Clinical trial NCT04907474
Conditions: Mydriasis
Sponsor: Eyenovia Inc.
Start date: 2021-05-24; Completion date: 2021-06-03
Enrollment: 61 participants; Allocation: RANDOMIZED; Intervention model: CROSSOVER

Brief summary:
1 mist or 2 mists of the study drug will be administered to both eyes according to a pre-specified randomization plan to determine the effect on pupil dilation

Detailed description:
After screening, eligible subjects will be scheduled for 2 treatment visits where either 1 mist or 2 mists of the study drug will be administered to both eyes according to a pre-specified randomization plan. Safety evaluations and efficacy measurements will be performed at specified time intervals thereafter.

Pupil dilation for each treatment will be compared at each time interval.

Arms: arm 1: Sequence AB: 1 mist of tropicamide-phenylephrine fixed combination solution administered to each eye with the Micro Array Print (MAP) Dispenser followed by 2 mists of tropicamide-phenylephrine fixed combination solution administered to each eye with the MAP Dispenser arm 2: Sequence BA: 2 mists of tropicamide-phenylephrine fixed combination solution administered to each eye with the Micro Array Print (MAP) Dispenser followed by 1 mist of tropicamide-phenylephrine fixed combination solution administered to each eye with the MAP Dispenser
Interventions: intervention 1: tropicamide-phenylephrine fixed combination ophthalmic solution
Intervention descriptions: intervention 1: Tropicamide-phenylephrine fixed combination ophthalmic solution is a tropical drug solution for mydriasis

Protocol documents:
Prot_SAP_000.pdf: 
TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-41 VERSION B  
 
 
 
CONFIDENTIAL  
06 MAY 2021 
 
  
 
 
  
 
 
PROTOCOL EYN-MYD-TP-41 
VERSION B 
 
 
A SINGLE-CENTER, ASSESSOR-MASKED, ACTIVE-CONTROLLED, PHASE 4 STUDY 
EVALUATING SPEED OF PUPIL DILATION WITH THE MICRO-ARRAY PRINT (MAP) 
DISPENSER WHEN COMPARING 2 DOSING REGIMENS OF TROPICAMIDE-
PHENYLEPHRINE FIXED COMBINATION OPHTHALMIC SOLUTION  
(THE SPEED STUDY) 
 
 
 
SPONSOR: 
 
EYENOVIA, INC. 
295 MADISON AVENUE, SUITE 2400 
NEW YORK NY  10017 
USA 
PHONE:  917-289-1117 
 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-41 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 2 OF 60 
06 MAY 2021 
 
TABLE OF CONTENTS 
 
LIST OF ABBREVIATIONS .................................................................................................................. 5 
1. PERSONNEL AND FACILITIES ....................................................................................................... 6 
2. STUDY SYNOPSIS ............................................................................................................................. 7 
2.1. Study Objective ............................................................................................................................ 7 
2.2. Study Drugs .................................................................................................................................. 7 
2.3. Study Population .......................................................................................................................... 7 
2.4. Study Design ................................................................................................................................ 7 
2.5. Inclusion Criteria .......................................................................................................................... 8 
2.6. Exclusion Criteria ......................................................................................................................... 8 
2.7. Primary Performance Endpoint .................................................................................................... 9 
2.8. Exploratory Outcomes ................................................................................................................. 9 
2.9. Safety Outcomes .......................................................................................................................... 9 
2.10. Schedule of Visits, Procedures and Evaluations ...................................................................... 10 
2.10.1. Study Medication Administration and Clinical Assessments ........................................... 10 
3. INTRODUCTION AND RATIONALE ............................................................................................ 11 
3.1. Introduction ................................................................................................................................ 11 
3.2. Pupil Dilation ............................................................................................................................. 11 
3.3. Existing Mydriatic Options ........................................................................................................ 12 
3.4. Tropicamide-Phenylephrine Fixed Combination Ophthalmic Microdosing Solutions for Pupil 
Dilation .............................................................................................................................................. 12 
3.5. Ophthalmic Microdosing ........................................................................................................... 13 
4. INVESTIGATIONAL PRODUCT DESCRIPTION ......................................................................... 16 
4.1 How Provided.............................................................................................................................. 16 
4.2 Route of Administration, Dosage, Regimen, Treatment Period(s) ............................................. 17 
5. STUDY OBJECTIVE ........................................................................................................................ 18 
6. STUDY DESIGN ............................................................................................................................... 18 
7. STUDY OUTCOMES ........................................................................................................................ 19 
7.1 Effectiveness Outcomes .............................................................................................................. 19 
7.2. 
Safety Outcomes .................................................................................................................. 19 
8. STUDY POPULATION ..................................................................................................................... 20 
8.1. Inclusion Criteria ........................................................................................................................ 20 
8.2. Exclusion Criteria ....................................................................................................................... 20 
9. STUDY METHODS .......................................................................................................................... 22 
9.1. Subject Informed Consent .......................................................................................................... 22 
9.2. Subject Selection and Screening Procedures ............................................................................. 22 
9.3. Randomization to Study Drug Administration Sequence .......................................................... 22 
9.3.1. 
Procedure for Unmasking Study Drug Assignment .................................................... 23 
9.4. Study Drug Packaging, Labeling and Storage............................................................................ 23 
9.5. Study Drug Administration ........................................................................................................ 24 
9.6. Study Visits and Clinical Assessments ...................................................................................... 24 
9.6.1. Screening Visit (-14 Days to Day 0) ................................................................................... 24 
9.6.2. Treatment Visit 1 (Day 1) ................................................................................................... 24 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
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CONFIDENTIAL - PAGE 3 OF 60 
06 MAY 2021 
 
9.6.3. Treatment Visit 2 (Day 5 to Day 8) .................................................................................... 26 
9.4.5. Unscheduled Visits ............................................................................................................. 27 
9.4.6. Missed Visits....................................................................................................................... 27 
9.7. Study Exit ................................................................................................................................... 27 
9.7.1. Subject Termination ............................................................................................................ 27 
9.7.2. Subject Completion ............................................................................................................. 27 
10. ADVERSE EVENTS ..................................................................................................................... 28 
10.1. Adverse Event Definitions ....................................................................................................... 28 
10.1.1. Adverse Event (AE) .......................................................................................................... 28 
10.1.2. Adverse Drug Reaction (ADR) ......................................................................................... 28 
10.1.3. Unexpected Adverse Event or Unexpected Suspected Adverse Reaction ........................ 28 
10.1.4. Suspected Adverse Reaction ............................................................................................. 28 
10.1.5. Life-Threatening Adverse Event or Life-Threatening Suspected Adverse Reaction ....... 28 
10.1.6. Serious Adverse Event (SAE) or Serious Suspected Adverse Reaction ........................... 28 
10.1.7. Serious Ocular Adverse Events ........................................................................................ 29 
10.1.8. Treatment-Emergent Adverse Event (TEAE) ................................................................... 29 
10.2. Adverse Event Assessment and Documentation ...................................................................... 29 
10.2.1. Adverse Event Severity..................................................................................................... 30 
10.2.2. Adverse Event Relationship to Study Drug ...................................................................... 30 
10.3. 
Adverse Event Reporting ..................................................................................................... 31 
10.3.1. 
Expedited Adverse Event Reporting ........................................................................... 32 
11. STATISTICAL METHODS .......................................................................................................... 33 
11.1. Effectiveness Endpoints ........................................................................................................... 33 
11.2. Sample Size .............................................................................................................................. 33 
11.3. Safety Outcomes ...................................................................................................................... 34 
11.4. Analysis Populations ................................................................................................................ 34 
11.4.1. Intent to Treat Population ................................................................................................. 34 
11.4.2. Per Protocol Population .................................................................................................... 34 
11.4.3. Modified Per Protocol Population .................................................................................... 34 
11.4.4. Safety Population .............................................................................................................. 35 
11.5. General Statistical Considerations ........................................................................................... 35 
11.5.1. Analysis of Primary Effectiveness .................................................................................... 35 
11.5.2. Additional Efficacy Analyses ........................................................................................... 36 
11.5.3. Sensitivity Analyses .......................................................................................................... 37 
11.5.4. Interim Analyses ............................................................................................................... 37 
11.5.5. Analysis of Baseline Data ................................................................................................. 37 
12. QUALITY ASSURANCE AND QUALITY CONTROL ............................................................... 37 
12.1. Study Monitoring ..................................................................................................................... 38 
12.2. Study Product Accountability .................................................................................................. 38 
13. ETHICAL AND REGULATORY CONSIDERATIONS ................................................................ 39 
13.1. Institutional Review ................................................................................................................. 39 
13.2. Informed Consent ..................................................................................................................... 39 
13.3. Subject Confidentiality ............................................................................................................. 39 
14. DATA HANDLING AND RECORD KEEPING............................................................................. 40 
14.1. Data Quality Control and Reporting ........................................................................................ 40 
14.2. Data Archiving ......................................................................................................................... 40 
14.3. Records Retention .................................................................................................................... 40 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
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06 MAY 2021 
 
14.4. Protocol Amendments .............................................................................................................. 40 
15. REFERENCES ................................................................................................................................. 41 
APPENDIX 1:  SCHEDULE OF MEDICATION ADMINISTRATION AND EXAMINATIONS .... 43 
APPENDIX 2:  STUDY MEDICATION PREPARATION PROCEDURE .......................................... 44 
APPENDIX 4:  EXAMINATION PROCEDURES ............................................................................... 47 
APPENDIX 5:  SPONSOR AND INVESTIGATOR OBLIGATIONS ................................................. 52 
APPENDIX 6:  WORLD MEDICAL ASSOCIATION DECLARATION OF HELSINKI .................. 55 
 
 
 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
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CONFIDENTIAL - PAGE 5 OF 60 
06 MAY 2021 
 
LIST OF ABBREVIATIONS 
 
Abbreviation 
Term 
AC 
Anterior chamber 
AE  
Adverse event 
ADR 
Adverse drug reaction 
AT 
Artificial tears 
BCDVA 
Best-corrected distance visual acuity 
CF 
Count fingers 
CFR 
Code of Federal Regulations 
eCRF 
Electronic case report form 
FDA 
United States Food and Drug Administration 
GCP 
Good Clinical Practice 
HM 
Hand motion 
IB 
Investigator brochure 
ICF 
Informed consent form 
ICH 
International Conference on Harmonization 
IND 
Investigational New Drug 
IOL 
Intraocular lens 
IOP 
Intraocular pressure 
IRB 
Institutional Review Board 
ITT 
Intent-to-treat  
LP 
Light perception 
MAP 
Eyenovia Micro-Array Print Dispenser 
NLP 
No light perception 
OU 
Both eyes (within this protocol, OU does not connote simultaneous treatment/evaluation) 
PETG 
Polyethylene terephthalate glycol 
PP 
Per-protocol 
SAE 
Serious adverse event 
SD 
Standard deviation 
SLE 
Slit lamp examination 
TEAE 
Treatment-emergent adverse event 
TLR 
Total letters read 
UCDVA 
Uncorrected distance visual acuity 
US 
United States 
VA 
Visual acuity 
WMA 
World Medical Association 
µD 
Microdose 
µL 
Microliter 
NOTE:  The first occurrence of some abbreviations is not spelled out in the document (e.g., units of 
measure). 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-41 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 6 OF 60 
06 MAY 2021 
 
 
PROTOCOL EYN-MYD-TP-41 
VERSION B 
 
 
A SINGLE-CENTER, ASSESSOR-MASKED, ACTIVE-CONTROLLED, PHASE 4 STUDY 
EVALUATING SPEED OF PUPIL DILATION WITH THE MICRO-ARRAY PRINT (MAP) 
DISPENSER WHEN COMPARING 2 DOSING REGIMENS OF TROPICAMIDE-
PHENYLEPHRINE FIXED COMBINATION OPHTHALMIC SOLUTION  
(THE SPEED STUDY) 
 
1. PERSONNEL AND FACILITIES 
 
Study Sponsor: 
Eyenovia, Inc. 
295 Madison Avenue, Suite 2400 
New York, NY  10017  USA 
Phone:  917-289-1117 
Email:  admin@eyenoviabio.com 
 
Chief Medical Officer: 
Tsontcho Ianchulev, MD, MPH 
Phone:  617-216-4369 
Email:  tianchul@eyenoviabio.com 
 
Medical Monitor: 
Beth Scott, OD, MS, Medical Affairs 
Phone:  214-578-4171 
Email:  bscott@eyenoviabio.com  
 
Overall Program Management: 
Beth Scott, OD, MS, Medical Affairs 
Phone:  214-578-4171 
Email:  bscott@eyenoviabio.com  
 
  
 
 
 
 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-41 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 7 OF 60 
06 MAY 2021 
 
2. STUDY SYNOPSIS 
 
2.1. Study Objective    
 
The primary objective of this study is to compare pupil dilation speed achieved with a single dose of 
MAP Dispenser-administered tropicamide-phenylephrine fixed combination to two doses of 
MAP Dispenser-administered tropicamide-phenylephrine fixed combination administered 5 
minutes apart.  
 
2.2. Study Drugs 
 
• Investigational Product, Dosage and Mode of Administration:  Commercially available 
tropicamide-phenylephrine fixed combination ophthalmic solution (T-P OFTENO SOLUCIÓN 50 
mg/8 mg/ ml) delivered by the MAP Dispenser. In this study, drug will be administered at separate 
visits as a single mist to each eye or as 2 mists administered 5 minutes apart to each eye. 
 
2.3. Study Population 
 
At least 60 volunteer participants will be randomized to study drug administration in order to complete 
required follow-up treatments and evaluations of 56 or more subjects.  Subjects will be enrolled at a 
single study site in Tijuana, Mexico.   
 
2.4. Study Design   
 
This trial is a single center, assessor-masked, active-controlled, cross-over noninferiority study evaluating 
tropicamide-phenylephrine fixed combination ophthalmic solution administered as a single mist to each 
eye and as two mists, administered 5 minutes apart in each eye.  
 
Volunteer participants will be screened for study eligibility during a Screening Visit and enrolled after 
signing the study-specific informed consent form (ICF).  Subjects meeting all inclusion/exclusion criteria 
will be scheduled for 2 treatment visits, which must occur at least 4 days, but no more than 7 days apart.  
At each of the two treatment visits, baseline measurements will be taken, then either 1 mist or 2 mists of 
the study drug will be administered to both eyes (OU).  In the instances of 2 mists, the separate dispensing 
will occur approximately 5 minutes apart.  Only 1 mist or 2 mists of the drug will be administered per 
treatment visit, and drug administration will occur in a randomized order using one of two sequences - 
AB and BA, where A is the dispensing of 1 mist of tropicamide-phenylephrine fixed combination and B 
is the dispensing of 2 mists of tropicamide-phenylephrine fixed combination. Afterwards, efficacy and 
safety assessments will be performed.   
 
