A Double-Masked, Active-Controlled Study of the Safety and Efficacy of Phenylephrine 2.5%-Tropicamide 1% Ophthalmic Solution Administered With a Microdose Dispenser for Dilation of the Pupil
Phase: PHASE3
Conditions: Mydriasis
Sponsor: Eyenovia Inc.

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL 
21 SEPTEMBER 2018 
 
  
 
 
  
 
 
PROTOCOL EYN-MYD-TP-31  
VERSION B 
 
 
A DOUBLE-MASKED, ACTIVE-CONTROLLED, PHASE 3 STUDY OF THE SAFETY AND 
EFFICACY OF FIXED COMBINATION PHENYLEPHRINE 2.5%-TROPICAMIDE 1% 
OPHTHALMIC SOLUTION ADMINISTERED WITH A MICRODOSE DISPENSER FOR 
DILATION OF THE PUPIL  
(THE MIST-1 STUDY) 
 
 
 
 
SPONSOR: 
 
EYENOVIA, INC. 
295 MADISON AVENUE, SUITE 2400 
NEW YORK NY  10017 
USA 
PHONE:  917-289-1117 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 2 OF 62 
21 SEPTEMBER 2018 
 
TABLE OF CONTENTS 
 
TABLE OF CONTENTS ......................................................................................................................... 2 
LIST OF ABBREVIATIONS .................................................................................................................. 5 
1. PERSONNEL AND FACILITIES ....................................................................................................... 6 
2. STUDY SYNOPSIS ............................................................................................................................. 7 
2.1. Study Objective ............................................................................................................................ 7 
2.2. Study Products ............................................................................................................................. 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 ........................................................................ 9 
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. Phenylephrine and Tropicamide Ophthalmic Microdosing Solutions for Pupil Dilation .......... 12 
3.5. Ophthalmic Microdosing ........................................................................................................... 13 
4. INVESTIGATIONAL PRODUCT DESCRIPTION ......................................................................... 16 
4.1. Findings from Non-Clinical and Clinical Studies ...................................................................... 16 
4.2. How Provided............................................................................................................................. 16 
4.3. 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 Code .................................................. 23 
9.4. Study Drug Packaging, Labeling and Storage............................................................................ 23 
9.5. Study Drug Administration ........................................................................................................ 24 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 3 OF 62 
21 SEPTEMBER 2018 
 
9.6. Study Visits and Clinical Assessments ...................................................................................... 24 
9.6.1. Screening Visit (-14 Days to -1 Day) ................................................................................. 24 
9.6.2. Treatment Visit 1 (Day 1) ................................................................................................... 25 
9.6.3. Treatment Visit 2 (Day +3 to Day +8) ................................................................................ 26 
9.6.4. Treatment Visit 3 (Day +5 to Day +15) .............................................................................. 27 
9.4.5. Unscheduled Visits ............................................................................................................. 28 
9.4.6. Missed Visits....................................................................................................................... 28 
9.7. Study Exit ................................................................................................................................... 28 
9.7.1. Subject Termination ............................................................................................................ 28 
9.7.2. Subject Completion ............................................................................................................. 28 
10.1. Adverse Event Definitions ....................................................................................................... 29 
10.1.1. Adverse Event (AE) .......................................................................................................... 29 
10.1.2. Adverse Drug Reaction (ADR) ......................................................................................... 29 
10.1.3. Unexpected Adverse Event or Unexpected Suspected Adverse Reaction ........................ 29 
10.1.4. Suspected Adverse Reaction ............................................................................................. 29 
10.1.5. Life-Threatening Adverse Event or Life-Threatening Suspected Adverse Reaction ....... 30 
10.1.6. Serious Adverse Event (SAE) or Serious Suspected Adverse Reaction ........................... 30 
10.1.7. Serious Ocular Adverse Events ........................................................................................ 30 
10.1.8. Treatment-Emergent Adverse Event (TEAE) ................................................................... 31 
10.2. Adverse Event Assessment and Documentation ...................................................................... 31 
10.2.1. Adverse Event Severity..................................................................................................... 31 
10.2.2. Adverse Event Relationship to Study Drug ...................................................................... 32 
10.3 Adverse Event Reporting .......................................................................................................... 32 
10.3.1. Expedited Adverse Event Reporting ................................................................................. 33 
11. STATISTICAL METHODS............................................................................................................. 34 
11.1. Efficacy Endpoints ................................................................................................................... 34 
11.2. Sample Size .............................................................................................................................. 34 
11.3. Safety Outcomes ...................................................................................................................... 35 
11.4. Analysis Populations ................................................................................................................ 35 
11.4.1. Intent to Treat Population ................................................................................................. 35 
11.4.2. Per Protocol Population .................................................................................................... 36 
11.4.3. Modified Per Protocol Population .................................................................................... 36 
11.4.4. Safety Population .............................................................................................................. 36 
11.5. General Statistical Considerations ........................................................................................... 36 
11.5.1. 
Analysis of Primary Efficacy ...................................................................................... 37 
11.5.2. 
Additional Efficacy Analyses ...................................................................................... 38 
11.5.3. 
Sensitivity Analyses .................................................................................................... 38 
11.5.4. Interim Analyses ............................................................................................................... 39 
11.5.5. Analysis of Baseline Data ................................................................................................. 39 
12. QUALITY ASSURANCE AND QUALITY CONTROL ............................................................... 40 
12.1. Study Monitoring ..................................................................................................................... 40 
12.2. Study Product Accountability .................................................................................................. 40 
13. ETHICAL AND REGULATORY CONSIDERATIONS ................................................................ 41 
13.1. Institutional Review ................................................................................................................. 41 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 4 OF 62 
21 SEPTEMBER 2018 
 
13.2. Informed Consent ..................................................................................................................... 41 
13.3. Subject Confidentiality ............................................................................................................. 41 
14. DATA HANDLING AND RECORD KEEPING............................................................................. 42 
14.1. Data Quality Control and Reporting ........................................................................................ 42 
14.2. Data Archiving ......................................................................................................................... 42 
14.3. Records Retention .................................................................................................................... 42 
14.4. Protocol Amendments .............................................................................................................. 42 
15. REFERENCES ................................................................................................................................. 43 
APPENDIX 1:  SCHEDULE OF MEDICATION ADMINISTRATION AND EXAMINATIONS .... 45 
APPENDIX 2:  STUDY MEDICATION PREPARATION PROCEDURE .......................................... 46 
APPENDIX 3:  STUDY MEDICATION ADMINISTRATION PROCEDURE .................................. 48 
APPENDIX 4:  EXAMINATION PROCEDURES ............................................................................... 50 
APPENDIX 5:  SPONSOR AND INVESTIGATOR OBLIGATIONS ................................................. 54 
APPENDIX 6:  WORLD MEDICAL ASSOCIATION DECLARATION OF HELSINKI .................. 57 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 5 OF 62 
21 SEPTEMBER 2018 
 
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 
EC 
Ethics committee 
eCRF 
Electronic case report form 
ETDRS 
Early Treatment Diabetic Retinopathy Study (Chart) 
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 
MiDD 
Eyenovia MicroDose Dispenser 
NLP 
No light perception 
OU 
Both eyes (in this protocol, OU does not connote simultaneous treatment/evaluation) 
PE 
Phenylephrine 
PETG 
Polyethylene terephthalate glycol 
PP 
Per-protocol 
SAE 
Serious adverse event 
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). 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
CONFIDENTIAL- PAGE 6 OF 62 
21 SEPTEMBER 2018 
 
