated every 3 days by 1 tablet 
(25 mg ENT-01 or placebo) 
• 
If the subject passes at least 1 CSBM/day on each of 
3 consecutive days the dose is “fixed.” Otherwise, the dose is 
escalated. 
• 
If, over the 2 days following the “fixed” dose, the 
subject passes at least one CSBM, the subject will remain on 
that dose for the remainder of the 25-day treatment period 
(until Visit 4). 
• 
If the subject fails to pass a CSBM on at least one of 
the 2 days following the “fixed” dose, the dose is then 
escalated by 1 tablet and CSBM rate evaluated as above. 
• 
Dose escalation can continue until either the MTD 
or the dosing limit of 250 mg (or 10 placebo tablets) is reached. 
 
 
 
Each subject will have 6 visits to the clinic: Visit 1/Screening Visit; Visit 2/Randomization 
Visit; Visit 3/Follow up Visit (during the treatment period); Visit 4/Single-Blind Placebo 
Withdrawal Period Visit, Visit 5/Washout Period; and Visit 6/ End of Study Visit. It is 
anticipated that the duration of study participation for each subject will be approximately 10 
weeks. 
For the duration of the study, subjects will discontinue opiates and all laxatives, bulking agents, 
softeners and suppositories. They may continue taking proton pump inhibitors and antacids 


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until the end of the screening period. Throughout the study, subjects may continue 
antihistamines, anti- cholinergic, anti-psychotic medication, dopamine and anti-depressant 
medication at stable doses. 
Each subject will be randomized to a pre-defined dose based upon their baseline weekly 
complete spontaneous bowel movements (CSBM) rate, then will be able to increase/adjust their 
dose every three days. This is called the Dose Adjustment period. After the subject documents 
three CSBMs in each of three consecutive days, the subject will “fix” at their current dose. Over 
the next 2 days, the subject will need to document a CSBM on at least 1 day. If this occurs, the 
“fixed” dose will continue for the remainder of the 25-day treatment period. If at least 1 CSBM 
does not occur over the next 2 days, then the subject will escalate the dose by 25mg. This is 
called the Fixed Dose period. In other words, the subject’s dose can’t be hard “fixed” until a 
CSBM has occurred on at least 4/5 consecutive days at a given dose. The subject’s total 
number of days on active study medication (Treatment Period) cannot exceed 25 days. 
Throughout the study, subjects will keep daily electronic stool diaries. The stool diaries will 
record information including stool frequency, consistency, ease of evacuation, completeness, 
and the use of rescue medication. Stool related information will be entered immediately upon 
completing the bowel movement. This information will be collected electronically through web- 
based questionnaires that the subjects and/or their caregivers complete. Site staff will monitor 
these entries regularly to ensure that subjects are not at risk of becoming impacted and are using 
rescue medication as directed. Site staff will contact subjects if the web-based questionnaires 
are not being completed on a daily basis or if there are any concerns about a subject’s safety. 
Rescue medication (suppositories and enemas) will be provided to subjects and will be initiated 
in the evening of the third day if a subject has had no bowel movement for three consecutive 
days. 
Symptoms of Parkinson’s Disease will be assessed using the Movement Disorder Society’s 
Unified Parkinson's Disease Rating Scale (MDS-UPDRS) during “ON” states, approximately 
one hour after levodopa intake (if applicable). 
If the “ON” time is not 60 minutes after L-Dopa, assessment has to occur at “ON” time or 
whenever the subject is “ON”. 


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Subjects who prematurely discontinue study drug for any reason will be asked to have an Early 
Termination (ET) visit within 3 days following the last study drug dose. If a subject discontinues 
study drug during a scheduled visit, then the Early Termination visit should be conducted at that 
time. These subjects will also be asked to complete the Wash-Out period and return for the End  
of Study (EoS) Visit.  
Study (EoS) Visit. The stool sample (if applicable) should be collected during the ET visit if 
the subject does not wish to return for the EoS Visit. 
 
Figure 1: Study Design 
 
 
 
Visit Schedule Overview 
Visit 1 
Screening 
Visit 2 
Randomization 
and Drug 
Dispensation 
Visit 3 
Follow 
up 
Visit 4 
End of 
Treatment – 
Beginning of 
Single Blind 
Placebo 
Withdrawal 
Visit 5 
End of 
Single 
blind 
Placebo – 
Beginning 
of Wash 
Out 
Visit 6 
End of 
Study 
Day -21 
Day 1 
+/- 8days 
Day 10 
+/- 3 day 
Day 26 
- 2 days 
Day 39 
+/- 8 
days 
Day 69 
+/- 8 
days 
 
 
4.1 
Visit 1: Screening Visit (Study days -21 to -1) 
 
The purpose of the baseline or “screening” period is to establish constipation severity off 
laxatives. At the end of the screening period the subject’s eligibility will be assessed by an 
Commented [A2]: Should this be +8days or +/- 8days 


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Evaluation Committee. 
During Visit 1, the following assessments and procedures will be performed: 
• Informed consent 
• Inclusion/Exclusion Criteria 
 
• Demographics/Medical History 
 
• Complete Physical Exam 
 
• Height 
 
• Weight 
 
• Orthostatic Vital Signs – 3 sets: Vital signs will include supine blood pressure and heart 
rate, sitting or standing blood pressure and heart rate, respiratory rate, and body 
temperature.  Sitting or standing measurements will be taken approximately 5 minutes after 
the subject has been sitting or standing. The sets should be taken throughout the Visit and not 
sequentially. 
• 12-lead ECG 
 
• Labs and urinalysis: as defined in APPENDIX 1. 
• Pregnancy Testing – For women of child-bearing potential, this is done on-site with the 
urine pregnancy kit (UPT) provided. If the on-site UPT is positive, then a serum pregnancy 
test will be done. For post-menopausal women less than 60 years or age, a serum FSH will 
be done at the screening visit only. 
• Instructions to Discontinue Laxatives and other Constipation medications. All prescription 
laxatives (Linzess, Amitiza), over the counter laxatives (Miralax, Ex-lax, 
Bisacodyl/Dulcolax), suppositories (Bisacodyl/Dulcolax), enemas (Fleets), and dietary 
supplements to increase bowel movements (chia seeds, bran, fiber supplements) will be 
discontinued. 
 
• Dispense Rescue Medication: Subjects will be provided with suppositories and enemas. 
Anytime during the study if a subject does not have any bowel movement for three 
consecutive days, rescue medication should be used to evacuate their bowel as directed by the 


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site. In the rare event that the above does not produce a bowel movement, the subject will 
be instructed to notify the site and ask for direction. 
• Stool and laxative Diary Instructions: Subjects will be taught how to use the I-Pad for stool 
and laxative diaries. They will be instructed to take the I-Pad to the toilet and fill it in 
immediately after a bowel movement. They will be taught the definition of a CSBM. If the 
subject “feels like he/she is completely emptied out” and that “they have no more stool left 
to pass”, and the stool is not just a few pebbles, then that will be a CSBM (a complete BM). 
If the subject leaves the house for a few hours, they will be told to take the I-Pad with them 
in case they have a BM elsewhere. Each BM will be entered immediately. If no entries have 
been made for the day, the subject will be sent 3 reminders starting at 7pm. Laxative use 
will also be entered immediately into the laxative diary. Multiple uses will be entered 
multiple times. 
• AE Review 
 
• Prior and Concomitant Medication 
 
• Rome IV questionnaire 
 
• MDS-UPDRS – The MDS-UPDRS should be administered while the subject is in an ON 
state, approximately one hour after taking levodopa (if applicable). If the “on” time is 
earlier or later than 60 minutes after L-Dopa, UPDRS will be performed at that time.  
• Subjects will be invited to participate in two sub-studies, consisting of  obtaining 
levodopa PK draws pre-dose, and 20, 40 and 60 minutes post-dose at Visits 2 and 3, 4, 5 
and 6 and (2) stool sample collection on Visits 2 and 4. This first twenty subjects to affirm 
consent will participate in the L-Dopa sub-study and the first 20 subjects to affirm 
consent will participate in the stool microbiome study. 
• Questionnaires: 
o C-SSRS 
o SAPS-PD and hallucination interview (only those subjects who have a SAPS-PD 
score > 0  at the Screening Visit and Randomization Visit). 
o BDI-II 
o MMSE 


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o Dispense Stool Collection Kit – Subjects participating in the sub-study will be given 
a stool collection kit so they can bring a stool sample at Visit 2 
 
For subjects who score > 0 on the SAPS-PD at Visit 1 and Visit 2, the SAPS-PD and hallucination 
interview are repeated at Visit 2. 
 
4.2 
Visit 2: Randomization (Study Day 1 + 8 days) 
 
The purpose of this period is to establish the prokinetic dose for subjects randomized to 
treatment (or placebo). The prokinetic dose is the dose that results in a CSBM on all three dosing 
days at any given dose. Subjects will also be required to have a CSBM on at least 1 of 2 
subsequent days at the “fixed” dose. In other words, the dose will only be “fixed” for the 
remainder of the treatment period if the subject has CSBMs on at least 4 of 5 consecutive 
days at a given dose. 
In this amended study, subjects will be randomized in a 1:1 ratio to receive either ENT-01 or 
placebo depending on baseline weekly CSBM rate: 
• Subjects with baseline CSBMs of 0-0.9 will be randomized to begin treatment with 150 mg 
of ENT-01 or 6 tablets of placebo. 
• Subjects with baseline CSBMs of 1.0-3.0 will be randomized to begin with 75 mg of 
ENT-01 or 3 tablets of placebo. 
Subjects will self-administer the study medication at home, beginning the day after Visit 2. The 
dose will be taken upon awakening on an empty stomach along with 8 oz. of water. The subject 
will be instructed to remain in the upright position for the next 60 minutes. This can be in the 
sitting or standing position, and the subject should be encouraged to walk around, go to the 
kitchen and make a cup of tea or coffee (with or without milk or sugar), to watch the news or 
read a paper in the upright position. The subject will also be instructed to not eat for at least 60 
minutes thereafter. 
After the first dose, the subject will be advised to stay home for 4-6 hours in case an emergency 
evacuation occurs. The subject can determine what to do after subsequent doses. Subjects are not 
allowed to miss a dose, however, in the event this does happen, subjects must wait a minimum of 5 
hours post ingestion of a meal and at least 1 hour prior to the next meal to take the missed dose. 
Any given dose is repeated for 3 consecutive days. If the subject has not produced a CSBM 


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within 24 hours of a dose on all three days at the initial dose, then they will increase their daily 
dose by 1 tablet (i.e., 25 mg for active treatment), repeating that dose for three consecutive 
days. This 3-day incremental dosing will continue until the subject: 
• Reaches the prokinetic dose. The prokinetic dose is determined when a subject has entered 
CSBMs on at least 4 of 5 consecutive days at a given dose. The subject then remains on 
that dose until Visit 4; 
• Reaches the maximum dose of 10 tablets (i.e., 250 mg for active treatment); or, 
• Reaches the dose limiting tolerability (DLT). 
 
If the subject has 4 or more episodes of diarrhea with a consistency of 6-7 (see APPENDIX 6) 
within 24 hours of a dose on 2 consecutive days at a given dose, the dose will be reduced by 25 mg 
and fixed at that dose assuming diarrhea subsides. 
Should a prokinetic response not occur at 250 mg (or 10 tablets), subjects will continue to 
take 10 tablets daily for the remainder of the 25-day treatment period. 
During this visit, the following assessments and procedures will be performed: 
 
• Review of Inclusion/Exclusion Criteria 
 
• Confirmation of eligibility: To be randomized to receive double-blind study medication in 
this study, subjects must meet the following criteria: 
• Have at least 11 ‘Available Data Days’ (days with e-Diary compliance sufficient for 
determining whether a CSBM occurred) (Subjects with fewer than 11 diary days will 
continue to fill in their diaries until 11 days’ worth of information is available and 
randomization will be delayed).  Have had fewer than 3 complete spontaneous bowel 
movements per week based upon the average CSBM rate reported during the 2-week 
Screening Period. This rate will be calculated centrally with a computer-based system and 
requested by the site the day before  randomization. 
• Be deemed eligible for randomization by the enrollment committee 
 
• Weight 
• Orthostatic Vital Signs 
 
• 12-lead ECG 
 


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• Pregnancy Testing – For women of child-bearing potential only. 
• Randomization: Sites will utilize a centralized, computer-based system (IWRS) to 
obtain the randomization bottle (s) assigned to each subject. 
• In-clinic Levodopa Dosing – Subjects will take their morning dose of levodopa in the 
clinic. 
• Subjects will be given study medication and will take their first dose the following day 
at home, on an empty stomach. The day after Visit 2 will be the first day of treatment. 
• Levodopa PK Draws – The first daily dose of L-Dopa will be administered in clinic. Blood 
samples will be obtained pre-L-Dopa dose and at 20, 40-and 60-minutes post-dose. 
(Applicable to the levodopa PK sub-study only.) 
• Dispense Rescue Medication, as needed. Subjects continue appropriate use of rescue 
medication, ensuring adequate bowel movements every 3 days. 
 
• AE Review 
• Prior and Concomitant Meds 
 
• C-SSRS 
 
• SAPS-PD (if SAPS-PD score is greater than 0 on Visit 1 and 2, subject will have SAPS- 
PD and hallucination questionnaire administered at each subsequent visit). 
 
• PAC-SYM 
 
• PAC-QOL 
 
• Collect Stool Sample (if participating in stool sub-study) 
 
• Dispense IP/placebo and instruct subjects to take their second dose of study drug the 
following morning. Subjects will be advised to stay at home for 4-6 hours in case they need 
to evacuate urgently. 
4.3 
Visit 3: Study Day 11 (+/- 3 days) 
 
The purpose of this visit is to evaluate the incidence of adverse events and to review the 
subject’s dosing experience by reviewing ePRO entries with the subject.


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During this visit, the following assessments and procedures will be performed: 
 
• Brief Physical Exam 
 
• Weight 
• Orthostatic Vital Signs 
• 12-lead ECG 
 
• Labs and urinalysis as defined in APPENDIX 1. 
 
• Pregnancy Testing - For women of child-bearing potential only. 
 
• For subjects participating in the L-Dopa sub-study, subjects will be instructed to come to  
clinic as early as possible and on an empty stomach. They will first take their study 
medication (ENT-01 or placebo) in clinic. Thirty minutes after the ENT-01/placebo dose, 
subjects will take their morning dose of levodopa.  
• Levodopa PK Draws - Blood samples will be obtained before the L-Dopa dose and at 20, 
40 and 60 minutes after the dose.  
• IP/Placebo Drug Accountability and Dispensation 
 
• Dispense Rescue Medication, as needed. Subjects continue appropriate use of rescue 
medication, ensuring adequate bowel movements every 3 days. 
• Stool Diary Review 
 
• AE Review 
 
• Prior and Concomitant Meds 
 
• MDS-UPDRS - The MDS-UPDRS should be administered while the subject is in an ON 
state, approximately one hour after taking levodopa (if applicable), or whenever the “ON” 
state occurs. 
• Questionnaires: 
 
o C-SSRS 
o BDI-II 
o MMSE 


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o PAC-SYM 
o PAC-QOL 
o SAPS-PD and hallucination questionnaire (if applicable) 
• Dispense Stool Collection Kit - Subjects participating in this sub-study will be given a 
stool collection kit so they can bring a stool sample at Visit 4. 
 
4.4 
Visit 4: End of treatment period/ Beginning of Placebo Period (Study Day 
26) (- 2 days) 
The purpose of the treatment period is to establish the impact of ENT-01 on CSBM rates. The 
purpose of the placebo period is to determine the duration of the treatment effect once study 
medication is discontinued. At this visit, all subjects will be given placebo and will continue 
taking the same number of tablets as they were taking at the maximum “fixed” dose period. The 
subject will not be made aware that the study medication provided is a placebo. 
During this visit, the following assessments and procedures will be performed: 
 
• Brief Physical Exam 
 
• Weight 
 
• Orthostatic Vital Signs – 3 sets: See screening description 
• 12-lead ECG 
 
• Labs and urinalysis as defined in APPENDIX 1. 
 
• Pregnancy Testing - For women of child-bearing potential only. 
 
• IP/Placebo Collection and Accountability 
• Single-blind Placebo Dispensation 
 
• For subjects participating in the L-Dopa sub-study, subjects will be instructed to come to 
clinic as early as possible and on an empty stomach. They will first take their study 
medication (ENT-01 or placebo) in clinic. Thirty minutes after the ENT-01/placebo dose, 
subjects will take their morning dose of levodopa. 
• Levodopa PK Draws - Blood samples will be obtained before the -L-Dopa dose and at 20, 


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40 and 60 minutes after the dose 
 
• IP/Placebo Collection and Accountability 
• Dispense Rescue Medication, as needed. Subjects continue appropriate use of rescue 
medication, ensuring adequate bowel movements every 3 days. 
• Stool Diary Review 
 
• AE Review 
 
• Prior and Concomitant Meds 
 
• MDS-UPDRS - The MDS-UPDRS should be administered while the subject is in an ON 
state, approximately one hour after taking levodopa (if applicable), or whenever “ON” 
occurs. 
• Questionnaires: 
 
o C-SSRS 
o BDI-II 
o MMSE 
o PAC-SYM 
o PAC-QOL 
o SAPS-PD and hallucination questionnaire (if applicable) 
• Collect Stool Sample (if applicable) 
 
 
4.5 
Visit 5: End of placebo period/ beginning of Wash-Out (Study Day 39) (+/- 8 
days) 
 
The purpose of the Wash-out period is to determine the duration of treatment effect. Subjects 
will be taken off placebo (study medication). They will continue to keep daily stool diaries for 
the duration of this period. During the first 3 weeks, subjects will continue to use rescue 
medication as needed. The following day, i.e., day 22 of the Wash-Out Period, subjects will be  
instructed to restart all their original medications, including oral laxatives. 


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During this visit, the following assessments and procedures will be performed: 
 
• Brief Physical Exam 
 
• Weight 
 
• Orthostatic Vital Signs 
 
• 12-lead ECG 
• Labs and urinalysis as defined in APPENDIX 1. 
 
• For subjects participating in the L-Dopa sub-study, subjects will be instructed to come to 
clinic as early as possible and on an empty stomach. They will first take their study 
medication (placebo) in clinic. Thirty minutes after the placebo dose, subjects will take 
their morning dose of levodopa. 
• Levodopa PK Draws - Blood samples will be obtained before the -L-Dopa dose and at 20, 
40 and 60 minutes after the dose. 
 