This study will be single-masked. There will be no differences in the presentation of the investigational 
drug administered and study personnel conducting ophthalmic assessments will be masked to study drug 
administration assignment.  Personnel who administer study drug on a given day will not be allowed to 
perform post-study drug administration ophthalmic assessments on that day. 
 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
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06 MAY 2021 
 
2.5. Inclusion Criteria 
 
Subjects eligible for study participation must meet each criterion listed below.  Ocular criteria must be 
met for both eyes. 
  
1. Male or female 18 years old or above. 
2. Female subjects must be either 1-year postmenopausal, surgically sterilized, or, if of 
childbearing potential, have a negative urine pregnancy test during the Screening Visit and 
agree to use an acceptable form of contraception throughout the study.  Acceptable methods 
include the use of at least one of the following: intrauterine (intrauterine device), hormonal 
(oral, injection, patch, implant, ring), barrier with spermicide (condom, diaphragm), or 
abstinence. 
3. Ability to provide signed written consent prior to participation in any study-related 
procedures. 
4. Ability to return for all study treatment visits. 
5. Photopic screening pupil diameter ≤ 3.5 mm in each eye. 
 
2.6. Exclusion Criteria 
 
Subjects with any of the following diseases, surgeries or conditions are ineligible for study participation.  
Subjects may not participate if either eye meets any of the ocular exclusion criteria. 
 
1. Pregnancy or lactation. 
2. Allergy to phenylephrine hydrochloride. 
3. Allergy to tropicamide. 
4. Allergy to benzalkonium chloride. 
5. History of benign prostatic hyperplasia. 
6. Use of benzodiazepines, monoamine oxidase inhibitors, tricyclic antidepressants, 
anticonvulsants, cholinergic drug  at screening or anticipated during the study period.  
7. Participation in any study of an investigational drug or device within 30 days prior to the 
Screening Visit, or at any time during the study period. 
8. History of closed-angle glaucoma. 
9. Anatomically narrow anterior chamber angles (Van Herrick grade ≤ 2 in either eye). 
10. Ocular surgery or laser treatment of any kind prior to the Screening Visit. 
11. History of chronic or acute uveitis. 
12. History of traumatic iritis or hyphema. 
13. History of traumatic mydriasis or angle recession. 
14. History of heterochromia. 
15. Irregularly-shaped pupil secondary to ocular trauma or congenital defect. 
16. History of neurogenic pupil disorder (e.g., Horner’s syndrome, third cranial nerve palsy, 
Adie’s pupil, Argyl Robertson syndrome, etc.). 
17. History of anterior chamber intraocular lens (IOL) or iris-fixated IOL. 
18. History of iris surgery of any kind (e.g., iridotomy, iridectomy, coreoplasty). 
19. History of iris atrophy. 
20. History of iris – cornea apposition/touch. 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
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21. Unwilling or unable to discontinue use of contact lenses at treatment visits. 
22. Current active eye disease for which topical or systemic ophthalmic medication is necessary, 
except for dry eye disease managed using artificial tears (AT).   AT’s must be 
discontinued on the day of each treatment visit. 
23. Presence of a severe/serious ocular condition, or any other unstable medical condition that, in 
the Investigator’s opinion, may preclude study treatment and/or follow-up. 
 
2.7. Primary Performance Endpoint 
 
The primary performance endpoint is mean change in pupil diameter at 35 minutes versus per-visit 
baseline, as measured by digital pupillometry in highly photopic conditions.  The highly photopic 
condition will be established using a fully-charged transilluminator (muscle light) at the brightest setting.  
 
2.8. Exploratory Outcomes 
 
• 
Proportion of eyes achieving pupil diameter of 6.0 mm or greater at 35 minutes 
• 
Proportion of eyes achieving pupil diameter of 7.0 mm or greater at 35 minutes 
• 
Mean change in pupil diameter at other timepoints (15, 55 minutes) 
• 
Distribution of pupil diameters at 15, 35, 55 minutes 
• 
Proportion of successful administrations on 1st attempt 
 
These exploratory outcomes will also be measured using digital pupillometry in highly photopic 
conditions. 
 
2.9. Safety Outcomes 
 
• 
Slit-lamp examination findings 
• 
Occurrence of AEs 
• 
Intraocular pressure (IOP) measured at 60 minutes post-administration 
• 
Visual acuity changes 
• 
Proportion of administrations without Micro-Array Print (MAP) Dispenser touch to the eye 
 
 
 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-41 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 10 OF 60 
06 MAY 2021 
 
2.10. Schedule of Visits, Procedures and Evaluations 
 
Study visits include a Screening Visit, which must occur between 1 and 14 days prior to treatment; 
followed by Treatment Day 1, then Treatment Day 2.  Treatment visits must occur at least 4 days and not 
more than 7 days apart.  The study visit schedule is presented in flow chart form in Figure 1. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Figure 1:  Study Visit Flow Chart 
 
 
2.10.1. Study Medication Administration and Clinical Assessments 
 
Study medication administration and clinical assessments will be performed at study visits as shown in 
APPENDIX 1:  SCHEDULE OF MEDICATION ADMINISTRATION AND EXAMINATIONS. 
Screening Visit 
Treatment Day 1 
Randomization 
Study drug administered 
Efficacy & Safety assessments 
 
Treatment Day 2 
Study drug administered 
Efficacy & Safety assessments 
 
-14 Days – 1 Day after Screening 
Visit 
4 – 7 Days after Treatment Day 1 
Study Exit 
Fails to meet 
inclusion/exclusion criteria 
Meets inclusion/exclusion criteria 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
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CONFIDENTIAL - PAGE 11 OF 60 
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3. INTRODUCTION AND RATIONALE  
  
3.1. Introduction 
 
Eyenovia, the Sponsor of this study, is a specialty pharmaceutical company pursuing development of a 
variety of ocular topical medications including mydriatics.  Eyenovia’s drug formulations are 
administered using a proprietary system called the Micro-Array Print (MAP) Dispenser, which is a 
container closure system designed to accurately deliver precise low-volume doses of topical ocular 
pharmaceuticals.  
 
Phenylephrine is an α1-adrenegic receptor sympathetic agonist that has been used for more than 70 years 
to dilate the pupil for diagnostic, therapeutic, and surgical procedures. Systemically, phenylephrine, when 
administered topically to the eye at higher doses, is known to cause serious cardiovascular reactions 
including ventricular arrhythmias, myocardial infarction, and elevated blood pressure in some patients1,2. 
These risks are especially relevant when more mydriasis is required, and phenylephrine 10% is used, 
rather than the lower 2.5% concentration.   Tropicamide is a synthetic derivative of tropic acid that 
became available for ocular use in 19593.  It is an anticholinergic mydriatic agent that acts by blocking the 
iris sphincter muscle and the ciliary muscle responses to acetylcholine.  As an anticholinergic related to 
atropine and scopolamine, systemically absorbed tropicamide in higher doses has the potential to cause 
rare but potentially serious adverse events, particularly in the pediatric population.  
 
The MAP Dispenser technology facilitates accurate topical ocular delivery of a controlled quantity of 
mydriatic agents in a precise manner to maximize therapeutic control and minimize systemic absorption 
and loss to washout. 
 
3.2. Pupil Dilation 
 
A variety of ophthalmic evaluations require pupil dilation to provide an unobstructed view of the lens, 
retina, and optic nerve. The degree of pupil dilation required depends on the procedure being performed.   
For routine diagnostic procedures, and in conditions where short term pupil dilation is desired, the goal is 
to achieve adequate mydriasis for visualization with minimal cycloplegic effect. 
 
 
 
 
1 Vaajanen A, Vapaatalo H. A single drop in the eye - Effects on the whole body? Open Ophthalmol J. 
2017;11:305‒314. doi: 10.2174/1874364101711010305 
2 Farkouh A, Frigo P, Czejka M. Systemic side effects of eye drops: a pharmacokinetic perspective. Clin 
Ophthalmol. 2016;10:2433‒2441. 
3 Merrill D L, Goldber, B, Zavell S. bis-Tropamide, A new parasympatholytic. Curr Ther Res. 1960;12:43‒50. 
 


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CONFIDENTIAL - PAGE 12 OF 60 
06 MAY 2021 
 
3.3. Existing Mydriatic Options 
 
Dilation of the pupil is currently produced with parasympathetic anticholinergics such as tropicamide, 
cyclopentolate, homatropine, and atropine; or with sympathetic agonists (sympathomimetics) such as 
phenylephrine4. 
 
Parasympathetic antagonists block acetylcholine activation of muscarinic receptors, thereby paralyzing 
the iris sphincter muscle. The result is an enlarged pupil; however, at the same time, the ciliary muscle 
that regulates visual accommodation can be paralyzed, resulting in blurry vision that can persist for hours. 
Additionally, rare but serious systemic side effects have been observed after ocular administration of 
anticholinergics such as tropicamide, particularly in infants and children5,6. 
 
Sympathetic agonists produce dilation by stimulating the iris dilator muscle and constricting ophthalmic 
blood vessels. Although phenylephrine does not induce blurry vision, at lower concentrations; e.g., 2.5%, 
it generally does not dilate the pupil as well as parasympathetic agonists. Where full mydriasis is required, 
higher concentrations of phenylephrine; e.g., 10%, are required.  Higher concentration phenylephrine, 
however, has been known to cause cardiovascular effects after systemic absorption, which include 
hypertension, tachycardia, and more rarely, arrhythmia and stroke, especially in patients with pre-existing 
heart conditions7.  
 
To maximize dilation efficacy and minimize side effects, most ophthalmologists employ a combination of 
both classes of mydriatic agents. Tropicamide-Phenylephrine fixed combination ophthalmic solution is 
approved for eyedrop delivery and currently marketed (T-P OFTENO SOLUCIÓN 50 mg/8 mg/ ml, 
AUGEX – MEDICAMENTO – PR VADEMECUM) for dilation of the pupil. 
 
3.4. Tropicamide-Phenylephrine Fixed Combination Ophthalmic Microdosing Solutions for 
Pupil Dilation 
 
Eyenovia’s MAP Dispenser is designed to deliver precise microvolumes of topical ophthalmic 
medications using a piezo-generated microdroplet spray. When the dispenser is actuated, a precisely 
calibrated single microdose is delivered.  It is anticipated that microdosing of ophthalmic drugs will 
enhance drug bioavailability and efficacy while minimizing systemic absorption and any associated side 
effects. Tropicamide-Phenylephrine fixed combination ophthalmic solution is approved for eyedrop 
delivery and currently marketed (T-P OFTENO SOLUCIÓN 50 mg/8 mg/ ml, AUGEX – 
MEDICAMENTO – PR VADEMECUM) for dilation of the pupil. The drug may be administered in 
approximately 8 µL microdoses using the MAP Dispenser.   
 
4 Grob SR, Gonzalez LA, Daly MK. Management of mydriasis and pain in cataract and intraocular lens surgery: 
review of current medications and future directions. Clin Ophthalmol. 2014;8:1281‒1289. doi: 
10.2147/OPTH.S47569 
5 Vaajanen A, Vapaatalo H. A single drop in the eye - Effects on the whole body? Open Ophthalmol J. 
2017;11:305‒314. doi: 10.2174/1874364101711010305 
6 Farkouh A, Frigo P, Czejka M. Systemic side effects of eye drops: a pharmacokinetic perspective. Clin Ophthalmol. 
2016;10:2433‒2441. 
7 Ibid. 


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CONFIDENTIAL - PAGE 13 OF 60 
06 MAY 2021 
 
 
Ocular microdosing with the MAP Dispenser has been evaluated in humans using phenylephrine 2.5%, 
phenylephrine 10%, and tropicamide 1.0% solutions.   
 
3.5. Ophthalmic Microdosing 
 
The rationale for Eyenovia’s mydriatic development is the desire to optimize pupil dilation while 
reducing systemic side effects, improving local tolerability, and decreasing the amount of medication lost 
to drainage from increased tear turnover rate.  
 
The average tear volume of an adult eye is 6−7 μL8,9. The estimated maximum volume of eyedrops that 
the cul de sac can contain, albeit momentarily, is approximately 30 μL; this is reduced to 10 µL with 
normal blinking10.  The typical eyedropper (either unit dose or multidose), however, provides drops that 
range from 25 μL to > 50 μL. Even among bottles of the same size and shape, eyedrop sizes vary 
significantly depending on the angle of the bottle, the amount of ophthalmic solution remaining, and even 
among manufacturers11.  
 
Much of the eyedrop volume overflows into the nasolacrimal canal where the active drug product 
becomes available systemically. Ocular drugs that are absorbed by the nasolacrimal mucosa mimic 
intravenous injection delivery insofar as they are not susceptible to first-pass hepatic metabolism. 
Additionally, ocular medication in swallowed nasolacrimal secretions is theoretically available for 
absorption in the gastrointestinal tract. Taken together, only a small fraction of the applied medication is 
absorbed directly into the eye, while there remain multiple opportunities for systemic absorption12. This 
can result in significant safety concerns, especially with drugs like phenylephrine and tropicamide that are 
known to produce dose-dependent systemic side effects13,14.  Additionally, excess drug in the eye may 
cause ocular surface toxicity and tolerability issues. 
 
Instillation of eyedrops stimulates lacrimation and can increase the tear turnover rate more than 4-fold, 
which additionally dilutes the drug product administered15.  If the eyedrop causes discomfort, the loss rate 
 
8 Mishima S, Gasset A, Klyce SD Jr, Baum JL. Determination of tear volume and tear flow. Invest. Ophthalmol. 
5(3), 264‒276 (1966). 
9 Scherz W, Doane MG, Dohlman CH. Tear volume in normal eyes and keratoconjunctivitis sicca. Albrecht Von 
Graefes Arch. Klin. Exp. Ophthalmol. 1974;192(2):141‒150. 
10 Shell JW. Pharmacokinetics of topically applied ophthalmic drugs. Surv Ophthalmol. 1982;26(4):207-18. 
11Mammo ZN, Flanagan JG, James DF, Trope GE.Generic versus brand-name North American topical glaucoma 
drops. Can J Ophthalmol. 2012;47(1):55‒61. doi: 10.1016/j.jcjo.2011.12.004. 
12 Urtti A, Salminen L. Minimizing systemic absorption of topically administered ophthalmic drugs. Surv. 
Ophthalmol. 37(6), 435−456 (1993). 
13Vaajanen A, Vapaatalo H. A single drop in the eye - Effects on the whole body? Open Ophthalmol J. 
2017;11:305‒314. doi: 10.2174/1874364101711010305 
14 Farkouh A, Frigo P, Czejka M. Systemic side effects of eye drops: a pharmacokinetic perspective. Clin Ophthalmol. 
2016;10:2433‒2441. 
15 Jordan A, Baum J. Basic tear flow. Does it exist? Ophthalmology. 1980 Sep;87(9):920‒930. 