 
PROTOCOL EYN-MYD-TP-31  
VERSION B 
 
 
A DOUBLE-MASKED, ACTIVE-CONTROLLED, PHASE 3 STUDY OF THE SAFETY AND 
EFFICACY OF FIXED COMBINATION PHENYLEPHRINE 2.5%-TROPICAMIDE 1% 
OPHTHALMIC SOLUTION ADMINISTERED WITH A MICRODOSE DISPENSER FOR 
DILATION OF THE PUPIL  
(THE MIST-1 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: 
Siddarth Rathi, MD 
Phone: 914 -714-8990 
Email:  siddarthrathi@gmail.com 
 
Clinical Study Manager: 
Greg Bennett, Sr. Director, Clinical Operations 
Phone: 650-504-2005 
Email: gbennett@eyenoviabio.com  
 
Clinical Data Manager: 
Lucinda Marinus, Director, Clinical Data Management 
Phone:  949-903-4325 
Email:  lmarinus@eyenoviabio.com 
 
Overall Program Management: 
Ginger Clasby, VP, Clinical Operations 
Phone:  714-269-8838 
Email:  gclasby@eyenoviabio.com  
 
  
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 7 OF 62 
21 SEPTEMBER 2018 
 
2. STUDY SYNOPSIS 
 
2.1. Study Objective    
 
The primary objective of this study is to evaluate the safety and efficacy of Eyenovia’s fixed combination 
of phenylephrine 2.5%-tropicamide 1% ophthalmic solution administered with Eyenovia’s microdose 
dispenser (MiDD) for dilation of the pupil as compared to the individual components. 
 
2.2. Study Products 
 
• Investigational Product, Dosage and Mode of Administration:  Fixed combination ophthalmic 
solution (phenylephrine 2.5%-tropicamide 1%) administered by Eyenovia’s MiDD. 
 
• Control Products, Dosage and Mode of Administration:  Phenylephrine 2.5% ophthalmic 
solution; Tropicamide 1% ophthalmic solution.  Each control product is administered by 
Eyenovia’s MiDD. 
 
2.3. Study Population 
 
Up to 90 volunteer participants will be enrolled at 1 – 2 study sites in the United States (US) and a 
minimum of 65 subjects will be randomized to study drug administration at Treatment Visit 1 in order to 
complete follow-up on 54 subjects.   
 
2.4. Study Design   
 
This trial is a double-masked, active-controlled, cross-over superiority study evaluating 3 study drugs 
administered by Eyenovia’s MiDD.  The drugs evaluated are: 
 
• 
Phenylephrine 2.5% ophthalmic solution,  
• 
Tropicamide 1% ophthalmic solution, and 
• 
Eyenovia’s fixed combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution   
 
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 3 treatment visits, which must occur at least 2 days, but no more than 7 days apart.  
At each treatment visit, baseline measurements will be taken, then 1 of the 3 study drugs will be 
administered to both eyes (OU) in two separate instances, approximately 5 minutes apart. Afterwards, 
efficacy and safety assessments will be performed at specific time intervals.  Study drug administration 
for each subject will be equally randomized to one of the 6 possible sequences of study drugs. 
 
This study will be double-masked.  There will be no differences in the presentation of study drug 
administered and all study personnel conducting ophthalmic assessments will be masked to study drug 
assignment.  Study drug administration will be performed by at least 3 different trained personnel during 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 8 OF 62 
21 SEPTEMBER 2018 
 
the study.  Personnel who administer study drug on a given day will not be allowed to perform post-drug 
administration ophthalmic assessments on that day. 
 
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 of any age. 
2. Female subjects must be either pre-menarche, 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.  Subjects under the age required for informed consent must have the ability to 
provide additional written assent and/or parental consent, as required by the reviewing 
Institutional Review Board (IRB). 
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 a benzodiazepine, monoamine oxidase inhibitor, tricyclic antidepressant, 
anticonvulsant, or 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. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 9 OF 62 
21 SEPTEMBER 2018 
 
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. 
 
2.7. Primary Performance Endpoint 
 
The primary performance endpoint is mean change in pupil diameter at 35 minutes from the time of first 
dose versus 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 (20, 50, 65, 80 120, and 180 minutes) 
• 
Distribution of pupil diameters at 20, 35, 50, 65, 80, 120, and 180 minutes 
• 
Time from baseline to maximal pupil dilation 
 
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 65 minutes post-administration 
• 
Visual acuity changes 
 
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, Treatment Day 2, and Treatment Day 3.  Treatment visits must occur at 
least 2 days and not more than 7 days apart.  The study visit schedule is presented in flow chart form in 
Figure 1. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 10 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
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 
1st drug administered 
Efficacy & Safety assessments 
Treatment Day 2 
2nd drug administered 
Efficacy & Safety assessments 
Treatment Day 3 
3rd drug administered 
Efficacy & Safety Assessments 
-14 Days to 1 Day prior to Treatment Day 1 
2 – 7 Days after Screening Visit 
2 – 7 Days after Treatment Day 1 
2 – 7 Days after Treatment Day 2 
Study Exit 
Fails to meet 
inclusion/exclusion criteria 
Meets inclusion/exclusion criteria 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 11 OF 62 
21 SEPTEMBER 2018 
 
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 a pupil dilation medication with a fixed combination 
formulation of phenylephrine 2.5% and tropicamide 1%, for microdose delivery.  Eyenovia’s drug 
formulations are administered using a proprietary system called the Microdose Delivery Dispenser 
(MiDD), which is 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.  
 
Eyenovia’s formulation (fixed combination phenylephrine 2.5%-tropicamide 1%) is a proprietary 
investigational product that utilizes microdosing technology to facilitate 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. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 12 OF 62 
21 SEPTEMBER 2018 
 
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.  
 
3.4. Phenylephrine and Tropicamide Ophthalmic Microdosing Solutions for Pupil Dilation 
 
Eyenovia’s MiDD 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. 
 
Phenylephrine ophthalmic solution is currently marketed with 2.5% and 10% concentrations (Paragon 
BioTeck; Akorn, Inc.) for dilation of the pupil.  Tropicamide ophthalmic solution is also approved for 
eyedrop delivery at 0.5% and 1.0% concentrations (Alcon Laboratories; Bausch & Lomb; Akorn, Inc.).  
Eyenovia’s fixed combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution formulation uses 
US Food and Drug Administration (FDA)-approved formulations of phenylephrine 2.5% and tropicamide 
1.0%.  The drug is administered in approximately 8 µL microdoses using the MiDD.   Eyedrops 
                                                 
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. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 13 OF 62 
21 SEPTEMBER 2018 
 
containing a combination of phenylephrine and tropicamide are only available outside of the US (Mydrin-
P, Japan; Phenyltrope, Canada); no similar drugs are formulated for microdosing. 
 
Ocular microdosing with the MiDD has been evaluated in humans using phenylephrine 2.5%, 
phenylephrine 10%, and tropicamide 1.0% solutions, individually.   
 
