Clinical trial NCT05646719
Conditions: Presbyopia
Keywords: Nyxol Presbyopia Pilocarpine
Sponsor: Ocuphire Pharma, Inc.
Start date: 2022-12-22; Completion date: 2023-10-11
Enrollment: 333 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
The objectives of this study are:

To evaluate the safety and efficacy of Nyxol alone and with adjunctive low dose pilocarpine to improve distance-corrected near visual acuity (DCNVA) in subjects with presbyopia.

Detailed description:
This is a Randomized, Double-Masked, Placebo-Controlled, Multicenter, Phase 3 Study of the Safety and Efficacy of Nyxol (Phentolamine Ophthalmic Solution 0.75%) as a Single Agent and With Adjunctive Low-Dose Pilocarpine Hydrochloride Ophthalmic Solution 0.4% in Subjects With Presbyopia

Arms: arm 1: Nyxol (phentolamine ophthalmic solution) 0.75% Pilocarpine (0.4%) arm 2: Nyxol (phentolamine ophthalmic solution) 0.75% Pilocarpine vehicle arm 3: Placebo Pilocarpine (0.4%) arm 4: Placebo Pilocarpine vehicle arm 5: Nyxol (phentolamine ophthalmic solution) 0.75% arm 6: Nyxol placebo
Interventions: intervention 1: Phentolamine Opthalmic Solution 0.75% intervention 2: Placebo intervention 3: Low dose pilocarpine intervention 4: Low dose pilocarpine vehicle
Intervention descriptions: intervention 1: phentolamine ophthalmic solution 0.75% (Nyxol), a non-selective alpha-1 and alpha-2 adrenergic antagonist intervention 2: Vehicle for Phentolamine Ophthalmic Solution intervention 3: Pilocarpine hydrochloride ophthalmic solution 0.4% intervention 4: Vehicle for low dose pilocarpine

Protocol documents:
Prot_000.pdf: 
 
 
 
 
CONFIDENTIAL 
 
Clinical Study Phase 3 Protocol 
OPI-NYXP-301 
VEGA-2 
 
Randomized, Double-Masked, Placebo-Controlled, Multicenter, Phase 3 Study of 
the Safety and Efficacy of Nyxol (Phentolamine Ophthalmic Solution 0.75%) as 
a Single Agent and With Adjunctive Low-Dose Pilocarpine Hydrochloride 
Ophthalmic Solution 0.4% in Subjects With Presbyopia 
 
Ocuphire Pharma, Inc. 
37000 Grand River Avenue, Suite 120 
Farmington Hills, MI 48335 
 
 
 
 
 
Original: 
July 08, 2022  
Amendment 1: 
October 31, 2022 
Amendment 2: 
November 15, 2022 
Amendment 3: 
September 29, 2023 
 
CONFIDENTIAL 
This document contains Ocuphire Pharma, Inc. (Ocuphire) information that is confidential, a 
trade secret, and/or proprietary in nature. It is loaned to you for your confidential use on behalf 
of Ocuphire and is not to be photocopied, disclosed, or transmitted to any other person or party 
who is not covered by a Confidential Disclosure Agreement with Ocuphire. As the Principal 
Investigator, you are responsible for the safekeeping and return of this document to Ocuphire 
upon request. You will be sent updated information and/or amendments as they become 
available. 
 
 
 
 
 
 


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Clinical Protocol 
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VEGA-2 
 
 
 
 
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SPONSOR SIGNATURE & CONTACTS 
Study Title: 
Randomized, Double-Masked, Placebo-Controlled, Multicenter, 
Phase 3 Study of the Safety and Efficacy of Nyxol (Phentolamine 
Ophthalmic Solution 0.75%) as a Single Agent and With 
Adjunctive Low-Dose Pilocarpine Hydrochloride Ophthalmic 
Solution 0.4% in Subjects With Presbyopia 
Study Number: 
OPI-NYXP-301 
Original Protocol: 
July 08, 2022 
Amendment 1: 
October 31, 2022 
Amendment 2: 
November 15, 2022 
Amendment 3: 
September 29, 2023 
Person authorized to sign the protocol and protocol amendment(s) for the Sponsor, Ocuphire 
Pharma, Inc. 
Richard Rodgers, MBA 
Interim CEO 
Ocuphire Pharma, Inc. 
Tel: 
248-957-9024 (Office) 
Tel: 
612-963-0118 (Mobile) 
Email: rrodgers@ocuphire.com 
 
 
____________________________________ 
Signature  
 
____________________________________ 
Date 
 
Charles Slonim, MD 
Medical Monitor 
Oculos Development Services, LLC  
Tel. 
813-690-0255 (Mobile) 
Email: chuck.slonim@oculoscr.com 
 
 
____________________________________ 
Signature  
 
____________________________________ 
Date 


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Clinical Protocol 
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VEGA-2 
 
 
 
 
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1. SYNOPSIS 
Study Number 
OPI-NYXP-301 
Clinical Phase 
Phase 3 
Type of Study 
Randomized, double-masked, placebo-controlled, multicenter 
study of the safety and efficacy of Nyxol® (Phentolamine 
Ophthalmic Solution [POS] 0.75%) as a single agent and with 
adjunctive Pilocarpine Hydrochloride Ophthalmic Solution 0.4% 
(Low-Dose Pilocarpine, LDP) in subjects with presbyopia 
Name of Investigational 
Products 
Nyxol Eye Drops – POS 0.75%   
LDP 0.4% 
Duration of Study  
22 to 36 days 
Study Objectives 
The primary objective of this study is: 
• To evaluate the efficacy of LDP as adjunctive therapy to 
Nyxol (Nyxol + LDP) to improve distance-corrected near 
visual acuity (DCNVA) without loss of best-corrected 
distance visual acuity (BCDVA) compared to placebo + 
LDP vehicle in subjects with presbyopia  
Additional objectives of this study are: 
• To evaluate the efficacy of Nyxol + LDP to improve 
DCNVA without loss of BCDVA compared to placebo + 
LDP (LDP alone) and Nyxol + LDP vehicle (Nyxol alone) 
in subjects with presbyopia 
• To evaluate the efficacy of Nyxol as a single agent to 
improve DCNVA without loss of BCDVA compared to 
placebo in subjects with presbyopia  
• To evaluate additional efficacy measures, such as visual 
acuity (VA) and pupil diameter (PD), across multiple 
timepoints and visits for Nyxol as a single agent and 
Nyxol + LDP 
• To evaluate the effect of iris color, Baseline PD, and 
Baseline DCNVA on efficacy of Nyxol as a single agent 
and Nyxol + LDP  
• To evaluate the ocular and systemic safety of Nyxol as a 
single agent and Nyxol + LDP 
• To evaluate subject-reported outcomes of Nyxol as a 
single agent and Nyxol + LDP 


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Design 
This is a randomized, double-masked, placebo-controlled, 
multicenter, Phase 3 study in approximately 320 subjects with 
presbyopia. The study will be performed in 2 stages. 
Subjects will be randomized 1:1:1:1 to treatments for both stages 
at Visit 1 (Screening/Baseline). Note that Visit 1 
(Screening/Baseline) assessments are recommended to be 
performed on the same day; however, in instances in which 
subjects are unable to complete the full day of assessments for 
Visit 1, subjects may be scheduled to return for the completion of 
Visit 1 within 1 week of Screening. If Visit 1 is split over 
2 separate days, all visit assessments up to and including 
randomization must occur on the first day, and then the subject 
should return to complete their post-randomization assessments 
on the second day.    
Treatment 1 assignment in Stage 1 will be maintained for Stage 2. 
Stage 1 will consist of 2 treatment groups (Nyxol or placebo [ie, 
Nyxol vehicle]), with approximately 160 subjects in each group. 
Stage 2 consists of 4 treatment groups (Nyxol + LDP, Nyxol + 
LDP vehicle, placebo + LDP, and placebo + LDP vehicle), with 
approximately 80 subjects per treatment group.  
Randomization will be stratified by light/dark irides. The number 
of subjects in each irides type should be as close as possible to 
1:1, with a minimum of 40% within each stratum. The subjects 
will be recruited from approximately 30 investigational sites in 
the United States. 
Stage 1: Dosing of Treatment 1 
In Stage 1, the safety and efficacy of Nyxol as a single agent will 
be evaluated following 1 day and 7 days of daily dosing, in which 
Treatment 1 (Nyxol or placebo) is dosed daily in the evenings 
near bedtime starting with the day after completion of Visit 1, 
except for Visit 1 [Stage 1 Day 1], when the subject is dosed 
during the day at the study site, and the evening before Visit 2, 
when Treatment 1 is dosed as close to 12 hours prior to Time 0 
min at Visit 2. 
Following Visit 2 (Stage 1 Day 8), subjects will discontinue 
dosing Treatment 1 for 7 to 14 days (Washout Period). During the 
Washout Period, subjects will return for Visit 3 (Stage 1 Day 10 ± 
1 day) to assess the resolution of drug treatment effects on PD. 
During Stage 1, subjects will dose with Treatment 1 (Nyxol or 
placebo [ie, Nyxol vehicle]) in the evenings near bedtime starting 
with the day after completion of Visit 1, except for Visit 1 [Stage 
1 Day 1], when the subject is dosed during the day at the study 


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site, and the evening before Visit 2, when Treatment 1 is dosed as 
close to 12 hours prior to Time 0 min at Visit 2.  
Stage 2: Dosing of Treatments 1 and 2 
Visit 4 (Stage 2 Day 1) is the start of Stage 2. In Stage 2, the 
efficacy and safety of Nyxol + LDP will be evaluated following 
1 day and 7 days of dosing, in which Treatment 1 (Nyxol or 
placebo) is dosed daily in the evenings near bedtime, except the 
evening before Visits 4 and 5, when Treatment 1 is dosed as close 
to 12 hours prior to the scheduled time for those visits. Treatment 
2 (LDP or LDP vehicle) is dosed daily in the mornings. Treatment 
2 will be dosed at the study site at the beginning of Visits 4 and 5 
(Stage 2 Days 1 and 8, respectively).  
Other design considerations: 
Study medications (Treatments 1 and 2) will each be dosed as a 
single drop in each eye, with Treatment 1 dosed in the evenings 
near bedtime and Treatment 2 dosed in the mornings. Baseline for 
the study is defined as pre-treatment (prior to Time 0 min) at 
Visit 1 (Screening/Baseline).  
During Visit 2 (Stage 1 Day 8), Time 0 min is defined as the 
timing of the first efficacy assessments at the visit, approximately 
12 hours after dosing with Nyxol the previous night (12-hour 
post-dose timepoint). During Visits 4 and 5 (Stage 2 Days 1 and 
8, respectively), Time 0 min is defined as the time of Treatment 2 
administration.  
The study eye is defined as the eligible eye with worse baseline 
DCNVA at Visit 1 (Screening/Baseline). In the case where both 
eyes have the same Baseline DCNVA, then the eye with the better 
BCDVA will be the study eye. If both DCNVA and BCDVA are 
equivalent between eyes, the study eye will be the right eye (OD). 
The study eye and fellow eye will both be evaluated at all 
assessments. The baseline values are determined during Screening 
assessments at Visit 1. 
Stage 1: Evaluation of Nyxol single agent 
At Visit 1 (Screening/Baseline), eligible subjects will be 
randomized, and the first dose of Treatment 1 will be 
administered (note: if Visit 1 is split into 2 separate days, then 
dosing occurs on the second day). Efficacy and safety 
measurements are made before dosing with Treatment 1 (Time 0 
min) and at multiple post-dose timepoints from 30 min to 8 hours. 
Prior to discharge following completion of Visit 1 (Stage 1 Day 
1), Treatment 1 study medication for Stages 1 and 2 will be 
dispensed for evening dosing by the subject before bedtime, to 
begin the night of Stage 1 Day 2.  