• Pregnancy Testing - For women of child-bearing potential only. 
 
• Dispense Rescue Medication, as needed. Subjects continue appropriate use of 
rescue  medication, ensuring adequate bowel movements every 3 days. 
• IP/Placebo Collection and Accountability 
 
• Stool Diary Review 
 
• AE Review 
 
• Prior and Concomitant Meds 
 
• MDS-UPDRS - The MDS-UPDRS should be administered while the subject is in an ON 
state, approximately one hour after taking levodopa (if applicable), or whenever “ON” 
occurs. 
• Questionnaires: 
 
o C-SSRS 
o BDI-II 


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o MMSE 
o PAC-SYM 
o PAC-QOL 
o SAPS-PD and hallucination questionnaire (if applicable) 
 
4.6 
Visit 6: End of Study Visit (Study Day 69) (+/- 8 days) 
 
This is the final visit of the trial. During this visit, the following assessments and procedures 
will be performed: 
• Brief Physical Exam 
 
• Weight 
 
• Orthostatic Vital Signs 
 
• 12-lead ECG 
 
• Labs and urinalysis as defined in APPENDIX 1 
 
• Pregnancy Testing - For women of child-bearing potential only. 
 
• In-clinic Study Medication and Levodopa Dosing - Subjects will be instructed to come to 
clinic as early as possible. They will take their first morning dose of levodopa in the clinic. 
• Levodopa PK Draws - Blood samples will be obtained before the -L-Dopa dose and at 20, 
40 and 60 minutes after the dose. 
 
• IP/Placebo Drug Accountability 
 
• Stool Diary Review 
 
• AE Review 
• Prior and Con Meds 
 
• MDS-UPDRS - The MDS-UPDRS should be administered while the subject is in an ON 
state, approximately one hour after taking levodopa (if applicable), or whenever “ON” 
occurs. 


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• Questionnaires: 
 
o C-SSRS 
o BDI-II 
o MMSE 
o PAC-SYM 
o PAC-QOL 
o SAPS-PD and hallucination questionnaire (if applicable) 
 
4.7 
Early Termination Visit (if appropriate) 
If a subject terminates the study prematurely for any reason, the following procedures will be 
performed: 
• Complete physical examination, including weight 
• Orthostatic vital signs 
• 12-lead ECG 
• Laboratory studies and urinalysis 
• Blood draw for L-Dopa PK sub-study only. Blood samples will be obtained pre-dose and at 
20, 40, and 60 minutes post-dose. (Applicable to the L-Dopa PK sub-study only. Subjects 
will take their dose of ENT-01/matching placebo 30 minutes prior to taking L-Dopa. 
• Pregnancy testing, for women of child-bearing potential only. 
• IP/Placebo collection and accountability 
• Electronic stool diary review 
• Review of AEs 
• Prior and concomitant medications 
• MDS-UPDRS should be administered while the subject is in an ON state, approximately one 
hour after taking L-Dopa (if applicable) 
• Subjects will complete the following questionnaires: 
• BDI-II 


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• MMSE 
• PAC-SYM 
• PAC-QoL 
• If subjects discontinue from the study prior to Visit 4, collect Stool Sample (as applicable) 
5 
STUDY POPULATION 
 
Subjects will have Parkinson’s Disease and constipation as defined by the inclusion criteria. 
Parkinson’s Disease will be diagnosed according to the UK Parkinson’s Disease Society Brain 
Bank criteria. 
5.1 
Number of Subjects in this amendment 
 
Approximately 80 subjects will be randomized to ensure approximately 60 subjects will 
complete all visits, assuming a 25% early termination rate. 
5.2 
Selection Criteria 
 
Subjects will be screened for the presence of constipation at baseline according to the Rome IV 
criteria and other inclusion criteria below. Onset of constipation before or after a diagnosis of 
Parkinson’s Disease will specifically be noted and its duration and severity established. The 
daily electronic stool diary will be analyzed at the end of the two-week screening period (Visit 
2) to establish the presence of constipation. Subjects with less than three CSBMs per week, 
without the use of rescue medication, will be randomized during Visit 2. 
5.3 
Inclusion Criteria 
 
The study population is defined as subjects who meet the following criteria: 
1. Subjects aged 30-90 years, both genders 
2. Subjects must provide written informed consent and be willing and able to comply with 
study procedures. 
3. Subjects must be diagnosed with Parkinson's Disease defined as the presence of at least 
three of the following cardinal features, in the absence of alternative explanations or 
atypical features: rest tremor, rigidity, bradykinesia and/or akinesia, postural and gait 
abnormalities. 
4. There are insufficient criteria for Irritable Bowel Syndrome (IBS) 


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5. Constipation which has been present for approximately 6 months  
6. Body mass index (BMI) of 18-40 kg/m2 
 
7. Subjects must fulfill Rome IV criteria for functional constipation which includes 2 or 
more of the following: 
a. Straining during at least 25% of defecations 
b. Lumpy or hard stools in at least 25% of defecations 
c. Sensation of incomplete evacuation for at least 25% of defecations 
d. Sensation of anorectal obstruction/blockage for at least 25% of defecations 
e. Manual maneuvers to facilitate at least 25% of defecations (e.g., digital 
evacuation, support of the pelvic floor) 
8. Self-report of 3 or fewer complete spontaneous bowel movements (CSBMs) per week 
9. Loose stools are rarely present without the use of laxatives 
10. Subjects must be able to read, understand, and accurately record data into the diary to 
guarantee full participation in the study. 
11. Female subjects must have negative serum or urine pregnancy tests and must not be 
lactating. For females able to bear children, hormonal (i.e., oral, implantable, or 
injectable), abstinence, same sex relationships, single-barrier method, or a double- 
barrier method of birth control must be used throughout the study. A vasectomized 
partner will be allowed as one in conjunction with another single-barrier method. 
12. Female subjects unable to bear children must have this documented in the CRF (i.e., 
tubal ligation, hysterectomy, or postmenopausal [defined as a minimum of one year 
since the last menstrual period]). Post-menopausal status will be confirmed by follicle 
stimulating hormone (FSH) in women less than 60 years of age 
5.4 
Exclusion Criteria 
 
1. Unable or unwilling to provide informed consent or to comply with study procedures. 
2. Diagnosis of secondary constipation beyond that of Parkinson’s Disease 


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3. Review of Screening Diaries indicates either of the following: 
 
• Fewer than 11 days of diary completion and/or 
 
• More than 3 complete spontaneous bowel movements per week based upon the 
average CSBM rate reported during the Screening Period. 
4. A compromised gastrointestinal system which includes: 
 
• Structural, metabolic, or functional GI diseases or disorders 
• Acute GI illness within 2 weeks of the screening visit 
• History of major GI surgery within 30 days of the screening visit (a history of 
cholecystectomy, polypectomy, hernia repair or appendicectomy are not 
exclusionary as long as they were performed more than 30 days before the 
screening visit) 
5. Unable or unwilling to withdraw from laxatives, opiates, clonazepam, or any 
medications which may cause constipation, 2 weeks prior to the dose adjustment period 
and throughout the rest of the study. 
6. Unable or unwilling to withdraw from proton pump inhibitors and antacids at the end 
of the screening period. 
7. Any clinically significant abnormalities on screening laboratories or physical 
examination requiring further evaluation or treatment. 
8. Neurological disorder other than Parkinson’s Disease that in the opinion of the 
investigator might interfere with the conduct of the study. 
9. On treatment with intra-jejunal dopamine or carbidopa/levodopa (i.e. Duopa). 
10. Subjects starting a new Parkinson’s Disease medication or modifying an 
existing  medication within 2 weeks prior to enrollment. 
11. Unable to maintain a stable diet regimen. 
12. Subjects with a cognitive impairment that preclude them from understanding the 
informed consent. 
13. Subjects placed under legal guardianship. 


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14. Females who are pregnant or breastfeeding. 
15. History of excessive alcohol use or substance abuse. 
16. Participation in an investigational drug trial within the month prior to dosing in the 
present study. 
 
17. Any other reason, which, in the opinion of the investigator, would confound proper 
interpretation of the study. 
5.5 
Screen Failures 
Screen failures are defined as subjects who consent to participate in the study but are not 
subsequently randomized to the study drug. Minimal information collected will include 
informed consent date, baseline subject characteristics, all the eligibility criteria violated, 
reasons for screen failure, AEs that led to screen failure, and any SAEs; these will be entered 
in the eCRF. 
Subjects who do not meet the criteria for participation in the study (screen failure) may / may 
not be re-screened. 
Any subjects who are re-screened should be assigned the same subject number given at 
the  initial screening. 
5.6 
Discontinuation Criteria and Early Termination Procedures 
 
Subjects may withdraw voluntarily from participation in the study at any time and for any 
reason. Subjects may also be withdrawn based on the Investigator’s clinical judgment. 
This study may be terminated at the discretion of the Enterin, Inc. or of any regulatory agency 
for reasons including safety and/or efficacy. 
When a subject withdraws or is withdrawn before completing the study, the date and reason for 
withdrawal are to be documented. Subjects who withdraw or who are withdrawn prematurely 
are to attend an early discontinuation visit, at which time they will complete all assessments as 
outlined in the Schedule of Events (APPENDIX 2). 
In the event that a subject is withdrawn prematurely due to an adverse event or serious adverse 
event, the adverse event or serious adverse event will be followed until it resolves or stabilizes, 
or until it is judged by the investigator to be no longer clinically significant. 


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5.7 
Concomitant and Prohibited Medication 
 
• Preventive laxatives will be discontinued during the screening period. This will include 
all prescription laxatives, over the counter laxatives and dietary supplements. 
Throughout the study, subjects will be allowed to take constipation rescue medication on 
demand, and only if they have not had a bowel movement in three consecutive days. 
Rescue medications will be recorded in the electronic daily diary every time a laxative 
is used. Bowel movement produced within 24 hours of rescue medication or accidental 
self-prescribed laxative use will not be counted as either spontaneous bowel movement 
or complete spontaneous bowel movement. 
• Proton pump inhibitors and antacids will be discontinued at the end of the screening 
period and reinstated during the wash-out period. 
• Barbiturates, and opiates will be discontinued at the end of the screening period and 2- 
3 days before dosing and reinstated during the wash-out period. All medications shall 
be reviewed and dis/approved by the investigator on a case-by-case basis. 
 
• Selective serotonin reuptake inhibitor (SSRI), SNRI, tricyclic antidepressant, anti- 
cholinergic, anti-histaminergic and anti-psychotic agents are permissible at stable 
doses. 
• Dopamine agonists and amantadine will be allowed if on a stable dose for at least 2 
weeks preceding the study. 
• Deep brain stimulation is allowed if present for more than 6 months and at stable 
stimulation parameters. 
Subgroup Analyses 
 
Subjects will be analyzed overall by their randomized treatment assignment (i.e., active versus 
placebo). Subgroup analyses will also be performed by a subject’s baseline CSBM rate (i.e., 0- 
0.9, or 1.0-3.0). The primary efficacy endpoint and key secondary, exploratory and safety 
endpoints will be summarized by these subgroups. Additional subgroup analyses may be 
defined in the statistical analysis plan (SAP). 


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6.0 
MATERIALS 
 
6.1 
Study Drug 
 
All study medication and placebo will be supplied by Enterin, Inc. as 25 mg tablets packaged 
in white opaque high-density polyethylene (HDPE) bottles with child-resistant closures. 
6.2 
Packaging and Labeling 
 
Study medication will be shipped to the investigational sites in blinded packaging. 
 


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6.3 
Storage, Dispensing and Reconciliation of Study Drug and Identity of 
Investigational Products 
All study medication is to be stored at “Controlled room temperature” as defined below. 
Controlled room temperature: The temperature maintained thermostatically that encompasses the 
usual and customary working environment of 20°–25° (68°–77° F). The following conditions also 
apply. Mean kinetic temperature not to exceed 25°. Excursions between 15° and 30° (59° and 86° F) 
that are experienced in pharmacies, hospitals, and warehouses, and during shipping are allowed. 
Provided the mean kinetic temperature does not exceed 25°, transient spikes up to 40° are permitted 
as long as they do not exceed 24 h. Spikes above 40° may be permitted only if the manufacturer so 
instructs. 
Articles may be labeled for storage at “controlled room temperature” or at “20°–25°”, or other 
wording based on the same mean kinetic temperature [see also Good Storage and Distribution 
Practices for Drug Products 〈1079〉, Quality Management System, Environmental Management 
System, Mean Kinetic Temperature (MKT) Calculation]. 
 
An article for which storage at Controlled room temperature is directed may, alternatively, be stored 
and shipped in a cool place or refrigerated, unless otherwise specified in the individual monograph or 
on the label. 
Storage on- site should be in a locked and secure location accessible only to site staff involved 
with this study. A temperature log must be kept to document temperature which should be 
recorded at least once each working day. If the temperature is not maintained, the sponsor should 
be contacted. 
If a site becomes aware that study medication has not been properly handled, the sponsor must 
be contacted immediately. In such an event, study medication should not be dispensed to any 
subject until the sponsor provides further direction. 
 
Neither the investigator nor any study personnel will distribute any study medication to any 
person not participating in this study. The study medication will be dispensed at the discretion 
and direction of the investigator in accordance with the conditions specified in this protocol. It 
Commented [A3]: Remaining comment from Carmen to 
be addressed:  Do you want to add the requirement to use 
min/max temperature measuring devise? 
 
 
 
We will discuss at Dev team tomorrow 
 


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is the investigator’s responsibility to ensure that accurate records of study medication issuance 
and return are maintained. 
 
Enterin is responsible for the tracking and accountability of study medication dispensed to sites 
and will inform sites how to return or destroy study medication once it is no longer needed at 
the site. 
Table 6.1: Identity of Investigational Products 
 
 
Active Dosage 
form 
25 mg Tablet 
White Tablet 
Placebo Dosage 
form 
25 mg tablet 
White tablet Identical in 
appearance but contains no 
active ingredient 


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Table 6.2: Route of Administration 
 
Route/dosage 
Oral (PO). Doses will require 
multiple tablets: 
75 mg=3 tablets 
100 mg=4 tablets 
125 mg=5 tablets 
150 mg=6 tablets 
175 mg=7 tablets 
200 mg=8 tablets 
225 mg=9 tablets 
250 mg=10 tablets 
Dosing 
Instructions 
Take 60 min before breakfast 
with 8 oz. water 
 
7 WARNINGS AND PRECAUTIONS 
 
Subjects will be advised to stay at home for 4-6 hours after taking the first at-home dose of 
medication in case they need to evacuate urgently. Beyond the first dose, subjects themselves 
will be in a position to know if and when to leave the house. 
 
8 
STUDY PROCEDURES 
 
8.0 Observations and Measurements 
 
Subject informed consent must be obtained prior to conducting any study-related procedures. 
The Principal Investigator at each center will ensure that the subject is given full and adequate 
oral and written information about the nature, purpose, possible risk and benefit of the study. 
Subjects must also be notified that they are free to discontinue from the study at any time. The 
subject should be given the opportunity to ask questions and allowed time to consider the 


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information provided. All assessments and procedures will be completed according to the 
Schedule of Events. 
 
8.1 
Instructions to Subjects 
 
Subjects and their spouse/companion will be taught how to fill in the daily diaries for stool 
and laxative. They will be instructed about the use of rescue medication and told to record it 
in the diary immediately. 
8.2 
NCI CTC AEv4.03 Definitions of Dose Limiting Adverse Events 
 
Adverse 
Event 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Grad 
e 5 
Diarrhea 
(watery 
stool) 
Increase of 
<4 stools a 
day over 
baseline 
Increase of 4 - 
6 stools per day 
over baseline; 
moderate 
increase in 
ostomy output 
compared to 
baseline 
Increase of >=7 
stools per day 
over baseline; 
incontinence; 
hospitalization 
indicated; severe 
increase in 
ostomy output 
compared to 
baseline; limiting 
self-care ADL 
Life 
threatening 
complications 
; urgent 
intervention 
indicated 
Death 
Vomiting 
1-2 distinct 
episodes per 
24 hrs 
3-5 distinct 
episodes 
per 24 hrs 
6 or more 
distinct episodes 
in 24 hrs; tube 
feeding, TPN or 
hospitalization 
indicated 
Life- 
threatening 
consequences 
; urgent 
intervention 
indicated 
Death 
Abdominal 
pain 
Mild pain 
Moderate pain; 
limiting 
instrumental1 
ADL 
Severe pain; 
limiting self-care 
ADL2 
- 
- 
Postural 
Mild, transient 
Moderate 
Severe 
Life- 
threatening 
Death 
hypotension3 
lightheadedn 
ess or 
lightheadedness 
, 
lightheadedness, 
and urgent 
dizziness upon 
dizziness or 
dizziness or 
fainting 
intervention 
rising from 
fainting upon 
upon rising from 
indicated 


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lying 
or sitting 
position 
rising from 
lying 
lying or sitting 
that does not 
or sitting 
position 
position that 
limits 
interfere with 
that limits 
self-care ADL; 
ADL 
instrumental 
Medical 
ADL; Non- 
urgent 
intervention or 
medical 
hospitalization 
intervention 
indicated 
indicated 
1Preparing meals, shopping for groceries or clothes, using the telephone, managing money, etc. 
2Bathing, dressing and undressing, feeding self, using the toilet, taking medications, and not bedridden 
3Definition adapted from combining CTCAE definitions for Hypotension and Dizziness 
 
8.3 
Criteria for Dose Limiting Toxicity (DLT) Endpoints: 
 
Recurrent vomiting: 3-5 episodes of vomiting within 24 hours of taking study 
medication/placebo (Grade 2 adverse event) 
Recurrent diarrhea: >7 episodes of diarrhea within 24 hours of taking study 
medication/placebo (Grade 3 adverse event) 
Abdominal pain: Moderate pain that limits instrumental activities of daily living within 24 
hours of taking study medication/placebo (Grade 2 adverse event) 
Dizziness: Moderate lightheadedness or fainting upon rising from lying to sitting or standing 
and severe enough to require medical intervention within 24 hours of taking study 
medication/placebo (Grade 2 adverse event) or either a systolic blood pressure less than 
80mmHg or diastolic blood pressure less than 40mm Hg. 
8.4 
Emergency Unblinding 
 
A treatment assignment should only be unblinded in a situation of urgent medical necessity, 
when the identity of the study medication must be known in order to select continuing 
therapy for the disease under study or an AE. The decision should be made only after 
 
consultation with the unblinded Medical Monitor, unless the urgency of a case requires 
immediate action. Treatment assignments can be obtained through a controlled transaction 


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with Interactive voice/web-response system (IVRS/IWRS). If a treatment code is  unblinded 
for any reason, the Investigator will notify Sponsor within 24 hours and will document the 
following in the eCRF: who unblinded the code, the reason for doing so, and the date of 
unblinding. 
8.5 
Pre-Existing Medical Conditions 
 
All subjects enrolled in the study will have Parkinson’s Disease. Specific pre-existing medical 
conditions that will be excluded are listed in subject exclusion criteria. 
 