3.5. Ophthalmic Microdosing 
 
The rationale for Eyenovia’s fixed combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution 
development is the desire to optimize mydriasis 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. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 14 OF 62 
21 SEPTEMBER 2018 
 
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 MiDD is designed to deliver a finely controlled microdroplet spray of 
phenylephrine 2.5%-tropicamide 1% 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. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 15 OF 62 
21 SEPTEMBER 2018 
 
 
 
Figure 1: Microdroplet Distribution Pattern from the MiDD  
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 MiDD nozzle. 
 
 
 
 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 16 OF 62 
21 SEPTEMBER 2018 
 
4. INVESTIGATIONAL PRODUCT DESCRIPTION 
 
This study will evaluate the safety and efficacy of topical ocular microdose administration of a fixed 
combination of phenylephrine 2.5%-tropicamide 1% ophthalmic solution for mydriasis in routine 
diagnostic procedures and in conditions where short-term pupil dilation is desired. 
 
4.1. Findings from Non-Clinical and Clinical Studies  
 
Detailed information on nonclinical and clinical studies of fixed combination phenylephrine 2.5%-
tropicamide 1% ophthalmic solution is provided in the Investigator Brochure (EYN-MYD-TP-
31/32). 
4.2. How Provided 
Eyenovia’s fixed combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution is contained 
sterile in the Eyenovia MiDD, which is designed to accurately deliver precise low-volume doses of 
topical ocular pharmaceutical ingredients.  As shown in Figure 2, the MiDD is composed of a “cartridge 
assembly” (shown in white in the the image below) and the base unit (shown in gray in the image 
below).   
 
 
 
Figure 2: Eyenovia MiDD Front (L) and Back (R) View 
 
Figure 3 shows a diagram of the MiDD base unit and cartridge assembly.  The cartridge assembly is the 
primary container closure system, which contains a vial filled with sterile fixed combination 
phenylephrine 2.5% - tropicamide 1% ophthalmic solution.  At the time of initial MiDD actuation, the 
seal on the vial is punctured to allow the drug to flow to the MiDD delivery system.  The cartridge 
assembly also contains a piezo-delivery system designed to drive delivery of the drug through the Spray 
Nozzle in discrete pulses repeated in a discrete cycle of duration ≤ 100 milliseconds, which is the average 
involuntary blink response time.  
 
 
Dose Button 
Load Button 
MiDD Base Unit 
MiDD Cartridge 
Spray Nozzle 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 17 OF 62 
21 SEPTEMBER 2018 
 
 
Figure 3:  Diagram of the MiDD Base Unit (L) and Cartridge Assemblies (R) 
 
Prior to initial MiDD use, fixed combination phenylephrine 2.5%-tropicamide 1% solution is aseptically 
transferred from the 2R vial in which it is received into the MiDD vial, the MiDD 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 MiDD Load Button.  Puncture of this seal creates a fluid path for the 
drug.  Depression and release of the Load Button 5 times primes the system for drug delivery.  At the time 
of dosing, the Load Button is again pressed and released to activate the piezo element to create the aerosol 
containing the drug dose.  When the Dose Button is pressed, approximately 8 µL of the drug is delivered 
through the Spray Nozzle to the cornea.   
4.3. Route of Administration, Dosage, Regimen, Treatment Period(s) 
The study medication will be administered in accordance with the treatment assignment schedule on a 
single day in the clinic by trained clinical study staff.  Two sprays of fixed combination phenylephrine 
2.5%-tropicamide 1% ophthalmic solution will be topically administered within a 5-minute period to 
both eyes.  The concentration of the active ingredients selected for this study is based on the standard 
regimen used for mydriasis where short term pupil dilation is required. The single-use regimen and 1-
day treatment period is selected based on the medication’s intended use for in-office mydriasis.  
 
 
 
 
Load Button 
Vial 
Cartridge Assembly 
Base Unit Assembly  
 
(Electrical mechanical 
components in top diagram) 
Spray Nozzle 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 18 OF 62 
21 SEPTEMBER 2018 
 
5. STUDY OBJECTIVE 
 
The primary objective of this study is to evaluate the safety and efficacy of Eyenovia’s fixed combination 
of phenylephrine 2.5%-tropicamide 1% ophthalmic solution administered with the MiDD for dilation of 
the pupil as compared to the individual components. 
 
 
6. STUDY DESIGN 
 
This trial is a double-masked, active-controlled, cross-over superiority study evaluating 3 study drugs 
administered by Eyenovia’s MiDD.  The drugs evaluated are: 
 
• 
Phenylephrine 2.5% ophthalmic solution 
• 
Tropicamide 1% ophthalmic solution, and  
• 
Eyenovia’s fixed combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution  
 
Volunteer participants from 1 – 2 US-based investigational sites 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 3 treatment visits, which must 
occur at least 2 days, but no more than 7 days apart.  At each treatment visit, baseline measurements will 
be taken, then 1 of the 3 study drugs will be administered in both eyes in two instances approximately 5 
minutes apart, after which efficacy and safety assessments will be performed at specific time intervals.  
The study drug administration schedule for subjects will be equally randomized across 6 sequences.    
 
This study will be double-masked.  There will be no differences in the presentation of study drug 
administered and all study personnel conducting ophthalmic assessments will be masked to study drug 
assignment.  Study drug administration will be performed by at least 3 different trained personnel during 
the study. Personnel who administer study drug on a given day will not be allowed to perform post-drug 
administration ophthalmic assessments on that day. 
 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 19 OF 62 
21 SEPTEMBER 2018 
 
7. STUDY OUTCOMES 
 
7.1. Effectiveness Outcomes 
 
The primary efficacy endpoint is mean change in pupil diameter at 35 minutes from time of first drug 
dose (T0) versus 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 for each drug 
(phenylephrine 2.5%, tropicamide 1%, and Eyenovia’s fixed combination phenylephrine 2.5%-
tropicamide 1%).   
 
The study’s sample size will provide 95% power to detect a 0.5 mm difference between drugs assuming a 
standard deviation of 1.0 mm using a paired t-test with a two-sided significance level of 0.05. 
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 (20, 50, 65, 80 120, and 180 minutes) 
• 
Distribution of pupil diameters at 20, 35, 50, 65, 80, 120, and 180 minutes 
• 
Time from baseline to maximal pupil dilation  
7.2. Safety Outcomes 
 
A safety analysis will be conducted to compare fixed combination of phenylephrine 2.5%-tropicamide 1% 
µD against two control therapies (individual components) for the following outcomes: 
 
• 
Slit-lamp examination (SLE) findings 
• 
Occurrence of AEs 
• 
IOP measured at 65 minutes post-administration 
• 
Visual acuity changes 
 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 20 OF 62 
21 SEPTEMBER 2018 
 
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 3 treatment visits.  It is assumed that up to 20% of subjects (n = 10) will fail to complete the 
study visits; therefore, to have 54 subjects evaluable for the primary efficacy analysis, a minimum of 65 
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 90 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 of any age. 
2. Female subjects must be either pre-menarche, 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.  Subjects under the age required for informed consent must have the ability to 
provide additional written assent and/or parental consent, as required by the reviewing IRB. 
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 a benzodiazepine, monoamine oxidase inhibitor, tricyclic antidepressant, 
anticonvulsant, or 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. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 21 OF 62 
21 SEPTEMBER 2018 
 
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. 
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. 
 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 22 OF 62 
21 SEPTEMBER 2018 
 
9. STUDY METHODS 
 
9.1. Subject Informed Consent 
 
The IRB-approved study ICF must be used for administration of informed consent.  Dependent on IRB 
requirements for subjects under 18 years of age, an IRB-approved study assent form may be supplemental 
to the study ICF.   
 