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On the evening prior to Visit 2, the subject will dose Treatment 1 
study medication as close to 12 hours prior to the scheduled time 
for that visit.  
At Visit 2 (Stage 1 Day 8), efficacy and safety measurements will 
be made at 12 hours after the dose of Treatment 1 study 
medication (12-hour endpoint). After completion of these 
assessments, the subject will take no study drug for 7 to 
14 days (during the Washout Period), after which Stage 1 is 
complete. Measurements will be made on Visit 3 (Stage 1 Day 10 
± 1 day) during the Washout Period to assess the absence of 
residual treatment effects. 
Stage 2: Evaluation of Nyxol + LDP 
Subjects will restart dosing daily with Treatment 1 in the evening 
immediately prior to Visit 4 (Stage 2 Day 1) as close to 12 hours 
prior to the scheduled time for Visit 4 for the 12-hour 
assessments. At this visit, after completion of the 12-hour 
assessments for Treatment 1, subjects will be administered their 
morning dose of Treatment 2 (at Time 0 min) at the study site, 
and efficacy and safety measurements will be made at multiple 
post-dose timepoints over the next 8 hours. During Stage 2, 
subjects will administer Treatment 1 in the evenings near bedtime 
and Treatment 2 in the mornings from the evening prior to Visit 4 
(Stage 2 Day 1) for the next 7 days or until the evening before 
their scheduled Visit 5 (Stage 2 Day 8). On the evening prior to 
Visit 5, Treatment 1 should be administered as close to 12 hours 
prior to the scheduled time for Visit 5. At Visit 5, measurements 
will be made before dosing with Treatment 2 at Time 0 min and 
at multiple post-dose timepoints over 8 hours after dosing with 
Treatment 2. 
Study Drug Dispensing 
and Treatment 
Adherence 
Dispensing: 
Following the completion of Visit 1 (Stage 1 Day 1), subjects will 
be dispensed Treatment 1 study drug for dosing at home in the 
evenings and a Treatment 1 subject diary to document dosing.  
At the end of Visit 4 (Stage 2 Day 1), subjects will be dispensed 
Treatment 2 study drug for dosing at home in the mornings and a 
Treatment 2 subject diary to document dosing.  
Treatment adherence: 
All subjects must be instructed on the importance of following the 
once-daily dosing regimen for each Treatment 1 and Treatment 2 
and maintaining their dosing diaries.  
Study staff will confirm treatment adherence by counting the used 
and unused vials at each study visit and by reviewing dosing in 
the diaries. Subjects must bring their used and unused study 


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medications and subject diaries with them to Visits 2 (Stage 1 
Day 8), Visit 4 (Stage 2 Day 1), and Visit 5 (Stage 2 Day 8).  
Subject Population  
Approximately 320 randomized subjects with presbyopia  
Inclusion Criteria 
1. Males or females ≥ 40 and ≤ 64 years of age. 
2. Able to comply with all protocol-mandated procedures 
independently and to attend all scheduled office visits. 
3. Able and willing to give signed informed consent. 
4. Able to self-administer study medication throughout the study 
period. 
5. BCDVA of 0.1 LogMAR (20/25 Snellen equivalent) or better 
in each eye in photopic conditions. 
6. DCNVA of 0.4 LogMAR (20/50 Snellen equivalent) or worse 
but not > 0.7 LogMAR (20/100 Snellen equivalent) in 
photopic conditions in each eye and binocularly. 
7. For subjects who depend on reading glasses or bifocals, 
binocular best-corrected near VA is 0.1 LogMAR (20/25 
Snellen equivalent) or better. 
8. Photopic PD of ≥ 3 mm in either eye.  
Exclusion Criteria 
Ophthalmic (in either eye): 
1. Use of any topical prescription (including Vuity™) or over-
the-counter (OTC) ophthalmic medications of any kind within 
7 days of Screening until study completion, with the exception 
of lid scrubs with OTC products (eg, OCuSOFT® lid scrub, 
SteriLid®, baby shampoo, etc.) and artificial tears as specified 
in Exclusion #2 below. 
2. Use of any OTC artificial tears (preserved or unpreserved) 
during Visit days or 15 min before or after instillation of 
Treatment 1 or Treatment 2. 
3. Current use of any dry eye product such as topical ophthalmic 
therapy for dry eye (eg, generic cyclosporine, Restasis, Xiidra, 
Cequa, and Eysuvis) or intranasal dry eye product (eg, 
Tyrvaya) or other devices. 
4. Tear break-up time of < 5 seconds or corneal fluorescein 
staining Grade ≥ 2 in the inferior zone or Grade ≥ 1 in the 
central zone using the National Eye Institute scale.  
5. Clinically significant ocular disease (eg, cataract, glaucoma, 
corneal edema, uveitis, retinal degeneration, loss of visual 
field, or any macular pathology) that might interfere with the 
study as deemed by the Investigator. 


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6. Recent or current evidence of ocular infection or 
inflammation in either eye (such as current evidence of 
clinically significant blepharitis, conjunctivitis, keratitis, etc.). 
Subjects must be symptom free for at least 7 days. 
7. Any history of herpes simplex or herpes zoster keratitis. 
8. Known allergy, hypersensitivity, or contraindication to any 
component of the phentolamine, pilocarpine, or vehicle 
formulations.  
9. Prior participation in a study involving the use of Nyxol for 
the treatment of presbyopia. 
10. History of cauterization of the punctum or punctal plug 
(silicone or collagen) insertion or removal. 
11. Ocular trauma within 6 months prior to Screening.  
12. Ocular surgery or any ocular laser treatment within 6 months 
prior to Screening. 
13. Subjects with surgical monovision, multifocal or extended 
depth of focus intraocular lenses (IOLs) are excluded 
(monofocal IOLs are accepted if in place > 6 months prior to 
Screening). 
14. History of any traumatic (surgical or nonsurgical) or 
nontraumatic condition affecting the pupil or iris (eg, 
irregularly shaped pupil, neurogenic pupil disorder, iris 
atrophy, iridotomy, iridectomy, iritis, etc.). 
15. Contact lens wear on the day of any study visit and contact 
lenses must be removed for home dosing and for at least 
10 minutes following dosing. 
Systemic: 
16. Known hypersensitivity or contraindication to alpha- and/or 
beta-adrenoceptor antagonists (eg, chronic obstructive 
pulmonary disease or bronchial asthma; abnormally low blood 
pressure [BP] or heart rate [HR]; second- or third-degree heart 
blockage or congestive heart failure). 
17. Known hypersensitivity or contraindication to any systemic 
cholinergic parasympathomimetic agent. 
18. Clinically significant systemic disease (eg, uncontrolled 
diabetes, myasthenia gravis, cancer, hepatic, renal, endocrine, 
or cardiovascular disorders) that might interfere with the study 
as deemed by the judgment of the Investigator. 
19. Initiation of treatment with, or any changes to, the current 
dosage, drug, or regimen of any systemic adrenergic or 
cholinergic drugs within 7 days prior to Screening or during 


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the study; however, Flomax (tamsulosin) is specifically 
excluded. 
20. Participation in any investigational study within 30 days prior 
to Screening. 
21. Females of childbearing potential who are pregnant, nursing, 
planning a pregnancy, or not using a medically acceptable 
form of birth control. Acceptable methods include the use of 
at least one of the following: intrauterine device (IUD), 
hormonal (oral, injection, patch, implant, ring), barrier with 
spermicide (condom, diaphragm), or abstinence. A female is 
considered to be of childbearing potential unless she is 1 year 
postmenopausal or 3 months post-surgical sterilization. All 
females of childbearing potential, including those with 
post−tubal ligation, must have a negative urine pregnancy test 
result at Visit 1 (Screening/Baseline). 
22. Resting HR outside the range of 50 to 110 beats per min 
following at least a 5-min rest period in the sitting position at 
Visit 1 (Screening/Baseline). Heart rate may be repeated only 
once if outside the specified range, following another 5-min 
rest period in the sitting position. 
23. Hypertension with resting diastolic BP > 105 mmHg or 
systolic BP > 160 mmHg following at least a 5-min rest 
period in the sitting position at Visit 1 (Screening/Baseline). 
Blood pressure may be repeated only once if outside the 
specified range, following another 5-min rest period in the 
sitting position. 
Study Assessments 
BCDVA (binocular and monocular photopic; binocular and 
monocular mesopic) 
Distance-corrected intermediate visual acuity (DCIVA) 
(binocular photopic) 
DCNVA (binocular and monocular photopic and binocular 
mesopic) 
PD (photopic and mesopic) 
Biomicroscopy  
Intraocular pressure (IOP)  
Direct or indirect ophthalmoscopy without dilation  
Urine pregnancy test for females of childbearing potential  
HR and BP  
Ocular tolerability 
Subject questionnaire 
Corneal fluorescein staining (for eligibility only) 


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Number of 
Investigational Sites 
Approximately 30 sites 
Estimated Total 
Sample Size 
Approximately 320 randomized subjects 
Sample Size 
Justification 
Using α = 0.05 significance and a two-tailed test, 300 total 
subjects (150 subjects each in the Nyxol and placebo arms) who 
are evaluable for efficacy will provide greater than 80% power to 
detect a difference of 17% between treatments in percent of 
subjects with ≥ 15 letters of improvement in photopic binocular 
DCNVA and with < 5 letters of loss in photopic binocular 
BCDVA from Baseline at 12 hours post-dose at Visit 2 (Stage 1 
Day 8). This calculation is based on a 12% placebo effect and a 
29% Nyxol treatment effect. An additional 20 subjects will be 
enrolled to account for dropouts. 
Primary Efficacy 
Endpoint 
The primary efficacy endpoint is the percent of subjects with 
≥ 15 letters of improvement in photopic binocular DCNVA and 
with < 5 letters of loss in photopic binocular BCDVA at 30 min 
post-LDP/vehicle comparing Nyxol + LDP to placebo + LDP 
vehicle at Visit 5 (Stage 2 Day 8).  
This comparison will be analyzed first using a hierarchical testing 
method. 
Key Secondary Efficacy 
Endpoints 
The key secondary endpoints are: 
• The percent of subjects with ≥15 letters of improvement 
in photopic binocular DCNVA and with < 5 letters of loss 
in photopic binocular BCDVA at 30 min post-
LDP/vehicle comparing Nyxol + LDP to LDP alone at 
Visit 5 (Stage 2 Day 8) 
• The percent of subjects with ≥ 15 letters of improvement 
in photopic binocular DCNVA and with < 5 letters of loss 
in photopic binocular BCDVA at 30 min post-
LDP/vehicle comparing Nyxol + LDP to Nyxol alone at 
Visit 5 (Stage 2 Day 8) 
These comparisons will be analyzed second and third using a 
hierarchical testing method. 
Other Secondary 
Efficacy Endpoints 
Other secondary endpoints measured at various timepoints in 
Stage 1 and Stage 2 include: 
• Percentage of subjects with ≥ 15 letters of improvement in 
binocular photopic DCNVA and with < 5 letters of loss in 


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binocular photopic BCDVA from Baseline (excluding 
primary and key secondary outcome timepoints) 
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of 
improvement in photopic DCNVA from Baseline 
(monocular and binocular) 
• Change in photopic DCNVA from Baseline (monocular 
and binocular) 
• 
Percent of subjects with photopic DCNVA ≤ 20/15, 
≤ 20/20, ≤ 20/25, ≤ 20/32, ≤ 20/40, ≤ 20/50, ≤ 20/63, 
≤ 20/80, ≤ 20/100, ≤ 20/125, and ≤ 20/160  
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of 
improvement in photopic binocular DCIVA from Baseline  
• Change in photopic binocular DCIVA from Baseline  
• Percentage of subjects with loss or improvement in 
photopic and mesopic BCDVA from Baseline (monocular 
and binocular) 
• Change in photopic and mesopic BCDVA from Baseline 
(monocular and binocular) 
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of 
improvement in mesopic binocular DCNVA from 
Baseline  
• Change in mesopic binocular DCNVA from Baseline  
• Change and percent change in photopic PD from Baseline  
• Percentage of subjects with photopic PD of < 3.5, < 3.0, 
< 2.5, < 2.0, and < 1.5 mm  
• Percentage of subjects with photopic PD of 1.5 to < 2.0 
mm, 2.0 to < 2.5 mm, 2.5 to < 3.0 mm, and 3.0 to < 3.5 
mm  
• Change and percent change in mesopic PD from Baseline  
• Percentage of subjects with mesopic PD of < 3.5, < 3.0, 
< 2.5, < 2.0, and < 1.5 mm  
• Percentage of subjects with mesopic PD of 1.5 to < 2.0 
mm, 2.0 to < 2.5 mm, 2.5 to < 3.0 mm, and 3.0 to < 3.5 
mm  
• Percentage of subjects with Baseline photopic DCNVA 
≤ 20/63, ≤ 20/80, ≤ 20/100, ≤ 20/125, and ≤ 20/160, with a 


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≥ 10-letter and ≥ 15-letter improvement in photopic 
DCNVA  
• Percentage of subjects with Baseline photopic DCNVA 
from 20/50 to 20/63, 20/50 to 20/80, 20/50 to 20/100, 
20/50 to 20/125, and 20/50 to 20/160, with a ≥ 10-letter 
and ≥ 15-letter improvement in photopic DCNVA  
• Subject questionnaire responses related to change in near 
vision and satisfaction with near vision 
All efficacy endpoints will be analyzed overall and by light/dark 
irides. Separate analyses will be performed for binocular, study 
eye, and fellow eye, where applicable. 
All efficacy endpoints will also be analyzed by the Modified 
Intention-to-Treat (mITT) Population. Selected analyses will be 
performed on the Per Protocol (PP) Population as specified in the 
Statistical Analysis Plan (SAP). 
A subset of the secondary pairwise comparisons will be included 
in the hierarchical analysis as specified in the SAP. 
Safety Endpoints 
Safety measures are adverse events, ocular tolerability, IOP, 
BCDVA, biomicroscopy, ophthalmoscopy, HR and BP, and 
subject questionnaire. Urine pregnancy tests for females of 
childbearing potential will be conducted.  
Study Medications, 
Dose, and Mode of 
Administration 
Treatment 1:  
• Nyxol® Eye Drops (Phentolamine Ophthalmic Solution 
0.75%), or 
• Placebo (ie, Nyxol vehicle)  
Treatment 1 is dosed during both Stages 1 and 2 of the study and is 
taken daily in the evenings near bedtime, except during the 
Washout Period.  
Treatment 2:  
• LDP (Pilocarpine Hydrochloride Ophthalmic Solution 
0.4%), or 
• LDP vehicle 
Treatment 2 is dosed during Stage 2 of the study and is taken daily 
in the mornings.  
 