8.6 
Treatment Emergent Adverse events 
 
A treatment-emergent adverse event (TEAE) is defined as any event not present prior to the 
initiation of the treatments or any event already present that worsens in either intensity or 
frequency following exposure to the treatments. 
An adverse event is typically collected after signing the informed consent form and could be 
related or unrelated to the study drug. A TEAE is for after the subject actually takes the study 
drug. 
Adverse events may be called “baseline-emergent adverse event” which is defined as any event 
which occurs or worsens during the staged screening process (after informed consent) Visit 2. 
We will include separate summaries for adverse events that occur prior to the initiation of the 
treatment and adverse events that occur after the initiation of the treatment (i.e., summary of 
treatment emergent adverse events). 
8.7 
Laboratory Abnormalities 
 
Clinical labs will be performed by a central laboratory. Labs to be drawn during the study 
include: serum chemistries and a hematology. A pregnancy test must be performed, and the 
result must be negative prior to the entry of women of child-bearing potential. The investigator 
must obtain verification that the local laboratory meets the standards for quality and consistency 
set by the College of American Pathologists. 
Clinical laboratory reports must be reviewed by a physician for out-of-range values. Out-of- 
range values will be evaluated using the following notations: 
• NCS: Not clinically significant 


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• LE: Laboratory Error 
 
• PT: Subject abnormal; relates to the subject's usual state of health 
 
• CS: Clinically Significant. This value cannot be explained by any of the other flags. 
 
By definition a lab value flagged as "CS" must be entered on the adverse clinical laboratory 
event page in the CRF. A laboratory test flagged "CS" must be repeated as soon as possible. 
The investigator may use his/her own judgment as to whether the abnormal finding is sufficient 
reason to immediately withdraw the subject from the study. 
 
If a laboratory value is considered to be serious and life-threatening the subject should be 
immediately discontinued from the study and appropriate therapy started. 
 
 
8.8 
Adverse Event Assessment and Recording 
 
All adverse events, exacerbations of concomitant illnesses, or events known to be related to 
underlying disease processes or concomitant medications are to be recorded on the CRF 
throughout the study. If a pre-existing condition worsens on study, the date on which the 
exacerbation began should be recorded. Onset dates for study treatment-related adverse events 
must be on or after the date of initial study treatment use. The need to record an adverse event 
on the CRF is not dependent on whether the adverse event is associated with the use of the study 
medication. In order to avoid vague, ambiguous or colloquial expressions, the adverse event 
should be recorded in standard medical terminology. 
Adverse event recording will include the date of onset, severity, duration, whether or not the 
study medication was discontinued, or its dosage changed because of the event, the treatment 
given and the outcome. The investigator must also assess whether the event was related to the 
study medication, concurrent drug therapy, underlying disease, a combination of these factors, 
or if it is unknown. Subjects with an adverse event should be carefully followed to determine 
outcome. 
The investigator will use the National Cancer Institute (NCI) Common Terminology Criteria 
for Adverse Events (CTCAE) V.4.03 definitions to grade the severity of the event. 


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Grade 1: Mild; asymptomatic or mild symptoms; clinical or diagnostic observations only; 
intervention not indicated 
Grade 2: Moderate; minimal, local, or non-invasive intervention indicated; limiting age- 
appropriate ADL 
Grade 3: Severe or medically significant but not immediately life-threatening; hospitalization 
or prolongation of hospitalization indicated; disabling; limiting self-care ADL. 
Grade 4: Life-threatening consequences; urgent intervention indicated 
 
Grade 5: Death 
 
The relationship or association of the study medication in causing or contributing to the adverse 
event will be characterized as remote, possible or probable as defined below: 
• Not related: Evidence indicates no plausible direct relationship to the study medication 
 
• Remote: Suggests other conditions are reasonably likely to account for the event 
including concurrent illness, progression or expression of the disease state, or reaction to 
concurrent medication 
• Possible: Suggests that the association of the event with the study medication is 
unknown; however, the adverse event is not reasonably supported by other conditions 
• Probable: Suggests that a reasonable temporal sequence of the event with medication 
administration exists and, based upon the investigator's clinical experience, the association 
of the event with study medication seems likely 
• Definite: Suggests that based upon the investigator’s experience, the association of the 
event with the study medication seems very certain. 
Procedures such as surgery should not be recorded as adverse events. However, the medical 
condition for which the procedure was performed should be reported if it meets the definition 
of adverse event. 
8.9 
Reporting Requirements 
 
Any adverse event, including both observed or volunteered problems, complaints, or symptoms 
that begins any time between the obtaining of informed consent and within 30 days after the 


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end of the last dose are to be recorded briefly on the appropriate CRF and in detail in the source 
documents. A check list of adverse events may not be used during this study. 
The following are specific definitions that are relevant to meeting your reporting obligations 
and which are included in the FDA Regulations, 21CFR Part 312.32, and International 
Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals 
for Human Use (ICH) Guidelines: 
Adverse Event: Any untoward medical occurrence in a subject administered a pharmaceutical 
product and which does not necessarily have to have a causal relationship with this treatment. 
An adverse event can be any unfavorable sign (including an abnormal laboratory finding), 
symptom, or disease temporally associated with the use of the investigational drug, whether or 
not considered related to the investigational drug. 
Serious Adverse Event: An untoward event or reaction that at any dose: 
 
• results in death 
 
• is life-threatening 
 
• requires in-patient hospitalization 
 
• prolongs existing hospitalization 
 
• results in permanent or significant disability or incapacity 
 
• is a congenital anomaly/birth defect 
• requires intervention to prevent permanent impairment/damage 
 
• other serious (Important Medical Events) that does not fit the other outcomes, but the 
event may jeopardize the subject and may require medical or surgical intervention 
(treatment) to prevent on of the other outcomes. 
 
Life-threatening adverse event: At the time of the event, a subject was at risk of death. 
There is a distinction between the severity and the seriousness of an adverse event. Severe is a 
measurement of intensity, thus a severe reaction is not necessarily a serious adverse event. For 
example, a headache may be severe in intensity, but would not be serious unless it met one of 
the criteria for serious adverse events listed previously. 


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8.10 
Serious Adverse Events 
 
Adverse events (AEs) and serious adverse events (SAEs) will be collected from signing of the 
informed consent until the follow-up contact. The investigator or site staff will be responsible 
for the detection and documentation of events meeting the criteria and definition of an adverse 
event or serious adverse event, as provided in the study protocol. 
 
However, any SAEs assessed as related to study participation (e.g., dosing, protocol mandated 
procedures, invasive tests, or change in existing therapy) or related to a concomitant medication 
will be recorded from the time a subject consent to participate in the study up to and including 
any follow-up contact. 
In the event of an AE or SAE, it will be the responsibility of the Investigator to review all 
documentation (e.g., hospital progress notes, laboratory, and diagnostics reports) relative to the 
event and attempt to establish a diagnosis of the event based on signs, symptoms, and other 
clinical information. The diagnosis was to be documented as the adverse event or serious 
adverse event and not the individual’s signs/symptoms. Once an Investigator becomes aware 
that a SAE has occurred in a study subject, they are' to report the information to Enterin within 
24hrs and provide an assessment of causality. 
8.11 
Notification of Serious Adverse Events 
 
Under IND regulations, 21 CFR Part 312.64, investigators are required to notify the 
Sponsor/representative immediately via a designated telephone number, within 24 hours of the 
sites’ notification of any SAEs, whether or not considered drug related.  
The Sponsor/representative will be notified and in turn, will report serious adverse events to 
regulatory agencies as required. In addition to the serious adverse events, other events that in 
the investigator's opinion suggest a significant hazard, contraindication, or precaution should 
be considered serious. This includes, but is not limited to, blood dyscrasias, endocrine 
disturbances, hemorrhage from any site, or severe skin disorder. Additional examples are 


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intensive treatment in an emergency room or at home for allergic bronchospasm, blood 
dyscrasias or convulsions that do not result in hospitalization, and development of drug 
dependency or drug abuse. Hospitalization for elective surgery is not considered an SAE. In 
addition, pregnancy is not an SAE, but is reportable and must be reported per IND regulations 
and timelines, 21 CFR Part 312.64, on the SAE reporting form and submitted to the 
sponsor/representative. 
Subjects who experience an SAE must be given appropriate examinations and treatment. The 
investigator must provide written information to the sponsor/representative as soon as possible. 
 
When an investigator is in doubt when to report an event, the investigator should err on the side 
of caution and contact the Sponsor/representative. 
8.12 
Reporting a Serious Adverse Event 
 
Any SAE, including death due to any cause that occurs during this study, whether or not 
believed to be related to the study medication, must be reported immediately (within one 
business day). 
The phone number the sites will use for reporting SAEs is shown below. 
 
NorthAmerica_Medical@parexel.com 
Safety Fax: 781-434-5957 
Safety Line: 781-434-5010 (Used for any reporting questions that may arise from the site 
 
Specific medical questions can be addressed to the Chief Medical Officer Denise Barbut, 
MD, FRCP d.barbut@enterininc.com Phone: 917-975-1377 
 
The initial report should contain as much information as is available concerning the event in 
order to permit the Sponsor/representative to file a report that satisfies regulatory requirements. 
Initial telephone reports of serious adverse events must be followed-up by a fax of a completed 
SAE report form or an appropriate event narrative. The event should also be entered into the 
source documents and CRF, as appropriate. When additional information is available, the 
investigator should send a follow-up SAE form or an appropriate supplementary event narrative 
to Enterin, Inc. 
All reportable SAEs will be reported to regulatory authorities by the  Sponsor/representative 
according to the required timelines. A copy of all FDA reportable serious adverse events will be 


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mailed to all investigators participating in ongoing clinical studies with the study medication in 
order to permit prompt notification of the appropriate institutional review board (IRB). 
 
8.13 
Departure from Protocol for Emergency or Adverse Event 
 
In medical emergencies, the investigator should use medical judgment and remove the subject 
from immediate hazard. As soon as possible after removing the subject from hazard, the 
investigator must contact Enterin Inc. by telephone to permit a decision as to whether the subject 
may continue in the study. The IRB should also be notified as to the type of emergency and the 
course of action. The CRF for the subject must describe the departure from the protocol and 
state the reason. 
8.14 
Safety and Quality Monitoring 
 
A Data Safety and Monitoring Board (DSMB) has been established to monitor the safety of the 
subjects during the course of the study. The DSMB (with the requisite Charter) includes 
members with relevant clinical expertise, including a good understanding of the safety of 
medications for Parkinson’s Disease. These members include a statistician, a gastroenterologist, 
and a neurologist. The DSMB will be called upon if and when necessary. In addition, subject 
appropriateness will be monitored by a committee consisting of neurologists, including the 
national PI and the CMO. The committee will review subject information half-way through the 
screening period and make recommendations about proceeding to randomization. 
8.15 
Stopping Rules 
 
It is anticipated that no more than 2 subjects out of 60 (3.3%) will have an AE of grade 4 or 5 
that is at least possibly related to ENT-01 in this Study. Should there be more than 2 subjects 
with an AE grade 4 or 5 that is at least possibly related to ENT-01 in the cohort of subjects 
randomized to ENT-01, the study will be put on an immediate clinical hold. 
In addition, individual safety stopping criteria will include: 
 
• Having a non-DLT gastrointestinal adverse event > grade 3 within 24 hours of taking 
ENT-01 that is at least possibly attributable to ENT-01 
 
• A fall in systolic blood pressure to < 80 mm Hg upon rising from lying to sitting or 
standing 


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• A fall in diastolic blood pressure to < 40 mm Hg upon rising from lying to sitting or 
standing 
• Elevation of liver function tests (LFTs) > 3 times the upper limit of normal (ULN) 
 
8.16 
Follow-Up and Final Reports 
 
The investigator shall provide Enterin, Inc. with an accurate final report within 1 month after 
completion, termination or discontinuation of the study. The final report may not precede 
retrieval of CRFs which have not been monitored. 
8.17 
Regulatory Aspects 
 
Neither the investigator nor Enterin Inc. shall modify this protocol without first obtaining 
concurrence of the other in writing. All changes must be submitted to the IRB. Protocol 
modifications which impact subject safety, or the validity of the study must be approved by the 
IRB and submitted to the FDA before implementation. In the case of a medical emergency to 
increase safety of subjects, a change may be made after discussion with the sponsor. In these 
instances, the IRB and FDA will be notified as soon as possible. 
 
9 
DATA MANAGEMENT AND STATISTICS 
 
9.0 
Populations for Analyses 
 
The following populations will be considered for statistical analyses. 
 
• Safety Population: The Safety Population will consist of all subjects who receive at least 
one dose of study medication during the study. 
• Modified ITT Population: The modified Intent-to-Treat (mITT) Population will include 
subjects who are randomized and have the stool diary assessments both at baseline and at least 
one post baseline time-point. 
• Fixed Dose Population: The Fixed Dose Population will include all subjects who are in 
mITT Population who enter the Fixed Dose period of the study. 
• Efficacy Evaluable Population: The Efficacy Evaluable Population will include 
subjects who are in the Fixed Dose Population, complete the Fixed Dose period, are 
compliant with study medication in the Fixed Dose period, and have no major protocol 
deviations. 


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9.1 
Statistical Analyses 
 
A separate statistical analysis plan (SAP) with additional details on the analyses to be 
performed and analysis methods will be developed and finalized prior to the interim analysis. 
9.2.1 
Analysis of Subject Disposition, History, and Baseline Characteristics 
 
Subject disposition, including analysis population allocation, subjects enrolled, completed each 
period, discontinued, and primary reason for discontinuation, will be summarized using 
frequency and percentage. 
Protocol deviations will be summarized using frequency and percentage. 
 
Medical history data and prior and concomitant medications will be summarized using 
frequency and percentage. 
Subjects’ age, height, weight, and baseline disease characteristics will be summarized using 
descriptive statistics. Gender, race, and other categorical variables will be provided using 
frequency and percentage. 
9.2.2 
Safety Analyses 
 
All safety analyses will be performed on the Safety Population. The safety data will be 
presented in individual listings and summary tables. 
Adverse Events 
 
AEs will be coded according to the latest version of the Medical Dictionary for Regulatory 
Activities (MedDRA). The intensity/severity of AEs will be graded according to NCI- 
CTCAE. 
TEAEs, AEs leading to study treatment discontinuation, AEs leading to dose interruption, AEs 
related to study medication, SAEs, and AEs leading to death will be summarized by system 
organ class, preferred term, and study period. A summary of AEs that are CTCAE Grade 3 or 
higher, as well as the most frequent preferred terms, will be provided. 
If a subject experience the same preferred term multiple times within a period, then the event 
will be counted only once within the period and by greatest severity. 


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Descriptive statistics will be used to summarize the safety data both by treatment group and 
overall. 
 
Clinical Laboratory Values 
All laboratory test results will be summarized by period together with the change from baseline. 
The frequency distribution for low/normal/high or normal/abnormal will be summarized as 
well. The denominators for calculating the percentages will be based on the number of subjects 
with non-missing values in the Safety Population. 
Vital Signs 
 
Vital sign results will be summarized by period, together with the change from baseline. 
 
Physical Examination 
 
Summaries of physical examinations will present frequency distribution of abnormal findings 
by body system and period. The denominators for calculating the percentages will be based on 
the number of subjects evaluated for a particular body system of each dose level in the Safety 
Population. 
Electrocardiogram (ECG) 
 
ECG findings will be classified as normal vs abnormal. The number and percentage of each 
category will be summarized using frequency table for each period. The denominators for 
calculating the percentages will be based on the number of subjects with non-missing values in 
each period. 
C-SSRS 
 
The C-SSRS is a questionnaire that assesses suicidal ideation and behavior using a semi- 
structured interview to probe subject responses. The “Baseline” version of the instrument will 
be administered to subjects during Screening, and the “Since Last Visit” version will be used at 
subsequent time points. Any subject who endorses suicidal ideation will be referred to a mental 
health professional for further assessment and/or treatment. 
9.2.3 
Tolerability Analyses 
 
Tolerability will be measured by DLT endpoints. The frequency of occurrence for each 
tolerability endpoint including recurrent vomiting, recurrent diarrhea, abdominal pain and 
dizziness will be summarized by treatment group and dose level for the Dose Adjustment 


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period. Percentages will be based on the number of subjects in each dose level in the Safety 
Population. 
 
9.2.4 
Efficacy Analyses 
 
9.2.4.1 Primary Efficacy Endpoint Analyses 
 
The primary efficacy endpoint is the change from baseline in weekly CSBM rate during the 
Fixed Dose period. The primary analysis will be based on the Efficacy Evaluable Population. 
The primary efficacy analysis will be performed via an analysis of covariance (ANCOVA) 
model with change from baseline in weekly CSBM rate as the dependent variable, treatment 
(active or placebo) as a factor, and a subject’s baseline weekly CSBM rate as a covariate. The 
comparison of active versus placebo treatment group will be tested at the two-sided 0.05 
significance level. Summary statistics will also be presented. 
As a supplemental analysis, the treatment effect for the primary efficacy endpoint will be 
summarized by the weekly CSBM rate subgroups (0-0.9, 1.0 – 3.0). The primary efficacy analysis 
will also be performed on the mITT and Fixed Dose Populations. 
 
As an additional supplemental analysis, the change from baseline in weekly CSBM rate using 
the subject’s last 2 weeks of the Treatment Period will be analyzed using the ANCOVA model 
described for the primary endpoint using the mITT population. 
 