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 (and assent, if applicable) for study 
participation must be present prior to initiation of any study-specific procedure; subjects will be 
considered enrolled in the study upon their (and/or their parent’s/guardian’s, if applicable) 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/assent 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 
IRB.   
 
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 6 sequences of drug administration 
where 1 of the 3 study medications will be administered in both eyes at each of the 3 treatment visits.  
After Treatment Visit 3, each study subject should have been treated with all 3 study medications.  The 
study drug administration schedule is equally distributed across 6 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 A, B or C) will be provided in paper format contained in a sealed envelope for each 
subject who randomized to treatment. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 23 OF 62 
21 SEPTEMBER 2018 
 
The study drug administered will be masked to the study subject, the investigator, and study staff 
administering the drug and/or performing clinical assessments.  The Sponsor (or designee) involved in 
day-to-day study management will also be masked to study drug assignments.  
 
9.3.1. Procedure for Unmasking Study Drug Assignment Code  
 
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 code 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 
 
Three drugs, each delivered in microdose form using the Eyenovia MiDD, will be evaluated in this study.  
Each of the drugs must be aseptically transferred by a licensed pharmacist affiliated with the study site 
from the container in which they are received to the sterile vial provided with the MiDD cartridge.  After 
aseptic transfer, the filled vial is re-inserted into the MiDD cartridge.   
 
To maintain masking of the study drug assignments for staff administering investigational product and/or 
performing clinical evaluations, study inventory storage and management will be performed by a 
pharmacy associated with the study site.  The Sponsor, or designee, will ship study drug kits to each site 
pharmacy.  Each kit will be labeled with the study protocol number, study drug code, product storage 
requirements, and the Caution – New Drug – Limited by Federal (US) Law to Investigational Use label.  
 
Instructions regarding study medication preparation are included in APPENDIX 2:  STUDY 
MEDICATION PREPARATION PROCEDURE.  Each designated site pharmacist 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, store at 20° 
to 25° C (68° to 77° F).  Protect product from light and excessive heat.   
 
New MiDDs must be prepared by the pharmacy for use by site personnel on each treatment day.  To 
maintain masking of site personnel, the MiDD containing each of the 3 study drugs will be labeled with 
the appropriate study randomization code, pharmacy preparation date and “use by” date, then forwarded 
to site personnel for use.  The date on which the MiDD is used will be added by site personnel.  At the 
end of the treatment day, each MiDD provided will be returned to the pharmacy for storage. Please note 
that each MiDD prepared must be used on the day of preparation; the MiDD may not be re-used on 
subsequent days.   
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 24 OF 62 
21 SEPTEMBER 2018 
 
9.5. Study Drug Administration 
 
Site personnel designated to administer study medication will be trained by the Sponsor on proper use of 
the MiDD prior to study initiation.  A single MiDD may be used for study medication administration to 
all subjects whose study drug assignment corresponds to the dispenser code 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 2 doses of study drug in each eye.  After the initial dosing of each eye, 
approximately 5 minutes should elapse before 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, Treatment Day 2 and Treatment Day 3 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 -1 Day) 
 
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 IRB-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 
• 
BCDVA25 – OU 
• 
Pupil diameter measurement – OU 
• 
Slit lamp biomicroscopy including Van Herrick angle assessment – OU 
• 
IOP measurement - OU   
• 
Determination of study eligibility 
 
                                                 
25 Pediatric subjects who are unable to perform ETDRS testing must have uncorrected visual acuity evaluated using 
an age-appropriate acuity testing method.   
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 25 OF 62 
21 SEPTEMBER 2018 
 
9.6.2. Treatment Visit 1 (Day 1) 
 
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 subject will be assigned to his/her study drug randomization schedule, and the following 
baseline measurements will be taken prior to study drug administration: 
 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
BCDVA26 – OU 
• 
Slit lamp biomicroscopy – OU 
• 
IOP measurement - OU   
 
The first study drug in the drug administration sequence will be administered OU, then administered 
again approximately 5 minutes later OU.  The 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 + 20 minutes (+/- 2 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 35 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
AE assessment 
T0 + 50 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 65 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
 IOP measurement – OU 
T0 + 80 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 120 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 180 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
BCDVA23 – OU 
• 
Slit lamp biomicroscopy – OU 
• 
AE assessment 
 
 
 
                                                 
26 Pediatric subjects who are unable to perform ETDRS testing must have uncorrected visual acuity evaluated using 
an age-appropriate acuity testing method.   

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 26 OF 62 
21 SEPTEMBER 2018 
 
9.6.3. Treatment Visit 2 (Day +3 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 
• 
Pupillary light reflex - OU 
• 
BCDVA27 – OU 
• 
Slit lamp biomicroscopy – OU 
• 
IOP measurement - OU   
 
The second study drug in the drug administration sequence will be administered OU, then administered 
again approximately 5 minutes later OU.  The 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 + 20 minutes (+/- 2 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 35 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
AE assessment 
T0 + 50 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 65 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
 IOP measurement – OU 
T0 + 80 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 120 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 180 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
BCDVA23 – OU 
• 
Slit lamp biomicroscopy – OU 
• 
AE assessment 
 
 
 
 
                                                 
27 Pediatric subjects who are unable to perform ETDRS testing must have uncorrected visual acuity evaluated using 
an age-appropriate acuity testing method.   

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 27 OF 62 
21 SEPTEMBER 2018 
 
9.6.4. Treatment Visit 3 (Day +5 to Day +15) 
 
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 
• 
Pupillary light reflex - OU 
• 
BCDVA28 – OU 
• 
Slit lamp biomicroscopy – OU 
• 
IOP measurement - OU   
 
The third study drug in the drug administration sequence will be administered OU, then administered 
again approximately 5 minutes later OU.  The completion of the first administration of medications will 
be considered T0.  The following assessments will be done at the timepoints specified below: 
 
Timepoint 
Clinical Assessment (to be performed in the specified order) 
T0 + 20 minutes (+/- 2 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 35 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
AE assessment 
T0 + 50 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 65 minutes (+/- 5 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
 IOP measurement – OU 
T0 + 80 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 120 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
T0 + 180 minutes (+/-10 minutes) 
• 
Pupil diameter measurement – OU 
• 
Pupillary light reflex - OU 
• 
BCDVA25 – OU 
• 
Slit lamp biomicroscopy – OU 
• 
AE assessment 
 
If applicable, a urine pregnancy test will be administered at this visit and, upon completion of the T0 + 
180-minute assessment, the subject will have completed the study.29 
 
 
 
                                                 
28 Pediatric subjects who are unable to perform ETDRS testing must have uncorrected visual acuity evaluated using 
an age-appropriate acuity testing method. 
29 Subjects whose pupil diameter is larger than baseline at the time of study exit will be followed until pupil size has 
returned to baseline.  