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can be even higher16.  Approximately 80% of a medication instilled as an eyedrop is lost to drainage 
during the first 15‒30 seconds17. 
 
Microdosed mydriatic medications have been demonstrated to provide similar or superior efficacy 
compared to traditional eyedropper systems. Microdosing can also reduce systemic absorption and side 
effects, improve patient comfort, and minimize drug dilution and spillage. Studies comparing microdosed 
phenylephrine and cyclopentolate to traditional eyedrops (26 ‒ 30 µL drop size) in premature babies18 
and in full-term infants19 have shown equivalent pupil dilation with drop sizes ranging from 5 ‒ 8 µL20,21 
while reducing systemic levels by more than 50%22. 
 
Tropicamide 1.0% spray application caused similar pupil dilation as the same solution delivered as an 
eyedrop, but with less discomfort23 .  The main reason microdosing has not become the accepted method 
for topical ocular dosing of ophthalmic medication is that no system has been developed that can 
consistently deliver accurate microdoses of these medications. 
 
As shown in Figure 1, the Eyenovia MAP Dispenser is designed to deliver a finely controlled 
microdroplet spray of tropicamide-phenylephrine fixed combination ophthalmic solution with precisely 
defined volume, velocity, and geometry.24 These characteristics facilitate high-precision, piezo-generated 
medication delivery that gently coats the ocular surface. 
 
 
16 Farris RL, Stuchell RN, Mandell ID. Basal and reflex human tear analysis. I Physical measurements: osmolarity, 
basal volumes, and reflex flow rate. Am Acad Ophthal 1981;88:852. 
17 Lallemand F, Daull P, Benita S, Buggage R, Garrigue JS. Successfully improving ocular drug delivery using the 
cationic nanoemulsion, novasorb. J Drug Deliv. 2012;2012:604204. 
18 Wheatcroft S, Sharma A, McAllister J. Reduction in mydriatic drop size in premature infants. Br J Ophthalmol. 
1993;77(6):364-5. 
19 Lynch MG, Brown RH, Goode SM, Schoenwald RD, Chien DS. Reduction of phenylephrine drop size in infants 
achieves equal dilation with decreased systemic absorption. Arch Ophthalmol. 1987;105(10):1364-5. 
20 Wheatcroft S, Sharma A, McAllister J. Reduction in mydriatic drop size in premature infants. Br J Ophthalmol. 
1993;77(6):364-5. 
21 Lynch MG, Brown RH, Goode SM, Schoenwald RD, Chien DS. Reduction of phenylephrine drop size in infants 
achieves equal dilation with decreased systemic absorption. Arch Ophthalmol. 1987;105(10):1364-5. 
22 Ibid. 
23Akman A, Aydin P. Comparison of mydriatic efficacy of spray application and drop instillation of tropicamide 
1%. Eye (Lond). 1999;13(5):653‒655. 
24 Ianchulev T, Chayet A, Kahook M, Packer M, Pasquale L, Weinreb RN. Pharmacodynamic profile of mydriatic 
agents delivered by ocular piezo-ejection microdosing compared with conventional eyedropper. Ther Deliv. 
2016;7(11):751-760. 
 


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Figure 1: Microdroplet Distribution Pattern from the MAP Dispenser 
Image analysis of high-speed photographs of the microdroplet plume quantitatively confirms precise 
medication microdroplet delivery. Shown are representative side (A) and cross-sectional (B) views of 
the droplet dispersal pattern. Color-coding indicates the percentage of the total medication dose 
present at different distances from the nozzle-to-target dead- center axis. The cross-sectional pattern on 
the right was captured 3 cm from the MAP Dispenser nozzle. 
 
 
 


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4. INVESTIGATIONAL PRODUCT DESCRIPTION 
 
This study will evaluate pupil dilation speed subsequent to dosing with 1 mist and 2 mists of tropicamide-
phenylephrine fixed combination ophthalmic solution (T-P OFTENO SOLUCIÓN 50 mg/8 mg/ ml) 
intended for mydriasis. 
4.1 How Provided 
The tropicamide-phenylephrine fixed combination ophthalmic solution to be studied in this protocol is 
contained sterile in the Micro-Array Print (MAP) dispenser, which is designed to deliver precise low-
volume (~ 8 µL) doses of topical ocular pharmaceutical ingredients.  As shown in Error! Reference s
ource not found., the Micro-Array Print (MAP) Dispenser is composed of a “cartridge” and a “base”. 
 
 
 
Figure 2: Micro-Array Print (MAP) Dispenser Front (L) and Back (R) View 
 
The cartridge is the primary container closure system, which contains a vial filled with sterile study 
ophthalmic solution.  At the time of initial Micro-Array Print (MAP) Dispenser actuation, the seal on the 
vial is punctured to allow the solution to flow to the Micro-Array Print (MAP) Dispenser delivery system.  
The cartridge also contains a piezo-delivery system designed to drive the solution through the Mist 
Opening.  There is ≤ 100 milliseconds time lapse between when the first solution droplet hits the corneal 
surface and completion of dose delivery, which is faster than the average involuntary blink response time 
(Kwon 2013). 
 
Dispense Button 
Fill Button 
Base Unit 
Cartridge 
Mist Nozzle 


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Figure 3:  Diagram of the Micro-Array Print (MAP) Dispenser Base Unit (L) and Cartridge 
Assemblies (R) 
 
Prior to initial Micro-Array Print (MAP) Dispenser use, tropicamide-phenylephrine fixed combination 
solution is aseptically transferred from the commercially available tropicamide-phenylephrine fixed 
combination ophthalmic solution bottle (T-P OFTENO SOLUCIÓN 50 mg/8 mg/ ml) in which it was 
received into the MAP Dispenser vial, the MAP Dispenser vial is inserted into the cartridge assembly, the 
cartridge and base are clipped together, and the seal on the vial containing the drug is punctured by 
depressing the MAP Dispenser Fill Button.  Puncture of this seal creates a fluid path for the drug.  
Depression and release of the Fill Button approximately 5 times primes the system for initial drug 
delivery.  At the time of dosing, the Fill Button is again pressed and released to activate the piezo element 
to create the aerosol containing the drug dose.  When the Dispense Button is pressed, approximately 8 µL 
of the drug is horizontally delivered through the Mist Nozzle to the cornea.  
4.2 Route of Administration, Dosage, Regimen, Treatment Period(s) 
The study medication will be administered by trained clinical study staff at two treatment visits in 
accordance with each subject’s treatment assignment schedule.  Either 1 mist in each eye or 2 mists 
administered approximately 5 minutes apart in each eye will be topically administered.    
 
 
  
 
Fill Button 
Vial 
Cartridge Assembly 
Base Unit Assembly  
 
(Electrical mechanical 
components in top diagram) 
Mist Nozzle 


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5. STUDY OBJECTIVE 
 
The primary objective of this study is to compare pupil dilation speed achieved with a single dose of 
MAP Dispenser-administered tropicamide-phenylephrine fixed combination to two doses of MAP 
Dispenser-administered tropicamide-phenylephrine fixed combination administered 5 minutes apart.  
 
 
6. STUDY DESIGN 
 
This trial is a single center, assessor-masked, active-controlled, cross-over noninferiority study evaluating 
tropicamide-phenylephrine fixed combination ophthalmic solution administered as a single mist to each 
eye and as two mists, administered 5 minutes apart in each eye.  
 
Volunteer participants will be screened for study eligibility during a Screening Visit and enrolled after 
signing the study-specific informed consent form (ICF).  Subjects meeting all inclusion/exclusion criteria 
will be scheduled for 2 treatment visits, which must occur at least 4 days, but no more than 7 days apart.  
At each of the two treatment visits, baseline measurements will be taken, then either 1 mist or 2 mists of 
the study drug will be administered to both eyes (OU).  In the instances of 2 mists, the separate dispensing 
will occur approximately 5 minutes apart.  Only 1 mist or 2 mists of the drug will be administered per 
treatment visit, and drug administration will occur in a randomized order using one of two sequences - 
AB and BA, where A is the dispensing of 1 mist of tropicamide-phenylephrine fixed combination and B 
is the dispensing of 2 mists of tropicamide-phenylephrine fixed combination. Afterwards, efficacy and 
safety assessments will be performed.   
 
This study will be single-masked. There will be no differences in the presentation of the investigational 
drug administered and study personnel conducting ophthalmic assessments will be masked to study drug 
administration assignment.  Personnel who administer study drug on a given day will not be allowed to 
perform post-study drug administration ophthalmic assessments on that day. 
 
 
 
 
 


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7. STUDY OUTCOMES 
 
7.1 Effectiveness Outcomes 
 
The primary performance endpoint is mean change in pupil diameter at 35 minutes versus visit-specific 
baseline, as measured by digital pupillometry in highly photopic conditions.  The highly photopic 
condition will be established using a fully-charged transilluminator (muscle light) at the brightest setting. 
The difference in mean change in pupil diameter will be compared between the single dose of MAP 
Dispenser-administered tropicamide-phenylephrine fixed combination and two doses of MAP Dispenser-
administered tropicamide-phenylephrine fixed combination administered 5 minutes apart. The primary 
objective is to demonstrate non-inferiority of the single dose to the two doses, with a non-inferiority 
margin of 0.5 mm. 
 
The study’s sample size will provide 90% power to demonstrate non-inferiority of the single dose 
regimen assuming a true difference of 0.0 mm, a standard deviation of 0.85 mm, and a non-inferiority 
margin of 0.5 mm. 
 
Additional exploratory outcomes (also to be measured in highly photopic conditions) include: 
 
• 
Proportion of eyes achieving pupil diameter of 6.0 mm or greater at 35 minutes 
• 
Proportion of eyes achieving pupil diameter of 7.0 mm or greater at 35 minutes 
• 
Mean change in pupil diameter at other timepoints (15, 55 minutes) 
• 
Distribution of pupil diameters at 15, 35, 55 minutes 
• 
Proportion of successful administrations on 1st attempt 
 
These exploratory outcomes will also be measured using digital pupillometry in highly photopic 
conditions. 
 
7.2. Safety Outcomes 
 
A safety analysis will be conducted to compare one dose vs. two doses of fixed combination of 
phenylephrine 2.5%-tropicamide-phenylephrine fixed combination µD for the following outcomes: 
 
• 
Slit-lamp examination (SLE) findings 
• 
Anterior segment exam by transilluminator findings 
• 
Occurrence of AEs 
• 
Intraocular pressure (IOP) measured at 60 minutes post-administration 
• 
Visual acuity changes 
• 
Proportion of administrations without Micro-Array Print (MAP) Dispenser touch to the eye 
 
 
 
 


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8. STUDY POPULATION   
 
Volunteer participants will be considered enrolled after signing the study-specific ICF(s) and screened for 
study eligibility during a Screening Visit.  Subjects meeting all inclusion/exclusion criteria will be 
scheduled for 2 treatment visits.  It is assumed that approximately 5% of subjects (n = 4) will fail to 
complete the study visits after randomization to treatment; therefore, to have 56 subjects evaluable for the 
primary efficacy analysis, a minimum of 60 subjects will be randomized to study drug administration at 
Treatment Visit 1 (Day 1).  To account for possible screen failures, the study will enroll up to 70 subjects. 
 
8.1. Inclusion Criteria 
 
Subjects eligible for study participation must meet each criterion listed below.  Ocular criteria must be 
met for both eyes. 
  
1. Male or female 18 years old or above. 
2. Female subjects must be either 1-year postmenopausal, surgically sterilized, or, if of 
childbearing potential, have a negative urine pregnancy test during the Screening Visit and 
agree to use an acceptable form of contraception throughout the study.  Acceptable methods 
include the use of at least one of the following: intrauterine (intrauterine device), hormonal 
(oral, injection, patch, implant, ring), barrier with spermicide (condom, diaphragm), or 
abstinence. 
3. Ability to provide signed written consent prior to participation in any study-related 
procedures.   
4. Ability to return for all study treatment visits. 
5. Photopic screening pupil diameter ≤ 3.5 mm in each eye. 
 
8.2. Exclusion Criteria 
 
Subjects with any of the following diseases, surgeries or conditions are ineligible for study participation.  
Subjects may not participate if either eye meets any of the ocular exclusion criteria. 
 
1. Pregnancy or lactation. 
2. Allergy to phenylephrine hydrochloride. 
3. Allergy to tropicamide. 
4. Allergy to benzalkonium chloride. 
5. History of benign prostatic hyperplasia. 
6. Use of benzodiazepines, monoamine oxidase inhibitors, tricyclic antidepressants, 
anticonvulsants at screening or anticipated during the study period.  
7. Participation in any study of an investigational drug or device within 30 days prior to the 
Screening Visit, or at any time during the study period. 
8. History of closed-angle glaucoma. 
9. Anatomically narrow anterior chamber angles (Van Herrick grade ≤ 2 in either eye). 
10. Ocular surgery or laser treatment of any kind prior to the Screening Visit. 
11. History of chronic or acute uveitis. 
12. History of traumatic iritis or hyphema. 


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13. History of traumatic mydriasis or angle recession. 
14. History of heterochromia. 
15. Irregularly-shaped pupil secondary to ocular trauma or congenital defect. 
16. History of neurogenic pupil disorder (e.g., Horner’s syndrome, third cranial nerve palsy, 
Adie’s pupil, Argyl Robertson syndrome, etc.). 
17. History of anterior chamber intraocular lens (IOL) or iris-fixated IOL. 
18. History of iris surgery of any kind (e.g., iridotomy, iridectomy, coreoplasty). 
19. History of iris atrophy. 
20. History of iris – cornea apposition/touch. 
21. Unwilling or unable to discontinue use of contact lenses at treatment visits. 
22. Current active eye disease for which topical or systemic ophthalmic medication is necessary, 
except for dry eye disease managed using artificial tears (AT).   AT’s must be discontinued on 
the day of each treatment visit. 
23. Presence of a severe/serious ocular condition, or any other unstable medical condition that, in 
the Investigator’s opinion, may preclude study treatment and/or follow-up. 
 
 
 
 


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9. STUDY METHODS 
 
9.1. Subject Informed Consent 
 
The EC-approved study ICF must be used for administration of informed consent.   
 
The investigator or designee will explain the study purpose, procedures and responsibilities to the 
potential participant and provide sufficient opportunity to ask questions, while allowing adequate time for 
consideration of the information provided.  Written consent for study participation must be present prior 
to initiation of any study-specific procedure; subjects will be considered enrolled in the study upon their  
signature on the consent form(s).  It is the responsibility of the Investigator to complete the informed 
consent process, maintain a copy of the signed consent forms in the subject’s medical records, and 
provide each subject with a copy of their fully-executed consent documents. 
 