 
 


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TABLE OF CONTENTS 
CONFIDENTIAL ........................................................................................................... 1 
SPONSOR SIGNATURE & CONTACTS ............................................................................ 2 
1. 
SYNOPSIS ......................................................................................................... 3 
2. 
ABBREVIATIONS AND TERMS.....................................................................16 
3. 
INTRODUCTION..............................................................................................18 
3.1. 
Findings From Nonclinical and Clinical Studies ................................................18 
3.2. 
Design Justification ............................................................................................19 
3.3. 
Route of Administration, Dosage Regimen, and Treatment Period ......................21 
3.4. 
Compliance ........................................................................................................21 
3.5. 
Study Population ................................................................................................21 
4. 
OBJECTIVES AND PURPOSE .........................................................................23 
5. 
STUDY DESIGN ...............................................................................................23 
5.1. 
Primary And Secondary Endpoints .....................................................................25 
5.2. 
Description and Schedule of Visits and Procedures ............................................27 
5.3. 
Measures Taken to Minimize/Avoid Bias ............................................................32 
5.4. 
Study Medications ..............................................................................................32 
5.4.1. 
Study Medication Identification .........................................................................32 
5.4.2. 
Packaging and Labeling .....................................................................................34 
5.4.3. 
Storage of Study Medication and Dispensing......................................................34 
5.4.4. 
Study Medication Administration .......................................................................35 
5.4.5. 
Study Medication Accountability .......................................................................35 
5.4.5.1. 
Receipt and Disposition of Study Medication .....................................................35 
5.4.5.2. 
Return of Study Medication................................................................................35 
5.5. 
Expected Duration of Subject Participation ........................................................35 
5.6. 
Randomization and Procedure for Breaking the Code ........................................36 
5.7. 
Collection of Data ..............................................................................................36 
5.8. 
Completed Subject ..............................................................................................36 
5.9. 
Non-completing Subject......................................................................................36 
5.9.1. 
Study Medication Discontinuation ......................................................................36 
5.9.2. 
Reasons for Withdrawal From Study ..................................................................37 
5.9.3. 
Entire Study Terminated .....................................................................................37 
5.9.4. 
Actions After Discontinuation ............................................................................37 
5.10. 
Completed Study ................................................................................................37 
5.11. 
Procedure After the Completion of the Study ......................................................37 
6. 
SUBJECT INCLUSION AND EXCLUSION CRITERIA ..................................38 
6.1. 
Subject Inclusion Criteria ...................................................................................38 


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6.2. 
Subject Exclusion Criteria ..................................................................................38 
7. 
TREATMENT OF SUBJECTS ..........................................................................40 
7.1. 
Treatment Adherence .........................................................................................40 
7.2. 
Concomitant Medications ...................................................................................40 
8. 
ASSESSMENT OF EFFICACY .........................................................................41 
8.1. 
Specification of the Efficacy Parameters ............................................................41 
8.2. 
Assessing, Recording, and Analyzing of Efficacy Parameters .............................41 
8.2.1. 
Screening/Baseline Visit 1 (Stage 1 Day 1) ........................................................41 
8.2.2. 
Visit 2 (Stage 1 Day 8) .......................................................................................43 
8.2.3. 
Visit 3 (Stage 1 Day 10 ± 1 day) – During Washout Period ................................43 
8.2.4. 
Visit 4 (Stage 2 Day 1 [Day 15-22]) ...................................................................43 
8.2.5. 
Visit 5 (Stage 2 Day 8 [Day 22-29]) ...................................................................44 
8.2.6. 
Unscheduled Visits .............................................................................................44 
8.3. 
Visit Variation ....................................................................................................45 
9. 
ASSESSMENT OF SAFETY .............................................................................45 
9.1. 
Specification of Safety Parameters .....................................................................45 
9.2. 
Assessing, Recording, and Analyzing Safety Parameters ....................................45 
9.3. 
Adverse Events and Serious Adverse Events .......................................................45 
9.3.1. 
Adverse Event Definitions ..................................................................................46 
9.3.2. 
Follow-Up of Subjects After Adverse Events .....................................................50 
10. 
STATISTICS .....................................................................................................50 
10.1. 
Sample Size ........................................................................................................50 
10.2. 
Analysis Populations ..........................................................................................50 
10.3. 
Statistical Methods .............................................................................................51 
10.3.1. 
General Considerations ......................................................................................51 
10.3.2. 
Demographic and Baseline Characteristics .........................................................51 
10.3.3. 
Subject Disposition ............................................................................................51 
10.3.4. 
Medical History and Prior/Concomitant Medications .........................................51 
10.3.5. 
Analysis of Efficacy ...........................................................................................51 
10.3.6. 
Analysis of Safety ..............................................................................................53 
10.4. 
Procedure for Accounting for Missing, Unused, or Spurious Data ......................53 
10.5. 
Procedure for Reporting Deviations From the Statistical Plan ...........................53 
11. 
DIRECT ACCESS TO SOURCE DATA AND DOCUMENTS .........................53 
12. 
QUALITY CONTROL AND QUALITY ASSURANCE ...................................53 
13. 
ETHICAL CONSIDERATIONS AND GOOD CLINICAL PRACTICE 
COMPLIANCE ..................................................................................................54 
13.1. 
Good Clinical Practice Compliance ...................................................................54 
13.2. 
Institutional Review Board .................................................................................54 


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13.3. 
Protocol Deviations/Violations ...........................................................................54 
13.4. 
Informed Consent Requirements .........................................................................55 
14. 
DATA HANDLING AND RECORD KEEPING ...............................................55 
14.1. 
Data Entry .........................................................................................................55 
14.2. 
Data Quality Control and Reporting ..................................................................55 
14.3. 
Archiving of Data ...............................................................................................55 
14.4. 
Records Retention ..............................................................................................56 
14.5. 
Amendments to the Protocol ...............................................................................56 
15. 
REFERENCES ..................................................................................................57 
16. 
SUMMARY OF CHANGES ..............................................................................58 
16.1. 
Protocol OPI-NYXP-301 Amendment 1 ..............................................................58 
16.2. 
Protocol OPI-NYXP-301 Amendment 2 ..............................................................62 
APPENDIX 1: IRIS COLOR CHART .............................................................................65 
APPENDIX 2: VISUAL ACUITY CHARTS ...................................................................68 
APPENDIX 3: ADRENERGIC AND CHOLINERGIC DRUGS ....................................69 
APPENDIX 4: STUDY DESIGN ......................................................................................71 
APPENDIX 5: SUBJECT QUESTIONNAIRE FOR BASELINE VISIT .......................72 
APPENDIX 6: SUBJECT QUESTIONNAIRE FOR VISITS 2, 4, AND 5 .....................74 
 
 
 
 
 
 
 


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2. ABBREVIATIONS AND TERMS 
Abbreviation 
Full term 
AE 
adverse event 
ANCOVA 
analysis of covariance 
ARP 
All Randomized Population 
BCDVA 
best-corrected distance visual acuity  
BP 
blood pressure 
cGMP 
current Good Manufacturing Practice 
CI 
confidence interval 
CRA 
clinical research associate 
CRO 
contract research organization 
DCNVA 
distance-corrected near visual acuity 
eCRF 
electronic Case Report Form 
ETDRS 
Early Treatment Diabetic Retinopathy Study 
FDA 
Food and Drug Administration 
GCP 
Good Clinical Practice 
HR 
heart rate 
IB 
Investigators’ Brochure 
ICH 
International Council for Harmonisation 
IOL 
intraocular lens  
IOP 
intraocular pressure 
IRB 
Institutional Review Board 
ITT 
Intention-to-Treat 
IUD 
intrauterine device 
LDP 
Low-Dose Pilocarpine Hydrochloride Ophthalmic Solution 0.4%; Low-Dose 
Pilocarpine 
LDPE 
low-density polyethylene 
LSM 
least-squares mean 
MedDRA 
Medical Dictionary for Regulatory Activities 


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mITT 
Modified Intention-to-Treat 
Nyxol 
Phentolamine Ophthalmic Solution 0.75%  
Nyxol + LDP 
Nyxol dosed in the evenings with LDP dosed during the mornings 
OD 
oculus dexter (right eye) 
OR 
odds ratio 
OS 
oculus sinister (left eye) 
OTC 
over-the-counter 
OU 
oculus uterque (both eyes) 
PCH 
pilocarpine hydrochloride 
PD 
pupil diameter 
POS 
Phentolamine Ophthalmic Solution 
PP 
Per Protocol  
PPS1 
PP Protocol Population Stage 1 
PPS2 
PP Population Stage 2 
SAE 
serious adverse event 
SAP 
Statistical Analysis Plan 
SOC 
system organ class 
SP 
Safety Population 
TEAE 
treatment-emergent adverse event 
US 
United States 
USP 
United States Pharmacopeia 
VA 
visual acuity 
 
 
 
 


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3. INTRODUCTION 
In this study, the efficacy of Nyxol (Phentolamine Ophthalmic Solution [POS] 0.75%) as a single 
agent and the combination of Nyxol and Pilocarpine Hydrochloride Ophthalmic Solution 0.4% 
(Low-Dose Pilocarpine, LDP) in improving distance-corrected near visual acuity (DCNVA) will be 
evaluated in presbyopic subjects.  
3.1. Findings From Nonclinical and Clinical Studies 
Nyxol 
Detailed findings from nonclinical and clinical studies for phentolamine mesylate or phentolamine 
ophthalmic solutions and potential risks are provided in the Investigators’ Brochure (IB) (2022).  
Nyxol has been studied in a total of 12 clinical trials (three Phase 1, five Phase 2, and four Phase 3) 
enrolling over 1100 subjects (including subjects 3 years of age and up, with over 650 
subjects treated with Nyxol) and has demonstrated promising clinical data for use in multiple 
ophthalmic indications.  
In prior clinical trials, Nyxol has demonstrated a consistent ability to decrease pupil diameter (PD) 
by approximately 20% - 30%  (~1-1.5 mm) in both mesopic and photopic conditions (1-3). Nyxol 
has been found to be efficacious for more than 24 hours, with an onset of action of approximately 
30 to 60 min. Nyxol has been observed to be well tolerated at single doses and multiple doses up to 
14 days. Safety of the subjects in these trials was evaluated by adverse event (AE) monitoring, 
physical examinations, and vital sign assessments. Across all trials, no serious adverse events 
(SAEs) have been reported. No deaths occurred in any of the trials. No clinically meaningful 
changes were observed in physical examinations or vital signs, including blood pressure (BP) and 
heart rate (HR). Adverse events reported were mild to moderate in intensity, with the most 
common being transient conjunctival hyperemia (eye redness) and ocular irritation; however, 
Nyxol dosing at or near bedtime was observed to mitigate or minimize these side effects during the 
daytime. 
The VEGA-1 study evaluated the efficacy and safety of Nyxol in combination with LDP and as a 
single agent in presbyopic subjects. The study met its primary and many secondary endpoints.  
Nyxol + LDP showed a favorable safety profile with no serious AEs. Most AEs were mild, with no 
headaches, no browaches, and no blurry vision AEs reported. Mild, transient conjunctival 
hyperemia AEs were reported in < 5% of subjects. 
Pilocarpine Hydrochloride 
A detailed report of pilocarpine, its product label, properties, and potential risk are provided in the 
IB (2022). 
Pilocarpine hydrochloride (PCH) has been studied in a prior clinical trial by Ocuphire. In addition, 
there is a recently approved PCH drug at 1.25% and extensive literature on pilocarpine’s ability to 
reduce PD across various concentrations. These pupil-constricting effects, however, are 
counteracted by a dose-dependent side effect profile, notably browache and stinging in the eye 
upon topical application of the drug.  
In an effort to better characterize an efficacious dose of PCH that would have minimal side effects, 
Ocuphire conducted an expansive review of the medical literature concerning experiments of 
various doses of PCH and their effects on PD and side effect profile. Leavitt et al. found that 