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 28 OF 62 
21 SEPTEMBER 2018 
 
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 via 
telephone, email, or regular mail to contact the subject.  If the subject does not reply to any of these 
attempts, site personnel must 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 3. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 29 OF 62 
21 SEPTEMBER 2018 
 
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 either CFR 312.32 – IND Safety Reporting or 
ICH E6 – Good Clinical Practice:  Consolidated Guidance.  Because of the differing sources for these 
definitions, some terminology; e.g., “adverse event”, “adverse reaction”, and “adverse experience”, 
differs.  For this study, these terms should be considered equivalent. 
 
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.30    
 
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.31 
 
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.32 
 
10.1.4. Suspected Adverse Reaction 
 
Any AE for which there is a reasonable possibility that the drug caused the event. For the purposes of 
Investigational New Drug (IND) safety reporting, "reasonable possibility" means there is evidence to 
                                                 
30 CFR 312.23 – IND Safety Reporting, 
https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/CFRSearch.cfm?fr=312.32 
31 ICH E6 – Good Clinical Practice:  Consolidated Guidance, Glossary, p 1. 
32 32 CFR 312.23 – IND Safety Reporting, 
https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/CFRSearch.cfm?fr=312.32 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 30 OF 62 
21 SEPTEMBER 2018 
 
suggest a causal relationship between the 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.33  
 
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.34 
 
10.1.6. Serious Adverse Event (SAE) or Serious Suspected Adverse Reaction 
 
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-threatening35  
• 
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.36  
 
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 ≥ 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 
                                                 
33 CFR 312.23 – IND Safety Reporting, 
https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/CFRSearch.cfm?fr=312.32 
34 CFR 312.23 – IND Safety Reporting, 
https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/CFRSearch.cfm?fr=312.32 
35 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). 
36 CFR 312.23 – IND Safety Reporting, 
https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/CFRSearch.cfm?fr=312.32 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 31 OF 62 
21 SEPTEMBER 2018 
 
• 
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 drug therapy, or an already present event that worsens either in intensity or 
frequency following initiation of therapy.  
 
10.2. Adverse Event Assessment and Documentation 
 
All AEs that occur from the time the subject receives the first dose of study drug37 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: 
 
                                                 
37 Events that occur between the Screening Visit and initial administration of study drug should be recorded as 
Medical History, and not as an AE.   
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 32 OF 62 
21 SEPTEMBER 2018 
 
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: 
 
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. 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 33 OF 62 
21 SEPTEMBER 2018 
 
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. 
 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 34 OF 62 
21 SEPTEMBER 2018 
 
11. STATISTICAL METHODS 
 
The primary objective of this study is to evaluate the safety and efficacy of Eyenovia’s fixed combination 
of phenylephrine 2.5%-tropicamide 1% ophthalmic solution administered with the MiDD for dilation of 
the pupil as compared to the individual components. 
 
11.1. Efficacy Endpoints 
 
Primary Efficacy Endpoint: An efficacy analysis will be conducted to compare fixed combination of 
phenylephrine 2.5%-tropicamide 1% microdose ophthalmic solution administered against its individual 
components with the following endpoint: 
 
The primary performance endpoint is mean change per visit in pupil diameter at 35 minutes versus 
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.  
 
Additional Efficacy Analyses: 
• 
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 (20, 50, 65, 80 120, and 180 minutes) 
• 
Distribution of pupil diameters at 20, 35, 50, 65, 80, 120, and 180 minutes 
• 
Time from baseline to maximal pupil dilation 
 
These additional efficacy analyses will also be measured using digital pupillometry in highly photopic 
conditions. 
 
11.2. Sample Size 
 
Up to 90 subjects who have provided informed consent for study participation will be enrolled in the 
study and a minimum of 65 subjects will be randomized to study drug administration at Treatment Visit 1 
with the goal of having 54 subjects evaluable for the primary efficacy endpoint.  
 
The study design is a digram-balanced Latin square with 3 treatments, 3 periods, 2 blocks and 6 
sequences.38  A sample of 9 replicates per sequence for a total size of 54 subjects will have 95% power to 
detect a 0.5 mm difference between A and B and A and C assuming a standard deviation of 1.0 mm using 
a paired t-test with a two-sided significance level of 0.05. 
 
 
                                                 
38 Ratkowsky, Evans and Alldredge, 1993. Cross-over experiments: design, analysis, and application 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 35 OF 62 
21 SEPTEMBER 2018 
 
Block 
Sequence 
Number 
Period 
1 
Period 
2 
Period 
3 
1 
1 
A 
B 
C 
2 
B 
C 
A 
3 
C 
A 
B 
2 
4 
A 
C 
B 
5 
B 
A 
C 
6 
C 
B 
A 
 
Power calculations are provided for additional exploratory outcomes below on an informational basis.  
 
When the sample size is 54, a two-sided 95% exact confidence interval for a single proportion will have a 
half-width of 13.9% or less.  
 
• 
For evaluation of the proportion of eyes achieving pupil size of 6.0 mm or greater at 35 minutes: if 
the observed proportion in the fixed combination treatment is 70.4% (38/54), then the exact 95% 
confidence interval for the proportion will be [56.4%, 82.0%], with a half-width of 12.8%. 
 
• 
For evaluation of the proportion of eyes achieving pupil size of 7.0 mm or greater at 35 minutes: if 
the observed proportion in the fixed combination treatment is 40.7% (22/54), then the exact 95% 
confidence interval for the proportion will be [27.6% 55.0%], with a half-width of 13.7%. 
 
11.3. Safety Outcomes 
 
A safety analysis will be conducted to compare fixed combination of phenylephrine 2.5%-tropicamide 1% 
µD against 2 control therapies (individual components) for the following outcomes: 
 
• 
Slit-lamp examination findings 
• 
Occurrence of AEs 
• 
IOP measured at 65 minutes post-administration* 
• 
Visual acuity changes* 
 
*IOP and VA line listings will be provided for pediatric subjects because of the variable testing methods 
appropriate for this sub-group  
 
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 
medication.   Subjects will be analyzed according to the planned sequence of treatments. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 36 OF 62 
21 SEPTEMBER 2018 
 
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 study drug assignment.  Subjects will be analyzed according to the sequence of 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 level, rather than at the subject 
level. Unlike the PP population, subjects who only complete 1 or 2 of the 3 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. Subjects to be excluded from the MPP population will be selected prior 
to unmasking treatment assignment. Subjects will be analyzed according to the sequence of treatments 
received. 
 
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.  Safety analyses will be performed 
using the safety analysis set.  All primary analyses will be performed using data from both eyes.  The 
pupil diameter measurement at each corresponding time point after administration will be compared 
between drugs. 
 
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 balanced crossover design used in this study allows for within-subject (i.e. within-eye) estimates of 
all study drug effects (A, B, and C) and all cross-drug first-order carryover effects (AB, AC, BA, BC, CA, 
CB).  Thus, there is no need to include random effects for subject or eye in the analysis model, we will 
use fixed effects ANOVA to analyze these data (Jones and Kenward, 2015, section 5.2) and tests will be 
based on within-eye variances.  A SAS REPEATED statement will be used to allow for correlation 
between fellow eyes. 
 
This study has both a “pre-randomization” covariate (iris color) and a “period-dependent” covariate 
(baseline pupil measurement) as discussed in Jones and Kenward (2015, section 5.4.4).  We are assuming 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 37 OF 62 
21 SEPTEMBER 2018 
 
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 Efficacy 
 
All time points for pupil diameter will be analyzed separately; however, only the 35-minute time point 
will support statistical inference.  Success is defined as the fixed combination being statistically 
significantly better than each individual component.   
 