9.2. Subject Selection and Screening Procedures 
 
Study participants will be recruited from the Investigator’s patient population, referrals, or other outreach 
methods.  Written recruitment materials directed to potential study participants must be approved by the 
EC.   
 
Where possible, candidate participants may be pre-screened via review of their medical charts to evaluate 
potential eligibility based on study inclusion/exclusion criteria.  During the informed consent discussion, 
the potential participant’s willingness and ability to meet the follow-up requirements will be evaluated.  
Those who elect to sign the study consent form(s) will be considered enrolled in the study and given a 
study identification code.   At/after the time of enrollment, the subject will be evaluated at the Screening 
Visit for study eligibility based on inclusion/exclusion requirements established in Sections 8.1 and 8.2.  
Subjects who fail to qualify for the study will be considered “screen failures” and exited from the study.  
Subjects who successfully complete the Screening Visit will be scheduled for their next visit at which the 
first study medication will be administered.   
 
9.3. Randomization to Study Drug Administration Sequence  
 
Each eligible subject will be randomly assigned to receive one of 2 sequences, AB and BA, where A is 
the dispensing of 1 mist of tropicamide-phenylephrine fixed combination and B is the dispensing of 2 
mists of tropicamide-phenylephrine fixed combination.  After Treatment Visit 2, each study subject 
should have received both 1 mist of tropicamide-phenylephrine fixed combination and the 2 mists of 
tropicamide-phenylephrine fixed combination. The study drug administration schedule is equally 
distributed across the 2 randomization sequences.   Subject drug administration assignments will be 
prepared using a computer-generated randomization scheme by an independent statistician who is not 
involved in the day-to-day conduct of the study.  The drug assignment sequence (Study Drug AB or BA) 
will be provided in paper format contained in a sealed envelope for each subject randomized to treatment. 
 
This study will be single-masked. There will be no differences in the presentation of the investigational 
drug administered and study personnel conducting ophthalmic assessments will be masked to study drug 
administration assignment.  Personnel who administer study drug on a given day will not be allowed to 


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perform post-study drug administration ophthalmic assessments on that day. 
 
9.3.1. Procedure for Unmasking Study Drug Assignment  
 
Unmasking of study drug assignments can only occur in the event of a medical emergency or occurrence 
of an AE that, in the opinion of the investigator, warrants such action.  In the absence of medical need, the 
study drug assignment will not be available to the investigator, site staff, or Sponsor (or designee) 
representative involved in day-to-day study management until study completion and database lock. 
 
In the event an investigator considers unmasking of a subject’s study drug assignment to be necessary, 
he/she must first contact the study Medical Monitor (or designee) to discuss the case.  Only after 
consultation with the study Medical Monitor will the decision be made regarding unmasking. Study drug 
assignments will be revealed only for the subjects approved for unmasking, thus leaving masking on 
remaining subjects intact. 
 
9.4. Study Drug Packaging, Labeling and Storage 
 
The investigational product will be delivered in microdose form using the Eyenovia MAP Dispenser.  The 
product must be aseptically transferred from the commercially available ophthalmic solution bottle (T-P 
OFTENO SOLUCIÓN 50 mg/8 mg/ ml for dilation of the pupil) in which it is received to the sterile vial 
provided with the MAP Dispenser cartridge.  After aseptic transfer, the filled vial is re-inserted into the 
MAP Dispenser cartridge. 
 
To maintain masking of the study drug assignments for study personnel performing clinical evaluations, 
study inventory storage and management will be performed by a sterile drug transfer technician not 
allowed to administer drug and not allowed to perform ophthalmic assessments.  The Sponsor, or 
designee, will provide study drug dispensers to the site.   
 
Instructions regarding study medication preparation are included in  
APPENDIX 2:  STUDY MEDICATION PREPARATION PROCEDURE.  Each designated sterile 
drug transfer technician will be trained and certified by the Sponsor, or designee, prior to study 
medication preparation. 
 
Prior to use, study medication must be stored in a secure area.  To maintain product stability, keep the 
bottle tightly closed in no more than 30 °C.  
 
New MAP Dispensers must be prepared by the sterile drug transfer technician for use by site personnel on 
each treatment day. To maintain masking of site personnel, the MAP Dispensers containing study drug 
will be labeled with the pharmacy preparation date and “use by” date, then forwarded to site personnel for 
use.  The date on which the MAP Dispenser is used will be added by site personnel. At the end of the 
treatment day, each MAP Dispenser provided will be returned to storage. Please note that each MAP 
Dispenser prepared must be used on the day of preparation; the MAP Dispenser may not be re-
used on subsequent days.   
 


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9.5. Study Drug Administration 
 
Site personnel designated to administer study medication will be trained by the Sponsor on proper use of 
the MAP Dispenser prior to study initiation.  A single MAP Dispenser should be used for study drug 
administration to all subjects as indicated by their study drug assignment on that treatment day. At each 
treatment visit, after collection of baseline BCDVA, IOP and SLE data, study medication will be 
administered by designated site personnel in accordance with the subject’s assigned sequence.  Subjects 
will be administered either 1 mist of tropicamide-phenylephrine fixed combination OU or 2 mists of 
tropicamide-phenylephrine fixed combination OU.  With the dispensing of 2 mists of tropicamide-
phenylephrine fixed combination, approximately 5 minutes should elapse between the first and the second 
dose administration.  Study drug administration procedures are further detailed in APPENDIX 3:  
STUDY MEDICATION ADMINISTRATION PROCEDURE. 
 
9.6. Study Visits and Clinical Assessments 
 
Study participants will be evaluated at a Screening Visit.  Subjects who qualify for further study 
participation will be scheduled for visits on Treatment Day 1 and Treatment Day 2 at which the assigned 
study drug is administered and examinations are performed.  Data collected from each visit will be 
documented in the source records and recorded on study electronic Case Report Forms (eCRF).   
 
The timing and frequency of assessments/procedures to be performed at each visit is outlined in 
APPENDIX 1:  SCHEDULE OF MEDICATION ADMINISTRATION AND EXAMINATIONS.  
Methodology for study examinations is presented in APPENDIX 4:  EXAMINATION 
PROCEDURES. 
 
9.6.1. Screening Visit (-14 Days to Day 0) 
 
The Screening Visit must occur no more than 14 days prior and no less than 1 day prior to Treatment 
Visit 1.  At the Screening Visit, the following activities will be performed:  
 
• 
Documentation of study informed consent/assent using EC-approved consent form(s) 
• 
Collection of demographic data (gender, age, ocular comorbidities) 
• 
Collection of medical/ocular history and medication use data 
• 
Administration of urine pregnancy test, if applicable 
• 
BCDVA – OU 
• 
Pupil diameter measurement – OU 
• 
Slit lamp biomicroscopy including Van Herrick angle assessment – OU 
• 
Dilated Fundus Exam – OU  
• 
IOP measurement - OU   
• 
Determination of study eligibility 
 
9.6.2. Treatment Visit 1 (Day 1) 
 


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Subjects will return to the site at least 1 day and no more than 14 days after the Screening Visit.  The 
subject will be queried regarding the occurrence of any adverse events since the time of the Screening 
Visit. The following baseline measurements will be taken prior to study drug administration: 
  
• 
Pupil diameter measurement – OU 
• 
BCDVA – OU 
• 
Slit lamp biomicroscopy – OU 
• 
IOP measurement - OU   
 
The first study drug dosing sequence will be either 1 mist or 2 mists of the study drug administered to 
both eyes (OU).  With the 2 mists sequence, the 2 dispensations will occur approximately 5 minutes apart.  
With the 2 mists sequence, the completion of the first administration of medication will be considered 
Time 0 (T0).  The following assessments will be done at the timepoints specified below: 
 
Timepoint 
Clinical Assessment (to be performed in the specified order) 
T0 + 15 minutes (+/- 2 minutes)* 
• 
Pupil diameter measurement – OU 
• 
AE assessment 
T0 + 35 minutes (+/- 5 minutes)** 
• 
Pupil diameter measurement – OU 
• 
AE assessment 
T0 + 55 minutes (+/-10 minutes)*** 
• 
Pupil diameter measurement – OU 
• 
Anterior segment exam by transilluminator – OU 
• 
AE assessment 
 
* The pupil diameter is the only measurement that must be within the window (  2 min) 
** The pupil diameter is the only measurement that must be within the window (  5 min) 
*** The pupil diameter is the only measurement that must be within the window (  10 min)  
 


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9.6.3. Treatment Visit 2 (Day 5 to Day 8) 
 
Subjects will return to the site.  Subjects will be queried regarding the occurrence of any adverse events 
since the previous visit and the following baseline measurements will be taken prior to study drug 
administration: 
 
• 
Pupil diameter measurement – OU 
• 
BCDVA – OU 
• 
Slit lamp biomicroscopy – OU 
• 
IOP measurement - OU   
 
The second study drug dosing sequence will be either 1 mist or 2 mists of the study drug administered to 
both eyes (OU).  With the 2 mists sequence, the 2 dispensations will occur approximately 5 minutes apart. 
With the 2 mists sequence, completion of the first administration of medications will be considered Time 
0 (T0).  The following assessments will be done at the timepoints specified below: 
 
Timepoint 
Clinical Assessment (to be performed in the specified order) 
T0 + 15 minutes (+/- 2 minutes)* 
• 
Pupil diameter measurement – OU 
• 
AE assessment 
T0 + 35 minutes (+/- 5 minutes)** 
• 
Pupil diameter measurement – OU 
• 
AE assessment 
T0 + 55 minutes (+/-10 minutes)*** 
• 
Pupil diameter measurement – OU 
• 
Anterior segment exam by transilluminator – OU 
• 
AE assessment 
 
* The pupil diameter is the only measurement that must be within the window (  2 min) 
** The pupil diameter is the only measurement that must be within the window (  5 min) 
*** The pupil diameter is the only measurement that must be within the window (  10 min) 
 
If applicable, a urine pregnancy test will be administered at this visit. 
 
 


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9.4.5. Unscheduled Visits 
 
Unscheduled visits are those which are not required by the study protocol, but which occur due to an 
ocular intervention or a subject complaint regarding their eye.  No specific testing is required at 
unscheduled visits; rather, the investigator and/or qualified study staff will perform the procedures 
necessary to treat/evaluate the subject at these visits.  Clinical data from these visits will be recorded on 
the relevant Unscheduled Visit eCRF.    
 
9.4.6. Missed Visits 
 
Subjects who miss a scheduled visit should be contacted by site personnel to encourage the subject to 
return to clinic as soon as possible after the missed visit.  In the event a subject does not return for 
multiple consecutive examinations, site personnel must make a minimum of 3 documented attempts to 
contact the subject.  If the subject does not reply to any of these attempts, site personnel should send a 
letter by certified mail (with request for notification of delivery receipt) to the subject.  If a subject is non-
responsive to each of these follow-up attempts, he/she will be terminated from the study and considered 
lost to follow-up. 
 
9.7. Study Exit 
 
9.7.1. Subject Termination 
 
Subjects may be terminated from the study due to: 
 
• 
Failure to meet protocol eligibility criteria  
• 
Investigator decision that termination is medically indicated 
• 
Voluntary withdrawal from the study 
• 
Loss to follow-up 
• 
Other administrative reasons (e.g., study terminated by Sponsor, technical problems, non-
compliance with study medication administration or other study requirements, etc.) 
 
Subjects who terminate due to the occurrence of an AE will be followed until resolution or stabilization of 
the event. 
 
Subjects who exit the study due to failure to meet eligibility criteria may be replaced.  Subjects who exit 
the study prematurely for other reasons will not be replaced.  The Sponsor (or designee) should be 
promptly notified of a subject’s early termination, and information regarding the termination (e.g., date 
and reason) should be documented in the source records and on the Study Exit eCRF. 
 
9.7.2. Subject Completion 
 
Subjects will be considered to have completed the study if they did not terminate study participation prior 
to completion of primary endpoint assessment at Treatment Visit 2. 
 


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10. ADVERSE EVENTS 
 
Throughout the course of the study, all efforts must be made by the investigator to remain alert to possible 
AEs or untoward findings.  All AEs must be assessed for severity and relationship to the study drug, and 
the investigator must take all appropriate and necessary therapeutic measures required for AE resolution.  
Adverse events must be evaluated until resolution or, if the AE is assessed as chronic, until stable.   
 
10.1. Adverse Event Definitions 
 
Most of the AE definitions provided are published in or ICH E6 – Good Clinical Practice:  
Consolidated Guidance.   
 
10.1.1. Adverse Event (AE) 
 
Any untoward medical occurrence associated with the use of a drug in humans, whether or not considered 
drug-related.  
 
10.1.2. Adverse Drug Reaction (ADR) 
 
Any noxious and unintended response to a medicinal product administered at any dose.  A reasonable 
possibility must exist that the adverse reaction is related to the medicinal product administered. 
 
10.1.3. Unexpected Adverse Event or Unexpected Suspected Adverse Reaction  
 
An AE or suspected adverse reaction is considered "unexpected" if it is not listed in the Investigator 
Brochure (IB) or is not listed at the specificity or severity that has been observed; or, if an IB is not 
required or available, is not consistent with the risk information described in the general investigational 
plan or elsewhere in the current application, as amended.  "Unexpected," as used in this definition, also 
refers to AEs or suspected adverse reactions that are mentioned in the IB as occurring with a class of 
drugs or as anticipated from the pharmacological properties of the drug but are not specifically mentioned 
as occurring with the particular drug under investigation. 
 
10.1.4.  Suspected Adverse Reaction 
 
Any AE for which there is a reasonable possibility that the drug caused the event.  "Reasonable 
possibility" means there is evidence to suggest a causal relationship between the study drug and the AE. 
Suspected adverse reaction implies a lesser degree of certainty about causality than adverse reaction, 
which means any AE caused by a drug. 
 
10.1.5. Life-Threatening Adverse Event or Life-Threatening Suspected Adverse Reaction 
 
An AE or suspected adverse reaction is considered “life-threatening” if, in the view of either the 
Investigator or Sponsor, its occurrence places the subject at immediate risk of death.  Life-threatening 
AEs do not include AEs or suspected adverse reactions that, had they occurred in a more severe form, 
might have caused death. 
 