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normal pupils constrict with dilute concentrations of 0.25% and 0.125% pilocarpine (1.7 and 
0.6 mm, respectively) but constrict insignificantly at concentrations of 0.0313% and 0.0625% (4). 
These decreases in PD were found to occur within 15 min of dosing, and peak pupil size reduction 
occurred at 30 to 60 min (4). On the other hand, 2% pilocarpine, although efficacious in decreasing 
PD by up to 3 mm, caused significant side effects, including worsened visual field and acuity (5). 
When considering the effects on PD and the potential side effects of higher doses of pilocarpine, 
Ocuphire found that a range of 0.3% to 0.6% PCH may provide moderate efficacy with potentially 
limited side effects. Moreover, based on these data, Ocuphire has shown in its Phase 2 VEGA-1 
study that the combination of Nyxol and LDP can have an additive to synergistic effect to produce 
a long duration of pupil reduction in an optimal range of PDs for treating presbyopia, without 
causing adverse side effects such as headache, browache, or accommodative spasm. 
3.2. Design Justification 
Presbyopia is an age-related condition, with onset most common in people over 40 years old (6). 
Patients experience blurred near vision, difficulty seeing in dim light, and eye strain. In young, 
healthy eyes, lenses are able to focus light from objects at different distances by a process called 
accommodation. During accommodation, the ciliary muscle surrounding the lens contracts, causing 
the lens to change shape, thus increasing the focusing power of the eye. This allows for dynamic, 
clear vision at both near and far distances. With increasing age, the lens becomes stiffer, which 
limits the eye’s ability to adjust its focus for reading or for other tasks that require clear vision at 
near distances.  
Presbyopia has a significantly negative impact on quality of life (7), interfering with daily activities 
such as reading, use of computers or hand-held devices, and seeing the dashboard of a car. It is 
estimated that 100 million Americans have presbyopia, and this number is expected to grow as the 
population above the age of 40 increases (8). 
Limitations of Existing Treatments for Presbyopia 
Vuity™ (PCH 1.25%) has recently been approved for the treatment of presbyopia. In two Phase 3 
studies, Vuity met its primary outcome measure, with 31% and 26% of Vuity-treated subjects 
gaining 3 or more lines of mesopic DCNVA 3 hours post-dose compared to 8% and 11% of 
placebo-treated subjects. However, limitations to Vuity include limited duration of efficacy, with a 
need for twice daily dosing to achieve efficacy throughout the day; headache, browache, and 
burning/stinging associated with drug administration; accommodative spasm and risk of angle-
closure glaucoma due to activation of the ciliary muscle; and risk of retinal detachment in subjects 
with high myopia (9).  
Other available treatments for presbyopia include reading glasses, bifocals, gradients, bifocal 
contact lenses, and multifocal intraocular lenses (IOLs). Reading glasses can be inconvenient and 
must be taken off and put on frequently throughout the day to see objects at far and near distances, 
respectively. Many patients express frustration with losing or forgetting their reading glasses. 
Moreover, some patients find glasses unflattering, and contact lenses for presbyopia have their own 
set of drawbacks regarding eye strain and limitation of use. 
A small portion of patients elect surgical intervention, including laser treatment, to achieve 
monovision. The risks of such interventions include a potential decrease in contrast sensitivity and 
stereopsis and the creation or worsening of dim light vision disturbances. 


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Nyxol Opportunity in Presbyopia 
Pharmacological treatment of presbyopia is a promising strategy and an intense area of focus by 
several pharmaceutical companies. Other pharmacological and device approaches have consistently 
demonstrated that reducing pupil size to a diameter of approximately 2 to 3 mm will lead to 
significant improvement in presbyopia symptoms by increasing depth of focus (10-12). In multiple 
trials, it has been shown that Nyxol reduces PD by approximately 20% to 30% and improves near 
visual acuity (VA) for at least 24 hours after a single application. OPI-NYXP-201 (VEGA-1) was a 
7-day (2 visits), double-masked, randomized, placebo-controlled, multicenter Phase 2 trial in 150 
presbyopic subjects. Results of this study suggest that treatment with Nyxol alone provides a 
≥ 3-line benefit in DCNVA in a similar percent of subjects to that observed with Vuity, with better 
durability and a better safety profile. Nyxol with adjunctive LDP therapy resulted in a higher 
percentage of presbyopic subjects achieving a gain of 3 or more lines of DCNVA, with increased 
durability and a better safety profile compared to those from the Vuity pivotal trials. A summary of 
the results of the VEGA-1 study are presented here. 
Efficacy Results From the VEGA-1 Study: 
Nyxol as a Single Agent 
Following 3 to 4 days of treatment with Nyxol or placebo, 30.1% of Nyxol-treated subjects had 
≥ 15 letters of improvement in photopic binocular DCNVA compared with 13.5% of placebo-
treated subjects 12 hours after the last dose (p=0.0265). This benefit remained stable over the next 
6 hours, with a mean of 36.7% of Nyxol-treated subjects improving by 15 or more letters DCNVA 
18 hours post-dose. In subjects with light irides, there was a 21.2% benefit of Nyxol treatment over 
placebo (p=0.0298) and subjects with dark irides had 10.0% more responders with Nyxol treatment 
compared with placebo (p=0.3771). Significantly more subjects treated with Nyxol achieved 
binocular DCNVA of 20/40 or better compared with placebo (56% vs 36%, respectively; 
p=0.0080).  
Photopic binocular mean DCNVA showed significantly greater improvement in mean number of 
letters read among Nyxol-treated subjects (10.1-letter improvement) compared with placebo-
treated subjects (5.6-letter improvement) at 12 hours post-dose (p=0.0006). Consistent with these 
results, mean photopic PD in the best eye was significantly reduced among Nyxol-treated subjects 
compared with placebo-treated subjects at 12 hours post-dose (p<0.0001). 
Nyxol + LDP as Adjunctive Therapy  
A statistically significantly greater number of subjects in the Per Protocol (PP) Population treated 
with Nyxol + LDP had ≥ 15 letters of improvement in photopic binocular DCNVA at 1 hour post-
LDP treatment (primary endpoint) compared with placebo alone (60.45% vs 27.9%, respectively; 
p=0.0037). In addition, the Nyxol + LDP arm had significantly more subjects with ≥ 15 letters of 
improvement in photopic binocular DCNVA compared with placebo alone at all timepoints from 
30 min through 4 hours post-LDP treatment (p≤0.0166 for each). This benefit was retained for all 
timepoints between 30 min and 4 hours when requiring that subjects with ≥ 15 letters of 
improvement in photopic binocular DCNVA also had to have < 5 letters of loss in photopic 
binocular best-corrected distance visual acuity (BCDVA), the accepted registration endpoint. The 
benefit of LDP adjunctive therapy to Nyxol was consistent regardless of iris color. 
The effect of Nyxol + LDP on DCNVA was rapid and clinically significant. By 30 min post-LDP 
treatment, 79% of subjects had photopic binocular DCNVA of 20/40 or better. By 1 hour post-LDP 


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treatment, 84% of subjects had binocular DCNVA of 20/40 or better. Significantly more subjects 
treated with LDP adjunctive to Nyxol had photopic binocular DCNVA 20/40 or better compared 
with placebo at 30 min through 2 hours (p≤0.0119 for each).  
Nyxol + LDP treatment resulted in more subjects achieving a ≥ 15-letter improvement in DCNVA 
compared with each of its components used individually at all timepoints from 30 min to 2 hours 
post-LDP treatment. Although the study was not statistically powered for this outcome, the percent 
of responders at 30 min with combination therapy (61%) was significantly greater than the 
response rate with LDP alone (26%; p=0.0076) or Nyxol alone (33%; p=0.0265). 
Safety Results From the VEGA-1 Study: 
Nyxol, when administered with LDP as adjunctive therapy or as a single agent, was well tolerated 
in this study. Fifty treatment-emergent adverse events (TEAEs) were reported in 15 subjects 
(34.1%) treated with Nyxol + LDP, 4 TEAEs were reported in 2 subjects (4.4%) treated with 
placebo alone, 18 TEAEs were reported in 6 subjects (20.0%) treated with Nyxol alone, and 
13 TEAEs were reported in 6 subjects (19.4%) treated with placebo + LDP. All subjects 
experienced mild TEAEs except for 3 subjects: 1 in the placebo alone arm and 2 in the Nyxol + 
LDP arm. All subjects had TEAEs that were considered possibly, probably, or definitely related to 
treatment, except for 2 subjects: 1 in the placebo alone arm had an unrelated TEAE and 1 in the 
Nyxol + LDP arm had a TEAE of unknown relatedness. One subject in the Nyxol + LDP arm had 
2 TEAEs (mild conjunctival hyperemia and mild eye irritation) leading both to study medication 
discontinuation and withdrawal from the study. No subjects had any serious TEAEs, and no 
subjects died during the study.  
3.3. Route of Administration, Dosage Regimen, and Treatment Period 
As the intended route of administration for Nyxol and its placebo and LDP and its placebo is 
topical ophthalmic, this is the route to be used in this study.  
The dose for 1 drop of Nyxol (POS 0.75%) selected for this study was based upon: 1) preclinical 
safety studies, 2) the results of the previous ophthalmic clinical studies described above and in the 
IB, and 3) clinical studies conducted with varying doses of drugs in the same class (13). 
Note that POS 0.75%, which expresses the phentolamine mesylate concentration as the free base, is 
the new nomenclature being used in place of Phentolamine Mesylate Ophthalmic Solution 1%, 
which was how Nyxol had been described in studies prior to 2020. 
The dose for 1 drop of Pilocarpine Hydrochloride Ophthalmic Solution selected for this study is 
0.4% (LDP). 
All treatments will be administered in both eyes (OU). 
3.4. Compliance 
This study will be conducted in compliance with the protocol and in accordance with Good 
Clinical Practice (GCP), the ethical principles set forth in the Declaration of Helsinki, and with the 
United States (US) Code of Federal Regulations. 
3.5. Study Population 
The study population for this trial will be comprised of males and females between 40 and 64 years 
of age with presbyopia. A sample size of approximately 320 subjects will be randomized into 1 of 


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4 treatment arms, with the expectation that approximately 300 subjects will be evaluable for 
efficacy.  
• Nyxol + LDP 
• Nyxol + LDP vehicle  
• Placebo (ie, Nyxol vehicle) + LDP 
• Placebo + LDP vehicle  
Subjects will be randomized 1:1:1:1 into the above groups. 
At Visit 1 (Screening/Baseline), eligible subjects will be randomized, and the first dose of 
Treatment 1 will be administered (note: if Visit 1 is split into 2 separate days, then dosing occurs 
on the second day). Efficacy and safety measurements are made before dosing with Treatment 1 
(Time 0 min) and at multiple post-dose timepoints from 30 min to 8 hours. If Visit 1 is split over 
2 separate days, all Screening assessments up to and including randomization must occur on the 
first day, and then the subject should return to complete their post-randomization assessments in 
the following order on the second day (ie, confirm concomitant medications, subject questionnaire, 
in-office Treatment 1 dosing, ocular tolerability, followed by all other assessments at time points 
0.5, 1, 3, 5, and 8 hours post-treatment). Prior to discharge following completion of Visit 1 (Stage 1 
Day 1), Treatment 1 study medication for Stages 1 and 2 will be dispensed for evening dosing by 
the subject before bedtime, to begin the night of Stage 1 Day 2. On the evening prior to Visit 2, the 
subject will dose Treatment 1 study medication as close to 12 hours prior to Time 0 min for that 
visit. 
At Visit 2 (Stage 1 Day 8), efficacy and safety measurements will be made at 12 hours after the 
dose of Treatment 1 study medication (12-hour endpoint). After completion of these assessments, 
the subject will take no study drug for 7 to 14 days (during the Washout Period), after which 
Stage 1 is complete. Measurements will be made on Visit 3 (Stage 1 Day 10 ± 1 day) during the 
Washout Period to assess the absence of residual treatment effects.  
Subjects will restart dosing daily with Treatment 1 in the evening immediately prior to Visit 4 
(Stage 2 Day 1) as close to 12 hours prior to the scheduled time for Visit 4 for the 12-hour 
assessments. At this visit, after completion of the 12-hour assessments for Treatment 1, subjects 
will be administered their morning dose of Treatment 2 (at Time 0 min) at the study site, and 
efficacy and safety measurements will be made at multiple post-dose timepoints over the next 8 
hours. During Stage 2, subjects will administer Treatment 1 in the evenings near bedtime and 
Treatment 2 in the mornings from the evening prior to Visit 4 (Stage 2 Day 1) for the next 7 days 
or until the evening before their scheduled Visit 5 (Stage 2 Day 8). On the evening prior to Visit 5, 
Treatment 1 should be administered as close to 12 hours prior to the scheduled time for Visit 5. At 
Visit 5, measurements will be made before dosing with Treatment 2 at Time 0 min and at multiple 
post-dose timepoints over 8 hours after dosing with Treatment 2. 
Randomization will be stratified by light/dark color irides (Appendix 1). The number of subjects in 
each irides type should be as close as possible to 1:1, with a minimum of 40% within each stratum. 
The subjects will be recruited from approximately 30 investigational sites in the US.  
 