9.2.4.2 Secondary Efficacy Endpoint Analyses 
 
Secondary efficacy analyses will be based on the Efficacy Evaluable Population. The 
comparison of active versus placebo treatment will be tested at the two-sided 0.05 
significance level. 
The following are the set of continuous secondary efficacy endpoints: 
 
• Change from baseline in the weekly SBM rate during the Fixed Dose period 
 
• Change from baseline in stool ease of passage during the Fixed Dose period 
• Change from baseline in stool consistency during the Fixed Dose period 
 
• Change from baseline in suppository/enema use during the Fixed Dose period 
 


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All continuous secondary efficacy endpoints will be analyzed in the same manner as the 
primary efficacy endpoint. 
The following are the set of dichotomous secondary efficacy endpoints: 
 
• Pro-kinetic bowel response during the Fixed Dose period 
 
• SBM bowel response to ENT-01 during the Fixed Dose period 
 
For the dichotomous secondary efficacy endpoints, the comparison of the ENT-01 versus 
placebo treatment groups will be performed by means of Fisher’s exact test. 
The following are the set of time to event secondary efficacy endpoints: 
 
• Time to first CSBM in the Fixed Dose period 
 
• Time to first SBM in the Fixed Dose period 
 
• Time to first rescue medication use in the Fixed Dose period 
 
• Time to event secondary efficacy endpoints will be summarized using Kaplan-Meier 
methods, comparisons between treatment groups will be made via log-rank tests. 
• For the frequency of dose adjustments in the Dose Adjustment period secondary endpoint, 
the frequency of the number of dose adjustments a subject has from the initial starting dose 
to the subject’s fixed dose will be summarized by treatment group and baseline CSBM 
strata. 
• For the PAC-SYM and PAC-QOL subjective assessments secondary efficacy endpoints, the 
overall scores from the Fixed Dose period will be analyzed via an ANCOVA model with 
treatment as a factor and the subject’s corresponding baseline score as a covariate. 
Additional summaries of the individual items will be performed. 
9.2.4.3 Exploratory Efficacy Endpoint Analyses: 
 
The analysis methods for the exploratory efficacy endpoints will be provided in the SAP. 
 
9.2.5 
Sample Size/Power Considerations 
 
Sample size for this study was determined based on the results from the phase 2a study. In the phase 2a 
study, efficacy was demonstrated in 34 subjects treated with ENT-01, with a mean change from baseline 
in CSBMs of 2.4. In that study, of the 72 randomized subjects, 64 (89%) were evaluable for the 
primary efficacy endpoint analysis. Randomization was 3:1 to active treatment or placebo; 64 


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subjects were randomized to active treatment and 18 subjects were randomized to placebo. The 
study assumed an increase from baseline in weekly CSBM rate in the Fixed Dose period of 2.4 
and 0.5 for the active drug and placebo-treated subjects, respectively, and an SD of 2.0. 
amended. 
 
In this protocol amendment, enrolling an additional 80 subjects in a 1:1 randomization will yield 
a conditional power of approximately 93% and an additional 60 subjects will yield a conditional 
power of 88%. This is based on the additional subjects having a least squares mean change from 
baseline for the Fixed Dose Period of 3.55 and 1.28 for ENT-01 and placebo, respectively, and 
a common standard deviation of 3.88. The power calculations incorporated a 10% drop-out rate.  
These parameter estimates and conditional power calculations are based on the results of the 
initial cohort of subjects in the Phase 2b study.  
 
 
9.2.6 
Data Cut Analyses 
 
An unblinded interim analysis was performed after the initial cohort of the study. An 
unblinded interim analysis may be performed after the 40th subject (in the 1:1 randomized 
cohort) completes the Fixed    Dose period (i.e., Visit 4). 
 
10 
ESTIMATED DURATION OF THE STUDY 
 
This study has an estimated maximum duration of up to 10 weeks for each subject. Maximum 
number of active dosing days for each subject will be no more than 25 days. The study duration 
from first subject in, to last subject out is expected to be 18 months. 
 
11 
ADMINISTRATIVE PROCEDURES AND ETHICAL CONSIDERATIONS 
 
11.0 
Subject Information and Informed Consent 
 
The Principal Investigator(s) at each center will ensure that the subject is given full and adequate 
oral and written information about the nature, purpose, possible risk and benefit of the study. 
Subjects must also be notified that they are free to discontinue from the study at any time. The 
subject should be given the opportunity to ask questions and allowed time to consider the 
information provided. 


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The subject’s signed and dated informed consent must be obtained before conducting any 
study procedures. 
The Principal Investigator(s) must maintain the original, signed Informed Consent Form. A 
copy of the signed Informed Consent Form must be given to the subject. 
11.1 
Study Monitoring 
 
During the study, a monitor from Enterin, Inc. or representative will have regular contacts with 
the investigational site, for the following: 
• Provide information and support to the investigator(s) 
 
• Confirm that facilities remain acceptable.  Confirm that the investigational team is adhering 
to the protocol, that data are being accurately recorded in the CRFs, and that investigational 
product accountability checks are being performed, and perform source data verification. 
This includes a comparison of the data in the CRFs with the subject’s medical records at the 
hospital or practice, and other records relevant to the study. This will require direct access to 
all original records for each subject (e.g. clinic charts). 
 
• Record and report any protocol deviations not previously sent to Enterin, Inc. 
 
• Confirm adverse events and serious adverse events have been properly documented on 
CRFs and confirm any serious adverse events have been forwarded to Enterin, Inc. and 
those serious adverse events that met criteria for reporting have been forwarded to the 
IRB. 
 
The CRA monitor will be available between visits if the investigator(s) or other staff 
needs information or advice.  
 
11.2 
Audits and Inspections 
 
Authorized representatives of Enterin, Inc., a regulatory authority, an Independent Ethics 
Committee (IEC) or an Institutional Review Board (IRB) may visit the site to perform audits 
or inspections, including source data verification. The purpose of an Enterin, Inc. audit or 


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inspection is to systematically and independently examine all study-related activities and 
documents to determine whether these activities were conducted, and data were recorded, 
analyzed, and accurately reported according to the protocol, Good Clinical Practice (GCP) 
guidelines of the International Conference on Harmonization (ICH), and any applicable 
regulatory requirements. The investigator should contact Enterin, Inc. immediately if contacted 
by a regulatory agency about an inspection. 
11.3 
Ethics Committee Review 
 
The final study protocol, including the final version of the Informed Consent Form, must be 
approved or given a favorable opinion in writing by an IRB or IEC as appropriate. The 
investigator must submit written approval to Enterin, Inc. before he or she can enroll any subject 
into the study. 
The Principal Investigator is responsible for informing the IRB or IEC of any amendment to the 
protocol in accordance with local requirements. In addition, the IRB or IEC must approve all 
advertising used to recruit subjects for the study. The protocol must be re-approved by the IRB or 
IEC upon receipt of amendments and annually, as local regulations require. 
 
The Principal Investigator is also responsible for providing the IRB with reports of any 
reportable serious adverse drug reactions from any other study conducted with the 
investigational product. Enterin, Inc. will provide this information to the Principal Investigator. 
Progress reports and notifications of serious adverse drug reactions will be provided to the IRB 
or IEC according to local regulations and guidelines. 
11.4 
Standards 
 
The study will be performed in accordance with ethical principles that have their origin in the 
Declaration of Helsinki and are consistent with ICH/GCP, applicable regulatory requirements. 
11.5 
Confidentiality 
 
Any research information obtained about the subject in this study will be kept confidential. A 
subject will not be identified by name, only by his/her initials. The subject’s name or any 
identifying information will not appear in any reports published as a result of this study. 


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However, information obtained from individual subject’s participation in the study may be 
disclosed with his/her consent to the health care providers for the purpose of obtaining 
appropriate medical care. The subject’s medical records/charts, tests with his/her name on them 
may be made available to the appropriate contract research organization (CRO), the sponsor, its 
potential eventual partners, and any other regulatory authorities. This is for the purpose of 
verifying information obtained for this study. Confidentiality will be maintained throughout the 
study within the limits of the law. 
A subject’s name will not be given to anyone except the researchers conducting the study, who 
have pledged an oath of confidentiality. All identifying information will be kept behind locked 
doors, under the supervision of the study Investigator and will not be transferred outside of the 
investigator site. 
A subject may take away his/her permission to collect, use and share information about him/her 
at any time. If this situation occurs, the subject will not be able to remain in the study. No new 
information that identifies the subject will be gathered after that date. However, the information 
about the subject that has already been gathered and transferred may still be used and given to 
others as described above in order to preserve the scientific integrity and quality of the study. 
11.6 
Protocol Adherence 
 
The site will maintain records of study treatment delivered to the study site; the inventory at the 
site; the distribution to and use by each subject; and the return of materials to Enterin, Inc. for 
storage or disposal. These records should include dates, quantities, batch/serial numbers, 
expiration dates, in-clinic temperature log, and unique code numbers assigned to the product 
and study subjects. 
At each visit after initiation of treatment, site staff will record compliance of subjects with their 
assigned regimen. Subjects will be instructed to return their diaries and unused/partially 
used/empty bottles for inspection at each study visit. Subjects are to be reminded of the 
importance of compliance with their assigned regimen, with an emphasis on taking their study 
drug on schedule and maintaining the prescribed interval between doses. 
Investigator will maintain records that document adequately that the subjects were provided 
with the correct study drug and will reconcile the products received from the drug dispensing 


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center. Investigational product will not be returned to Enterin, Inc. until accountability has been 
fully monitored. 
Medication containers must be returned at each visit, as compliance will be assessed by tablet 
counts. Noncompliance is defined as taking less than 80% or more than 120% of study drug 
during any out-patient evaluation period (visit to visit). Discontinuation for noncompliance is 
at the Investigator’s discretion and is to be noted on the CRF. 
11.7 
Amendments to the Protocol 
 
Modifications to the protocol are only possible by approved protocol amendments authorized 
by the study sponsor. All protocol amendments will be approved by the appropriate regulatory 
authorities as well as each institutional review board prior to implementation. The Investigator 
must not implement any deviations from, or changes to the protocol, except where it is necessary 
to eliminate an immediate hazard to the study subject. 
 
11.8 
Protocol Deviations 
 
The protocol must be read thoroughly, and the instructions followed exactly. The sponsor and/or 
designee will not grant waivers for protocol deviations. Any deviation to the protocol must be  
reported as soon as possible to the sponsor and to the IRB per IRB reporting requirements. 
These reporting guidelines for protocol deviations must be adhered to by the Investigator. 
11.9 
Study Termination 
 
This study may be prematurely terminated, if in the opinion of the Investigator or Enterin, there 
is sufficient reasonable cause. Written notification documenting the reason for study termination 
will be provided to the Investigator or Enterin by the terminating party. Circumstances that may 
warrant termination include, but are not limited to: 
• Determination of unexpected, significant, or unacceptable risk to subjects. 
 
• Failure to enroll subjects at an acceptable rate. 
 
• Insufficient adherence to protocol requirements. 
 
• Insufficient complete and/or evaluable data. 
 
• Plans to modify, suspend, or discontinue the development of ENT-01. 


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Should the study be closed prematurely, all study materials must be returned to Enterin. 
 
12 
DATA HANDLING AND RECORD KEEPING 
 
12.0 
Inspection of Records 
 
Enterin, Inc. will be allowed to conduct site visits to the investigation facilities for the purpose 
of monitoring any aspect of the study. The Investigator agrees to allow the monitor to inspect 
the drug storage area, study drug stocks, drug accountability records, subject charts and study 
source documents, and other records relative to study conduct. 
12.1 
Data Management 
 
All data relating to study procedures will be entered on to electronic CRFs provided by the 
sponsor. All forms must be completed electronically. All requested information must be entered 
on the electronic CRF. If an item is not available or not applicable this fact should be indicated 
by the use of "NA". Spaces should not be left blank. Electronic CRF will be reviewed during 
the monitoring visits. Data will be entered into a database. Data entry and management and the 
production of the clinical study report will be the responsibility of the sponsor or a designated 
agent. 
The data cut-off for the study will be 60 days after entry of the last subject. If subjects remain 
on study thereafter, a revised final study report will be issued after the last subject has completed 
the final study evaluation. 
12.2 
Data Capture and Management 
 
The sponsor will provide the study sites with an electronic case report system. 
 
Electronic CRFs will be completed for each subject. It is the Investigator’s responsibility to 
ensure the accuracy, completeness, and timeliness of the data entered in each subject’s eCRF. 
Source documentation supporting the eCRF data should indicate the subject’s participation in 
the study and document the dates and details of study procedures, adverse events, and subject 
status. 
The Investigator, or designated representative, should complete the eCRF as soon as possible 
after information is collected, preferably on the same day that a subject is seen for an 


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examination, treatment, or any other study procedure. Any outstanding entries must be entered 
immediately after the final examination. An explanation should be given for all missing data. 
12.3 
Liability and Insurance 
 
The sponsor has subscribed to an insurance policy covering, in its terms and conditions, its legal 
liability for certain injuries to participating persons arising out of this research performed strictly 
in accordance with the scientific protocol as well as with applicable law and professional 
standards. 
12.4 
Retention of Records 
 
Study records and source documents must be preserved for the longer of (i) two (2) years 
following completion of the study; or (ii) two (2) years following the termination or withdrawal 
of the Investigational New Drug application under which this study was conducted; or (iii) the 
period required by local, state, and federal laws, regulations and FDA Guidance. 
 
The Investigator agrees to comply with all applicable federal, state, and local laws and 
regulations relating to the privacy of subject health information, including, but not limited to, 
the Standards for Individually Identifiable Health Information, 45 CFR, Parts 160 and 164 (the 
Health Insurance Portability Accountability Act of 1996 [HIPAA] Privacy Regulation). The 
Investigator shall ensure that study subjects authorize the use and disclosure of protected health 
information in accordance with HIPAA Privacy Regulation and in a form satisfactory to the 
Enterin, Inc. 
12.5 
Data Quality Assurance 
 
As per GCP guidelines, the sponsor or designee will be responsible for implementing and 
maintaining quality assurance and quality control systems for this study. 
Participating sites, the study database, and study documentation including subject medical 
records may be subject to a quality assurance audit during the course of the study. In addition, 
inspections may be conducted by regulatory bodies at their discretion. 
If sites receive a request for an inspection or written or oral inquiries regarding any aspect of 


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institution’s or Investigator’s activities related to this study from a regulatory authority, the 
Investigator must immediately notify the sponsor and the appropriate CRO of the request. 
Following this inspection and/or audit, the Investigator must notify the sponsor of any violation 
or deficiency noted by the regulatory authority. 
 
13 
USE OF INFORMATION 
 
Both the use of data and the publication policy are detailed within the clinical study agreement. 
Intellectual property rights (and related matters) generated by the Investigator and others 
performing the clinical study will be subject to the terms of a clinical study agreement that will 
be agreed between the Institution and Enterin, Inc. or their designee. With respect to such rights, 
Enterin, Inc. or its designee will solely own all rights and interests in any materials, data, and 
intellectual property rights developed by Investigators and others performing the clinical study 
described in this protocol, subject to the terms of any such agreement. In order to facilitate such 
ownership, Investigators will be required to assign all such inventions directly to Enterin, Inc. 
or its designee, as will be set forth in the clinical study agreement. 


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14 
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22. Perni, M., Galvagnion, C., Maltsev, A., Meisl, G., Muller, M.B., Challa, P.K., Kirkegaard, J.B., 
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APPENDIX 1 
LABORATORY TESTS 
 
Hematology 
Serum Chemistry 
Urinalysis 
White Blood Cell (WBC) Count 
Albumin 
Appearance 
Red Blood Cell (RBC) Count 
Alanine Aminotransferase (ALT) 
pH 
Hemoglobin (Hb) 
Alkaline Phosphatase (ALP) 
Protein 
Hematocrit (Hct) 
Aspartate Aminotransferase (AST) 
Glucose 
Mean Corpuscular Volume 
(MCV) 
Blood Urea Nitrogen (BUN) 
Ketone Bodies 
Red Blood Cell Distribution 
Width 
Ratio 
Indicators of Blood and WBCs 
(RDW) 
Calcium 
Specific Gravity 
Platelet Count 
Carbon Dioxide 
Urobilinogen 
Differential - absolute and 
percent 
Chloride 
of: 
Creatinine 
Neutrophils 
Glucose 
Lymphocytes 
Potassium 
Monocytes 
Sodium 
Eosinophils 
Total Bilirubin 
Basophils 
PT 
PTT 
INR 
Total Protein 
Urea Nitrogen 
Lactate Dehydrogenase (LDH) 
Creatine Kinase, Total 
Pregnancy tests 
A serum pregnancy test will be performed on all female subjects of child-bearing potential at the screening visit. 
Urine Pregnancy Test (on site): Urine human chorionic gonadotropin (HCG) (pre-menopausal females only) 
Follicle Stimulating Hormone (FSH) for post-menopausal women under age 60. 