10.1.6. Serious Adverse Event (SAE) or Serious Suspected Adverse Reaction 


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An AE or suspected adverse reaction that, in the view of either the Investigator or Sponsor, results in any 
of the following outcomes:  
 
• 
Death 
• 
Is life-threatening25  
• 
Requires in-patient hospitalization or prolongation of existing hospitalization 
• 
Persistent or significant disability/incapacity, meaning there is a substantial disruption of the 
subject’s ability to conduct normal life functions 
• 
A congenital anomaly or birth defect 
 
Important medical events that may not result in death, be life-threatening, or require hospitalization may 
be considered serious when, based upon appropriate medical judgment, they may jeopardize the subject 
and may require medical or surgical intervention to prevent one of the outcomes listed in this 
definition.26 
 
10.1.7. Serious Ocular Adverse Events 
 
Serious ocular AEs include, but are not limited to: 
 
• 
A decrease in BCDVA of ≥ 30 total letters read (TLR) or an increase of ≥ 0.6 logMar from 
the most recent previous measurement of BCDVA 
• 
A decrease in BCDVA to light perception (LP) or worse that lasts > 1 hour 
• 
An ocular event that, in the opinion of the investigator, requires medical or surgical 
intervention to prevent permanent loss of sight 
• 
Severe intraocular inflammation; e.g., 4+ anterior chamber cell/flare or 4+ vitritis 
• 
Corneal decompensation 
• 
Retinal tear or detachment 
• 
Central or branch retinal vein occlusion 
 
10.1.8. Treatment-Emergent Adverse Event (TEAE) 
 
An AE not present prior to initiation of therapy, or an already present event that worsens either in 
intensity or frequency following initiation of therapy.  
 
10.2. Adverse Event Assessment and Documentation 
 
 
25 A life-threatening SAE is any AE or adverse reaction that places the subject, in the view of the Investigator, at 
immediate risk of death from the reaction as it occurred (i.e., it does not include a reaction that, had it occurred in a 
more severe form, might have caused death). 
26 CFR 312.23 – IND Safety Reporting, 
https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/CFRSearch.cfm?fr=312.32 
 


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All AEs that occur from the time the subject receives the first dose of study drug27 until study 
participation has been completed (e.g., TEAEs) must be documented.  Any medical condition present 
prior to the first study drug dose that remains unchanged or improves will not be recorded as an AE; 
however, a worsening of the condition after dosing with the study drug, will be considered an AE.   
 
Adverse events may be determined by evaluating the following in relation to the study subject: 
 
• 
Observed or volunteered problems 
• 
Complaints 
• 
Physical signs and symptoms  
• 
The occurrence of a medical condition during the study, which was absent at baseline 
• 
The worsening of a baseline medical condition during the study 
 
Treatment-emergent AEs, regardless of causal relationship, must be assessed by the investigator and 
recorded in the source documentation and the appropriate eCRF.  Each AE must be described as ocular or 
non-ocular along with the following information: date of onset, date of resolution, severity, frequency of 
the event (single episode, intermittent, continuous), action taken (none, medical and/or surgical), 
relationship to study drug, and seriousness criteria.  Any medication necessary for the treatment of an AE 
must be recorded on the Concomitant Medication eCRF and any surgical intervention necessary for AE 
treatment must be recorded on the Concomitant Procedure eCRF.  If more than one distinct AE occurs, 
each event should be recorded separately. 
 
Adverse events will be documented beginning at the time of onset, and documentation must continue 
until recovery is noted.  Events that are ongoing at the time of study exit must be followed until resolution 
or stabilization. 
 
10.2.1. Adverse Event Severity 
 
Adverse event severity will be assessed by the investigator using the following definitions: 
 
Mild 
Subject is aware of sign or symptom, but it is easily tolerated 
Moderate 
Subject experiences discomfort enough to cause interference with 
usual activity 
Severe 
AE is incapacitating to subject, causing inability to work or do usual 
activity 
 
10.2.2. Adverse Event Relationship to Study Drug 
 
The relationship of the AE to study drug will be assessed by the investigator using the following 
definitions: 
 
 
27 Events that occur between the Screening Visit and initial administration of study drug should be recorded as 
Medical History, and not as an AE.   
 


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Not Related 
Evidence exists that the AE has a cause other than the study 
drug (e.g. pre-existing condition or underlying disease, 
intercurrent illness, or concomitant medication) and does not 
meet any other criteria listed.  
  
Possibly Related 
A temporal relationship exists between event onset and 
administration of study drug.  Although the AE may appear 
unlikely to be related to the study drug, it cannot be ruled out 
with certainty; and/or the event cannot be readily explained by 
the patient’s clinical state or concomitant therapies.  
 
Probably Related 
A temporal relationship exists between the event onset and 
administration of study drug; it appears with some degree of 
certainty to be related based on known therapeutic and 
pharmacologic actions of the study drug.  It cannot be readily 
explained by the patient’s clinical state or concomitant 
therapies. 
 
Definitely Related 
Strong evidence exists that the study drug caused the AE.  
There is a temporal relationship between the event onset and 
administration of the study drug.  There is strong therapeutic 
and pharmacologic evidence that the event was caused by the 
study drug.  The patient’s clinical state and concomitant 
therapies have been ruled out as a cause.  
 
10.3. 
Adverse Event Reporting 
 
Any treatment-emergent AE occurring in the study must be reported on the AE eCRF.  Expedited 
reporting of an AE is required if the event is considered “serious”, regardless of relationship to the study 
drug or whether the event is expected or unexpected. 
 
 
 


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10.3.1. Expedited Adverse Event Reporting 
 
Any SAE that occurs during the study, regardless of relationship to the study drug, or whether the event is 
expected or unexpected, must be reported to the Sponsor, or the Sponsor’s designee, within 24 hours of 
the investigator’s becoming aware of the event.   
 
Reports should be made by completing the SAE eCRF.  Information provided on the SAE eCRF should 
be supplemented with hospitalization records, death certificate, clinic notes from specialists evaluating the 
subject’s condition, etc. as applicable for the event.  The urgency for reporting SAEs is 3-fold:  
 
• 
to facilitate discussion [and implementation, if necessary] by the Sponsor and the Investigator of 
appropriate follow-up measures;  
• 
To facilitate Investigator reporting of unanticipated problems involving risk to human subjects to 
the IRB/EC; and  
• 
To enable the Sponsor to fulfill the reporting requirements to the appropriate regulatory authority. 
 
 
 
 


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11. STATISTICAL METHODS 
 
The primary objective of this study is to compare pupil dilation at 35 minutes achieved with a single dose 
of MAP Dispenser-administered tropicamide-phenylephrine fixed combination to the pupil dialation 
achieved with two doses of MAP Dispenser-administered tropicamide-phenylephrine fixed combination 
administered 5 minutes apart.  
 
11.1. Effectiveness Endpoints 
 
Primary Effectiveness Endpoint:  
 
The primary effectiveness endpoint is mean change in pupil diameter at 35 minutes versus visit-specific 
baseline, as measured by digital pupillometry in highly photopic conditions.  The highly photopic 
condition will be established using a fully-charged transilluminator (muscle light) at the brightest setting.  
 
Exploratory Effectiveness Outcomes: 
 
• 
Proportion of eyes achieving pupil diameter of 6.0 mm or greater at 35 minutes 
• 
Proportion of eyes achieving pupil diameter of 7.0 mm or greater at 35 minutes 
• 
Mean change in pupil diameter at other timepoints (15, 55 minutes) 
• 
Distribution of pupil diameters at 15, 35 and 55 minutes 
• 
Proportion of successful administrations on 1st attempt 
 
These additional efficacy analyses will also be measured using digital pupillometry in highly photopic 
conditions. 
 
11.2. Sample Size 
 
Sixty (60) subjects who have provided informed consent for study participation will be enrolled and 
randomized with the goal of having 56 subjects evaluable for the primary efficacy endpoint.  
 
Sample size can be calculated using methods for cluster randomized trials28.  The sample size calculated 
assuming independence is multiplied by the design effect (𝐷𝐸= 1 + (𝑚−1)𝜌, where m is the cluster 
size and  the intraclass correlation) to account for correlation between eyes. In this situation the 
correlation between eyes in an individual is quite high, approximately 0.8 from previous studies, thus 
with two eyes per person the DE is 1.8. 
 
Assuming a standard deviation of 0.85 mm for change-from-baseline pupil diameter (from previous 
studies) and a true difference of 0.0 mm. a sample of 62 independent eyes would provide 90% power to 
show non-inferiority (NI) of one dose to two, using a NI margin of 0.5 mm and a one-sided t-test at alpha 
= 0.025. 
 
 
28 Campbell, M.J., and S.J. Walters, 2014. How to Design, Analyse and Report Cluster Randomized Trials in Medicine 
and Health Related Research. Wiley, NY. 


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Adjusting for correlation between eyes results in a required sample of 62*1.8=112 “clustered” eyes, or 56 
enrolled subjects. 
 
Allowing for approximately 5% for loss to follow-up results in an enrolled sample size of 60 subjects. 
 
11.3. Safety Outcomes 
 
A safety analysis will be conducted to compare a single dose of MAP Dispenser-administered 
tropicamide-phenylephrine fixed combination to two doses of MAP Dispenser-administered tropicamide-
phenylephrine fixed combination administered 5 minutes apart  
for the following outcomes: 
• 
Slit-lamp examination findings 
• 
Anterior segment exam by transilluminator findings 
• 
Occurrence of AEs 
• 
IOP measured at 60 minutes post-administration 
• 
Visual acuity changes 
• 
Proportion of administrations without Micro-Array Print (MAP) Dispenser touch to the eye 
 
11.4. Analysis Populations 
 
Subjects who are missing efficacy assessments at any of the time points or are otherwise unevaluable for 
efficacy considerations will not be replaced.  
 
11.4.1. Intent to Treat Population 
 
The intent to treat (ITT) population will consist of all randomized subjects who received a dose of study 
drug.   Subjects will be analyzed according to the planned sequence of treatments. 
 
11.4.2. Per Protocol Population 
 
The per protocol (PP) population will consist of all ITT subjects who completed all planned assessments 
without major protocol violations.  Subjects to be excluded from the PP population will be selected prior 
to unmasking drug assignment.  Subjects will be analyzed according to the sequence of study treatments 
received. 
 
11.4.3. Modified Per Protocol Population 
 
The modified per protocol (MPP) population is defined similarly to the PP population, except that 
classification will be made for each subject at the individual treatment visit level, rather than at the subject 
level. Unlike the PP population, subjects who only complete 1 of the 2 treatment visits may be included.  
 
The MPP will consist of all ITT subjects who completed all planned assessments without major protocol 
violations for the given treatment visit. Subjects to be excluded from the MPP population will be selected 
prior to unmasking treatment 
 


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11.4.4. Safety Population 
 
The safety population will consist of all ITT subjects.  Subjects will be analyzed according to the 
sequence of study drugs received 
 
11.5. General Statistical Considerations 
 
Primary efficacy analyses will be conducted on the PP population. The primary performance endpoint and 
exploratory outcomes will be summarized with descriptive statistics, and point estimates with 95% 
confidence intervals will be calculated for each outcome.  
 
Safety analyses will be performed using the safety population. The number of subjects reporting at least 1 
AE of a given type will be summarized. Each AE will be summarized by incidence and percentage and by 
eye. Slit lamp (including hyperemia assessment) findings at scheduled follow-up visits will be tabulated 
such that the number and percentage of subjects in each category will be summarized.  
 
All primary analyses will be performed in both eyes.  The pupil diameter measurement at each 
corresponding time point after administration will be compared between study treatments. 
 
Descriptive statistics will be used to summarize continuous outcomes (number of subjects [N], mean, 
standard deviation or standard error of the mean, median, maximum, and minimum) and categorical 
variables, i.e. race, sex, iris color (frequency and percentage) at each assessment time point.  
 
The AB/BA crossover design used in this study allows for within-eye estimates of both treatment effects 
(A and B) and carryover (AB, BA).  Thus, there is no need to include random effects for eye in the 
analysis model. However, to account for the clustering of eyes within individuals the analysis will include 
a specification of repeated measurements per person (e.g. using a REPEATED statement in SAS or an 
appropriate correlation structure in the R nlme package). 
 
This study has a “period-dependent” covariate (baseline pupil measurement) as discussed in Jones and 
Kenward29 (section 5.4.4).  We are assuming that the time between visits is sufficiently long that the 
baseline pupil measurement at each visit is unaffected by carryover. 
 
Unless explicitly stated otherwise, all statistical tests and confidence intervals will be two-sided, alpha = 
0.05. 
 
11.5.1. Analysis of Primary Effectiveness 
 
All time points for pupil diameter will be analyzed separately; however, only the 35-minute time point 
will support primary effectiveness. 
 
 
29 Jones, B, and M.G. Kenward, 2015.  Design and Analysis of Cross-Over Trials, 3rd ed.  CRC Press, London. 


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Success is defined as non-inferiority of the single-dose MAP Dispenser-administered tropicamide-
phenylephrine fixed combination compared to the two-dose MAP Dispenser-administered tropicamide-
phenylephrine fixed combination administered 5 minutes apart. 
 
Comparisons of mean pupil diameter between treatments will be based on an ACOVA model for 
longitudinal data.  The model will contain an effect due to subject x eye, period (1 or 2), direct effect of 
drug dosing, AB/BA carryover, and baseline pupil diameter.  The model will include a compound 
symmetric correlation structure between eyes.  Hypothesis tests will be based on confidence intervals for 
least squares mean (expected marginal mean, see Dean et. al., 201730) contrasts between treatments.  
 
Let 𝑀1 denote the expected marginal mean from the single-dose MAP Dispenser treatment change-from-
baseline pupil diameter and 𝑀2 denote the expected marginal mean from the two-dose MAP Dispenser 
treatment, both calculated at the observed mean baseline pupil diameter. 
 
The null and alternative hypotheses are defined as follows: 
 
𝐻0: 𝑀1 −𝑀2 ≤−0.5 𝑚𝑚
𝐻𝐴: 𝑀1 −𝑀2 > −0.5 𝑚𝑚 
 
The non-inferiority null hypothesis will be tested by constructing a 95% confidence interval for the 
contrast for the difference between 𝑀1 and 𝑀2, based on the fitted model. If the lower bound of the 95% 
confidence interval is above -0.5 mm then the null hypothesis will be rejected and the single-dose MAP 
Dispenser treatment will be declared non-inferior to the two-dose MAP Dispenser treatment. 
 
11.5.2. Additional Efficacy Analyses 
 
The proportion of eyes achieving pupil size of 6.0 mm or greater and 7.0 mm or greater at 35 minutes 
post-dose will be summarized by eye and by treatment group with descriptive statistics.  
 
For each of these proportions, a comparison of the single-dose MAP Dispenser treatment to the two-dose 
MAP Dispenser treatment will be conducted using logistic regression, with generalized estimating 
equation (GEE) methods used to account for correlation between eyes.   
 