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4. OBJECTIVES AND PURPOSE 
The VEGA-2 study is a randomized, placebo-controlled, double-masked study of the safety and 
efficacy of Nyxol (POS 0.75%) in combination with LDP 0.4% in subjects with presbyopia. 
The objectives of this study are: 
Primary objective 
• To evaluate the efficacy of LDP as adjunctive therapy to Nyxol (Nyxol + LDP) to 
improve DCNVA without loss of BCDVA compared to placebo + LDP vehicle in 
subjects with presbyopia  
Additional objectives 
• To evaluate the efficacy of Nyxol + LDP to improve DCNVA without loss of BCDVA 
compared to placebo + LDP (LDP alone) and Nyxol + LDP vehicle (Nyxol alone) in 
subjects with presbyopia 
• To evaluate the efficacy of Nyxol as a single agent to improve DCNVA without loss of 
BCDVA compared to placebo in subjects with presbyopia   
• To evaluate additional efficacy measures, such as VA and PD, across multiple 
timepoints and visits for Nyxol as a single agent and Nyxol + LDP  
• To evaluate the effect of iris color, Baseline PD, and Baseline DCNVA on efficacy of 
Nyxol as a single agent and Nyxol + LDP 
• To evaluate the ocular and systemic safety of Nyxol as a single agent and Nyxol + LDP  
• To evaluate subject-reported outcomes of Nyxol as a single agent and Nyxol + LDP 
he Sponsor intends to use this Phase 3 pivotal study to evaluate Nyxol as a single agent and Nyxol 
+ LDP for the indication of “treatment of presbyopia.” 
 
5. STUDY DESIGN 
This is a randomized, double-masked, placebo-controlled, multicenter, Phase 3 study in 
approximately 320 subjects with presbyopia. The study will be performed in 2 stages, as shown in 
Appendix 4.  
Subjects will be randomized 1:1:1:1 to treatments for both stages at Visit 1 (Screening/Baseline). 
Treatment 1 assignment in Stage 1 will be maintained for Stage 2. Stage 1 will consist of 
2 treatment groups (Nyxol or placebo [ie, Nyxol vehicle]), with approximately 160 subjects in each 
group. Stage 2 consists of 4 treatment groups (Nyxol + LDP, Nyxol + LDP vehicle, placebo + 
LDP, and placebo + LDP vehicle), with approximately 80 subjects per treatment group. 
Randomization will be stratified by light/dark irides (Appendix 1). The number of subjects in each 
irides type should be as close as possible to 1:1, with a minimum of 40% within each stratum. The 
subjects will be recruited from approximately 30 investigational sites in the US. 
Stage 1: Dosing of Treatment 1 


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In Stage 1, the safety and efficacy of Nyxol as a single agent will be evaluated over the following 
1 day and 7 days of daily dosing in the evenings near bedtime, except the evening before Visit 2, 
when Treatment 1 is dosed as close to 12 hours prior to Time 0 min for that visit. Following Visit 2 
(Stage 1 Day 8), subjects will discontinue dosing Treatment 1 for 7 to 14 days (Washout Period). 
During the Washout Period, subjects will return for Visit 3 (Stage 1 Day 10 ± 1 day) to assess the 
resolution of drug treatment effects.  
During Stage 1, subjects will dose with Treatment 1 (Nyxol or placebo [ie, Nyxol vehicle]) in the 
evenings near bedtime (except for Visit 1 [Stage 1 Day 1], when the subject is dosed during the day 
at the study site).  
Measurements will be made at multiple visits and timepoints, and analysis will consist of 
comparison across the 2 arms: 
• Nyxol 
• Placebo 
Stage 2: Dosing of Treatments 1 and 2 
Visit 4 (Stage 2 Day 1) is the start of Stage 2. In Stage 2, the efficacy and safety of Nyxol + LDP 
will be evaluated following 1 day and 7 days of dosing, in which Treatment 1 (Nyxol or placebo) is 
dosed daily in the evenings near bedtime, except the evening before Visits 4 and 5, when 
Treatment 1 is dosed as close to 12 hours prior to the scheduled time for those visits. Treatment 2 
(LDP or LDP vehicle) is dosed daily in the mornings. Treatment 2 will be dosed at the study site at 
the beginning of Visits 4 and 5 (Stage 2 Days 1 and 8, respectively).  
Measurements will be made at multiple visits and timepoints, and analysis will consist of 
comparison across the 4 arms:  
• Nyxol + LDP 
• Nyxol + LDP vehicle  
• Placebo + LDP 
• Placebo + LDP vehicle  
Other design considerations: 
Study medications (Treatments 1 and 2) will each be dosed as a single drop in each eye, with 
Treatment 1 dosed in the evenings near bedtime and Treatment 2 dosed in the mornings. Baseline 
for the study is defined as pre-treatment up to and including randomization (prior to Time 0 min) at 
Visit 1 (Screening/Baseline).  
During Visit 2 (Stage 1 Day 8), Time 0 min is defined as the timing of the first efficacy 
assessments at the visit, approximately 12 hours after dosing with Nyxol the previous night (12-
hour post-dose timepoint). During Visits 4 and 5 (Stage 2 Days 1 and 8, respectively), Time 0 min 
is defined as the time of Treatment 2 administration.  
The study eye is defined as the eligible eye with worse Baseline DCNVA at Visit 1 
(Screening/Baseline). In the case where both eyes have the same Baseline DCNVA, then the eye 
with the better BCDVA will be the study eye. If both DCNVA and BCDVA are equivalent 
between eyes, the study eye will be the right eye (OD). The study eye and fellow eye will both be 


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evaluated at all assessments. The baseline values are determined during Screening assessments at 
Visit 1. 
5.1. Primary And Secondary Endpoints 
Efficacy: 
The primary efficacy endpoint is the percent of subjects with ≥ 15 letters of improvement in 
photopic binocular DCNVA and with < 5 letters of loss in photopic binocular BCDVA at 30 min 
post-LDP/vehicle comparing Nyxol + LDP to placebo + LDP vehicle at Visit 5 (Stage 2 Day 8).    
This comparison will be analyzed first using a hierarchical testing method.  
The key secondary endpoints are:  
• The percent of subjects with ≥15 letters of improvement in photopic binocular DCNVA 
and with < 5 letters of loss in photopic binocular BCDVA at 30 min post-LDP/vehicle 
comparing Nyxol + LDP to LDP alone at Visit 5 (Stage 2 Day 8) 
• The percent of subjects with ≥ 15 letters of improvement in photopic binocular DCNVA 
and with < 5 letters of loss in photopic binocular BCDVA at 30 min post-LDP/vehicle 
comparing Nyxol + LDP to Nyxol alone at Visit 5 (Stage 2 Day 8)  
These comparisons will be analyzed second and third using a hierarchical testing method. 
Other secondary endpoints measured at various timepoints and visits by treatment arms in Stage 1 
and Stage 2 include: 
• Percentage of subjects with ≥ 15 letters of improvement in binocular photopic DCNVA 
and with < 5 letters of loss in binocular photopic BCDVA from Baseline (excluding 
primary and key secondary outcome timepoints) 
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of improvement in photopic 
DCNVA from Baseline (monocular and binocular) 
• Change in photopic DCNVA from Baseline (monocular and binocular) 
• Percent of subjects with photopic DCNVA ≤ 20/15, ≤ 20/20, ≤ 20/25, ≤ 20/32, ≤ 20/40, 
≤ 20/50, ≤ 20/63, ≤ 20/80, ≤ 20/100, ≤ 20/125, and ≤ 20/160  
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of improvement in photopic 
binocular distance-corrected intermediate visual acuity (DCIVA) from Baseline  
• Change in photopic binocular DCIVA from Baseline  
• Percentage of subjects with loss or improvement in photopic and mesopic BCDVA 
from Baseline (monocular and binocular) 
• Change in photopic and mesopic BCDVA from Baseline (monocular and binocular) 
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of improvement in mesopic 
binocular DCNVA from Baseline  
• Change in mesopic binocular DCNVA from Baseline  
• Change and percent change in photopic PD from Baseline  
• Percentage of subjects with photopic PD of < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 mm  


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• Percentage of subjects with photopic PD of 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 2.5 to 
< 3.0 mm, and 3.0 to < 3.5 mm  
• Change and percent change in mesopic PD from Baseline  
• Percentage of subjects with mesopic PD of < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 mm  
• Percentage of subjects with mesopic PD of 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 2.5 to 
< 3.0 mm, and 3.0 to < 3.5 mm  
• Percentage of subjects with Baseline photopic DCNVA ≤ 20/63, ≤ 20/80, ≤ 20/100, 
≤ 20/125, and ≤ 20/160, with a ≥ 10-letter and ≥ 15-letter improvement in photopic 
DCNVA  
• Percentage of subjects with Baseline photopic DCNVA from 20/50 to 20/63, 20/50 to 
20/80, 20/50 to 20/100, 20/50 to 20/125, and 20/50 to 20/160, with a ≥ 10-letter and 
≥ 15-letter improvement in photopic DCNVA  
• Subject questionnaire responses related to change in near vision and satisfaction with 
near vision 
Measurements: 
• Best-corrected distance VA will be measured in photopic conditions by a high-contrast 
Standard Early Treatment Diabetic Retinopathy Study (ETDRS) illuminated chart (on 
wall or stand) at 4 m  
• Distance-corrected intermediate VA will be measured in photopic conditions by a high-
contrast Near Visual Acuity Chart in the Precision Vision Small 914 Illuminator 
Cabinet (light box) at 26 inches (66 cm) 
• Distance-corrected near VA will be measured in photopic and mesopic conditions by a 
high-contrast Near Visual Acuity Chart in the Precision Vision Small 914 Illuminator 
Cabinet (light box) at 16 inches (~40 cm) (Appendix 2) 
• Pupil diameter will be measured with a NeurOptics pupillometer (mm)  
All of the efficacy endpoints will be analyzed overall and by light/dark irides at all timepoints. 
Subjects will also be analyzed by binocular, study eye, and fellow eye, where applicable. 
In photopic lighting conditions, the distance and near illuminated charts will be at a luminance 
level of approximately 85 to 160 cd/m2 (85 to 160 nits or 267 to 502 lux). In mesopic conditions, 
the distance and near illuminated charts will be at a luminance level of approximately 3 cd/m2 (3 
nits or 9.42 lux).  
The subject will be in the same room for all assessments, and every effort will be made to have the 
same person perform the measurements at all visits. 
For VA, measurements will be made in letters and will be converted to LogMAR and lines, as 
appropriate. 
All efficacy endpoints will also be analyzed by the Modified Intention-to-Treat (mITT) Population 
and selected efficacy endpoints will be analyzed using the PP Population as specified in the 
Statistical Analysis Plan (SAP). 


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Safety: 
Safety measures are AEs, ocular tolerability, intraocular pressure (IOP), BCDVA, biomicroscopy, 
ophthalmoscopy, HR and BP, and subject questionnaire. Urine pregnancy tests for females of 
childbearing potential will also be conducted. 
Measurements:                                                                                                                                                                
• Biomicroscopy, ophthalmoscopy, and HR and BP measured in accordance with the 
site’s standard practice  
• Ocular tolerability will be measured on a 4-point scale (0-3) 
• Intraocular pressure will be measured with a Tono-Pen or Goldmann Applanation 
tonometry 
• Subject questionnaire will be completed by the subject (Appendix 5 and Appendix 6) 
5.2. Description and Schedule of Visits and Procedures 
Study procedures are shown in detail in the Schedule of Events tables (Table 1 and Table 2). 