The null and alternative hypotheses for comparison of Eyenovia’s fixed combination phenylephrine 
2.5%-tropicamide 1% ophthalmic solution with the first individual component, phenylephrine 2.5% 
ophthalmic solution, based on the primary variable, is defined as follows: 
 
• 
H0: The mean change per visit in pupil diameter at 35 minutes versus baseline, as measured by 
digital pupillometry in highly photopic conditions, is not different between Eyenovia’s fixed 
combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution and phenylephrine 2.5% 
ophthalmic solution  
• 
HA: The mean change per visit in pupil diameter at 35 minutes versus baseline, as measured by 
digital pupillometry in highly photopic conditions, is different between Eyenovia’s fixed 
combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution and phenylephrine 2.5% 
ophthalmic solution  
 
The null and alternative hypotheses for comparison of Eyenovia’s fixed combination phenylephrine 
2.5%-tropicamide 1% ophthalmic solution with the second individual component, Tropicamide 1% 
ophthalmic solution, based on the primary variable, is defined as follows: 
 
• 
H0: The mean change per visit in pupil diameter at 35 minutes versus baseline, as measured by 
digital pupillometry in highly photopic conditions, is not different between Eyenovia’s fixed 
combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution and Tropicamide 1% 
ophthalmic solution 
• 
HA: The mean change per visit in pupil diameter at 35 minutes versus baseline, as measured by 
digital pupillometry in highly photopic conditions, is different between Eyenovia’s fixed 
combination phenylephrine 2.5%-tropicamide 1% ophthalmic solution and Tropicamide 1% 
ophthalmic solution 
 
Both null hypotheses must be rejected in order to claim success for the study. 
 
Comparisons among drugs on mean pupil diameter will be based on a fixed-effects analysis of variance 
model.  The model will contain an effect due to subject, eye, subject x eye, period (1, 2, or 3), direct 
effect of drug, first order carryover, baseline pupil diameter, and iris color (dark vs light).  The SAS 
PROC MIXED procedure will be used to fit the ANOVA model.  Hypothesis tests will be based on 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 38 OF 62 
21 SEPTEMBER 2018 
 
confidence intervals produced by the SAS LSMEANS option for the pairwise differences between 
treatment. Details are provided in the statistical analysis plan. 
 
The SAS PROC MIXED procedure uses all available data to produce maximum likelihood parameter 
estimates, in such a setting the missing observations are ignorable if data are missing at random (MAR).   
 
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, pairwise comparisons of the fixed combination of phenylephrine 2.5%-
tropicamide 1% microdose ophthalmic solution administered against its individual components will be 
conducted with a generalized estimating equation (GEE) model for binomial outcome, allowing for 
correlation between eyes.   
 
The SAS PROC GENMOD procedure will be used to fit the model, using the same fixed effects as the 
primary analysis.  Once again, comparisons between treatment groups will be based on SAS LSMEANS 
for the pairwise differences.  Details are provided in the statistical analysis plan. 
 
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 endpoints expressed as proportion of responders will be compared among drugs using the Mantel-
Haenszel statistic stratified on iris color.  The Mantel-Fleiss criterion will be employed to assess the 
appropriateness of the chi-square approximation for the Mantel-Haenszel statistic. 
 
The distribution of time to maximum pupil diameter will be described by Kaplan-Meier plots.   
 
11.5.3. Sensitivity Analyses 
 
The primary efficacy analysis described above will be rerun using the MPP population as a sensitivity 
analysis.  
 
As a further sensitivity analysis, multiple imputation based on reference-based multiple imputation will be 
used to impute missing data at the 35-minute time point based on a missing not at random (MNAR) 
assumption. Since the PP population contains only subjects with non-missing values, this analysis will be 
conducted using the ITT population, which may contain subjects who have only partially available 
information. Placebo treatment will serve as reference group for this approach. Each subject will be 
included for all three treatment visits for this approach, regardless of how many treatment visits were 
actually completed.  Details are provided in the statistical analysis plan. 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 39 OF 62 
21 SEPTEMBER 2018 
 
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. 
 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 40 OF 62 
21 SEPTEMBER 2018 
 
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, site study 
staff, and pharmacy:  
 
• 
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 will perform periodic visits to study sites. 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 IRB and Sponsor-specified reporting requirements, and product 
accountability will be checked.   
 
12.2. Study Product Accountability 
 
Accurate records of each MiDD base unit and cartridge shipped and medication administered must be 
maintained by the Sponsor, or designee, and each study investigator.  Each of the 3 study medications 
will be provided to the site pharmacy in a kit labeled with the medication code.  Study kits will be 
accompanied by a shipper listing the number of kits provided and the contents of each kit.  Pharmacy 
personnel must review the shipper at the time of kit receipt and notify the Sponsor, or designee, of any 
discrepancies.  The Sponsor, or designee, will provide a study kit accountability log to the pharmacy for 
documentation of kits received, kits used for site drug administration, and kits returned or discarded.    
 
The study investigator is responsible for documenting each MiDD used, the date of use, the identifiers for 
subjects who received study medication with the MiDD, and the number of sprays administered to each 
subject eye.    
 
MiDD product accountability will be performed by reconciling all dispensers provided to the pharmacy 
with dispensers used for study drug administration, subjects for whom the dispenser was used, and 
dispensers returned to the Sponsor or designee. Additionally, for the 2 control drugs, each commercial 
bottle provided will be reconciled with vials prepared by the pharmacy, the associated dispensers used in 
study drug administration, subjects for whom the dispenser was used, and the dispenser returned to the 
Sponsor or designee.  
 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 41 OF 62 
21 SEPTEMBER 2018 
 
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)39.  Sponsor and Investigator responsibilities associated with 
adherence to GCP and CFR 312 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/assent form(s) must be 
reviewed and approved by an IRB operating in accordance with 21 CFR Part 50.  Any changes to the 
study protocol or consent forms must be approved by the IRB prior to implementation.  Materials for 
study patient recruitment and study-specific written materials provided to the subject must also be 
approved by the IRB prior to use.   
 
Ongoing study progress reports will be submitted to the IRB at least annually, and more frequently if 
specified by the IRB.  Reports of safety events and any protocol deviations that affect the safety and 
welfare of a study subject will be submitted to the IRB in accordance with FDA and IRB requirements. 
 
13.2. Informed Consent 
 
The study consent form has been developed in compliance with 21 CFR Part 50.25.  Study consent 
form(s) must be submitted to and approved by the IRB prior to implementation.  Any IRB-requested 
modifications to the consent form must remain in compliance with 21 CFR Part 50.25. 
 
As discussed in Section 9.1, each participant must be provided with a copy of the IRB-approved consent 
form(s) 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 
FDA and a reviewing IRB; however, to the extent possible, the subject’s identity will not be disclosed. 
 