The logistic regression model will use the same fixed effects as the primary analysis.  Comparisons 
between treatment groups will be based on a contrast between estimated marginal means for the log odds 
ratios.  Confidence intervals will be produced for odds ratios. In the event that the logistic regression fails 
to converge, the model may be simplified to obtain convergence.  In the event of complete separation (no 
responses in one or more groups), the Firth method (Firth 1993, Heinze, 200631), will be used to produce 
 
30 Dean, A., D. Voss, D. Draguljić, 2017. Design and Analysis of Experiments, Springer. 
31 Firth D. Bias reduction of maximum likelihood estimates. Biometrika 1993; 80:27-38. 
 


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estimates, with bootstrap confidence intervals.  If possible (if the fitting is numerically stable) then a 
similar model will be fit to the proportions and estimated marginal means for the difference in proportions 
will also be produced. 
 
In addition, a cumulative proportion of responders analysis (CPRA) will be created to graphically display 
the proportions of subjects with a pupil size at 35 minutes post-dose over the entire range of possible 
pupil size cut-offs (Farrar 2006), with separate lines for each treatment and eye combination.  
 
The same model used for the primary endpoint will be fitted to the mean change in pupil diameter at other 
timepoints (15, 55 minutes).  For each of these time points, estimated marginal means and associated 95% 
confidence intervals will be produced, as in the primary analysis. 
 
The distribution of pupil diameters at 15, 35, 55 minutes will be summarized for each dosing group with 
descriptive statistics, including quantiles (at q = 0.05, 0.10, 0.25, 0.50, 0.75, 0.90, 0.95).  Plots will be 
created to show change in the distribution through time.  Pupil diameters through time will also be 
graphically compared to results from the earlier MIST-1 and MIST-2 studies. 
 
The proportion of successful administrations on 1st attempt will be summarized overall and for each 
dosing group, with associated (Wilson score) 95% confidence intervals. 
 
11.5.3. Sensitivity Analyses 
 
The primary efficacy analysis described above will be rerun using the MPP population as a sensitivity 
analysis.  No further imputation of missing data will be attempted. 
 
11.5.4. Interim Analyses 
 
No interim analyses are planned for this study. 
 
11.5.5. Analysis of Baseline Data 
 
No separate analyses of baseline data are planned for this study. 
 
 
12. QUALITY ASSURANCE AND QUALITY CONTROL 
 
The Sponsor or designee will perform quality control and quality assurance checks for this study.  Before 
initiation of study enrollment, the following documents will be reviewed with the investigator and site 
study staff:  
 
 
Heinze, G. A comparative investigation of methods for logistic regression with separated or nearly 
separated data., Statistics in Medicine 2006; 25:4216-4226 
 


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• 
Study protocol 
• 
Study medication preparation procedures 
• 
Study medication administration procedures 
• 
Electronic Case Report Forms (eCRF) and procedures for completion 
• 
Informed consent process 
• 
Adverse event (AE) reporting procedures 
 
12.1. Study Monitoring 
 
During the study, the Sponsor or designee may perform periodic on-site or remote site visits. During these 
visits, information recorded on the study eCRFs will be verified against source documents to confirm data 
capture completeness, accuracy and logical consistency.  Study documents will be reviewed to confirm 
protocol compliance and adherence to EC and Sponsor-specified reporting requirements, and product 
accountability will be checked.   
 
12.2. Study Product Accountability 
 
Accurate records of each MAP Dispenser base unit and cartridge shipped must be maintained by the 
Sponsor, or designee, and study investigator. Study drug will be purchased in Mexico on-site. Study drug 
purchased and used will be documented and the dispensers received and used will be documented.  
 
The study investigator is responsible for documenting each MAP Dispenser used, the date of use, the 
identifiers for subjects who received study drug with the MAP Dispenser, and the number of mists 
administered to each subject eye.    
 
MAP Dispenser product accountability will be performed by reconciling all dispensers provided to the 
site with dispensers used for study drug administration, subjects for whom the dispenser was used, and 
dispensers returned to the Sponsor or designee. Additionally, for tropicamide-phenylephrine fixed 
combination product, each commercial bottle purchased will be reconciled with vials prepared, the 
associated dispensers used in study drug administration, subjects for whom the dispenser was used, and 
the dispenser returned to the Sponsor or designee.  
 
 


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13. ETHICAL AND REGULATORY CONSIDERATIONS 
 
This protocol was designed and will be conducted, recorded, and reported in compliance with the 
principles of Good Clinical Practice (GCP)32.  Sponsor and Investigator responsibilities associated with 
adherence to GCP are specified in APPENDIX 5:  SPONSOR AND INVESTIGATOR 
OBLIGATIONS.  The study will be performed in accordance with the recommendations guiding 
physicians in biomedical research involving human subjects adopted by the 18th World Medical 
Assembly, Helsinki, Finland, 1964 and later amendments (see APPENDIX 6:  WORLD MEDICAL 
ASSOCIATION DECLARATION OF HELSINKI). 
 
13.1. Institutional Review 
 
Before enrollment of study subjects, this protocol and the study-specific consent form must be reviewed 
and approved by an Ethics Committee (EC) operating in accordance with local regulatory requirements.  
Any changes to the study protocol or consent form must be approved by the EC prior to implementation.  
Materials for study patient recruitment and study-specific written materials provided to the subject must 
also be approved by the EC prior to use.   
 
Ongoing study progress reports will be submitted to the EC at least annually, and more frequently if 
specified by the EC.  Reports of safety events and any protocol deviations that affect the safety and 
welfare of a study subject will be submitted to the EC in accordance with EC requirements. 
 
13.2. Informed Consent 
 
The study consent form has been developed in compliance with ICH E(6).  Study consent form must be 
submitted to and approved by the EC prior to implementation.  Any EC-requested modifications to the 
consent form must remain in compliance with ICH E(6). 
 
As discussed in Section 9.1, each participant must be provided with a copy of the EC-approved consent 
form for their review, and the participants written approval(s) must be provided prior to initiation of study 
procedures.   
 
13.3. Subject Confidentiality 
 
The Investigator will maintain the confidentiality of the identity of subjects enrolled in the study and the 
information contained in the study records.  The records will be made available as required for review by 
a reviewing EC; however, to the extent possible, the subject’s identity will not be disclosed.  
 
32 Good Clinical Practice: Integrated Addendum to ICH E6(R1) Guidance for Industry, March 2018. 


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14. DATA HANDLING AND RECORD KEEPING 
 
Procedures for the handling and analysis of data will be conducted using good computing practices 
meeting ICH guidelines for the handling and analysis of data for clinical trials. 
 
14.1. Data Quality Control and Reporting 
 
After data have been entered into the study database, a system of computerized data validation checks 
will be applied and written queries pertaining to data omissions and discrepancies will be forwarded to 
study sites for resolution. Study staff will update the database as appropriate to resolve queries 
generated.  All changes to the study database will be documented. 
 
14.2. Data Archiving 
 
Archived versions of the database will be saved by the Sponsor or designee consistent with ICH GCP 
Guidelines, complying with whichever of the requirements is longer. The Sponsor will notify the 
investigator when documents should be returned. 
 
14.3. Records Retention 
 
The Investigator’s site will retain all records related to the study in compliance with ICH GCP 
Guidelines. 
 
14.4. Protocol Amendments 
 
Modifications to the approved protocol are only possible using approved protocol amendments and 
with the agreement of all responsible persons. The procedure for approval of a protocol amendment is 
identical to that for approval of the protocol. The EC must be informed of all protocol amendments and 
should be asked for its opinion as to whether a full re-evaluation of the ethical aspects of the study is 
necessary.  This should be fully documented. 
 
The investigator must not implement any deviation from or change to the protocol, without 
discussion with, and agreement by the Sponsor and prior review and documented approval/favorable 
opinion of the amendment from the relevant EC, except where it is necessary to eliminate an 
immediate hazard to study patients, or where the change(s) involves only logistical or administrative 
aspects of the study (e.g., change in monitor(s), change of telephone number(s)). 
 
Protocol amendments will be submitted to the appropriate authority(ies) as required by the 
applicable regulatory requirement(s). 
 
 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
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06 MAY 2021 
 
15. REFERENCES 
 
• 
Akman A, Aydin P. Comparison of mydriatic efficacy of spray application and drop instillation of 
tropicamide-phenylephrine fixed combination. Eye (Lond). 1999;13(5):653‒655. 
• 
Farkouh A, Frigo P, Czejka M. Systemic side effects of eye drops: a pharmacokinetic perspective. 
Clin Ophthalmol. 2016;10:2433‒2441. 
• 
Farris RL, Stuchell RN, Mandell ID. Basal and reflex human tear analysis. I Physical measurements: 
osmolarity, basal volumes, and reflex flow rate. Am Acad Ophthal 1981;88:852. 
• 
Good Clinical Practice: Integrated Addendum to ICH E6(R1) Guidance for Industry, March 2018. 
• 
Grob SR, GonzalezGonzalez LA, Daly MK. Management of mydriasis and pain in cataract and 
intraocular lens surgery: review of current medications and future directions. Clin Ophthalmol. 
2014;8:1281‒1289. doi: 10.2147/OPTH.S47569 
• 
Ianchulev T, Chayet A, Kahook M, Packer M, Pasquale L, Weinreb RN. Pharmacodynamic profile of 
mydriatic agents delivered by ocular piezo-ejection microdosing compared with conventional 
eyedropper. Ther Deliv. 2016;7(11):751-760. 
• 
Jordan A, Baum J. Basic tear flow. Does it exist? Ophthalmology. 1980 Sep;87(9):920‒
930.Lallemand F, Daull P, Benita S, Buggage R, Garrigue JS. Successfully improving ocular drug 
delivery using the cationic nanoemulsion, novasorb. J Drug Deliv. 2012;2012:604204. 
• 
Lallemand F, Daull P, Benita S, Buggage R, Garrigue JS. Successfully improving ocular drug 
delivery using the cationic nanoemulsion, novasorb. J Drug Deliv. 2012;2012:604204. 
• 
Lynch MG, Brown RH, Goode SM, Schoenwald RD, Chien DS. Reduction of phenylephrine drop 
size in infants achieves equal dilation with decreased systemic absorption. Arch Ophthalmol. 
1987;105(10):1364-5. 
• 
Madsen JT, Andersen KE. Phenylephrine is a frequent cause of periorbital allergic contact dermatitis. 
Contact Dermatitis. 2015;73(1):64-5. doi: 10.1111/cod.12394. 
• 
Mammo ZN, Flanagan JG, James DF, Trope GE.Generic versus brand-name North American topical 
glaucoma drops. Can J Ophthalmol. 2012;47(1):55‒61. doi: 10.1016/j.jcjo.2011.12.004. 
• 
Merrill D L, Goldber, B, Zavell S. bis-Tropamide, A new parasympatholytic. Curr Ther Res. 
1960;12:43‒50. 
• 
Mishima S, Gasset A, Klyce SD Jr, Baum JL. Determination of tear volume and tear flow. Invest. 
Ophthalmol. 5(3), 264‒276 (1966). 
• 
Petursson G, Cole R, Hanna C. Treatment of Glaucoma Using Minidrops of Clonidine. Arch 
Ophthalmol. 1984;102(8):1180-1181. 
• 
Ratkowsky, Evans and Alldredge, 1993. Cross-over experiments: design, analysis, and application  
• 
Scherz W, Doane MG, Dohlman CH. Tear volume in normal eyes and keratoconjunctivitis sicca. 
Albrecht Von Graefes Arch. Klin. Exp. Ophthalmol. 1974;192(2):141‒150. 
• 
Shell JW. Pharmacokinetics of topically applied ophthalmic drugs. Surv Ophthalmol. 
1982;26(4):207-18. 
• 
Urtti A, Salminen L. Minimizing systemic absorption of topically administered ophthalmic drugs. 
Surv. Ophthalmol. 37(6), 435−456 (1993). 


TROPICAMIDE-PHENYLEPHRINE FIXED COMBINATION MICRODOSE OPHTHALMIC SOLUTION  
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CONFIDENTIAL - PAGE 42 OF 60 
06 MAY 2021 
 
• 
Vaajanen A, Vapaatalo H. A single drop in the eye - Effects on the whole body? Open Ophthalmol J. 
2017;11:305‒314. doi: 10.2174/1874364101711010305 
• 
Wheatcroft S, Sharma A, McAllister J. Reduction in mydriatic drop size in premature infants. Br J 
Ophthalmol. 1993;77(6):364-5. 
 


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CONFIDENTIAL - PAGE 43 OF 60 
06 MAY 2021 
 
 
 
 
APPENDIX 1:  SCHEDULE OF MEDICATION ADMINISTRATION AND EXAMINATIONS 
Assessment/Procedure 
Screening Visit 
(Day -14 to Day -1) 
Treatment Visits 1 and 21 
Baseline0 
Time 02 
Time 1 
T0+15 min ( 2 min) 8 
Time 2 
T0+35 min ( 5 min)9 
Time 3 
T0+55 min ( 10 min) 
10 
Informed consent 
X 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
Medical history 
X 
 
 
 
 
 
Ocular history 
X 
 
 
 
 
 
Prior/concomitant medication use 
X 
 
 
 
 
 
Urine pregnancy test3 
X3 
Manifest refraction (OU) 
X 
 
 
 
 
 
BCDVA (OU)4 
X 
X 
 
 
 
 
Study drug administration sequence 
determination (OU) 
 
X (at Treatment 
Visit 1 only) 
 
 
 
 
Study drug administration (OU) 
 
 
X 
 
 
 
Slit lamp biomicroscopy (OU) 
X 
X 
 
 
 
 
Anterior segment exam by transilluminator (OU) 
 
 
 
 
 
X 
Van Herrick Angle Assessment (OU)5 
X 
 
 
 
 
 
IOP (OU)6 
X 
X 
 
 
 
 
Pupil diameter assessment (OU)7 
X 
X 
 
X 
X 
X 
Study eligibility determination 
X 
 
 
 
 
 
Dilated fundus exam (OU) 
X 
 
 
 
 
 
AE assessment 
 
X 
 
X 
X 
X 
0 Baseline refers to evaluations made at each Treatment Day prior to study medication administration. 
1 Treatment Visit 1 and 2 must be separated by at least 4 days; but may be up to 7 days apart to allow for scheduling flexibility. 
2 Time 0 starts at the point that either the 1 mist dose or first of two mists of study medication doses is administered in second eye of the study subject 
3 A urine pregnancy test will be conducted in females of childbearing potential at the Screening Visit and at the last visit (Treatment Visit 2). The test may be administered at any time during the Screening and 
Treatment Day 2 Visits.  If the subject terminates study participation prior to Treatment Visit 2, this test must be administered promptly at the time of termination. 
4 BCDVA to be measured using Snellen methods.  BCDVA to be assessed using Screening Visit manifest refraction at each Treatment Visit. 
5 Performed as part of slit lamp examination. 
6 IOP to be measured without anesthesia using the Oculus Orvis tonometer.  
7 Performed using Neuroptics pupillometer – VIP 300.   
8 The pupil diameter is the only measurement that must be within the window (  2 min)  
9 The pupil diameter is the only measurement that must be within the window (  5 min)  
10 The pupil diameter is the only measurement that must be within the window (  10 min)  


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APPENDIX 2:  STUDY MEDICATION PREPARATION PROCEDURE  
 
To maintain masking of the drug assignments for study staff performing clinical evaluations, study 
medication preparation and inventory management will be performed by a sterile transfer technician.  
The Sponsor, or designee, will provide the following to the site: 
 
MAP Components 
• 
6 x MAP Dispenser base units 
• 
12 x MAP Dispenser cartridges containing empty 2 mL vial (sterile) + loose adhesive label 
to be applied to cartridge 
• 
Accessories for sterile transfer 
• 
8 x Micro USB charging cable/connector plug 
 
 
Prior to initial MAP Dispenser use, the study drug must be aseptically transferred from the commercial 
product bottle in which it is received into the MAP Dispenser vial.  The vial is then inserted into the 
MAP Dispenser cartridge assembly, the cartridge and base are clipped together, and the seal on the vial 
containing the drug is punctured by depressing the MAP Dispenser Fill Button.  Puncture of this seal 
creates a fluid path for the drug.  Depression and release of the Fill Button approximately 5 times 
primes the system for drug delivery.  At the time of dosing, the Fill Button is again pressed and released 
to activate the piezo element to create the aerosol containing the dose.  When the Dispenser Button is 
pressed, approximately 8 µL of the ophthalmic solution is delivered horizontally through the Spray 
Nozzle to the cornea.  
 