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Table 1: 
Schedule of Events for Stage 1 
 
Screening/Baseline Visit 1[a] 
Visit 2[b] 
 
Visit 3[b] 
Day  
Stage 1 Day 1  
Stage 1 Day 8 
Washout Period 
7 – 14 days 
Stage 1 Day 10 
(± 1 day) 
Time (hr)[c]  
0 
0.5  
1  
3 
5 
8  
0 
0.5 
1 
3 
5 
8 
 
 
Informed consent   
X  
  
  
  
  
  
 
  
  
  
  
  
  
 
Subject ID # assigned   
X  
  
  
  
  
  
  
  
  
  
  
  
  
 
Medical/Ophthalmic history   
X  
  
  
  
  
  
 
  
  
  
  
  
  
 
Demographics    
X  
  
  
  
  
  
  
  
  
  
  
  
 
 
Prior/Concomitant medications  
X* 
  
 
 
 
 
X 
 
 
 
 
 
 
X 
Urine pregnancy test[d]  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
HR/BP  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
Manifest refraction and near add  
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
PD (photopic)[e]  
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
DCNVA (photopic)[f]  
X[g] 
 
X 
X 
X 
X 
X[g] 
X 
X 
X 
X 
X[g] 
 
X 
DCIVA (photopic)[f] 
X 
 
 
X 
X 
 
X 
 
X 
X 
 
X 
 
 
BCDVA (photopic)[f]  
X[g] 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X[g] 
 
X 
PD (mesopic)[e]  
X 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
DCNVA (mesopic)[f]  
X 
 
X 
X 
X 
X 
X 
 
 
X 
X 
X 
 
 
BCDVA (mesopic)[f]  
X[g] 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X[g] 
 
X  
Biomicroscopy  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
Corneal fluorescein staining and TBUT 
X 
 
 
 
 
 
 
 
 
 
 
 
 
  
IOP[h]  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
Ophthalmoscopy[i]  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
AEs  
  X---------------------------------------------------------------------------------------------------------------------X 
Randomization[a] 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
Subject questionnaire[l]  
X* 
 
 
 
 
 
 
 
 
X 
 
 
 
 
In-office Treatment 1: Nyxol or placebo[j]  
X* 
 
 
 
 
 
 
 
 
 
 
 
 
 
Ocular tolerability[k]  
X* 
 
 
 
 
 
X 
 
 
 
 
 
 
 
Treatment 1: Nyxol or placebo and Subject 
diary dispensing/review[j]  
 
 
 
 
 
X 
 
 
 
 
 
X 
 
 
AE, adverse event; BCDVA, best-corrected distance visual acuity; BP, blood pressure; DCNVA, distance-corrected near visual acuity; HR, heart rate; ID, identification; 
IOP, intraocular pressure; OD, right eye; OS, left eye; PD, pupil diameter; VA, visual acuity. 
[a] If the subject meets all the inclusion criteria and none of the exclusion criteria, this Screening Visit becomes the Baseline Visit. Visit 1 (Screening/Baseline) 


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assessments are recommended to be performed on the same day; however, in instances in which subjects are unable to complete the full day of assessments for Visit 1, 
subjects may be scheduled to return for the completion of Visit 1 within 1 week of Screening. If Visit 1 is split over 2 separate days, all Screening assessments up to and 
including randomization must occur on the first day, and then the subject should return to complete their post-randomization assessments on the second day. *Post-
randomization assessments to be performed on the second day (ie, confirm concomitant medications, subject questionnaire, in-office Treatment 1 dosing, ocular 
tolerability, followed by all other assessments at time points 0.5, 1, 3, 5, and 8 hours post-treatment).    
[b] Time of visits will be scheduled relative to the prior evening’s dose of Treatment 1. Visit 2 will be scheduled such that the 0 hour (Time 0 min) assessments on Visit 2 
will occur approximately 12 hours (± 1 hour) after the prior evening’s dose of Treatment 1. Visit 3 occurs at Stage 1 Day 10 (±1 day) during the Washout Period. 
[c] An additional ± 5 min is permitted for measurements at 0.5 and 1 hour; an additional ± 10 min is permitted at 3, 5, and 8 hours for Visits 1 and 2.   
[d] Urine pregnancy test is for females of childbearing potential only.   
[e] PD performed with NeurOptics VIP-300 pupillometer.    
[f] VA performed binocularly, except where specifically noted [g]. 
[g] VA performed monocularly (OD, OS) following binocular assessment. 
[h] IOP performed with Tono-Pen or Goldmann applanation tonometry.   
[i] Direct or indirect ophthalmoscopy performed without dilation. Use of 90 D lens (indirect) is allowed.   
[j] Treatment 1: At Visit 1 (either first or second day of Visit 1, see * above), one drop of Nyxol or placebo (ie, Nyxol vehicle) will be dosed at the study site after the 
completion of 0 hour (Time 0 min) ocular assessments. Subject will dose with Treatment 1 each evening near bedtime, starting the evening of Stage 1 Day 2 through the 
evening prior to Visit 2 (Stage 1 Day 8), except the evening before Visit 2, when Treatment 1 is dosed as close to 12 hours prior to Time 0 min for that visit. Subjects will 
discontinue dosing during the 7- to 14-day Washout Period. 
[k] Ocular tolerability performed using a 4-point scale; it will be assessed at the beginning of each specified timepoint after dosing at Visits 1 and 2. 
[l] Subject questionnaire (Appendix 5 and Appendix 6) will be completed by the subject at specified visits and timepoints. 
 
 


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Table 2: 
Schedule of Events for Stage 2  
Visit 4[a] 
Visit 5[a] 
 
Stage 2 Day 1 (Day 15 to 22) 
Stage 2 Day 8 (7 days after Stage 2 Day 1) 
Time (hr)[b] 
12 hr post 
Treatment 1  
0  
0.5  
1  
3 
5  
8  
12 hr post 
Treatment 1  
0  
0.5  
1  
3 
5  
8  
Prior/Concomitant medications  
X 
X 
Urine pregnancy test[c]  
X 
  
X 
HR/BP  
X 
PD (photopic)[d]  
X 
  
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
DCNVA (photopic)[e]  
X 
 
X 
X 
X 
X 
X 
X 
 
X[f] 
X 
X 
X 
X[f] 
DCIVA (photopic)[e]  
X 
 
X 
X 
X 
X 
 
X 
X 
X  
BCDVA (photopic)[e]  
X 
 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X[f] 
PD (mesopic)[d]  
X 
 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X 
DCNVA (mesopic)[e]  
X 
 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X  
BCDVA (mesopic)[e]  
X 
  
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X[f] 
Biomicroscopy  
 
X 
IOP[g]  
 
X 
Ophthalmoscopy[h]  
 
X 
AEs  
           X------------------------------------------------------------------------------------------------------------------X 
Subject questionnaire[k] 
 
 
 
 
X 
 
 
 
 
 
 
X 
 
 
In-office Treatment 2: LDP or LDP vehicle[i] 
X 
X 
Ocular tolerability[j]  
X 
X 
Treatment 1: Nyxol or placebo and Subject 
diary dispensed / reviewed[l] 
  
 
 
 
 
 
X  
 
 
 
 
 
 
X 
Treatment 2: LDP or LDP vehicle and Subject 
diary dispensed / collected[i] 
 
 
 
 
 
 
X 
 
 
 
 
 
 
X 
AE, adverse event; BCDVA, best-corrected distance visual acuity; BP, blood pressure; DCNVA, distance-corrected near visual acuity; HR, heart rate; ID, identification; 
IOP, intraocular pressure; LDP, Low-Dose Pilocarpine; OD, right eye; OS, left eye; PD, pupil diameter; VA, visual acuity. 
[a] Time of visits will be scheduled relative to the prior evening’s dose of Treatment 1. Visits 4 and 5 will be scheduled such that the 12 hour post-dose assessments on 
Visits 4 and 5 will occur approximately 12 hours (± 1 hour) after the prior evening’s dose of Treatment 1.  
[b] An additional ± 5 min is permitted for measurements at 0.5 and 1 hour; an additional ± 10 min is permitted at 3, 5, and 8 hours for Visits 4 and 5. 
[c] Urine pregnancy test is for females of childbearing potential only. 
[d] PD performed with NeurOptics VIP-300 pupillometer. 
[e] VA performed binocularly, except where specifically noted [f]. 


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[f] VA performed monocularly, following binocular assessment. 
[g] IOP performed with Tono-Pen or Goldmann applanation tonometry.   
[h] Direct or indirect ophthalmoscopy performed without dilation. Use of 90 D lens (indirect) is allowed.  
[i] Treatment 2: One drop of LDP or LDP vehicle in each eye dosed daily in the mornings beginning on the day after Visit 4 and daily through the day prior to Visit 5. At 
Visits 4 and 5, subject will be dosed with Treatment 2 at the study site. Subject will bring Treatment 2 kit and diary to Visits 4 and 5 for review and accountability. 
[j] Ocular tolerability performed using a 4-point scale; it will be assessed at the beginning of each specified timepoint after dosing at Visits 4 and 5.  
[k] Subject questionnaire (Appendix 5 and Appendix 6) will be completed by the subject at specified visits and timepoints.  
[l] Treatment 1: One drop of Nyxol or placebo in each eye dosed daily in the evenings near bedtime, except the evening before Visits 4 and 5, when Treatment 1 is dosed 
as close to 12 hours prior to the scheduled time for those visits. Subjects will bring their Treatment 1 kit and diary to Visits 4 and 5 for review and accountability. 
 
 
 


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5.3. Measures Taken to Minimize/Avoid Bias 
This is a placebo-controlled, double-masked, randomized, 4-arm Phase 3 study.  
5.4. Study Medications 
5.4.1. 
Study Medication Identification 
The POS treatment medication is phentolamine mesylate; its chemical name is 3-[N-(4,5-
dihydro-1H-imidazol-2-ylmethyl)-4-methylanilino]phenol;methanesulfonic acid. It is a white to 
off-white, odorless crystalline powder with a molecular weight of 377.46 g/mol. Solutions of 
phentolamine mesylate are acid to litmus. It is freely soluble in water and in ethanol, and slightly 
soluble in chloroform. It melts at approximately 178ºC. 
The properties of phentolamine mesylate are described in Table 3. 
Table 3: 
Phentolamine Mesylate Drug Substance and Drug Product Identifiers 
Established name 
Phentolamine mesylate – parent phentolamine 
CAS registry number 
65-28-1 – parent 50-60-2 
Chemical class 
An alpha-adrenergic receptor antagonist, it is a 
member of the following classes: imidazoles, of 
phenols, is a tertiary amino compound and a 
substituted aniline 
Chemical name 
3-[N-(4,5-dihydro-1H-imidazol-2-ylmethyl)-4-
methylanilino]phenol;methanesulfonic acid 
Molecular formula 
C18H23N3O4S– parent C17H19N3O 
Molecular weight (g/mol) 
377.140 – parent 281.352 
Drug name/formulation 
Nyxol/aqueous isotonic solution  
Concentration active 
Mesylate salt 1% – parent 0.75% 
Manufacturer drug substance 
Industriale Chimica s.r.l. (Italy) 
Manufacturer drug product, placebo 
Woodstock Sterile Solutions  
Storage requirements 
Shipped and stored at the study site refrigerated 
(2°C to 8°C, 36°F to 46°F) until provided to the 
subject; then subject stores at room temperature 
(15°C to 25°C, 59°F to 77°F) Stored at the site, 
drug will be placed in a secured location (locked) 
with no access for unauthorized personnel. 
Nyxol (POS 0.75%) is a clear, colorless to slightly brown, sterile, non-preserved, isotonic, 
buffered aqueous solution containing 1% phentolamine mesylate (equivalent to 0.75% 
phentolamine free base), mannitol, sodium acetate, and water for injection. Placebo for Nyxol is 
a clear, colorless, sterile, non-preserved, isotonic, buffered aqueous solution containing mannitol, 
sodium acetate, and water for injection. The pH of both Nyxol and its placebo is adjusted with 
NaOH (United States Pharmacopeia [USP]) and/or HCl (USP) to a pH range between 4.5 to 5.3.  


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For the proposed study, a second treatment medication, Pilocarpine Hydrochloride Ophthalmic 
Solution 0.4% (LDP) will be used. Pilocarpine Hydrochloride Ophthalmic Solutions 1%, 1.25%, 
2%, and 4% are approved by the US Food and Drug Administration (FDA) and available 
generically by multiple pharmaceutical manufacturers, including Sandoz Inc., Somerset LLC, 
Akorn Inc, and Alcon Laboratories.  
Pilocarpine hydrochloride is the hydrochloride salt of (+)-pilocarpine (parent compound), a 
medication used to treat increased IOP, dry mouth, and presbyopia. Its chemical name is 
(3S,4R)-3-ethyl-4-[(3-methylimidazol-4-yl)methyl]oxolan-2-one hydrochloride, and it has a 
molecular weight of 244.72 g/mol. Pilocarpine hydrochloride is a hygroscopic, odorless, bitter-
tasting white crystal or powder which is soluble in water and alcohol and virtually insoluble in 
most non-polar solvents. It melts at approximately 204.5°C. Its aqueous solubility in buffer at pH 
7.4 is > 36.7 μg/mL. The parent (+)-pilocarpine is a natural alkaloid extracted from plants of the 
genus Pilocarpus with cholinergic agonist activity. 
The properties of PCH are described in Table 4. 
Table 4: 
Pilocarpine Hydrochloride Drug Substance and Drug Product Identifiers 
Established name 
Pilocarpine hydrochloride – parent (+)-pilocarpine 
CAS registry number 
54-71-7 – parent 92-13-7 
Chemical class 
A cholinergic parasympathomimetic agent, pilocarpine 
predominantly binds to muscarinic receptors, thereby inducing 
exocrine gland secretion and stimulating smooth muscle in the 
bronchi, urinary tract, biliary tract, and intestinal tract. When 
applied topically to the eye, this agent stimulates the sphincter 
pupillae to contract, resulting in miosis; stimulates the ciliary 
muscle to contract, resulting in spasm of accommodation; and 
may cause a transitory rise in IOP followed by a more persistent 
fall due to opening of the trabecular meshwork and an increase 
in the outflow of aqueous humor. 
Chemical name 
(3S,4R)-3-ethyl-4-[(3-methylimidazol-4-yl)methyl]oxolan-2-one 
hydrochloride  
or  
(3S-cis)-2(3H)-Furanone, 3-ethyl-dihydro-4-[(1-methyl-1H-
imidazol-5-yl)methyl] mono-hydrochloride 
Molecular formula 
C11H17ClN2O2 – parent C11H16N2O2 
Molecular weight (g/mol) 
244.72 – parent 208.261 
Investigational product 
name/formulation 
Low-Dose Pilocarpine Hydrochloride Ophthalmic Solution 0.4% 
(LDP) 
Aqueous isotonic solution, compounded by Pine 
Pharmaceuticals LLC and released by Iuvo BioScience LLC  
Concentration active 
0.4% 