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APPENDIX 2 
SCHEDULE OF EVENTS 
 
Visit 1 
Screening 
Visit 2 
Randomization 
 
Visit 3 
Follow-Up 
Visit 4 
End of 
Treatment 
Visit 5 
End of 
Placebo 
Visit 6 
End of 
Study 
Early 
Termination 
Day -21 
Day 1  
(+/-8 days) 
 
Day 11 
(+/- 3 day) 
Day 26 
(- 2 days) 
Day 39 
(+/- 8 days) 
Day 69  
(+/- 8 days) 
Informed Consent 
X 
 
Inclusion/Exclusion Criteria a 
X 
X 
 
Demographics/Medical History 
X 
 
Complete Physical Exam 
X 
 
X 
Brief Physical Exam 
X 
X 
X 
X 
Height 
X 
 
Weight 
X 
X 
X 
X 
X 
X 
X 
Orthostatic Vital Signsb 
X 
X 
X 
X 
X 
X 
X 
12-lead ECG 
X 
X 
X 
X 
X 
X 
X 
Labs and Urinalysis 
X 
X 
X 
X 
X 
X 
Pregnancy Testingc 
X 
X 
X 
X 
X 
X 
X 
In-clinic IP/Placebo Dosingd 
X 
X 
X 
In-clinic Levodopa Dosing 
X 
X 
X 
X 
X 
X 
Levodopa PK Drawse 
X 
X 
X 
X 
X 
X 
IP/Placebo Dispense 
X 
X 
X 
IP/Placebo Collection and 
Accountability 
X 
X 
X 
X 
X 


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Visit 1 
Screening 
Visit 2 
Randomization 
Visit 3 
Follow- 
Up 
Visit 4 
End of 
Treatment 
Visit 5 
End of 
Placebo 
Visit 6 
End 
of Study 
 
 
 
 
Early 
Termination 
Day -21 
Day 1  
(+/-8 days) 
Day 11 
(+/- 3 day) 
 
Day 26 
(- 2 days) 
 
Day 39 
(+/- 8 days) 
 
Day 69  
(+/- 8 days) 
Instructions to Discontinue 
Laxatives and other BM 
Medications 
X 
 
 
 
 
Dispense Rescue Medicationsf 
X 
X 
X 
X 
X 
 
 
Stool Diary Instruction/Reviewg 
X 
X 
X 
 
X 
 
X 
 
X 
 
X 
AE Review 
X 
X 
X 
X 
X 
X 
X 
Prior and Con Meds 
X 
X 
X 
X 
X 
X 
X 
Rome IV questionnaire 
X 
 
 
 
 
MDS-UPDRSh 
X 
X 
X 
X 
X 
X 
C-SSRS 
X 
X 
X 
X 
X 
X 
X 
BDI-II 
X 
X 
X 
X 
X 
X 
MMSE 
X 
X 
X 
X 
X 
X 
SAPS-PD, 
Hallucination/Delusion 
Questionnairei 
X 
X 
X 
 
X 
 
X 
 
X 
 
X 
PAC-QOL and PAC-SYM 
X 
X 
X 
X 
X 
X 
Dispense Stool Collection Kits 
X 
X 
            
 
 
 
Collect Stool Samplej 
X 
X 
          
 
           X 


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a. 
Confirmation of eligibility: To be randomized to receive double-blind study medication in this study, subjects must meet the following criteria: 
• 
Have had at least 11 “Available Data Days” (days with electronic diary compliance sufficient for determining whether a BM occurred) 
• 
Have had <3 CSBMs/week based upon the average CSBM rate during the first 2 screening weeks (baseline period). 
• 
Be approved for randomization by the enrollment committee 
 
b. 
Three sets will be taken. Time points to be defined. Orthostatic vital signs will include supine BP and HR, sitting or standing BP and HR, RR, and body temperature. 
Supine measurements will be taken after the subject has been in the supine position for approximately 5 minutes. Sitting or standing measurements will be taken within 
approximately 5 minutes  of sitting or standing.  
c. 
For women of child-bearing potential, on-site with urine pregnancy test kit. If the on-site UPT is positive, a serum pregnancy test will be done. For post- menopausal 
women <60 years or of age, a serum FSH will be done at the screening visit only. 
 
d. 
Sites will use a centralized computer-based system to obtain the randomization bottle(s) assigned to each subject. 
 
• 
Subjects with baseline CSBMs from 0 to 0.9 will be randomized to begin treatment with ENT-01 150 mg or 6 tablets of placebo. 
• 
Subjects with baseline CSBMs from 1.0 to 3.0 will be randomized to begin treatment with ENT-01 75 mg or 3 tablets of placebo. 
• 
Subjects will self-administer study medication at home. The dose will be taken upon awakening, on an empty stomach with 8 oz. of water. 
• 
Subjects will be advised to stay at home for 4 to 6 hours after taking the first dose of medication in case there is a need to evacuate urgently. After the 
first dose, subjects will know if and when to leave the house. 
 
e. 
A subset of 20 subjects from this study population will participate in an L-Dopa PK Study, as follows: 
• 
Visits 2 and 6: Blood draws for L-dopa levels (PK) will be drawn at Time zero. 
• 
The first daily dose of L-Dopa will be administered on site after the first blood draw. There will be another blood draw at 20, 40, and 60 minutes post- L- 
Dopa dose 
• 
Visits 3. 4, and 5: Immediately upon arrival at the study site, the subject will take study medication (ENT-01 or placebo) on an empty stomach. Blood will 
be drawn at Time zero. 30 to 60 minutes after taking ENT-01 or placebo, the subject will take their first daily dose of L-Dopa. Blood will be drawn again 
20, 40, and 60 minutes post-L-Dopa dose. The subject may have food 60 minutes after taking ENT-01. 
 
f. 
Rescue medications will be dispensed as needed and instructions given concerning laxatives and other medications to treat constipation. 
 
g. 
Subjects will be educated concerning use of the electronic diary to report stools immediately after a bowel movement 
 
h. 
The first 3 parts of the MDS-UPDRS should be administered while the subject is in an ON state, approximately one hour after taking L-Dopa (if applicable). In 
some cases, patients will reach the “on” state earlier or later than at 60 minutes. In those cases, MDS-UPDRS should be performed when the patient is “on”. 
 
i. 
Those subjects who have a SAPS-PD score >0 on Visits 1 and 2 will have the SAPS-PD and hallucination/delusion questionnaire at all subsequent visits. 
• 
If only Visit 1 or Visit 2 has a score >0, then SAPS-PD will not be repeated in subsequent visits.  
j. 
The stool sample should be collected during the EoT visit.  If the subjects discontinues from the study prior to the EoT visit, a stool sample should be collected, as applicable, at 
the Early Termination visit.  


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AE=adverse event; BDI-II= Beck Depression Inventory II; BM=bowel movement; BP=blood pressure; C-SSRS= Columbia-Suicide Severity Rating Scale; ECG=electrocardiogram; 
EoT=end of treatment; FSH=follicle-stimulating hormone; HR=heart rate; L-Dopa=levodopa; MDS-UPDRS=Movement Disorder Society’s Unified Parkinson’s Disease Rating Scale; 
MMSE=Mini-mental State Examination; PAC-QoL=Patient Assessment of Constipation - Quality of Life; 
PAC-SYM=Patient Assessment of Constipation Symptoms; PK=pharmacokinetic(s); RR=respiratory rate; SAPS-PD=Scale for Assessment of Positive Symptoms – Parkinson’s 
Disease; UPT=urine pregnancy test. 


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75 75  75  100 100 100 125 125 125 150 150 150 175 175 175 175 175 175 175 175 175 175 175 175 175 7pill 7pills 7pill 7pi 7pil 7pil7pill7pil 7pil 7pil 7pil 7pil 7pil 4wk WO 
75   75   75   100 100 100 125 125 125 150 150 150 150 150 150 150 150 150 150 150 150 150 150 150 150 6 pill6 pill 6 pil 6 p 6 pi 6 pi 6 pil6 pi 6 pi 6 pi 6 pi 6 pi 6 pi 6 pi 4wk WO 
75   75   75   100 100 100 125 125 125 125 125 125 125 125 125 125 125 125 125 125 125 125 125 125 125 5 pill5 pill 5 pil 5 p 5 pi 5 pi 5 pil5 pi 5 pi 5 pi 5 pi 5 pi 5 pi 5 pi 4wk WO 
1‐3 CSBM/wk 
75    75     75 
100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 4 pill4 pill 4 pil 4 p 4 pi 4 pi 4 pil4 pi 4 pi 4 pi 4 pi 4 pi 4 pi 4 pi 4wk WO 
75    75     75 
75 75 75 75 75 75 75 75 75 75 75 75 75 75 75 75 75 75 75 75 75 75 3 pill3 pill 3 pil 3 p 3 pi 3 pi 3 pil3 pi 3 pi 3 pi 3 pi 3 pi 3 pi 3 pi 4wk WO 
 
 
APPENDIX 3 
DOSING SCHEDULE 
 
 
 
 
 
 
 
 
week 1 and 2 d 1 2d d3 d4 d5 d6 d7 d8 d9 d10 d11 d12 d13 d14 d15 d16 d17 d18 d19 d20 d21 d22 d23 d24 d25 d26 d27 d28 d29d30d31d32 d33 d34 d35 d36 d37 d38 d39 d40 
KARMET DOSING SCHEDULE 
 
ENT‐01 cohort 
Screening 
Placebo cohort 


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APPENDIX 4 
ROME-IV CRITERIA FOR CONSTIPATION 
 
Must include two or more of the following: 
Straining during at least 25% of defecations 
Lumpy or hard stools in at least 25% of defecations 
Sensation of incomplete evacuation for at least 25% of defecations 
Sensation of anorectal obstruction/block age for at least 25% of defecations 
Manual maneuvers to facilitate at least 25%of defecations (e.g., digital evacuation, support of 
the pelvic floor) 
Fewer than 3 defecations per week 
Loose stools are rarely present without the use of laxatives 


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APPENDIX 5 
EASE OF PASSAGE 
 
 
1.Manual disimpaction 
2.Enema needed 
3.Straining needed 
4.Normal 
5.Urgent without pain 
6.Urgent with pain 
7.Incontinent 


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APPENDIX 6      BRISTOL STOOL CHART 
 
The assessment of stool consistency is a validated surrogate of intestinal motility (Lewis and 
Heaton, 1997). We have chosen the Bristol Stool Chart as a subject-friendly means of 
categorizing stool characteristics. 


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APPENDIX 7          SUBJECT DIARIES 
 
Participants will complete a stool diary on an IPAD tablet on a daily basis throughout the study. 
The diaries will include information about whether the stool was a CSBM as well information 
about consistency, ease, laxative and rescue medication use. 
 
 
 


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APPENDIX 8 
PD HALLUCINATION QUESTIONNAIRE 
 
Many patients with Parkinson’s disease suffer from what we call hallucinations or apparitions. 
We would like to ask you some questions regarding hallucinations. 
a. Do you sometimes feel someone is present who isn’t actually there? 
 
b. Do you sometimes think someone went by who didn’t? If so, any one in particular? 
 
c. Do you see things not seen by others? If so, how many months or years have you been 
seeing things? what do you see? People? Animals? Relatives? Children? When do you see 
them? Daytime? Evening? At night? Are they bothersome? Do they upset you? How often 
does this occur, many times a day? every day? Twice a week? Once a month? Can you 
provide a narrative description of them below? 
d. Do you mistake objects for animals, like a white cloth for a white cat? 
 
e. Do you feel like someone is going to harm you or is out to get you? 
 
f. Do you feel things such as insects crawling up your legs on your hands or strange 
sensations such as scratching or burning or tingling on your hands? 
g. Do you hear voices that aren’t actually there? Do the voices talk to you? To each other? 
Talk about you? 
h. Do you smell any abnormal smells not smelled by others? Are they unpleasant smells? 
 
Please describe your experience in detail (i.e. what do you hear, see, smell or feel? Make sure 
you note how many times a day or night or week any of these episodes occur) 
 
 
 
 
 
 
 
 
 


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i. 
Have you been diagnosed with any eye disease? How good is your eyesight? If it’s poor, 
how bad is it? How long have you had poor eyesight? How fast has it been getting 
worse? 
ii. 
Do you experience hallucinations while “on” or “off”? 
 
iii. 
Has your dopamine dose been changed recently? 
 
iv.  
v. 
 
Interviewer’s Signature: 
 
Date: 
/ 
/  
 
 
 
Title:  
 


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PI Review of Interview 
 
Once completed by or reviewed by the PI, please enter any observations or notes of 
significance regarding this patient. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
PI Signature: 
 Date: 
/ 
/  
 
 
DD 
MMM 
YYYY 

SAP_001.pdf: 
 
Document No.: ENT-01-30 Statistical Analysis Plan  Final 
Version 2.0 
 
STATISTICAL ANALYSIS PLAN  
ENT-01-30 
A Multicenter, Randomized, Double-Blind, Placebo-Controlled, Multiple Dose Study to 
Evaluate Safety, Tolerability and Efficacy or Orally Administered ENT-01 for the 
Treatment of Parkinson’s Disease-Related Constipation (KARMET) 
 
Protocol Number: 
 
ENT-01-30 
 
Name of Test Drug: 
ENT-01 
Phase: 
2b 
Methodology: 
Double-Blind, Placebo-Controlled, Randomized Trial 
Sponsor: 
Enterin Inc. 
2005 Market Street, Philadelphia, PA 19103 
Sponsor Representative: 
Denise Barbut, MD FRCP  
President and CMO  
Document Date: 
28MAY2020 
Document Version: 
Final v2.0 
 


 
Document No.: ENT-01-30 Statistical Analysis Plan  Final 
Version 2.0 
 
CONFIDENTIAL 
2 
SIGNATURE PAGE 
Protocol Title: 
A Multicenter, Randomized, Double-Blind, Placebo-
Controlled, Multiple Dose Study to Evaluate Safety, 
Tolerability and Efficacy of Orally Administered ENT-
01 for the Treatment of Parkinson’s Disease-Related 
Constipation (KARMET) 
Sponsor: 
Enterin Inc. 
2005 Market Street, Philadelphia, PA 19103 
Protocol Number: 
ENT-01-30 
Document Date/Version: 
31DEC2018 Version 2.0 
Cytel, Inc. Author: 
Adam Hamm 
Cytel, Inc. 
460 Totten Pond Road, Suite 640 
Waltham, MA 02451 
 
Signature:____________________________________ 
Date:________________________________________ 
Sponsor Approval 
By signing this document, I acknowledge that I have read the document and approve of the 
planned statistical analyses described herein.  I agree that the planned statistical analyses are 
appropriate for this study, are in accordance with the study objectives, and are consistent with 
the statistical methodology described in the protocol, clinical development plan, and all 
applicable regulatory guidances and guidelines. 
I have discussed any questions I have regarding the contents of this document with the 
biostatistical author. 
I also understand that any subsequent changes to the planned statistical analyses, as described 
herein, may have a regulatory impact and/or result in timeline adjustments.  All changes to the 
planned analyses will be described in the clinical study report (CSR). 
Sponsor Signatory: 
Denise Barbut, MD FRCP  
Enterin Inc. 
2005 Market Street 
Philadelphia, PA 19103 
 
Signature:____________________________________ 
Date:________________________________________ 
 
 
 
 
 


 
Document No.: ENT-01-30 Statistical Analysis Plan  Final 
Version 2.0 
 
CONFIDENTIAL 
3 
REVISION HISTORY 
 
Date 
Version number 
Reason for change 
Authored by 
21AUG2018 
0.1 
First Draft 
Jianfen Shu 
26APR2019 
0.2 
Second Draft – to be 
consistent with 
updated analyses 
Adam Hamm 
7MAY2019 
Final 
Final Version 
Adam Hamm 
28May2020 
Final v2 
Updated due to 
additional patient 
cohort added to study 
Adam Hamm 
 
 
 


 
Document No.: ENT-01-30 Statistical Analysis Plan  Final 
Version 2.0 
 
CONFIDENTIAL 
4 
TABLE OF CONTENTS 
ABBREVIATIONS ................................................................................................................. 6 
1. 
INTRODUCTION AND OBJECTIVES OF ANALYSIS .......................................... 9 
1.1. 
Introduction ...................................................................................................... 9 
1.2. 
Objectives of Statistical Analysis ................................................................... 10 
1.2.1 Primary Objective .................................................................................... 10 
1.2.2 Secondary Objectives .............................................................................. 10 
1.2.3 Exploratory Objectives ............................................................................ 10 
2. 
STUDY DESIGN ....................................................................................................... 11 
2.1. 
Synopsis of Study Design ............................................................................... 11 
2.2. 
Randomization Methodology ......................................................................... 11 
2.3. 
Stopping Rules and Unblinding ..................................................................... 12 
2.4. 
Study Procedures ........................................................................................... 13 
2.5. 
Study Periods .................................................................................................. 17 
2.6. 
Efficacy, Pharmacokinetic, Safety and Tolerability Endpoints .................... 17 
2.6.1 Efficacy Endpoints .................................................................................. 17 
2.6.4 Pharmacokinetic (PK) Endpoints ............................................................. 20 
2.6.5 Safety Endpoints ...................................................................................... 20 
2.6.6 Tolerability Endpoints ............................................................................. 20 
3. 
SUBJECT POPULATIONS ...................................................................................... 22 
3.1. 
Population Definitions .................................................................................... 22 
3.2. 
Protocol Violations ......................................................................................... 22 
4. 
STATISTICAL METHODS ...................................................................................... 23 
4.1. 
Sample Size Justification ................................................................................ 23 
4.2. 
General Statistical Methods and Data Handling........................................... 23 
4.3. 
Interim Analyses............................................................................................. 25 
4.4. 
Subject Disposition ......................................................................................... 25 
4.5. 
Demographic and Baseline Characteristics ................................................... 26 
4.6. 
Efficacy Analyses ............................................................................................ 26 
4.6.1 Derivation of Weekly CSBM and SBM Rates .......................................... 26 
4.6.2 Stool Consistency and Ease of Passage Analysis Conventions ................. 29 
4.6.3 Analysis of the Primary Efficacy Endpoint .............................................. 30 
4.6.4 Analysis of Secondary Efficacy Endpoints ............................................... 30 


 
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4.6.5 Analysis of Exploratory Efficacy Endpoints ............................................ 31 
4.7. 
Pharmacokinetic Analysis .............................................................................. 32 
4.8. 
Safety Analyses ............................................................................................... 33 
5. 
CHANGES TO PLANNED ANALYSES .................................................................. 37 
6. 
References .................................................................................................................. 38 
 
 
 


 
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ABBREVIATIONS 
Abbreviation 
Definition of Terms 
AE 
Adverse Events  
ADL 
Activities of Daily Living 
ANCOVA 
Analysis of Covariance 
ATC 
Anatomic Therapeutic Class 
BDI 
Beck Depression Inventory 
CMH 
Cochran-Mantel-Haenszel 
CSBM 
Complete Spontaneous Bowel Movement 
CSR 
Clinical Study Report 
C-SSRS 
Columbia Suicide Severity Rating Scale 
DLT 
Dose limiting Toxicity 
ECG 
Electrocardiogram 
eCRF 
Electronic Case Report Form 
ENS 
Enteric Nervous System 
EOS 
End of Study 
ET 
Early Termination 
GI 
Gastrointestinal 
ICH 
International Conference on Harmonization 
ID 
Identification 
IRT 
Interactive Response Technology 
IXRS 
Interactive voice/web-response system 
MedDRA 
Medical Dictionary for Regulatory Activities 
MMRM 
Mixed Model Repeated Measures 


 
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Abbreviation 
Definition of Terms 
MMSE 
Mini Mental State Examination  
mITT 
Modified Intention to Treat 
NCI-CTCAE 
National Cancer Institute- Common Terminology Criteria for Adverse 
Events 
PAC-QoL 
Patient Assessment of Constipation Quality of Life 
PAC-SYM 
Patient Assessment of Constipation Symptoms 
PD 
Parkinson’s Disease 
PK 
Pharmacokinetics 
PT 
Preferred Term 
QOL 
Quality of Life 
RASMET 
Evaluation of Safety and Tolerability of ENT-01 for the Treatment of 
Parkinson’s Disease Related Constipation 
SAE 
Serious Adverse Event 
SAP 
Statistical Analysis Plan 
SAPS-PD 
The Scale for the Assessment of Positive Symptoms -Parkinson's Disease 
SBM 
Spontaneous Bowel Movement 
SD 
Standard Deviation 
SOC 
System Organ Class 
TEAEs 
Treatment Emergent Adverse Events 
ULN 
Upper limit of normal 
UPDRS 
Unified Parkinson’s Disease Rating Scale 
US  
United States 
WHO 
World Health Organization  


 
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Abbreviation 
Definition of Terms 
WHO-DD 
World Health Organization Drug Dictionary  
 


 
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1. 
INTRODUCTION AND OBJECTIVES OF ANALYSIS 
1.1. 
Introduction 
Constipation is a common problem worldwide, affecting 2% to 27% of the population, with 
most estimates varying from 12% to 20% (Bouras and Tangalos, 2009). The prevalence of 
constipation increases to 30%-40% among people aged >65 years and women are 
disproportionately affected. Constipation is much more common among patients with 
Parkinson’s Disease (PD) than in the general population. There are 1 million people suffering 
from Parkinson’s Disease in the US, of which roughly 60%, or 600,000 suffer from chronic 
constipation and in most, the condition is chronic, severe and unresponsive to standard therapy 
(Ondo et al., 2012; Zangaglia et al., 2007).  Constipation not only constitutes a major economic 
burden, but it also significantly affects the quality of life of the individual with Parkinson’s 
disease. An effective prokinetic medication for individuals with PD would be a useful addition 
to the currently available treatments for this disabling condition. 
 