 
 
                                                 
39 Good Clinical Practice: Integrated Addendum to ICH E6(R1) Guidance for Industry, March 2018. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 42 OF 62 
21 SEPTEMBER 2018 
 
14. DATA HANDLING AND RECORD KEEPING 
 
Procedures for the handling and analysis of data will be conducted using good computing practices 
meeting ICH and US FDA 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 IRB 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 IRB, 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). 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 43 OF 62 
21 SEPTEMBER 2018 
 
15. REFERENCES 
 
• 
Akman A, Aydin P. Comparison of mydriatic efficacy of spray application and drop instillation 
of tropicamide 1%. Eye (Lond). 1999;13(5):653‒655. 
• 
CFR 312.23 – IND Safety Reporting, 
https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/CFRSearch.cfm?fr=312.32 
• 
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. 
• 
Jones, B., and M.G. Kenward, 2015.  Design and analysis of cross-over trials.  CRC Press. 
• 
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. Marcel Dekker. 
• 
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. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
 
CONFIDENTIAL - PAGE 44 OF 62 
21 SEPTEMBER 2018 
 
• 
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). 
• 
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. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
CONFIDENTIAL- PAGE 45 OF 62 
21 SEPTEMBER 2018 
 
APPENDIX 1:  SCHEDULE OF MEDICATION ADMINISTRATION AND EXAMINATIONS 
 
Assessment/Procedure 
Screening 
Visit 
(Day -14 to 
Day -1) 
Treatment  
Visit 1 (Day 1)1, Visit 21 and Visit 31 
Baseline0 
Time 02 
Time 1 
T0+20 min 
(± 2 min) 
Time 2 
T0+35 min 
(± 5 min) 
Time 3 
T0+50 min 
(± 5 min) 
Time 4 
T0+65 min 
(± 5 min) 
Time 5 
T0+80 min 
(± 10 min) 
Time 6 
T0+120 min 
(± 10 min) 
Time 7 
T0+180 min 
(± 10 min) 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
 
 
 
Ocular history 
X 
 
 
 
 
 
 
 
 
 
Prior/concomitant medication use 
X 
 
 
 
 
 
 
 
 
 
Urine pregnancy test3 
X 
X3 
Manifest refraction (OU) 
X 
 
 
 
 
 
 
 
 
 
BCDVA (OU)4 
X 
X 
 
 
 
 
 
 
 
X 
Study drug administration sequence 
determination (OU) 
 
X 
 
 
 
 
 
 
 
 
Study drug administration (OU) 
 
 
X 
 
 
 
 
 
 
 
Slit lamp biomicroscopy (OU)5 
X 
X 
 
 
 
 
 
 
 
X 
Van Herrick Angle Assessment (OU)6 
X 
 
 
 
 
 
 
 
 
 
IOP (OU)7 
X 
X 
 
 
 
 
X 
 
 
 
Pupil diameter assessment (OU)8 
X 
X 
 
X 
X 
X 
X 
X 
X 
X 
Pupillary light reflex (OU) 
 
X 
 
 
X 
 
X 
 
 
X 
Study eligibility determination 
X 
 
 
 
 
 
 
 
 
 
Dilated fundus exam (OU) 
X 
 
 
 
 
 
 
 
 
 
AE assessment 
 
X 
 
 
X 
 
 
 
 
X 
0 Baseline refers to evaluations made at each Treatment Day prior to study medication administration. 
1 Treatment Visit 1, 2 and 3 must be separated by at least 2 days; but may be up to 7 days apart to allow for scheduling flexibility. 
2 Time 0 starts at the point the first of two 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 3). The test may be administered at any time during 
the Screening and Treatment Day 3 Visits.  If the subject terminates study participation prior to Treatment Visit 3, this test must be administered promptly at the time of termination. 
4 BCDVA to be measured using ETDRS methods.  For younger pediatric subjects, UCDVA may be measured using age-appropriate methods per investigator’s usual practice. 
5 For younger pediatric subjects who cannot cooperate with a traditional SLE, a portable slit lamp model may be used, if necessary. 
6 Performed as part of slit lamp examination. 
7 IOP to be measured using Goldmann Applanation tonometry. For younger pediatric subjects, IOP may be measured using age-appropriate methods per investigator’s usual practice. 
8 Performed using Neuroptics pupillometer – VIP 300.  For younger pediatric subjects for whom the pupillometer cannot be successfully used, a ruler or pupil gauge may be used. 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B  
 
 
CONFIDENTIAL- PAGE 46 OF 62 
21 SEPTEMBER 2018 
 
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 pharmacy associated with 
the study site.  The Sponsor, or designee, will ship the following 3 study drug kits to each site’s 
pharmacy: 
 
Study Kit Components 
• 2 x MiDD base units 
• 2 x MiDD cartridges containing empty 2 mL vial (sterile) + loose adhesive label to be applied to 
cartridge 
• 2 x Site transport boxes 
• 2 x 10cc syringes (sterile) 
• 2 x 21g needles (sterile) 
• 2 x 2 mL injection vials of phenylephrine 2.5%-tropicamide 1% ophthalmic solution for sterile fill 
• 1 x Micro USB charging cable/connector plug 
• 2 x MiDD base units 
• 2 x MiDD cartridges containing empty 2 mL vial (sterile) + loose adhesive label to be applied to 
cartridge 
• 2 x Site transport boxes 
• 2 x 10cc syringes (sterile) 
• 2 x 21g needles (sterile) 
• 2 x 15mL bottles of tropicamide 1% ophthalmic solution for sterile fill (commercial container) 
• 1 x Micro USB charging cable/connector plug 
• 2 x MiDD base units 
• 2 x MiDD cartridges containing empty 2 mL vial (sterile) + loose adhesive label to be applied to 
cartridge 
• 2 x Site transport boxes 
• 2 x 10cc syringes (sterile) 
• 2 x 21g needles (sterile) 
• 2 x 6cc bottles of phenylephrine 2.5% ophthalmic solution for sterile fill (commercial container) 
• 1 x Micro USB charging cable/connector plug 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 47 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
Each kit will be labeled with the following information: 
• 
Study protocol number 
• 
Study drug code (A, B, or C) 
• 
Statement that kit contains either phenylephrine 2.5%-tropicamide 1%, or phenylephrine 2.5%, or 
tropicamide 1% ophthalmic solution 
• 
Product storage requirement 
• 
Caution – New Drug – Limited by Federal (US) Law to Investigational Use 
 
The MiDD base unit and cartridge are supplied in separate boxes within each study drug kit.  The MiDD 
base unit is contained in a polyethylene terephthalate glycol (PETG) plastic tray inside the box.  The 
MiDD cartridge, which includes the empty vial into which the drug is transferred, is supplied sterile in a 
PETG plastic tray with a peel lid.   Prior to initial MiDD use, the relevant study ophthalmic solution must 
be aseptically transferred from the container in which it is received into the MiDD vial.  The vial is then 
inserted into the MiDD cartridge assembly, the cartridge and base are clipped together, and the seal on the 
vial containing the drug is punctured by depressing the MiDD Load Button.  Puncture of this seal creates 
a fluid path for the drug.  Depression and release of the Load Button 5 times primes the system for drug 
delivery.  At the time of dosing, the Load Button is again pressed and released to activate the piezo 
element to create the aerosol containing the dose.  When the Dose Button is pressed, approximately 8 µL 
of the ophthalmic solution is delivered through the Spray Nozzle to the cornea.  
 
The site study pharmacist will be trained and certified by the Sponsor, or designee, regarding aseptic 
drug transfer and MiDD cartridge assembly processes prior to their performance.   
 
New study drug MiDDs must be prepared for each treatment day.  MiDD cartridges containing each of 
the 3 study drugs and labeled with the appropriate study drug code, pharmacy preparation date and “use 
by” date will be forwarded with the accompanying base unit by the pharmacy to site personnel for use.   
 