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Drug used in 
compounding the 
investigational product 
PILOCARPINE HYDROCHLORIDE 4% - pilocarpine 
hydrochloride solution/ drops  
NDC 61314-0206-15 
Manufacturer drug 
product used for 
compounding 
Sandoz, Inc. 
Storage requirements 
Stored at room temperature (15°C to 25°C, 59°F to 77°F) 
Stored in a secured location (locked) with no access for 
unauthorized personnel 
Pilocarpine hydrochloride 0.4% investigational drug is compounded in pharmacy, starting with 
pilocarpine hydrochloride 4% produced by Sandoz, Inc. In addition to the active drug, the 
formulation also comprises the following inactive excipients: benzalkonium chloride NF, 
hypromellose 2910 USP, sodium citrate USP, boric acid NF, sodium chloride USP and sterile 
water for injection. The pH is adjusted with sodium hydroxide NF (10%) and hydrochloric acid 
NF (37%) to achieve a target pH of 3.5 to 5.5. 
Low-Dose Pilocarpine is primary-packaged in multi-use, low-density polyethylene (LDPE) 
dropper bottles for investigational use by subjects, and it is stored at room temperature (15°C to 
25°C, 59°F to 77°F). 
5.4.2. 
 Packaging and Labeling 
Nyxol and its placebo (Treatment 1) are packaged in a 0.5-mL LDPE Blow-Fill-Seal vial 
containing 0.3 mL solution for single-dose use. Five individual vials are included in a molded 
strip. Each strip containing 5 vials is wrapped with a multicolor “rainbow” aluminum foil 
overwrap that has been purged with nitrogen. The foil is impermeable to water and oxygen and 
will be labeled with an investigational label showing the study protocol number and other 
relevant information, including a statement “Caution – New Drug – Limited by Federal (US) 
Law to Investigational Use”.  
Low-Dose Pilocarpine and its placebo (Treatment 2) is provided in 15 mL LDPE, multi-use 
dropper bottles. They will be labeled with an investigational label showing the study protocol 
number and other relevant information, including a statement “Caution – New Drug – Limited 
by Federal (US) Law to Investigational Use.” 
5.4.3. 
 Storage of Study Medication and Dispensing 
Prior to dispensing, all investigational material must be stored in a secure location with strictly 
limited access documented by signature of authorized persons who may dispense investigational 
materials.  
Treatment 1 study medication (Nyxol and its placebo) should be shipped and stored at the study 
site refrigerated (2°C to 8°C, 36°F to 46°F) until provided to the subject; then the subject should 
store Treatment 1 at room temperature (15°C to 25°C, 59°F to 77°F). Treatment 1 study 
medication must not be frozen and must be protected from light.  
Treatment 2 study medication (LDP and its placebo) should be stored at room temperature (15°C 
to 25°C, 59°F to 77°F). Treatment 2 study medication must not be frozen. 


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Prior to dispensing, all investigational study medication must be stored in a secure facility, with 
access limited to the Investigator and authorized staff. 
5.4.4. 
 Study Medication Administration 
Subjects will restart dosing daily with Treatment 1 in the evening immediately prior to Visit 4 
(Stage 2 Day 1). The evening doses prior to Visit 4 and Visit 5 (Stage 2 Day 8) should be as 
close to 12 hours prior to the scheduled time for those visits. At both of these visits, after 
completion of the 12-hour assessments for Treatment 1, subjects will be administered their 
morning dose of Treatment 2 (at Time 0 min) at the study site, and efficacy and safety 
measurements will be made at multiple post-dose timepoints over the next 8 hours.  
5.4.5. 
 Study Medication Accountability 
5.4.5.1. 
Receipt and Disposition of Study Medication 
The Investigator or designee (eg, study coordinator or pharmacist) will maintain a full 
accountability record for both Treatment 1 and Treatment 2 study medications and will be 
responsible for recording the receipt, dispensing, and return of all supplies of the study 
medication and subject diaries using the inventories supplied by Ocuphire. The Investigator or 
designee will account for both Treatment 1 and Treatment 2 study medications and subject 
diaries. The monitor will review dispensing and study medication accountability records during 
site visits and at the completion of the study and note any discrepancies.  
5.4.5.2. 
 Return of Study Medication 
When the study is completed or is terminated by Ocuphire, all study material including used and 
unused study medication bottles will be returned to Ocuphire (or its designee) or destroyed under 
the direction of same. All study medication accounting procedures must be completed before the 
study is considered completed. A final study medication and subject diaries disposition will be 
completed by the study coordinator. 
5.5. Expected Duration of Subject Participation 
The total length of subject participation is approximately 22 to 36 days (accommodating for 
weekends), with 5 clinic visits as summarized below: 
• Screening/Baseline Visit 1 (Stage 1 Day 1). Note that Visit 1 (Screening/Baseline) 
assessments are recommended to be performed on the same day; however, in 
instances in which subjects are unable to complete the full day of assessments for 
Visit 1, subjects may be scheduled to return for the completion of Visit 1 within 
1 week of Screening. If Visit 1 is split over 2 separate days, all Screening assessments 
up to and including randomization must occur on the first day, and then the subject 
should return to complete their post-randomization assessments on the second day.    
• Visit 2 (Stage 1 Day 8) 
• Visit 3 (Stage 1 Day 10 ± 1 day) during the Washout Period of 7 to 14 days 
• Visit 4 (Stage 2 Day 1 [Day 15-22]) 
• Visit 5 (Stage 2 Day 8 [Day 22-29]) 


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The execution of the entire study (first subject screen through last randomized subject 
completed) is expected to take approximately 9 months.  
5.6. Randomization and Procedure for Breaking the Code 
A randomization code for allocating subjects to each treatment group will be prepared by an 
unmasked biostatistician not connected with the study. Subjects will be then randomized 1:1:1:1 
into 1 of 4 treatment arms at Visit 1:  
• Nyxol + LDP 
• Nyxol + LDP vehicle  
• Placebo + LDP 
• Placebo + LDP vehicle  
Randomization will be stratified by iris color (light/dark irides). The number of subjects in each 
irides type should be as close as possible to 1:1, with a minimum of 40% within each stratum. 
5.7. Collection of Data 
Study-specific data that have been outlined in the protocol will be entered into the clinical 
database by individual(s) designated by the Investigator. Data will be verified electronically 
using a series of online programmed edit checks that have been created by the Clinical Data 
Manager and programmed by the Clinical Data Programmer or designee. Data discrepancies will 
be brought to the attention of the clinical team and investigated by the Study Monitor and Site 
Staff. Study Monitors will review and verify all data collected in the electronic Case Report 
Form (eCRF) against any applicable source documentation during remote review or scheduled 
monitoring visits. The Study Monitor will work closely with the Site Staff to address any 
discrepancies that have been found so that proper resolutions can be made and documented in the 
clinical database. An audit trail within the system will track all changes made to the data. 
5.8. Completed Subject 
A completed subject is defined as one who completes all planned procedures through the end of 
Visit 5. 
5.9. Non-completing Subject 
A non-completing subject is defined as one who exits the study by their own volition or at the 
discretion of the Investigator and/or the Medical Monitor prior to completing all of the study 
procedures required in this protocol. Any subject may decide to voluntarily withdraw from the 
study at any time without prejudice.  
5.9.1. 
 Study Medication Discontinuation 
The study medication may be discontinued for the following reasons: 
• Adverse events: Adverse events include clinically significant laboratory 
abnormalities and intercurrent diseases reported by the subject or observed by the 
Investigator, with documentation on the eCRF 
• Death: If a subject dies, the AE(s) that caused the death should be documented on the 
eCRF and be noted as serious and fatal 


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• Disallowed concurrent medication: Any medication not allowed by the protocol 
would be a protocol violation 
• Lack of efficacy: A subject may elect to discontinue participation in the study for a 
perceived lack of efficacy 
• Investigator decision: A subject may be discontinued for reasons other than those 
bulleted previously if the Investigator thinks it is not in the best interest of the subject 
to continue 
• Pregnancy: A subject may be discontinued from study medication if pregnancy 
occurs while on study 
• Other: If there is any other reason for subject discontinuation 
The reason for premature study medication discontinuation should be entered onto the 
appropriate eCRF.  
5.9.2. 
 Reasons for Withdrawal From Study 
• Subject withdraws consent  
• Subject withdraws due to an AE 
• Subject is lost to follow-up  
• Subject withdraws for other reason  
5.9.3. 
 Entire Study Terminated 
The entire study may be terminated by Investigators or Ocuphire. Prompt, written notice of 
reasonable cause to the other party (Ocuphire or Investigators, respectively) is required. Prompt 
notice to the Institutional Review Board (IRB) and to regulatory authorities is also required. 
5.9.4. 
Actions After Discontinuation 
All subjects who discontinue study medication due to a report of an AE must be followed up and 
provided appropriate medical care until their signs and symptoms have remitted or stabilized or 
until medical assessments have returned to acceptable or pre-study limits. 
For any subject who chooses to withdraw consent or who is non-compliant, every possible effort 
should be made by the Investigator to assure the 8-hour measurements are assessed prior to 
discontinuation, in addition to a follow-up telephone call that includes assessments for AEs and 
concomitant medications.  
5.10. Completed Study 
The study is completed when all randomized subjects have completed or discontinued the study, 
all eCRFs have been completed, and all eCRF data have been entered into the database. Final 
database lock will occur after the last randomized subject completes last visit, all data have been 
entered, and all queries are resolved.  
5.11. Procedure After the Completion of the Study 
When the study is completed, the contract research organization (CRO) will provide Ocuphire 
and the Investigator with a brief (ie, 1-3 pages) report, containing a description of the study, the 


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number of subjects enrolled, the number of subjects completed, the number of subjects who 
dropped out and why, efficacy findings, and AEs. 
 
6. SUBJECT INCLUSION AND EXCLUSION CRITERIA 
6.1. Subject Inclusion Criteria 
1. Males or females ≥ 40 and ≤ 64 years of age. 
2. Able to comply with all protocol-mandated procedures independently and to attend all 
scheduled office visits. 
3. Able and willing to give signed informed consent. 
4. Able to self-administer study medication throughout the study period. 
5. Best-corrected distance VA of 0.1 LogMAR (20/25 Snellen equivalent) or better in each 
eye in photopic conditions. 
6. Distance-corrected near VA of 0.4 LogMAR (20/50 Snellen equivalent) or worse but not 
> 0.7 LogMAR (20/100 Snellen equivalent) in photopic conditions in each eye and 
binocularly. 
7. For subjects who depend on reading glasses or bifocals, binocular best-corrected near VA 
is 0.1 LogMAR (20/25 Snellen equivalent) or better. 
8. Photopic PD of ≥ 3 mm in either eye. 
6.2. Subject Exclusion Criteria 
Subjects excluded from the study will be individuals with the following characteristics: 
Ophthalmic (in either eye): 
1. Use of any topical prescription (including Vuity) or over-the-counter (OTC) ophthalmic 
medications of any kind within 7 days of Screening until study completion, with the 
exception of lid scrubs with OTC products (eg, OCuSOFT® lid scrub, SteriLid®, baby 
shampoo, etc.) and artificial tears as specified in Exclusion #2 below. 
2. Use of any OTC artificial tears (preserved or unpreserved) during Visit days or 15 min 
before or after instillation of Treatment 1 or Treatment 2. 
3. Current use of any dry eye product such as topical ophthalmic therapy for dry eye (eg, 
generic cyclosporine, Restasis, Xiidra, Cequa, and Eysuvis) or intranasal dry eye product 
(eg, Tyrvaya) or other devices.  
4. Tear break-up time of < 5 seconds or corneal fluorescein staining Grade ≥ 2 in the inferior 
zone or Grade ≥ 1 in the central zone using the National Eye Institute scale.  
5. Clinically significant ocular disease (eg, cataract, glaucoma, corneal edema, uveitis, retinal 
degeneration, loss of visual field, or any macular pathology) that might interfere with the 
study as deemed by the Investigator. 