The individual with Parkinson’s Disease suffers from a form of constipation that is believed to 
be caused principally by delayed transit through the colon. In addition, defecation is often 
impaired by dysfunction of the Parkinson’s Disease subject’s anorectal reflex.  
 
The pathophysiology of the gastrointestinal (GI) dysfunction in Parkinson’s Disease involves 
deposition of alpha-synuclein within both the enteric nervous system (ENS) as well as within 
the brainstem. For reasons that remain unknown alpha-synuclein, which is a protein normally 
produced in neurons, forms neurotoxic intracellular aggregates in Parkinson’s Disease.  
Squalamine is a small molecule, originally discovered in the liver of the dogfish shark, where it 
is delivered following a meal via the biliary tract into the intestine (Moore et al., 1993). In a 
recent study, squalamine has been shown to prevent the formation of toxic alpha-synuclein 
aggregates (Perni et al., 2017). Squalamine both excites the neurons of the ENS and protects 
them from alpha-synuclein induced cytotoxicity. We will evaluate squalamine as a therapeutic 
to treat the constipation associated with Parkinson’s Disease and we will explore its effect on 
sleep architecture and hallucinations/delusions. 
 
A Phase 2a study, Evaluation of Safety and Tolerability of ENT-01 for the Treatment of 
Parkinson’s Disease Related Constipation (RASMET) was conducted. The purpose of the study 
was to assess the safety, tolerability, pharmacokinetics, and pharmacodynamics of an orally 
administered medication to relieve symptoms of constipation associated with Parkinson's 
Disease. The results of the study indicated that orally administered ENT-01 was safe and 
significantly improved bowel function and neurologic symptoms. Minimal systemic absorption 
suggests that improvements result from local stimulation of the ENS. Data from this study 
informed the development of the 2b study. 
 
 


 
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1.2. 
Objectives of Statistical Analysis 
This statistical analysis plan (SAP) is designed to outline the methods to be used in the analysis 
of study data in order to answer the study objectives. Populations for analysis, data handling 
rules, statistical methods, and formats for data presentation are provided. The statistical analyses 
and summary tabulations described in this SAP will provide the basis for the results sections of 
the clinical study report (CSR) for this trial. 
1.2.1 Primary Objective 
The primary objective is to evaluate the safety, tolerability, and efficacy of repeated oral doses 
of ENT-01 for up to 25 days in subjects with Parkinson’s Disease related constipation in a 
randomized, double-blind, placebo-controlled study. 
• Primary Safety Objective: The primary safety objective of this study will be evaluation 
of the safety of repeated daily oral doses of ENT-01 for up to 25 days in patients with 
PD- related constipation. 
• Primary Tolerability Objective: The primary tolerability objective will be evaluation 
of recurrent vomiting, recurrent diarrhea, abdominal pain and dizziness. 
• Primary Efficacy Objective: The primary efficacy objective will be evaluation of the 
efficacy of repeated daily oral doses of ENT-01 for up to 25 days in improving CSBM 
rate. 
1.2.2 Secondary Objectives 
The secondary objectives include: 
• To determine if the correct starting dose (75mg or 150mg) can be predicted from the 
baseline weekly CSBM rate. 
• To collect information on the effect of ENT-01 on CSBM, SBM, ease of passage, 
consistency, suppository/enema use and subjective bowel function and quality of life 
(QOL). 
1.2.3 Exploratory Objectives 
The exploratory objectives include: 
• To determine the decrease in efficacy in the blinded placebo withdrawal and washout 
periods. 
• To collect information on the effect of ENT-01 on motor and non-motor symptoms of 
Parkinson’s disease including mood, memory, sleep, and hallucinations/delusions. 
• To collect information on the effect of ENT-01 on levodopa absorption. 
• To collect information on the effect of ENT-01 on the stool microbiome. 
 
 


 
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2. 
STUDY DESIGN 
2.1. 
Synopsis of Study Design 
This study will be conducted as a multicenter, randomized, double-blind, placebo-controlled 
study in patients with Parkinson’s Disease and constipation.  
 
This study has two parts. The first part of this randomized approximately 72 subjects in a 3:1 to 
treatment or placebo ratio (double-blind), with approximately 54 subjects allocated to the 
treatment groups and approximately 18 to placebo. In the second part of the study, an additional 
80 patients (approximately) will be randomized 1:1 to treatment or placebo (double-blind) with 
approximately 40 subjects allocated to each group.  
 
In both parts of the study, the study was conducted as a multicenter, randomized, double-blind, 
placebo-controlled study. A subject’s total study duration was no more than 10 weeks and total 
active dosing days was a maximum of 25 days. 
 
During the active dosing period, patients will be able to increase/adjust their dose every three 
days. This is called the Dose Adjustment period. After the subject documents two complete 
spontaneous bowel movements in three days, the subject will fix at the dose for the remainder of 
the study. This is called the Fixed Dose period. After completing up to 25 days of dosing, all 
subjects will be given placebo for 2 weeks (single-blind) followed by 4 weeks of wash-out.  
 
 
2.2. 
Randomization Methodology 
Randomization occurs after all screening assessments have been completed and after the 
Investigator has verified that they are eligible. In the first part of the study, patients will be 


 
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randomized 3:1 to receive either ENT-01 or placebo; in the second part of the study patients are 
randomized 1:1 to receive either ENT-01 or placebo. Any patient identification numbers that are 
assigned will not be reused, even if the patient does not receive treatment. The patient’s 
identification (ID) number will be used on that patient’s eCRFs and serious adverse event (SAE) 
forms.  
Randomization will be conducted centrally via an Interactive voice/web-response system 
(IXRS). The randomization will be stratified by the baseline weekly CSBM rate. Subjects with a 
baseline CSBM rate of 0-0.9 will begin treatment with 150 mg of ENT-01 or 6 tablets of 
placebo and Subjects with a baseline CSBM rate of 1.0-2.9 will begin with 75 mg of ENT-01 or 
3 tablets of placebo. Note that in the second part of the study, the baseline CSBM criteria of 1.0 
– 2.9 was modified to 1.0 – 3.0 (i.e., including subjects with a baseline CSBM rate of 3.0).  
Instructions for patient screening, registration, number assignment, and enrollment are 
referenced in the Interactive Response Technology (IRT) vendor manual. 
2.3. 
Stopping Rules and Unblinding 
This study may be terminated at the discretion of the Enterin, Inc. or of any regulatory agency 
for reasons including safety and/or efficacy. 
It is anticipated that no more than 2 subjects out of 60 (3.3%) will have an AE of grade 4 or 5 
that is at least possibly related to ENT-01 in this Study. Should there be more than 2 subjects 
with an AE grade 4 or 5 that is at least possibly related to ENT-01 in the cohort of subjects 
randomized to ENT-01, the study will be put on an immediate clinical hold.  
In addition, individual safety stopping criteria will include:  
• Reaching DLT before a prokinetic effect  
• Having a non-DLT gastrointestinal adverse event > grade 3 within 24 hours of taking 
ENT-01 that is at least possibly attributable to ENT-01 
• A fall in systolic blood pressure to < 80 mm Hg upon rising from lying to sitting or 
standing  
• A fall in diastolic blood pressure to < 40 mm Hg upon rising from lying to sitting or 
standing  
• Elevation of liver function tests (LFTs) > 3 times the upper limit of normal (ULN) 
 
A treatment assignment should only be unblinded in a situation of urgent medical necessity, 
when the identity of the study medication must be known in order to select continuing therapy 
for the disease under study or an AE. The decision should be made only after consultation with 
the unblinded Medical Monitor, unless the urgency of a case requires immediate action. 
Treatment assignments can be obtained through a controlled transaction with IXRS. If a 
treatment code is unblinded for any reason, the Investigator will notify Sponsor within 24 hours 
and will document the following in the eCRF: who unblinded the code, the reason for doing so, 
and the date of unblinding. 


 
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2.4. 
Study Procedures 
The schedule of events, as outlined in the study protocol, is provided in Table 1. 
 
 
 


 
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Table 1. Schedule of Events 
 
 


 
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a. Confirmation of eligibility: To be randomized to receive double-blind study medication in this study, subjects must meet the 
following criteria:  
• 
Have had at least 11 “Available Data Days” (days with electronic diary compliance sufficient for determining whether 
a BM occurred)  
• 
Have had <3 CSBMs/week based upon the average CSBM rate during the first 2 screening weeks (baseline period).  
• 
Be approved for randomization by the enrollment committee  
 
b. Three sets will be taken. Time points to be defined. Orthostatic vital signs will include supine BP and HR, sitting or standing 
BP and HR, RR, and body temperature. Supine measurements will be taken after the subject has been in the supine position 
for at least 5 minutes. Sitting or standing measurements will be taken within 5 minutes of sitting or standing from the supine 
position. Respiration rate and body temperature will be taken after the subject has been sitting or standing for at least 5 
minutes.  
  
c. For women of child-bearing potential, on-site with urine pregnancy test kit. If the on-site UPT is positive, a serum pregnancy 
test will be done. For post- menopausal women <60 years or of age, a serum FSH will be done at the screening visit only.  
 
d. Sites will use a centralized computer-based system to obtain the randomization bottle(s) assigned to each subject.   
• 
Subjects with baseline CSBMs from 0 to 0.9 will be randomized to begin treatment with ENT-01 150 mg or 6 tablets 
of placebo.  
• 
Subjects with baseline CSBMs from 1.0 to 3.0 will be randomized to begin treatment with ENT-01 75 mg or 3 tablets 
of placebo.  
 
• 
Subjects will self-administer study medication at home. The dose will be taken upon awakening, on an empty stomach 
with 8 oz. of water.  
• 
Subjects will be advised to stay at home for 4 to 6 hours after taking the first dose of medication in case there is a need 
to evacuate urgently. After the first dose, subjects will know if and when to leave the house.  
 


 
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e. A subset of 20 subjects from this study population will participate in an L-Dopa PK Study, as follows:  
• 
Visits 2 and 6: Blood draws for L-dopa levels (PK) will be drawn at Time zero.  
• 
The first daily dose of L-Dopa will be administered on site after the first blood draw. There will be another blood draw 
at 20, 40, and 60 minutes post- L-Dopa dose  
• 
Visits 3. 4, and 5: Immediately upon arrival at the study site, the subject will take study medication (ENT-01 or 
placebo) on an empty stomach. Blood will be drawn at Time zero. 30 to 60 minutes after taking ENT-01 or placebo, 
the subject will take their first daily dose of L-Dopa. Blood will be drawn again 20, 40, and 60 minutes post-L-Dopa 
dose. The subject may have food 60 minutes after taking ENT-01.  
 
 
f. Rescue medications will be dispensed as needed and instructions given concerning laxatives and other medications to treat 
constipation.  
 
g. Subjects will be educated concerning use of the electronic diary to report stools immediately after a bowel movement   
 
h. The first 3 parts of the MDS-UPDRS should be administered while the subject is in an ON state, approximately one hour after 
taking L-Dopa (if applicable). In some cases, patients will reach the “on” state earlier or later than at 60 minutes. In those 
cases, MDS-UPDRS should be performed when the patient is “on”.  
 
i. Those subjects who have a SAPS-PD score >0 on Visits 1 and 2 will have the SAPS-PD and hallucination/delusion 
questionnaire at all subsequent visits.  
• 
If only Visit 1 or Visit 2 has a score >0, then SAPS-PD will not be repeated in subsequent visits.  
 
j. The stool sample should be collected during the EoT visit if the subject does not wish to return for the Washout Visit.  


 
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2.5. 
Study Periods  
The study consists of the following periods: 
1. Screening Period: This period lasts from Visit 1 to the day before Visit 2 (Day of 
randomization) 
2. Treatment Period: This period lasts from Visit 2 (day of randomization) to the day before 
Visit 4 (start of single blind placebo period) and is divided into two sub-periods 
2.1. Dose Escalation Period: This period starts at Visit 2 and ends when the subject is on a 
Fixed Dose of study medication. 
2.2. Fixed Dose Period: This period starts when the subject is on a Fixed Dose of study 
medication and lasts until Visit 4. The fixed dose period is only defined if the subject 
was in the treatment period for at least seven days after the date of randomization. In the 
event that a subject is on the same dose until the start of the single blind placebo period 
or until withdrawal from treatment after being in the treatment period for at least seven 
days after randomization, then the fixed dose period will be the entire treatment period 
and the dose escalation period will not be defined. If the subject was not on their fixed 
dose for at least seven days, then the fixed dose period will extend to the subject’s last 
seven days of treatment (including fixed dose) in order to derived rate variables during 
the fixed dose period, as described in Section 4.7. Missing diary entries in the middle of 
treatments of the same dose are not considered when determining the start of the fixed 
dose period. 
3. Single Blind Placebo Withdrawal Period: This period starts at Visit 4 and ends at Visit 5. 
Weekly rate derivations during the single blind placebo withdrawal period will not include 
assessments on the Visit 4 date. 
4. Wash Out Period: This period starts at Visit 5 and ends at Visit 6.   
 
2.6. 
Efficacy, Pharmacokinetic, Safety and Tolerability Endpoints 
2.6.1 Efficacy Endpoints 
Primary Efficacy Endpoint 
The primary efficacy endpoint is the change from baseline in the weekly CSBM rate during the 
Fixed Dose period.  
 
Baseline is defined as the weekly CSBM rate during the 2-week screening period. If a subject 
has more than two weeks of entries during the screening period, consecutive missing entries 
during the screening period will be excluded from baseline calculations, and the last fourteen 
days with entries during the screening period will be used to derive baseline weekly rates. 
 
Subjects with fewer than 11 days of diary data during screening would be excluded from the 
study. The primary efficacy endpoint will be obtained as the weekly CSBM rate during the 
Fixed Dose period with exceptions as noted in section 4.7. The frequency of CSBMs will be 


 
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determined via a web-based electronic diary where the passage of a bowel movement or absence 
thereof will be recorded daily throughout the study by the subject.   
 
2.6.2 Secondary Efficacy Endpoints 
The secondary efficacy endpoints include:  
• Frequency of dose adjustments in the Dose Adjustment period: The number of dose 
adjustments a subject has from the initial starting dose to the subject’s fixed dose. 
• Prokinetic bowel response during the Fixed Dose period: A prokinetic response is 
defined as a weekly CSBM rate increase of 1 or more compared to baseline or an 
absolute rate of 3 or more CSBMs per week.  
• Change from baseline in the weekly SBM rate during the Fixed Dose period: An SBM is 
a bowel movement which did not occur within 24 hours after taking rescue medication. 
• SBM bowel response to ENT-01 during the Fixed Dose period: An SBM bowel response 
is defined as a weekly SBM rate increase of 1 or more compared to baseline or an 
absolute rate of 3 or more SBMs per week.  
• Change from baseline in stool ease of passage during the Fixed Dose period: Stool ease 
of passage is rated on a 1-7 ordinal point scale (see APPENDIX 4 of the Protocol). A 
subject’s ease of passage score for a given time period is the average of the ease of 
passage scores associated with SBMs during that time period. 
• Change from baseline in stool consistency during the Fixed Dose period: Stool 
consistency is measured using the 7-point Bristol Stool Chart (see APPENDIX 5 of the 
Protocol). A subject’s stool consistency score for a given time period is the average of 
the stool consistency scores associated with SBMs during that time period.  
• Change from baseline in suppository/enema use during the Fixed Dose period: 
Suppository/enema use is measured as a weekly average during the Fixed Dose period. 
This is captured in the subject stool diary.  
• Time to first CSBM in the Fixed Dose period: Time to first CSBM in the Fixed Dose 
period is defined as the time from the subject taking the first dose of study medication in 
the Fixed Dose period until the subject has a CSBM.  
• Time to first SBM in the Fixed Dose period: Time to first SBM in the Fixed Dose period 
is defined as the time from the subject taking the first dose of study medication in the 
Fixed Dose period until the subject has an SBM.  
• Time to first rescue medication use in the Fixed Dose period: Time to first rescue 
medication use in the Fixed Dose period is defined as the time from the subject taking 
the first dose of study medication in the Fixed Dose period until the subject first uses 
rescue medication.  
• Stool-related subjective assessments using the PAC-SYM: Subjective bowel function 
data is captured using the Patient Assessment of Constipation-Symptoms: The PAC-
SYM is a 12-item questionnaire, and each question has a 0 (Absent) – 4 (Very Severe) 
point response scale. The PAC-SYM overall score is the score obtained by averaging the 
response from the 12 items. The PAC-SYM scores and change from baseline will be 
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• 
Stool-related subjective assessments using the PAC-QOL: Constipation-related quality 
of life is assessed using the Patient Assessment of Constipation-Quality of Life. The 
PAC-QOL is a 28-item questionnaire, and each question has a 5-point ordinal response 
scale. The PAC-QOL endpoints consist of an overall average score and the four subscale 
scores (i.e., physical discomfort, psychosocial discomfort, worries/concerns, and 
satisfaction). The PAC-QOL scores and change from baseline will be calculated for each 
scheduled assessment time-point. 
2.6.3 Exploratory efficacy endpoints 
The exploratory efficacy endpoints include:  
• A weekly CSBM rate increase of 1 or more compared to baseline and an absolute 
rate of 3 or more CSBMs per week in the Fixed Dose period (i.e., this is a 
modification of the Prokinetic Response secondary endpoint using “AND” instead of 
“OR” as the criterion). 
• A weekly SBM rate increase of 1 or more compared to baseline and an absolute rate 
of 3 or more CSBMs per week in the Fixed Dose period. 
• Change from baseline in the weekly CSBM rate during the Placebo Withdrawal and 
Washout periods. 
• Change from baseline in the weekly SBM rate during the Placebo Withdrawal and 
Washout periods. 
• Change from baseline in stool ease of passage during the Placebo Withdrawal and 
Washout periods.  
• Change from baseline in stool consistency during the Placebo Withdrawal and 
Washout periods. 
• Change from baseline in suppository/enema use during Placebo Withdrawal and 
Washout periods.  
• Improvement in frequency and/or severity of hallucinations/delusions during the 
Fixed Dose period over baseline: Improvement is defined as an improvement of 2.33 
points or greater reduction in score from baseline on the Scale for the Assessment of 
Positive Symptoms -Parkinson's Disease (SAPS-PD). Up to 40% of subjects are 
expected to have minor or major hallucinations and/or delusions. The SAPS-PD will 
be administered at Screening, and subsequently will only be administered to those 
subjects with hallucinations and/or delusions (as determined by an SAPS-PD score 
>0 at the Screening visit).  
• Improvement of the symptoms of Parkinson’s Disease as assessed by the UPDRS: 
The standard four-part UPDRS will be administered to determine whether there is 
any deterioration in overall scores during the treatment period. Motor deterioration 
will be assessed from Part 3 of the UPDRS, but total score will also be reported. The 
assessment will be performed during the “ON” phase, approximately 60 minutes 
after taking levodopa. 
• Improvement in mood as assessed by the Beck Depression Index (BDI-II): The 21-
item BDI–II assesses the intensity of depression. Each item is a list of four 
statements arranged in increasing severity about a particular symptom of depression. 