The site will be trained and certified by the Sponsor, or designee, regarding MAP Dispenser cartridge 
assembly processes prior to their performance.   
 
MAP Dispenser cartridges and base units will be provided by Eyenovia to site personnel for use.  A 
cartridge and base unit will be assembled to be used that day and the “date used” for the MAP 
Dispenser will be added to the product preparation/use label.  At the end of the treatment day, each 
MAP Dispenser provided will be returned to storage. Please note that each MAP Dispenser must be 
used on the day of preparation; the MAP Dispenser may not be re-used on subsequent days.


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APPENDIX 3:  STUDY MEDICATION ADMINISTRATION PROCEDURE  
 
This study will be single-masked. There will be no differences in the presentation of the investigational 
drug administered and study personnel conducting ophthalmic assessments will be masked to study 
drug administration assignment.  Personnel who administer study drug on a given day will not be 
allowed to perform post-study drug administration ophthalmic assessments on that day. Site personnel 
designated to administer study medication will be trained by the Sponsor on proper use of the MAP 
Dispenser prior to study medication administration.  In order to reduce the potential for operational 
bias, study personnel who administer study medication on a given day will not be allowed to perform 
post-study drug administration ophthalmic assessments on that day.  Study drug must be administered 
in location unattended by examining staff so that examiners are not aware of which drug dose the 
subject received. 
 
For each treatment day, one MAP Dispenser base units and 2 cartridges containing the study drug to be 
administered should be provided for use.  Each MAP Dispenser will appear the same.  The cartridge 
and base unit will be assembled, then the study drug administration day will be added to the MAP 
Dispenser label and the system will be primed for use by depressing and releasing the Fill Button, then 
depressing and releasing the Dispense Button.  This sequence must be performed up to 5 times to 
complete the priming process.   
 
 
 
 
Figure 1: Eyenovia MAP Dispenser Front (L) and Back (R) View 
 
At the time of study medication administration, the following steps should be followed sequentially: 
 
1. Fill MAP Dispenser dose by depressing the Fill BUTTON once. 
2. Hold MAP Dispenser such that that the thumb and 3rd finger are positioned on each side of the 
cartridge and index finger is positioned on Dispense BUTTON at the back of the MAP 
Dispenser.  Note:  Holding MAP Dispenser with fingers around the front of the dispenser may 
affect dispenser alignment with the eye, resulting in MIST administration outside of the 
targeted corneal area.   
Dispense 
Button 
Fill Button 
MAP Dispenser 
Base Unit 
MAP Dispenser 
Cartridge 
Mist Nozzle 


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3. Begin to position the MAP Dispenser over the patient’s eye such that the MIST NOZZLE is 
facing the cornea and the DISPENSE BUTTON is facing the medication administrator.  While 
positioning, it is suggested to use the little finger of the hand holding the MAP Dispenser to 
open the lower lid and the thumb of the opposite hand to hold the upper lid open.  
4. Lower the MAP Dispenser toward the eye such that it is almost touching the patient’s brow and 
cheek.  The MAP Dispenser may be rotated slightly to avoid interference that might affect 
stability during spray administration; however, it is important that the MIST NOZZLE be 
aligned directly toward the patient’s cornea. 
5. Ask patient to first look up, then look at the blue light around the MIST NOZZLE 
6. When blue light is centered on the patient’s iris, depress, then quickly release DISPENSE 
BUTTON to administer medication.  Visually confirm medication delivery to the cornea.   
7. If medication delivery missed the cornea, withdraw MAP Dispenser from eye, then reload by 
depressing the FILL BUTTON once and repeat Steps 1-6. 
8. Repeat steps 1-7 for the contralateral eye. 
 
 
 
. 
 
 


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CONFIDENTIAL - PAGE 47 OF 60 
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APPENDIX 4:  EXAMINATION PROCEDURES 
 
All study personnel conducting ophthalmic assessments will be masked to the study drug administered.  
 
1. Visual Acuity 
 
The evaluation of VA should be performed before the subject’s pupils have been dilated.  Distance VA 
measurements should be obtained at a testing distance of 4 meters by trained personnel using Snellen 
lighted charts.  Chart background luminance should be between 80 cd/m2 and 160 cd/m2, inclusive (85 
cd/m2 is recommended).    
  
Subjects’ BCDVA will be determined using subjective manifest refraction. 
 
The subject should be seated so that the distance from the subject's eyes to the Snellen chart is 4 meters 
(157.5 inches).  The middle line of letters to approximately one-third of the distance from the top line of 
letters should be at eye level for the subject.  The subject should be asked to start reading letters at the 
top line of the chart.   Document the number of letters correctly read the line.  Move to the next line 
down if 4 or more letters are read correctly and document the number of letters correctly read on this 
line.  Continue moving down the lines (documenting the number of letters read for each line) until the 
subject reads 2 or more letters incorrectly, then stop.  Note:  If the total number of letters read (TLR) 
correctly is less than 20 letters, position the subject 1 meter from the chart and add +0.75 D sphere, 
then resume testing in the same manner.  If the subject is unable to read letters correctly at both 4.0 
meters and 1.0 meters, then the subject should be tested to determine if vision is count fingers (CF), 
hand motion (HM), light perception (LP), or no light perception (NLP).  All visual acuities will be 
expressed in standard notation for Snellen. 
 
2. Pupil Diameter Measurement 
 
Pupil diameter should be measured prior to additional study-specified examinations using the 
Neuroptics VIP 300 pupillometer; the same pupillometer should be used for each evaluation on any 
study visit day for a given subject.  The subject should be asked to focus straight ahead at a target 
placed at 3 meters to avoid accommodation.  A fully-charged transilluminator, or muscle light, will be 
shone into one eye while the pupillometer is used to measure the pupil diameter in the fellow eye.  After 
a brief rest period, the process will be repeated to measure pupil diameter in the other eye.  
 
 Pupillometry – Suggestions for Obtaining Consistent Readings 
 
As pupil diameter is the primary endpoint in this study, it is very important to have consistent 
pupillometry measurements at baseline (prior to medication administration) and at each 
measurement timepoint subsequent to medical administration.  Two factors that can affect 
pupil dilation during the measurement process include the brightness of the muscle light and 
the distance of the light from the eye.  The following steps should be taken to minimize 
variability in these factors. 
 


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06 MAY 2021 
 
 
MUSCLE LIGHT 
 
1. Make sure the muscle light is stored in the charger between subject exams in order to 
maintain full charge. 
2. Make sure the muscle light is adjusted to the brightest setting during the pupil 
measurement. 
 
PUPILLOMETER (VIP-300) 
 
1. Bring the VIP-300 Pupillometer in position on the eye to be measured. 
2. When the “circle” appears in the screen bring the fully charged muscle light into the 
visual field of the contralateral eye. 
3. The bend in the muscle light should be aligned with the handle of the pupillometer 
and shining directly in the pupil of the contralateral eye. (See Figure 1) 
4. Release the measurement button on the pupillometer to obtain the measurement. 
 
 
 
Figure 1:  Proper alignment and position of the muscle light 
 
 
4. Intraocular Pressure Measurement   
 
After completion of slit lamp biomicroscopy, IOP measurements with the Oculus Corvis Tonometer. 
Recent tonometer calibration must be documented. 
 
Two consecutive IOP measurements of each eye must be obtained. If the 2 measurements differ by 
more than 2 mmHg, a third measurement must be obtained. IOP will be analyzed as the mean of these 
2 measurements, or as the median of the 3 measurements. 


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5. Slit Lamp Biomicroscopy or Anterior segment exam by transilluminator 
 
The eyelids, conjunctiva, sclera, cornea, anterior chamber, lens, iris and anterior vitreous of the eye will 
be examined with the aid of a table-mounted binocular microscope, called a slit lamp or a 
transilluminator.  The patient will be seated during this examination and fluorescein dye should be 
instilled into the ocular cul-de-sac to facilitate the examination.  Grading will be assigned as follows: 
 
 
 


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LID 
Erythema 
None (0) 
 
Normal, without any redness, or less than mild 
Mild (+1) 
 
A low grade flushed reddish color 
Moderate (+2) 
Diffused redness encompassing entire lid margin 
Severe (+3)  
Deep diffused reddish color of lid margins and superior or inferior eyelid 
Edema 
None (0) 
 
Normal, no swelling of the lid tissue, or less than mild 
Mild (+1) 
 
Slight diffuse swelling above normal 
Moderate (+2) 
General swelling 
Severe (+3)  
Extensive swelling of the eyelid, with/without eversion of upper or lower lids 
 
CONJUNCTIVA 
Hyperemia 
None (0) 
Normal.  Appears white with a small number of conjunctival blood vessels easily 
observed 
Mild (+1) 
 
Prominent, pinkish-red color of both the bulbar and palpebral conjunctiva 
Moderate (+2) 
Bright, scarlet red color of the bulbar and palpebral conjunctiva 
Severe (+3) 
“Beefy red” with petechiae.  Dark red bulbar and palpebral conjunctival with evidence 
of subconjunctival hemorrhage 
Edema 
None (0) 
 
Normal, no swelling of the conjunctiva or less than mild 
Mild (+1) 
 
Slight diffuse or regional swelling of the conjunctiva 
Moderate (+2) 
General swelling of the conjunctiva 
Severe (+3)  
Extensive swelling of the conjunctiva 
 
SCLERA 
Normal 
Abnormal 
 
CORNEA 
Edema 
None (0) 
 
Transparent and clear, or less than mild 
Mild (+1) 
 
Dull glassy appearance 
Moderate (+2) 
Dull glassy appearance of epithelium with large number of vacuoles 
Severe (+3)  
Stromal edema, localized or diffuse, with stromal striae 
 
Staining 
None (0) 
 
No fluorescein staining of epithelium, or less than mild 
Mild (+1) 
 
Slight punctate fluorescein staining 
Moderate (+2) 
Regionally dense coalescent fluorescein staining 
Severe (+3) 
Marked fluorescein staining with immediate stromal leakage as a result of epithelial 
loss 
 
 
 


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ANTERIOR CHAMBER 
Cells 
None (0) 
 
No cells seen, or less than mild 
Mild (+1) 
 
+ cells 
Moderate (+2) 
++ cells 
Severe (+3)  
+++ cells 
Hypopyon (+4) 
++++ cells, hypopyon formation (indicate hypopyon size) 
Flare 
None (0) 
 
No Tyndall effect, or less than mild 
Mild (+1) 
 
Tyndall beam in the AC has mild intensity 
Moderate (+2) 
Tyndall beam in the AC is of strong intensity 
Severe (+3)  
Tyndall beam is very intensive.  Aqueous has a white, milky appearance 
 
IRIS 
Normal 
Abnormal 
 
LENS 
Lens Status 
Phakic* 
 
*Indicate Lens Opacity Grade: 
 
None (0) 
None present, or less than mild 
 
 
 
 
 
 
Mild (+1) 
Subtle 
 
 
 
 
 
 
Moderate (+2) 
Moderate 
 
 
 
 
 
 
Severe (+3) 
Dense 
Pseudophakic 
Aphakic 
 
ANTERIOR VITREOUS 
Normal 
Abnormal 
 
 
6. Dilated Fundus Examination 
 
Using ophthalmoscopy, the fundus will be examined and evaluated by the Investigator to be either 
normal or abnormal at the Screening Visit.   
 
 


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APPENDIX 5:  SPONSOR AND INVESTIGATOR OBLIGATIONS  
 
SPONSOR OBLIGATIONS 
 
The Sponsor is committed to: 
 
1. 
Complying with all applicable health authority regulations governing the conduct of clinical 
research studies. 
2. 
Protecting the rights, health, safety and welfare of study subjects. 
3. 
Informing clinical investigators of any new information about the study which may affect the 
health, safety or welfare of the subjects, or may influence their decision to continue 
participation in the study. 
4. 
Providing clinical investigators with the study protocol, and CRFs on which to document the 
study evaluation variables for each subject entered into the study. 
5. 
Providing the statistical analysis and study report writing resources necessary to complete 
reporting of study results. 
6. 
Ensuring equity of consideration among all investigators in multicenter studies in all matters of 
publications, meeting presentations, etc. 
7. 
Certifying that IRB/EC approval of the protocol and Investigator Agreement will be completed 
prior to treatment at an investigational site.  
 
The Sponsor shall have the right to terminate the study at any time by written notice to the investigator.  
Without limiting this right, the Sponsor will normally only terminate the study under the following 
circumstances: 
 
• 
If severe and/or SAEs associated with the study medication in human and/or animal studies 
indicate discontinuation of the study 
• 
If the Sponsor wishes to discontinue the study for commercial reasons 
• 
If the Sponsor has reasons to believe that the study cannot be satisfactorily completed due to 
insufficient patient enrollment, or an insufficient number of participating study sites identified 
within a reasonable time frame. 
 
 
 


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INVESTIGATOR OBLIGATIONS 
 
Each Investigator must be a licensed physician who has completed a residency or preceptorship in 
ophthalmology.  The Investigators have the following responsibilities: 
 
1. Subject Selection 
The Investigator is responsible for assuring that all subjects entering the study conform to the 
patient selection criteria. 
 