Site personnel will assemble the cartridge and base unit for each study drug to be used that day and the 
“date used” for the MiDD will be added to the product preparation/use label by site personnel.  At the end 
of the treatment day, each MiDD provided will be returned to the pharmacy for storage. Please note that 
each MiDD prepared must be used on the day of preparation; the MiDD may not be re-used on 
subsequent days. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 48 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
APPENDIX 3:  STUDY MEDICATION ADMINISTRATION PROCEDURE  
 
This study will be double-masked so that there are no differences in presentation (Eyenovia’s fixed 
combination mydriasis agent is formulated as a multi-dose microdroplet spray and the components are 
also administrated as a microdroplet spray).  Site personnel designated to administer study medication 
will be trained by the Sponsor on proper use of the MiDD 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 the subject received. 
 
For each treatment day, one or more new MiDD base units and cartridges containing the study drug(s) to 
be administered will be provided by the pharmacy for use.  Each MiDD will appear the same except for a 
label indicating the study drug code.  Study site personnel will assemble the cartridge and base unit, then 
add the study drug administration day to the MiDD label and prime the system for use by depressing and 
releasing the Load Button, then depressing and releasing the Dose Button.  This sequence must be 
performed 5 times to complete the priming process.   
 
 
 
 
Figure 1: Eyenovia MiDD Front (L) and Back (R) View 
 
At the time of study medication administration, the following steps should be followed sequentially: 
 
1. Load MiDD dose by depressing the LOAD BUTTON once. 
2. Hold MiDD such that that the thumb and 3rd finger are positioned on each side of the cartridge 
and index finger is positioned on DOSE BUTTON at the back of the MiDD.  Note:  Holding 
MiDD with fingers around the front of the device may affect dispenser alignment with the eye, 
resulting in spray administration outside of the targeted corneal area.   
Dose Button 
Load Button 
MiDD Base Unit 
MiDD Cartridge 
Spray Nozzle 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 49 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
3. Begin to position the MiDD over the patient’s eye such that the SPRAY NOZZLE is facing the 
cornea and the DOSE BUTTON is facing the medication administrator.  While positioning, it is 
suggested to use the little finger of the hand holding the MiDD to open the lower lid and the 
thumb of the opposite hand to hold the upper lid open.  
4. Lower the MiDD toward the eye such that it is almost touching the patient’s brow and cheek.  
The MiDD may be rotated slightly to avoid interference that might affect stability during spray 
administration; however, it is important that the SPRAY NOZZLE be aligned directly toward the 
patient’s cornea. 
5. Ask patient to first look up, then look at the blue light around the SPRAY NOZZLE 
6. When blue light is centered on the patient’s iris, depress, then quickly release DOSE BUTTON to 
administer medication.  Visually confirm medication delivery to the cornea.   
7. If medication delivery missed the cornea, withdraw MiDD from eye, then reload by depressing 
the LOAD BUTTON once and repeat Steps 1-6. 
8. Repeat steps 1-7 for the contralateral eye. 
 
 
 
. 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 50 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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 a physician or trained technician 
using ETDRS lighted charts.  Chart background luminance must be between 80 cd/m2 and 160 cd/m2, 
inclusive (85 cd/m2 is recommended).    
  
Adult 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 ETDRS chart (Chart 1 for 
right eye and Chart 2 for left eye) 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 logMar units. 
 
For younger pediatric subjects, uncorrected distance visual acuity UCDVA should be measured using 
age-appropriate methods per the Investigator’s usual practice. 
 
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.   
 
For younger pediatric subjects in whom the Neuroptics pupillometer cannot be used to obtain a Time 0 
measurement, the pupil diameter may be measured using a ruler or pupil gage using the instructions that 
follow.  Only a single measurement method can be used on any study visit day for a given subject. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 51 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
a. Pupil Diameter Determination by Ruler or Pupil Gauge 
 
For younger pediatric subjects in whom the Neuroptics pupillometer cannot be used to obtain a 
Time 0 measurement, the pupil diameter may be measured using a ruler or pupil gage using the 
instructions that follow.  A ruler or pupil gauge requires special care to ensure the measurement is 
accurate. Begin by placing the ruler as close as possible alongside the eye, making certain not to 
tilt or bend the device. 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 pupil diameter in the fellow eye is measured. For optimal accuracy, take 
the measurement at the patient’s eye level and repeat each measurement 2-3 times to ensure the 
result is consistent.  The pupil diameter recorded should be either the mean (if 2 measurements 
are taken) or the median (if 3 measurements are taken) values. 
 
3. Pupillary Light Reflex 
 
Pupillary light reflex will be evaluated using a fully-charged transillumator, or muscle light.  The 
response will be recorded on a scale ranging from 0 (non-responsive) to 3 (brisk). 
 
4. Intraocular Pressure Measurement   
 
After completion of slit lamp biomicroscopy, local anesthetic will be applied to facilitate IOP 
measurements with the Goldmann Applanation 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. 
 
For pediatric subjects, IOP should be measured using age-appropriate methods per the Investigator’s 
usual practice. 
 
5. Slit Lamp Biomicroscopy 
 
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.  The patient will be 
seated during this examination and fluorescein dye should be instilled into the ocular cul-de-sac to 
facilitate the examination.  For pediatric subjects who cannot cooperate with a traditional SLE, a portable 
slit lamp model may be used according to the Investigator’s usual practice.  Grading will be assigned as 
follows: 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 52 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 53 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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.   
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 54 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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, including the US FDA. 
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. 
 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 55 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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 21 CFR 50 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. Institutional Review Board/Ethics Committee Approval 
The Investigator must obtain approval for participation in this protocol from the IRB/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 IRB/EC for approval prior to initiation of the study.  
Assurance that IRB/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 whichever of the following periods is shortest: 
a. A period of 2 years after the date on which the FDA approves the marketing of the drug for the 
purpose that was the subject of the study. 
b. A period of 5 years after the date on which the results of the study are submitted to the FDA in 
support of the marketing of the drug for the purpose that was the subject of the study. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 56 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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. 
 
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. 
 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 57 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 58 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
(methods, procedures and treatments). Even the best current interventions must be 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 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 59 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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 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. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 60 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
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 
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. 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 61 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
 
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 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. 
 

PHENYLEPHRINE 2.5%-TROPICAMIDE 1% MICRODOSE OPHTHALMIC SOLUTION  
EYENOVIA CLINICAL STUDY PROTOCOL EYN-MYD-TP-31 VERSION B 
 
 
 
 
 
 
CONFIDENTIAL- PAGE 62 OF 62 
21 SEPTEMBER 2018 
 
 
 
 
 
PROTOCOL EYN-MYD-TP-31  
VERSION B 
 
 
A DOUBLE-MASKED, ACTIVE-CONTROLLED, PHASE 3 STUDY OF THE SAFETY AND 
EFFICACY OF FIXED COMBINATION PHENYLEPHRINE 2.5%-TROPICAMIDE 1% 
OPHTHALMIC SOLUTION ADMINISTERED WITH A MICRODOSE DISPENSER FOR 
DILATION OF THE PUPIL 
(THE MIST-1 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 by FDA and my reviewing Institutional Review Board (IRB); 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 IRB 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