 
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• Improvement in cognition as assessed by the MMSE: The MMSE is an 11-item 
examination with a score ranging from 0 – 30 with a higher score indicating better 
memory. The MMSE will be summarized over time.  
• Improvement in sleep as assessed by sleep diaries.  
• 
Change in plasma L-Dopa levels 20, 40, and 60 minutes after L-Dopa and ENT-01 
dose: An increase of >30% in peak plasma level during treatment versus baseline 
would indicate clinically significant increase in absorption attributable to ENT-01 
(10 subjects only).  
• 
Change in stool microbiome during treatment: Since ENT-01 has antimicrobial 
properties in vitro, it may alter the intestinal microbiome; the subject’s stool will be 
analyzed for common intestinal bacteria and viruses at Baseline, Fixed Dose, and 
Washout periods (10 subjects only). 
2.6.4 Pharmacokinetic (PK) Endpoints 
PK samples for levodopa will be collected from trial patients according to the instructions 
detailed in the lab manual and in accordance with the protocol schedule. A separate SAP will be 
prepared for the PK analysis. 
2.6.5 Safety Endpoints 
Safety assessments performed during the study include: 
• Adverse Events collection and reporting 
• Physical Examinations 
• Vital sign assessment 
• Laboratory assessment including Hematology, Serum chemistry, and Urinalysis 
• Electrocardiogram (ECG). 
• The Columbia Suicide Severity Rating Scale (C-SSRS) 
AEs will be coded according to the latest version of the Medical Dictionary for Regulatory 
Activities (MedDRA). The intensity/severity of AEs will be graded according to NCI-CTCAE 
(Common Terminology Criteria for Adverse Events.).  
A treatment-emergent adverse event (TEAE) is defined as any event not present prior to the 
initiation of the treatments or any event already present that worsens in either intensity or 
frequency following exposure to the treatments. 
The C-SSRS is a questionnaire used to measure of suicidal ideation and behavior. 
2.6.6 Tolerability Endpoints 
The tolerability endpoint is referred to as the dose limiting toxicity (DLT). These adverse events 
are considered as DLT: 
• Recurrent vomiting defined as 3-5 episodes of vomiting within 24 hours of taking study 
medication/placebo  


 
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• Recurrent diarrhea defined as 4 or more episodes of diarrhea within 24 hours of taking 
study medication/placebo 
• Abdominal pain defined as moderate pain that limits instrumental activities of daily 
living (ADL) (e.g., preparing meals, shopping for groceries or clothes, using the 
telephone, managing money, etc.) within 24 hours of taking study medication/placebo 
• Dizziness defined as lightheadedness or fainting on rising from lying to sitting or 
standing that limits instrumental ADL and severe enough to indicate non-urgent medical 
intervention within 24 hours of taking study medication/placebo. 
It is anticipated that all subjects receiving active treatment will exhibit a prokinetic effect at a 
dose lower than DLT and less than or equal to 250 mg. Subjects reaching DLT before a 
prokinetic effect will be treated at the highest dose at which they did not experience a DLT. 


 
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3. 
SUBJECT POPULATIONS 
3.1. 
Population Definitions 
The following subject populations will be evaluated and used for presentation and analysis of 
the data: 
• Safety Population: The Safety Population will consist of all subjects who receive at least 
one dose of study medication during the study. Subjects will be analyzed based on the 
treatment that they actually received.  All safety analyses will be based on this 
population. 
• Modified ITT Population: The modified Intent-to-Treat (mITT) Population will include 
subjects who are randomized and have the stool diary assessments both at baseline and at 
least one post baseline time-point. Subjects who are found later to have inclusion/exclusion 
violations will be excluded from the mITT population. 
• Fixed Dose Population: The Fixed Dose Population will include all subjects who are in 
mITT Population who enter the Fixed Dose period of the study and are in the treatment 
period for at least seven days, not including the date of randomization.  
• Efficacy Evaluable Population: The Efficacy Evaluable Population will include 
subjects who are in the Fixed Dose Population, complete the Fixed Dose period, are 
compliant with study medication in the Fixed Dose period, and have no major protocol 
deviations.    
• Pharmacokinetics Population: Pharmacokinetics Population will include subjects with 
a least one L-Dopa PK result. 
The precise reasons for excluding subjects from the Efficacy Evaluable Population will be fully 
defined and documented prior to the unblinded interim analysis.  
3.2. 
Protocol Violations 
At the discretion of the sponsor, major protocol violations as determined by a review of the data 
prior to the database locks or unblinded interim analysis and the conduct of statistical analyses 
may result in the removal of a subject’s data from the Efficacy Evaluable Population. The 
sponsor, or designee, will be responsible for producing the final protocol violation file 
(formatted as a Microsoft Excel file), in collaboration with Cytel and the data monitoring group 
as applicable; this file will include a description of the protocol violation, and clearly identify 
whether or not this violation warrants exclusion from the Efficacy Evaluable Population.  This 
file will be finalized prior to the database locks. 
All protocol violations will be presented in the data listings. 
 
 


 
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4. 
STATISTICAL METHODS 
4.1. 
Sample Size Justification 
Sample size for this study was determined based on the results from the phase 2a study. In the 
phase 2a study, efficacy was demonstrated in 34 subjects treated with ENT-01, with a mean 
change from baseline in CSBMs of 2.4.  
For the first part of the study  it was anticipated that, out of the 72 randomized subjects, 64 
(89%) will be evaluable for the primary efficacy endpoint analysis. Assuming an increase from 
baseline in weekly CSBM rate in the Fixed Dose period of 2.4 and 0.5 for the ENT-01 and 
placebo-treated subjects, respectively, and a standard deviation (SD) of 2.0, the study has 90% 
power to detect a difference between the treatment groups. This power estimate is based on a 2-
sided t-test at the 0.05 significance level. 
For the second part of the study, enrolling an additional 80 patients in a 1:1 randomization will 
yield a conditional power of approximately 90%. This is based on the additional subjects having 
a mean (SD) change from baseline for the Fixed Dose Period of 3.6 (4.11) and 1.3 (2.38) for 
ENT-01 and placebo, respectively. These parameter estimates are based on the results of the 
initial cohort of patients in the Phase 2b study. These parameter estimates are based on results of 
the initial cohort of patients in the Phase 2b study. Eighty additional patients will be randomized 
to ensure 60 evaluable patients. 
 
4.2. 
General Statistical Methods and Data Handling 
General Methods 
All outputs will be incorporated into Microsoft Excel or Word files, sorted and labeled 
according to the International Conference on Harmonisation (ICH) recommendations, and 
formatted to the appropriate page size(s). 
Tabulations will be produced for appropriate demographic, baseline, efficacy, pharmacokinetic 
and safety parameters. For categorical variables, summary tabulations of the frequency and 
percentage within each category (with a category for missing data) of the parameter will be 
presented.  For continuous variables, the mean, median, SD, minimum and maximum values 
will be presented.  Time to event data will be summarized using Kaplan-Meier Methodology 
using 25th, 50th (median), and 75th percentiles with associated 2-sided 95% confidence intervals, 
as well as percent of observed event rate and summary statistics for the follow-up time.   
Formal statistical hypothesis testing will be performed on the efficacy endpoints with all tests 
conducted at the 2-sided 0.05 level of significance. Summary statistics will be presented, as well 
as confidence intervals on selected parameters, as described in the sections below. 
For all analyses, baseline will be defined as the most recent measurement prior to the date and 
time of the administration of ENT-01 or placebo. Therefore, baseline may be on the same day as 
the dose of study treatment, but prior to the administration of the study treatment.   
Computing Environment 


 
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All descriptive statistical analyses will be performed using SAS statistical software 
(Version 9.4), unless otherwise noted.  Medical History and Adverse Events will be coding 
using the latest MedDRA version available upon the study start.  Concomitant medications will 
be coded using the latest World Health Organization (WHO) Drug version available upon the 
study start.  
Methods of Pooling Data 
Not applicable to the present study. 
Adjustments for Covariates 
Statistical analyses for the efficacy endpoints will be adjusted by the randomization strata, the 
baseline CSBM rate (0 – 0.9 vs. 1.0 – 3.0). 
Multiple Comparisons/Multiplicity 
Adjustments for multiplicity will not be made for this study.  
Subpopulations 
The primary and secondary efficacy endpoints will be summarized by the subgroups based on 
the baseline CSBM rate (i.e., 0 - 0.9 or 1.0 – 3.0). Additionally, the primary and secondary 
endpoints will be presented by first and second “cohort” (i.e., the 1st cohort of subjects enrolled 
under the first part via the 3:1 randomization scheme; and the 2nd cohort of subjects randomized 
via the 1:1 randomization scheme).   
Withdrawals, Dropouts, Loss to Follow-up 
Subjects who withdrew from the study were not to be replaced. 
Missing, Unused, and Spurious Data 
When deriving endpoints that use dates, for dates that include missing values, the following 
conventions will be used. 
Value  
Imputation  
Missing day, month, and year  
Not imputed  
Missing day and month  
Day imputed to June 30th  
Missing day  
Day imputed to the 15th  
When using dates from drug administration pages for the purposes of deriving treatment 
exposure, the following conventions will apply:  
• If the start date is missing, it will be assumed that the first drug intake was given at the 
date of randomization. This date will replace all first administration missing dates.  
• If the last administration date is missing, the date will be imputed with the date taken 
from the “Status of the Patient at the End of the Treatment” eCRF page (Date of last 
treatment) or if missing, with the date of last contact taken from the “Status of the 
Patient” eCRF page.  
For missing or partial missing onset dates for adverse events, the following conventions will 
apply: 
The start dates for AEs are important for the following:  


 
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1. Treatment-emergent adverse event (TEAE) algorithm. A TEAE is an event that first 
occurred or worsened in severity after baseline.  
2. Designation of unique AE occurrences.  
3. Completely missing or partially missing AE onset dates will be imputed after due 
diligence to obtain accurate AE information has failed. The imputation will consider the 
start and stop dates relative to the each other, and relative to the date of consent and date 
of first dose.  
AEs with start dates that are completely or partially missing will be analyzed as follows:  
• If the start date of an event is completely missing, then the event is assumed to be 
treatment-emergent.  
• If the start date has the month and year but day is missing, the event will be considered 
treatment-emergent if the month and year of the start date of the event are equal to, or 
greater than, the month and year of the date of first dose of study drug.  
If the start date has the year, but day and month are missing, the event will be considered 
treatment-emergent if the year of the start date of the event is  the same as, or later than, the year 
of the date of the first dose of study drug. 
Visit Windows 
No visit windows will be applied. 
4.3. 
Interim Analyses 
An unblinded interim analysis will be performed after the last subject completes the Fixed Dose 
period. 
4.4. 
Subject Disposition 
Patient disposition, including analysis population allocation, patients enrolled, completed, and 
discontinued, and primary reason for discontinuation, will be summarized using frequency and 
percent.  For the purposes of reporting, patients who complete the Fixed Dose period are 
considered to have completed the trial.  
Analysis populations, patients who completed each period (fixed dose, single blind, and 
washout), and patients who discontinued the study and reasons for discontinuation will be 
summarized by randomized treatment (placebo or ENT-01). The disposition of subjects in the 
mITT and safety populations up to the end of the fixed dose period will also be summarized by 
summarizing the number of subjects who completed the fixed dose period or discontinued study 
during the fixed dose period, along with the reason for discontinuation. This summary will be 
done by dose during the fixed dose period. Patients will be counted in every dose that the 
received during the fixed dose period. 
Patients who give informed written consent but are not randomized are considered screen 
failures. Minimal data, such as demographic information and the reason for screen failure, for 
patients who fail screening will be recorded on the IXRS. Reasons for screen failure will be 
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A by-subject listing of study completion information, including the reason for premature 
treatment and study discontinuation, if applicable, will be presented. 
4.5. 
Demographic and Baseline Characteristics 
Demographics, baseline characteristics and medical history will be summarized for the safety, 
mITT and fixed dose populations by treatment group (placebo and fixed dose ENT-01) and data 
listings will be provided as well. Demographic variables, including age, gender, race, ethnicity 
height, weight, and BMI will be summarized in tabular form.  Age (years) will be calculated as 
(date of informed consent – date of birth +1) / 365.25. Baseline characteristics include age at 
diagnosis of Parkinson’s disease, years with Parkinson’s disease, years of constipations, Hoehn 
and Yahr stage, and constipation severity using baseline CSBM rate. 
4.6. 
Efficacy Analyses 
Efficacy analyses to evaluate the treatment effect will be conducted using the Fixed Dose 
population as the primary population. The analyses will also be performed using the mITT and 
Efficacy Evaluable populations. Summary statistics will also be presented. 
The comparison of active versus placebo treatment will be tested at the two-sided 0.05 
significance level. 
 
The date of randomization will not be included for either baseline or post-baseline weekly 
efficacy endpoints (i.e., rates or mean values).  Similarly, the date of the single-blind period visit 
will also not be used in derivation of weekly efficacy endpoints (rates or mean values).  Also, the 
visit 5 Date (ie, the date the patient begins the Washout period) will be included in the single-blind 
period (i.e., the single-blind period lasts from the day after Visit 4 up to and including Visit 5). 
4.6.1 Derivation of Weekly CSBM and SBM Rates 
There are 4 CSBM Definitions, each of which are based on the set of questions from the 
patient’s daily diary. The rates from each of the CSBM definitions will be presented. Note: since 
the assessments are different between the 2 cohorts the definitions are different. 
CSBM Definition 1:  
A CSBM is a BM which meets each of the following 3 criteria. 
For the 1st Cohort of patients: 
1) For this BM the patient responded “No rescue medication used” to the query “Have you 
used rescue medication within 24 hours prior to this bowel movement?”   
2) For this BM the patient responded  
• “Yes” to the query “Did you feel like you completely emptied your bowels?”  
OR 
• “No” to the query “Did you feel like you completely emptied your bowels?” but the 
BSFS Ease of Passage score is 6 or 7.  
3) The bowel movement has a BSFS Ease of Passage score of 3 or greater (or missing). 
 


 
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For the 2nd Cohort of patient: 
1) No rescue mediation was taken within 24 hours of the Bowel Movement (note: the date 
and time of the bowel movements and rescue medication (i.e., laxative) use are included 
in the subjects daily diary data). 
2) For this BM the patient responded : 
• “No” to the query “Do you feel like you have more poop left to pass?”  
OR  
•  “Yes” to the query “Do you feel like you have more poop left to pass?” and the 
BSFS Ease of Passage score is 6 or 7.  
3) The bowel movement has a BSFS Ease of Passage  score of 3 or greater (or missing). 
 
CSBM Definition 2:  
The 2nd CSBM definition removes the BSFS criteria from the CSBM definition. A CSBM is a 
BM which meets each of the following 2 criteria 
For the 1st Cohort of patients: 
1) For this BM the patient responded “No rescue medication used” to the query “Have you 
used rescue medication within 24 hours prior to this bowel movement?”   
2) For this BM the patient responded  “Yes” to the query “Did you feel like you completely 
emptied your bowels?”  
 
For the 2nd Cohort of patient: 
1) No rescue mediation was taken within 24 hours of the Bowel Movement (note: the date 
and time of the bowel movements and rescue medication (i.e., laxative) use are included 
in the subjects daily diary data). 
2) For this BM the patient responded “No” to the query “Do you feel like you have more 
poop left to pass?”  
 
CSBM Definition 3:  
The 3rd CSBM definition broadens the definition of a CSBM for the 2nd cohort. A CSBM is a 
BM which meets each of the following 2 criteria 
For the 1st Cohort of patients: 
1) For this BM the patient responded “No rescue medication used” to the query “Have you 
used rescue medication within 24 hours prior to this bowel movement?”   
2) For this BM the patient responded  “Yes” to the query “Did you feel like you completely 
emptied your bowels?”  
 
For the 2nd Cohort of patient: 


 
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1) No rescue mediation was taken within 24 hours of the Bowel Movement (note: the date 
and time of the bowel movements and rescue medication (i.e., laxative) use are included 
in the subjects daily diary data). 
2) For this BM the patient responded  
• “No” to the query “Do you feel like you have more poop left to pass?”  
OR 
• “Yes” to the query “Are you Done for the Day” 
OR 
• “Yes” to the query “Did you feel like you completely emptied your bowels?”  
CSBM Definition 4:  
This is a more stringent definition for CSBMs for Cohort 2, requiring each of the 3 
completeness questions to be appropriately answered. A CSBM is a BM which meets each of 
the following 3 criteria. 
For the 1st Cohort of patients: 
1) For this BM the patient responded “No rescue medication used” to the query “Have you 
used rescue medication within 24 hours prior to this bowel movement?”   
2) For this BM the patient responded  
• “Yes” to the query “Did you feel like you completely emptied your bowels?”  
OR 
• “No” to the query “Did you feel like you completely emptied your bowels?” but the 
BSFS Ease of Passage score is 6 or 7.  
3) The bowel movement has a BSFS Ease of Passage score of 3 or greater (or missing). 
 