2. Informed Consent 
The Investigator is responsible for fully reviewing the nature of the study, the possible risks, and 
alternative treatments with prospective patients prior to their enrollment in the study.  The 
Investigator is responsible for obtaining written informed consent in compliance with ICH E(6) for 
each patient, prior to enrollment in the trial.  A copy of the signed Informed Consent Form will be 
maintained in the patient's medical record. 
 
3. Ethics Committee Approval 
The Investigator must obtain approval for participation in this protocol from the EC for the 
institution at which the procedure will be performed prior to entering any patients in the study.  The 
ICF to be used will also be submitted to the EC for approval prior to initiation of the study.  
Assurance that EC approval of the study protocol and ICF has been obtained will be provided to the 
Sponsor, or Sponsor designee, prior to initiation of the study. 
 
4. Subject Evaluations and Data Reporting 
The Investigator is responsible for complying with the requirements of the study protocol and any 
amendment or clarification as published by the Sponsor, or Sponsor designee.  Patient evaluations 
will be performed as described in the study protocol.  All information generated by the patient 
evaluation will be recorded using eCRFs with access provided by the Sponsor or Sponsor designee.     
 
Investigator(s) will not deviate from the study protocol without prior approval of the Sponsor, or 
Sponsor designee, unless the protection of health, safety or welfare of study subjects requires 
prompt action. 
 
5. Record Retention 
The Investigator shall maintain all patient records for at least 2 years after completion of the study. 
 
6. Investigational Material Accountability 
The Investigator must maintain accurate records of the receipt of all investigational material 
shipped by the Sponsor, or Sponsor designee, including the date, and identification numbers of the 
product received.  In addition, accurate records must be kept on the amount and date that 
investigational material was dispensed or returned to the Sponsor.  The Investigator must assure 
that study supplies be dispensed only to subjects enrolled in the study per study inclusion / 
exclusion criteria and under the direct supervision of the Investigator or his/her Co-Investigators. 
 


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Records of all investigational supplies received, used and returned must be kept by the Investigator.  
All unused investigational supplies must be returned to the Sponsor, or Sponsor designee, as soon 
as practical upon completion of the trial.  Investigational material accounting procedures must be 
completed before the study is considered terminated. 
 
 


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APPENDIX 6:  WORLD MEDICAL ASSOCIATION DECLARATION OF HELSINKI 
 
Ethical Principles for Medical Research Involving Human Subjects 
 
Adopted by the 18th WMA General Assembly, Helsinki, Finland, June 1964, and amended by the: 
29th WMA General Assembly, Tokyo, Japan, October 1975 
35th WMA General Assembly, Venice, Italy, October 1983 
41st WMA General Assembly, Hong Kong, September 1989 
48th WMA General Assembly, Somerset West, Republic of South Africa, October 1996 
52nd WMA General Assembly, Edinburgh, Scotland, October 2000 
53th WMA General Assembly, Washington 2002 (Note of Clarification on paragraph 29 added) 
55th WMA General Assembly, Tokyo 2004 (Note of Clarification on Paragraph 30 added) 
59th WMA General Assembly, Seoul, October 2008 
 
A.  INTRODUCTION 
 
1.  The World Medical Association (WMA) has developed the Declaration of Helsinki as a statement 
of ethical principles for medical research involving human subjects, including research on 
identifiable human material and data. The Declaration is intended to be read as a whole and each of 
its constituent paragraphs should not be applied without consideration of all other relevant 
paragraphs. 
 
2.  Although the Declaration is addressed primarily to physicians, the WMA encourages other 
participants in medical research involving human subjects to adopt these principles. 
 
3.  It is the duty of the physician to promote and safeguard the health of patients, including those who 
are involved in medical research. The physician's knowledge and conscience are dedicated to the 
fulfillment of this duty. 
 
4.  The Declaration of Geneva of the WMA binds the physician with the words, “The health of my 
patient will be my first consideration,” and the International Code of Medical Ethics declares that, 
“A physician shall act in the patient's best interest when providing medical care.” 
 
5.  Medical progress is based on research that ultimately must include studies involving human 
subjects. Populations that are underrepresented in medical research should be provided appropriate 
access to participation in research. 
 
6.  In medical research involving human subjects, the well-being of the individual research subject 
must take precedence over all other interests. 
 
7.  The primary purpose of medical research involving human subjects is to understand the causes, 
development and effects of diseases and improve preventive, diagnostic and therapeutic 
interventions (methods, procedures and treatments). Even the best current interventions must be 


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evaluated continually through research for their safety, effectiveness, efficiency, accessibility and 
quality. 
 
8.  In medical practice and in medical research, most interventions involve risks and 
 
burdens. 
 
9.  Medical research is subject to ethical standards that promote respect for all human subjects and 
protect their health and rights. Some research populations are particularly vulnerable and need 
special protection. These include those who cannot give or refuse consent for themselves and those 
who may be vulnerable to coercion or undue influence. 
 
10. Physicians should consider the ethical, legal and regulatory norms and standards for research 
involving human subjects in their own countries as well as applicable international norms and 
standards. No national or international ethical, legal or regulatory requirement should reduce or 
eliminate any of the protections for research subjects set forth in this Declaration. 
 
B.  PRINCIPLES FOR ALL MEDICAL RESEARCH 
 
11. It is the duty of physicians who participate in medical research to protect the life, health, dignity, 
integrity, right to self-determination, privacy, and confidentiality of personal information of 
research subjects. 
 
12. Medical research involving human subjects must conform to generally accepted scientific 
principles, be based on a thorough knowledge of the scientific literature, other relevant sources of 
information, and adequate laboratory and, as appropriate, animal experimentation. The welfare of 
animals used for research must be respected. 
 
13.  Appropriate caution must be exercised in the conduct of medical research that may harm the 
environment. 
 
14. The design and performance of each research study involving human subjects must be clearly 
described in a research protocol. The protocol should contain a statement of the ethical 
considerations involved and should indicate how the principles in this Declaration have been 
addressed. The protocol should include information regarding funding, institutional affiliations, 
other potential conflicts of interest, incentives for subjects and provisions for treating and/or 
compensating subjects who are harmed as a consequence of participation in the research study. The 
protocol should describe arrangements for post-study access by study subjects to interventions 
identified as beneficial in the study or access to other appropriate care or benefits. 
 
15. The research protocol must be submitted for consideration, comment, guidance and approval to a 
research ethics committee before the study begins. This committee must be independent of the 
researcher, the sponsor and any other undue influence. It must take into consideration the laws and 
regulations of the country or countries in which the research is to be performed as well as 
applicable international norms and standards but these must not be allowed to reduce or eliminate 


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any of the protections for research subjects set forth in this Declaration. The committee must have 
the right to monitor ongoing studies. The researcher must provide monitoring information to the 
committee, especially information about any serious AEs. No change to the protocol may be made 
without consideration and approval by the committee. 
 
16. Medical research involving human subjects must be conducted only by individuals with the 
appropriate scientific training and qualifications. Research on patients or healthy volunteers 
requires the supervision of a competent and appropriately qualified physician or other health care 
professional. The responsibility for the protection of research subjects must always rest with the 
physician or other health care professional and never the research subjects, even though they have 
given consent. 
 
17. Medical research involving a disadvantaged or vulnerable population or community is only justified 
if the research is responsive to the health needs and priorities of this population or community and 
if there is a reasonable likelihood that this population or community stands to benefit from the 
results of the research. 
 
18. Every medical research study involving human subjects must be preceded by careful assessment of 
predictable risks and burdens to the individuals and communities involved in the research in 
comparison with foreseeable benefits to them and to other individuals or communities affected by 
the condition under investigation. 
 
19.  Every clinical trial must be registered in a publicly accessible database before recruitment of the 
first subject. 
 
20. Physicians may not participate in a research study involving human subjects unless they are 
confident that the risks involved have been adequately assessed and can be satisfactorily managed. 
Physicians must immediately stop a study when the risks are found to outweigh the potential 
benefits or when there is conclusive proof of positive and beneficial results. 
 
21. Medical research involving human subjects may only be conducted if the importance of the 
objective outweighs the inherent risks and burdens to the research subjects. 
 
22. Participation by competent individuals as subjects in medical research must be voluntary. Although 
it may be appropriate to consult family members or community leaders, no competent individual 
may be enrolled in a research study unless he or she freely agrees. 
 
23. Every precaution must be taken to protect the privacy of research subjects and the confidentiality of 
their personal information and to minimize the impact of the study on their physical, mental and 
social integrity. 
 
24. In medical research involving competent human subjects, each potential subject must be adequately 
informed of the aims, methods, sources of funding, any possible conflicts of interest, institutional 
affiliations of the researcher, the anticipated benefits and potential risks of the study and the 


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discomfort it may entail, and any other relevant aspects of the study. The potential subject must be 
informed of the right to refuse to participate in the study or to withdraw consent to participate at 
any time without reprisal. Special attention should be given to the specific information needs of 
individual potential subjects as well as to the methods used to deliver the information. After 
ensuring that the potential subject has understood the information, the physician or another 
appropriately qualified individual must then seek the potential subject’s freely-given informed 
consent, preferably in writing. If the consent cannot be expressed in writing, the non-written 
consent must be formally documented and witnessed. 
 
25. For medical research using identifiable human material or data, physicians must normally seek 
consent for the collection, analysis, storage and/or reuse. There may be situations where consent 
would be impossible or impractical to obtain for such research or would pose a threat to the validity 
of the research. In such situations the research may be done only after consideration and approval of 
a research ethics committee. 
 
26. When seeking informed consent for participation in a research study the physician should be 
particularly cautious if the potential subject is in a dependent relationship with the physician or may 
consent under duress. In such situations the informed consent should be sought by an appropriately 
qualified individual who is completely independent of this relationship. 
 
27. For a potential research subject who is incompetent, the physician must seek informed consent from 
the legally authorized representative. These individuals must not be included in a research study 
that has no likelihood of benefit for them unless it is intended to promote the health of the 
population represented by the potential subject, the research cannot instead be performed with 
competent persons, and the research entails only minimal risk and minimal burden. 
 
28. When a potential research subject who is deemed incompetent is able to give assent to decisions 
about participation in research, the physician must seek that assent in addition to the consent of the 
legally authorized representative. The potential subject’s dissent should be respected. 
 
29. Research involving subjects who are physically or mentally incapable of giving consent, for 
example, unconscious patients, may be done only if the physical or mental condition that prevents 
giving informed consent is a necessary characteristic of the research population. In such 
circumstances the physician should seek informed consent from the legally authorized 
representative. If no such representative is available and if the research cannot be delayed, the study 
may proceed without informed consent provided that the specific reasons for involving subjects 
with a condition that renders them unable to give informed consent have been stated in the research 
protocol and the study has been approved by a research ethics committee. Consent to remain in the 
research should be obtained as soon as possible from the subject or a legally authorized 
representative. 
 
30. Authors, editors and publishers all have ethical obligations with regard to the publication of the 
results of research. Authors have a duty to make publicly available the results of their research on 
human subjects and are accountable for the completeness and accuracy of their reports. They should 


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adhere to accepted guidelines for ethical reporting. Negative and inconclusive as well as positive 
results should be published or otherwise made publicly available. Sources of funding, institutional 
affiliations and conflicts of interest should be declared in the publication. Reports of research not in 
accordance with the principles of this Declaration should not be accepted for publication. 
 
C.   ADDITIONAL PRINCIPLES FOR MEDICAL RESEARCH COMBINED WITH 
MEDICAL CARE 
 
31. The physician may combine medical research with medical care only to the extent that the research 
is justified by its potential preventive, diagnostic or therapeutic value and if the physician has good 
reason to believe that participation in the research study will not adversely affect the health of the 
patients who serve as research subjects. 
 
32. The benefits, risks, burdens and effectiveness of a new intervention must be tested against those of 
the best current proven intervention, except in the following circumstances: 
• 
The use of placebo, or no treatment, is acceptable in studies where no current proven 
intervention exists; or 
• 
Where for compelling and scientifically sound methodological reasons the use of placebo is 
necessary to determine the efficacy or safety of an intervention and the patients who receive 
placebo or no treatment will not be subject to any risk of serious or irreversible harm. Extreme 
care must be taken to avoid abuse of this option. 
 
33. At the conclusion of the study, patients entered into the study are entitled to be informed about the 
outcome of the study and to share any benefits that result from it, for example, access to 
interventions identified as beneficial in the study or to other appropriate care or benefits. 
 
34. The physician must fully inform the patient which aspects of the care are related to the research. 
The refusal of a patient to participate in a study or the patient’s decision to withdraw from the study 
must never interfere with the patient-physician relationship. 
 
35. In the treatment of a patient, where proven interventions do not exist or have been ineffective, the 
physician, after seeking expert advice, with informed consent from the patient or a legally 
authorized representative, may use an unproven intervention if in the physician's judgment it offers 
hope of saving life, re-establishing health or alleviating suffering. Where possible, this intervention 
should be made the object of research, designed to evaluate its safety and efficacy. In all cases, new 
information should be recorded and, where appropriate, made publicly available. 
 


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PROTOCOL EYN-MYD-TP-41 
VERSION B 
 
 
A SINGLE-CENTER, ASSESSOR-MASKED, ACTIVE-CONTROLLED, PHASE 4 STUDY 
EVALUATING SPEED OF PUPIL DILATION WITH THE MICRO-ARRAY PRINT (MAP) 
DISPENSER WHEN COMPARING 2 DOSING REGIMENS OF TROPICAMIDE-
PHENYLEPHRINE FIXED COMBINATION OPHTHALMIC SOLUTION  
(THE SPEED STUDY) 
 
 
SPONSOR: 
 
EYENOVIA, INC. 
295 MADISON AVENUE, SUITE 2400 
NEW YORK NY  10017 
USA 
PHONE:  917-289-1117 
 
 
I have read this protocol in its entirety.  I agree to: 
 
• 
Implement and conduct this study in compliance with this study protocol; conditions of 
approval imposed my reviewing Ethics Committee (EC); Good Clinical Practice (GCP); and 
any other applicable laws and regulations.   
• 
Maintain all study-related information supplied by Eyenovia in a confidential manner.  
 
Protocol Amendment 
 
Modification of the study protocol is prohibited without prior written agreement in the form of a 
protocol amendment. All amendments will be created by the Sponsor and must be approved 
by the reviewing EC prior to implementation at your site, except when required to mitigate immediate 
safety risks or when the changes involve only logistical or administrative revisions.  
 
 
 
 
___________________________        ___________________________          ______________                       
Print Name of Investigator 
 
       
Signature 
 
 
      Date