For the 2nd Cohort of patient: 
1) No rescue mediation was taken within 24 hours of the Bowel Movement (note: the date 
and time of the bowel movements and rescue medication (i.e., laxative) use are included 
in the subjects daily diary data). 
2) For this BM the patient met either of these 2 criteria: 
2a) The patient responded: 
• “No” to the query “Do you feel like you have more poop left to pass?”  
AND 
• “Yes” to the query “Are you Done for the Day” 
AND 
• “Yes” to the query “Did you feel like you completely emptied your bowels?”  
OR 
2b) The BSFS Ease of Passage score is 6 or 7.  


 
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3) The bowel movement has a BSFS Ease of Passage  score of 3 or greater (or missing). 
 
An SBM is a BM which meets the following criteria: 
• For this BM the patient responded “No rescue medication used” to the query “Have you 
used rescue medication within 24 hours prior to this bowel movement”   
A patient’s CSBM weekly rate over a particular period will be derived using the following 
algorithm: 
Weekly CSBM Rate = 7×{# of CSBMs in the period}/{# of days in the period} 
The weekly SBM rate calculations use the same algorithm. 
If a patient is missing BM information for a day, it will be assumed that the patient had 0 SBMs 
and 0 CSBMs for that period (with the exceptions in calculating Baseline CSBM and SBM rates 
noted in Section 2.6).  
If a subject diary has “repeat” stool data (eg, a BM repeated with the same date/time of BM) 
only the latest duplicate observation relative to when the patient entered the data into the diary 
will be used. 
For the calculation of the Baseline CSBM (SBM) weekly rate: 
• If the patient has more than 14 days of diary data during the Screening Period, only the 
last 14 days with non-missing diary entries prior to the randomization visit will be used 
(i.e., the days which are more than 14 days prior to the randomization visit will be 
excluded) 
• If a patient has 11- 14 days of diary data, all available diary days will be used.  
• If a subject has more than two weeks of entries during the screening period, consecutive 
missing entries during the screening period will be excluded from baseline calculations, 
and the last fourteen days with entries during the screening period will be used to derive 
baseline weekly rates. 
For analyses of the CSBM (SBM) in the Fixed dose period, if a patient has not been in the Fixed 
Dose period for at least seven days, then the number of days used for calculating the weekly 
CSBM (SBM) rate will be adjusted to seven days by using the last “n” days of the Dose 
Escalation period leading up to the fixed dose period to get to 7 days in the Fixed Dose period. 
This derivation will be used for all efficacy endpoints for the Fixed Dose Period (e.g., stool ease 
of passage and stool consistency). 
For analyses of the CSBM (SBM) rate during the single-blind placebo and washout periods, all 
available data in those periods, excluding observations on the date of the single-blind placebo 
visit as described before, will be used in the derivations of weekly rates described at the 
beginning of this section. 
  
4.6.2 Stool Consistency and Ease of Passage Analysis Conventions 
Only stool consistency and ease of passage scores associated with SBMs will be used in 
analyses (i.e., excluding BMs where it was indicated that rescue medication had been used 
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4.6.3 Analysis of the Primary Efficacy Endpoint 
The primary efficacy endpoint is the change from baseline in weekly CSBM rate during the 
Fixed Dose period. The primary analysis will be based on the Fixed Dose Population. The 
primary efficacy analysis will be performed via an analysis of covariance (ANCOVA) model 
with change from baseline in weekly CSBM rate as the dependent variable, treatment (ENT-01 
or placebo) as the independent variable, and baseline weekly CSBM rate as a covariate.  
 
As a supplemental analysis, the treatment effect for the primary efficacy endpoint will be 
summarized by the weekly CSBM rate subgroups (0 – 0.9, 1.0 – 3.0). The analysis will be 
performed for both Efficacy Evaluable population and Fixed Dose population. 
 
The study will be considered a success if the p-value for the comparison of treatment groups 
from the ANCOVA model using the Fixed Dose population is significant (p<0.05). 
 
Change from baseline in weekly CSBM rate will also be summarized by the specific fixed dose 
for patients. These summaries will be descriptive in nature only and no inferential statistics will 
be performed.  
 
A linear trend test for weekly CSBM rate within each treatment group across baseline, fixed 
dose, and single blind periods will be conducted by using mixed model repeated measures 
(MMRM) methods, where the Baseline, Fixed Dose, and Single Blind periods are assigned 
values of 0,1, and 2 respectively for modeling purposes. Baseline CSBM rate (0-0.9/1.0-3.0) is 
included as a factor, with subject intercept and slope as random effects in the mixed model. The 
trend test will also be conducted across all subjects, including both Placebo and ENT-01 
treatment groups in the model as a factor (fixed effect).    
 
4.6.4 Analysis of Secondary Efficacy Endpoints 
All secondary efficacy endpoints will be analyzed based on both the Efficacy Evaluable 
population and Fixed Dose population. 
Continuous Secondary Efficacy Endpoints 
The following are the set of continuous secondary efficacy endpoints: 
• Change from baseline in the weekly SBM rate during the Fixed Dose period 
• Change from baseline in stool ease of passage during the Fixed Dose period 
• Change from baseline in stool consistency during the Fixed Dose period 
• Change from baseline in suppository/enema use during the Fixed Dose period 
• Change from baseline in the overall scores and individual items of PAC-SYM and 
PAC-QOL subjective assessments during the Fixed Dose period 
All continuous secondary efficacy endpoints will be analyzed in the same manner as the primary 
efficacy endpoint with treatment, baseline CSBM strata, and corresponding baselines as the 
independent variables in the ANCOVA model and also by summarizing change from baseline 


 
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by the specific fixed doses. The same linear trend tests will also be performed on these 
endpoints. 
Categorical Secondary Efficacy Endpoints 
The following are the set of categorical secondary efficacy endpoints: 
• Dichotomous prokinetic bowel response during the Fixed Dose period 
• Dichotomous SBM bowel response during the Fixed Dose period 
• Frequency of dose adjustments in the Dose Adjustment period from the initial 
starting dose to the subject’s fixed dose  
The dichotomous endpoints will be summarizes using count data, frequencies, and odds ratios 
for comparisons between groups, and analyzed using Fisher’s exact test. If there are sufficient 
numbers in each strata, the analysis may also be performed using the Cochran-Mantel-Haenszel 
(CMH) test adjusting for the baseline CSBM strata, to compare ENT-01 versus placebo.  The 
frequency of dose adjustment will be summarized by treatment groups and baseline CSBM 
strata and no formal test will be performed to compare the treatment groups. 
Summary statistics will also be presented by specific fixed dose groups for descriptive purposes 
only. 
Time to Event  Secondary Efficacy Endpoints 
The following are the set of time to event secondary efficacy endpoints: 
• Time to first CSBM in the Fixed Dose period 
• Time to first SBM in the Fixed Dose period 
• Time to first rescue medication use in the Fixed Dose period 
Time to event endpoints will be summarized using Kaplan-Meier methods and comparisons 
between treatment groups will be made via log-rank tests, stratified by the baseline CSBM rate. 
The start date for time to event analyses will be the date of first dose in the fixed dose period. 
4.6.5 Analysis of Exploratory Efficacy Endpoints 
Continuous Exploratory Efficacy Endpoints 
Continuous endpoints include change from baseline of the following outcomes: 
• Weekly CSBM rate during the Placebo Withdrawal and Washout periods 
• Weekly SBM rate during the Placebo Withdrawal and Washout periods 
• Stool ease of passage during the Placebo Withdrawal and Washout periods 
• Stool consistency during the Placebo Withdrawal and Washout periods 
• Suppository/enema use during Placebo Withdrawal and Washout periods 
• Total score and 3 subdomain scores from PAC-SYM during the Placebo Withdrawal 
and Washout periods 
• Total Score and 4 subdomain scores from PAC-QoL during the Placebo Withdrawal 
and Washout periods 
• Stool microbiome during each study period, defined as the number of species found 
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• Total sleep time (hours) per day during each study period 
•  Delay (minutes)  in sleep onset per night during each study period 
• Total awake time (hours) per night during each study period 
• Number of wakes per night during each study period 
• Total score from UPDRS and individual part scores to assess the symptoms of 
Parkinson’s Disease during each study period 
• Total score from BDI-II to assesses the intensity of depression during each study 
period 
• Total score from MMSE to assess cognition during each study period 
All continuous exploratory efficacy endpoints will be analyzed in the same manner as the 
primary efficacy endpoint with treatment, baseline CSBM strata, and corresponding baselines as 
the independent variables in the ANCOVA model. Changes to the single blind placebo and 
washout periods will also be repeated using the fixed dose rate or end of fixed dose 
measurement as baseline. Summary statistics will also be presented by specific fixed dose 
groups for descriptive purposes only. 
Categorical Exploratory Efficacy Endpoints 
The following is the lone categorical exploratory efficacy endpoint: 
 
• Improvement in frequency and/or severity of hallucinations/delusions as measured 
by SAPS-PD score during the Fixed Dose period over baseline for subjects with 
SAPS-PD score >0 at the Screening visit. Improvement is defined as ≥2.33 points 
reduction in score from baseline on the SAPS-PD 
The endpoint will be analyzed using Fisher’s exact test, and also by the Cochran-Mantel-
Haenszel (CMH) test adjusting the baseline CSBM strata, if numbers permit, to compare ENT-
01 versus placebo. Summary statistics will also be presented by specific fixed dose groups for 
descriptive purposes only. 
 
4.7. 
Pharmacokinetic Analysis 
The plasma concentration data and PK parameters of L-Dopa will be summarized based on the 
PK population by time point (pre-dose, 20, 40 and 60 minutes post-dose) and dose level. These 
measures will be presented in listings as well. 
An increase of >30% in peak plasma level of L-Dopa during treatment from baseline would 
indicate clinically significant increase in absorption attributable to ENT-01. The frequency and 
percentage of subject with clinically significant increase will be tabulated without formal 
statistical analysis for the PK population. 


 
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4.8. 
Safety Analyses 
All safety analyses will be conducted on the Safety Population. The safety data will be 
presented in individual listings and summary tables. Descriptive statistics will be used to 
summarize the safety data by period and both by treatment group and overall. The denominator 
for calculating the percentages will be based on the number of subjects on each treatment arm 
(ENT-01 or placebo) and overall in the Safety Population. The summary for the ENT-01 arm 
during the Fixed Dose period will also be broken down to the fixed dose levels. 
No formal hypothesis-testing analysis of safety data will be performed.   
4.8.1 Extent of Exposure 
The extent of exposure will be summarized by the number of tablets received and number of 
days on treatment during the Dose Adjustment period and Fixed Dose period.  
4.8.2 Adverse Events 
Adverse events (AEs) will be coded by system organ class (SOC) and preferred term (PT) 
according to the latest version of the Medical Dictionary for Regulatory Activities (MedDRA).  
The intensity/severity of AEs will be graded according to the latest NCI-CTCAE. Treatment-
emergent AEs (TEAEs) are defined as any AE occurring or worsening on or after the first dose 
of study treatment. AEs that have a possible, probable or definite relationship to study drug will 
be defined to be related to the study drug while others will be defined as “not related”. AEs with 
the closest relationship to the study drug will be used for summary. 
The following types of summaries will be provided: 
• Overview of TEAEs 
• TEAEs by SOC and PT 
• TEAEs by SOC, PT, and Maximum CTCAE toxicity grade 
• TEAEs by SOC, PT, and Maximum CTCAE toxicity grade >= 3 
• TEAEs by SOC, PT, and closest relationship to study drug 
• TEAEs by PT in decreasing frequency 
• Drug-related TEAEs by SOC and PT 
• Drug-related TEAEs by PT in decreasing frequency 
• Serious TEAEs by SOC and PT 
• TEAEs leading to study treatment discontinuation by SOC and PT 
• TEAEs leading to dose interruption by SOC and PT 
 
All adverse event summaries will be presented by the dose the subject was on at the time of the 
AE.  Adverse event summaries will also be presented during the following time periods: 
 
• Dose Escalation 
• Fixed Dose 
• Dose Escalation and Fixed Dose (Treatment Period) 
• Single Blind Placebo 


 
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• Washout 
• Entire Study 
 
In these tabulations, if a subject experiences the same preferred term multiple times within a 
period, then the event will be counted only once within the period. 
In tabulation by severity (i.e., CTCAE toxicity grade),  
• For a given preferred term, only the most severe preferred term for each subject will be 
included.  
• For a given system organ class, only the most severe system organ class for each subject 
will be included.  
Similarly, in tabulation by relationship, 
• For a given preferred term, the most closely related preferred term to the study drug for 
each subject will be included.  
• For a given system organ class, the most closely related system organ class to the study 
drug for each subject will be included.  
TEAEs, serious TEAEs, and AEs leading to discontinuation from the study treatment will be 
provided in separate data listings.  
4.8.3 Deaths 
All deaths and reasons for death will be provided in a listing.   
4.8.4 Physical Examinations 
Summaries of physical examinations will present frequency distribution of abnormal findings by 
body system and visit. The denominators for calculating the percentages will be based on the 
number of subjects evaluated for a particular body system. All physical examination findings will 
be presented in a data listing. 
4.8.5 Laboratory Data 
Clinical labs will be performed by a central laboratory. Labs to be drawn during the study 
include serum chemistries, a hematology panel, coagulation panel and urine analysis. Clinical 
laboratory values will be expressed using conventional units. 
Descriptive statistics by period (or visit) and treatment group will be presented for each 
laboratory parameter. Changes from Baseline as well as shift tables will be presented. All 
laboratory values will be classified as normal, below normal, or above normal based on normal 
ranges supplied by the central laboratory. Frequencies of abnormal values and frequencies of 
clinically significant abnormal values will be presented in tabular form. In the event of repeat 
values, the last non-missing value per study period will be used. 
All laboratory data will be provided in data listings. 


 
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4.8.6 Vital Signs 
The actual value and change from baseline to each on study evaluation will be summarized for 
vital signs.  By-subject listings of vital sign measurements will be presented in data listings. 
4.8.7 Electrocardiogram 
ECG results will be summarized descriptively, including the number and percent of subjects 
with normal, abnormal and clinically significant abnormal results at Baseline and each study 
visit. The denominators for calculating the percentages will be based on the number of subjects 
with non-missing values in each assessment. 
All ECG data for each subject will be provided in data listings. 
4.8.8 Columbia-Suicide Severity Rating Scale (C-SSRS)  
The number and percentage of patients with Suicidal Ideation, Suicidal Behavior, and Self-
Injurious Behavior without Suicidal Intent based on the Columbia-Suicide Severity Rating Scale 
(C-SSRS) during the treatment period will be summarized by treatment group. Treatment-
emergent Suicidal Ideation, Suicidal Behavior, and Self-Injurious Behavior without Suicidal 
Intent will be based on Visit 3 through End of Study (EOS)/Early Termination (ET). In this 
summary, the number and percentage of patients who experience the particular event at least 
once during the treatment period will be summarized. The particular events are: 
• Suicidal Ideation (1-5) 
1. Wish to be Dead 
2. Non-specific active suicidal thoughts 
3. Active suicidal ideation with any methods (not plan) without intent to act 
4. Active suicidal ideation with some intent to act, without specific plan 
5. Active suicidal ideation with specific plan and intent 
• Suicidal Behavior (6-10) 
6. Preparatory acts or behavior 
7. Aborted attempt 
8. Interrupted attempt 
9. Non-fatal suicide attempt 
10. Completed suicide 
• Self-injurious behavior without suicidal intent 
For the composite endpoint of suicidal ideation (Categories 1-5), the number and percentage of 
patients who experience any one of the five suicidal events at least once during the treatment 
period will be summarized by treatment group. For the composite endpoint of suicidal behavior 
(Categories 6-10), the number and percentage of patients who experience any one of the five 


 
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suicidal behavior events at least once during the treatment period will be summarized by 
treatment group.  
Two shift tabulations from baseline for the C-SSRS will be produced by treatment group during 
the treatment period. The Baseline C-SSRS assessment includes both (A) Lifetime Assessment 
(for both the suicidal behavior and for the suicidal ideation sections) and (B) Recent 
Assessment: 1 year prior to study start for the suicidal behavior section, and 1 month prior to the 
study start for the suicidal ideation section. A separate shift tabulation will be produced using 
each of these two baselines. The three groupings for the shift tables are: (a) No suicidal ideation 
or behavior, (b) Suicidal Ideation, and (c) Suicidal Behavior. Suicidal Ideation includes any one 
of the five suicidal ideation events (Categories 1-5). Suicidal behavior includes any one of the 
five suicidal behavior categories (Categories 6-10). Each patient is counted in one cell only for 
each of the two tabulations. Patients with both Suicidal Ideation and Suicidal Behavior are 
included in the Suicidal Behavior category for the particular tabulation.  
Patient listings for the C-SSRS will also be produced. 
4.8.9 Concomitant Medications 
Prior medications are defined as medications taken prior to the first dose of study drug (ENT-01 
or placebo). Concomitant medications are defined as medications taken on or after the first dose 
study drug.  
Prior and Concomitant medications will be coded using the WHO Drug dictionary. Results will 
be tabulated by Anatomic Therapeutic Class (ATC) and preferred term. 
The use of prior and concomitant medications will be included in by-subject data listing. 
4.8.10 Tolerability 
Tolerability will be measured by DLT endpoints. The frequency of occurrence for each 
tolerability endpoint including recurrent vomiting, recurrent diarrhea, abdominal pain and 
dizziness will be summarized by treatment group and dose level for the Dose Adjustment 
period. Percentages will be based on the number of subjects in each dose level in the Safety 
Population. Since this occurs during the dose adjustment period, the dose level is not fixed for 
each patient. The number of patients in a dose level will be calculated based on the highest dose 
level a patient ever took during the Dose Adjustment Period. For example if a patient with low 
baseline CSBMs had the highest dose of 200 mg, then this patient would be included in the 
doses of 150, 175, and 200 mg.   


 
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5. 
CHANGES TO PLANNED ANALYSES 
As of this date, there have been no changes between the protocol-defined statistical analyses and 
those presented in this statistical plan. 


 
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6. 
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