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6. Recent or current evidence of ocular infection or inflammation in either eye (such as 
current evidence of clinically significant blepharitis, conjunctivitis, keratitis, etc.). 
Subjects must be symptom free for at least 7 days. 
7. Any history of herpes simplex or herpes zoster keratitis. 
8. Known allergy, hypersensitivity, or contraindication to any component of the 
phentolamine, pilocarpine, or vehicle formulations.  
9. Prior participation in a study involving the use of Nyxol for the treatment of presbyopia. 
10. History of cauterization of the punctum or punctal plug (silicone or collagen) insertion or 
removal. 
11. Ocular trauma within 6 months prior to Screening.  
12. Ocular surgery or any ocular laser treatment within 6 months prior to Screening. 
13. Subjects with surgical monovision, multifocal or extended depth of focus IOLs are 
excluded (monofocal IOLs are accepted if in place > 6 months prior to Screening). 
14. History of any traumatic (surgical or nonsurgical) or nontraumatic condition affecting the 
pupil or iris (eg, irregularly shaped pupil, neurogenic pupil disorder, iris atrophy, 
iridotomy, iridectomy, iritis, etc.). 
15. Contact lens wear on the day of any study visit, and contact lenses must be removed for 
home dosing and for at least 10 minutes following dosing. 
Systemic: 
16. Known hypersensitivity or contraindication to alpha- and/or beta-adrenoceptor antagonists 
(eg, chronic obstructive pulmonary disease or bronchial asthma; abnormally low BP or 
HR; second- or third-degree heart blockage or congestive heart failure). 
17. Known hypersensitivity or contraindication to any systemic cholinergic 
parasympathomimetic agent. 
18. Clinically significant systemic disease (eg, uncontrolled diabetes, myasthenia gravis, 
cancer, hepatic, renal, endocrine, or cardiovascular disorders) that might interfere with the 
study as deemed by the judgment of the Investigator. 
19. Initiation of treatment with, or any changes to, the current dosage, drug, or regimen of any 
systemic adrenergic or cholinergic drugs within 7 days prior to Screening or during the 
study; however, Flomax (tamsulosin) is specifically excluded. 
20. Participation in any investigational study within 30 days prior to Screening. 
21. Females of childbearing potential who are pregnant, nursing, planning a pregnancy, or not 
using a medically acceptable form of birth control. Acceptable methods include the use of 
at least one of the following: intrauterine device (IUD), hormonal (oral, injection, patch, 
implant, ring), barrier with spermicide (condom, diaphragm), or abstinence. A female is 
considered to be of childbearing potential unless she is 1 year postmenopausal or 3 months 
post-surgical sterilization. All females of childbearing potential, including those with 
post−tubal ligation, must have a negative urine pregnancy test result at Visit 1 
(Screening/Baseline).  


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22. Resting HR outside the range of 50 to 110 beats per minute following at least a 5-min rest 
period in the sitting position at Visit 1 (Screening/Baseline). Heart rate may be repeated 
only once if outside the specified range, following another 5-min rest period in the sitting 
position. 
23. Hypertension with resting diastolic BP > 105 mmHg or systolic BP > 160 mmHg 
following at least a 5-min rest period in the sitting position at Visit 1 (Screening/Baseline). 
Blood pressure may be repeated only once if outside the specified range, following 
another 5-min rest period in the sitting position. 
 
7.  TREATMENT OF SUBJECTS 
Approximately 320 eligible subjects will be randomized into 1 of 4 treatment arms:  
• Nyxol + LDP 
• Nyxol + LDP vehicle 
• Placebo + LDP 
• Placebo + LDP vehicle 
Subjects will be randomized 1:1:1:1 into the above groups. Randomization will be stratified by 
iris color (light/dark irides). The number of subjects in each irides type should be as close as 
possible to 1:1, with a minimum of 40% within each stratum. 
7.1. Treatment Adherence 
Following the completion of Visit 1 (Stage 1 Day 1), subjects will be dispensed Treatment 1 
study drug for dosing at home in the evenings and a Treatment 1 subject diary to document 
dosing.  
At the end of Visit 4 (Stage 2 Day 1), subjects will be dispensed Treatment 2 study drug for 
dosing at home in the mornings and a Treatment 2 subject diary to document dosing.  
Subjects must bring their used and unused study medications and subject diaries with them to 
Visit 2 (Stage 1 Day 8), Visit 4 (Stage 2 Day 1), and Visit 5 (Stage 2 Day 8). 
All subjects must be instructed on the importance of following the once-daily dosing 
regimen for each Treatment 1 and Treatment 2 and maintaining their dosing diaries.  
Treatment adherence will be measured by counting the used and unused vials at each study visit 
and by reviewing dosing in the subject diaries for both Treatments 1 and 2.  
7.2. Concomitant Medications 
As noted in the exclusion criteria (Section 6.2), use of any topical prescription or OTC 
ophthalmic medications of any kind within 7 days of Screening is prohibited, with the exception 
of lid scrubs with OTC products (eg, OCuSOFT® lid scrub, SteriLid®, baby shampoo, etc.) and 
artificial tears, except the use of any OTC artificial tears (preserved or unpreserved) during Visit 
days or 15 min before or after instillation of Treatment 1 or Treatment 2. 
Additionally, initiation of treatment with or any changes to the current dosage, drug, or regimen 
of any systemic adrenergic or cholinergic drugs (Appendix 3) within 7 days prior to Screening, 


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or during the study, is prohibited. However, a subject can be treated with a systemic 
adrenoceptor antagonist, for example, as long as the particular agent and its dose and regimen 
had been consistent for the 7 days prior to Screening, and there was no reason to believe that 
alteration would be necessary at some point later during the study. However, Flomax 
(tamsulosin) is specifically excluded 
If there is any question about whether a medication is acceptable, the Medical Monitor should 
be consulted before proceeding. 
Use of all medications should be documented on the appropriate eCRF. Investigators are 
encouraged to contact the Medical Monitor for any questions regarding allowed medications. 
Judgment of continued study participation by the subject, and inclusion of this subject’s 
subsequent visits in the safety and efficacy analysis, will be made by Ocuphire. 
All medications that the subject has taken within 30 days prior to the Screening Visit and during 
the study will be recorded in the eCRF. The name of the drug, dose, route of administration, 
duration of treatment, and indication will be recorded for each medication. For combination 
products (eg, Contac®, Cosopt®) and non-combination products, the generic name is desired. The 
use of routine ophthalmic diagnostic pharmaceutical agents (eg, fluorescein and local anesthetic) 
will be allowed and should be documented. Any change in dosing parameters should also be 
recorded in the eCRF. 
 
8. ASSESSMENT OF EFFICACY 
8.1. Specification of the Efficacy Parameters 
The primary efficacy endpoint is the percent of subjects with ≥ 15 letters of improvement in 
photopic binocular DCNVA and with < 5 letters of loss in photopic binocular BCDVA at 30 min 
post-LDP/vehicle comparing Nyxol + LDP to placebo + LDP vehicle at Visit 5 (Stage 2 Day 8). 
This comparison will be analyzed first using a hierarchical testing method. 
Key secondary efficacy endpoints can be found in Section 5.1.  
All of the efficacy endpoints will be analyzed overall and by light/dark irides at all timepoints. 
All of the efficacy endpoints will also be analyzed by mITT and PP Populations. 
8.2. Assessing, Recording, and Analyzing of Efficacy Parameters 
Visual acuity and PD assessments will be measured as described in Section 5.1. 
The subject should be in the same room for all assessments, and every effort will be made to 
have the same person perform the measurements at all visits. 
The timing for recording efficacy parameters may be found in Table 1 and Table 2. 
8.2.1. 
Screening/Baseline Visit 1 (Stage 1 Day 1)  
Individuals who are potential subjects will be identified by the study center to schedule the 
Screening Visit. If a subject successfully completes their Screening Assessments, then Visit 1 
becomes the Baseline Visit. Note that Visit 1 (Screening/Baseline) assessments are 
recommended to be performed on the same day; however, in instances in which subjects are 
unable to complete the full day of assessments for Visit 1, subjects may be scheduled to return 


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for the completion of Visit 1 within 1 week of Screening. If Visit 1 is split over 2 separate days, 
all Screening assessments up to and including randomization must occur on the first day, and 
then the subject should return to complete their post-randomization assessments on the second 
day.    
Once a subject arrives at the study center, a member of the Site Staff will interview the 
individual as to their qualifications for participation in the study, and if the subject wishes to 
continue, the informed consent form (ICF) is signed, and a subject number is assigned.  
The start of Screening includes an explanation of the study, a medical and ophthalmic history, 
demographics, HR and BP, urine pregnancy test (females of childbearing potential) and 
eligibility assessments.  
The subject will then undergo manifest refraction and near add, and several VA measurements, 
including photopic BCDVA as well as mesopic and photopic DCNVA (Inclusion Criteria #5-7). 
In addition, subjects will undergo measurement of photopic DCIVA and mesopic BCDVA 
The Screening assessment will also include an ophthalmic examination that includes assessment 
of PD, biomicroscopy, dry eye examination with tear break-up time testing and corneal 
fluorescein staining, IOP measurement (using a Tono-Pen or Goldmann Applanation tonometry), 
and direct or indirect ophthalmoscopy without dilation.  
If all eligibility criteria are met, the subject will be randomized into the study.  
At Visit 1 (Stage 1 Day 1), eligible subjects will be randomized, and the first dose of Treatment 1 
will be administered at the study site at Time 0 min. Efficacy and safety measurements are made 
before dosing with Treatment 1 (Time 0 min) and at multiple post-dose timepoints from 30 min 
to 8 hours. If Visit 1 is split over 2 separate days, all Screening assessments up to and including 
randomization must occur on the first day, and then the subject should return to complete their 
post-randomization assessments in the following order on the second day (ie, confirm 
concomitant medications, subject questionnaire, in-office Treatment 1 dosing, ocular tolerability, 
followed by all other assessments at time points 0.5, 1, 3, 5, and 8 hours post-treatment). The 
following assessments are conducted during Visit 1: 
Prior/Concomitant medication 
Heart rate and BP 
Pupil diameter (photopic and mesopic) 
Distance-corrected near VA (photopic and mesopic), BCDVA (photopic and mesopic), 
DCIVA (photopic); all VA measurements will be made binocularly, except selected 
measurements at 0 min (DCNVA photopic, BCDVA photopic and mesopic), which will 
be measured binocularly followed by OD and OS measurements 
Biomicroscopy 
Intraocular pressure 
Ophthalmoscopy 
Ocular tolerability 
Subject questionnaire 


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Adverse events 
Treatment 1/Diary dispensing 
8.2.2. 
Visit 2 (Stage 1 Day 8)  
At Visit 2 (Stage 1 Day 8), efficacy and safety measurements will be made at 12 hours after the 
dose of Treatment 1 study medication (12-hour endpoint): 
Heart rate and BP  
Urine pregnancy test (females of childbearing potential) 
Concomitant medications  
Pupil diameter (photopic and mesopic) 
Distance-corrected near VA (photopic and mesopic), BCDVA (photopic and mesopic), 
DCIVA (photopic); all VA measurements will be made binocularly, except selected 
measurements at 0 min (DCNVA photopic) and 8 hours (DCNVA photopic, BCDVA 
photopic and mesopic), which will be measured binocularly followed by OD and OS 
measurements 
Biomicroscopy 
Intraocular pressure 
Ophthalmoscopy 
Ocular tolerability 
Subject questionnaire 
Adverse events 
After completion of these assessments, the subject will take no study drug for the next 7 to 14 
days (Washout Period), after which Stage 1 is complete.  
8.2.3. 
Visit 3 (Stage 1 Day 10 ± 1 day) – During Washout Period  
Measurements will be made on Visit 3 (Stage 1 Day 10 ± 1 day) during the Washout Period to 
assess the absence of residual treatment effects: 
 
Concomitant medications  
Pupil diameter (photopic and mesopic) 
Distance-corrected near VA (photopic), BCDVA (photopic and mesopic) 
Adverse events 
At the conclusion of the Washout Visit measurements, Stage 1 of the study is complete. 
8.2.4. 
Visit 4 (Stage 2 Day 1 [Day 15-22]) 
Visit 4 occurs after completion of the Washout Period. During Visit 4, efficacy and safety 
measurements will be made approximately 12 hours after the dose of Treatment 1 study 
medication and at multiple timepoints after the dose of Treatment 2 (at Time 0 min): 
Urine pregnancy test (females of childbearing potential) 


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Concomitant medications  
Pupil diameter (photopic and mesopic) 
Distance-corrected near VA (photopic and mesopic), BCDVA (photopic and mesopic), 
DCIVA (photopic). All VA measurements will be made binocularly 
Ocular tolerability 
Subject questionnaire 
Adverse events 
Treatment 2/Diary dispensing 
8.2.5. 
 Visit 5 (Stage 2 Day 8 [Day 22-29]) 
Visit 5 occurs 7 days after Visit 4. During Visit 5, efficacy and safety measurements will be 
made approximately 12 hours after the dose of Treatment 1 study medication and at multiple 
timepoints after the dose of Treatment 2 (at Time 0 min) [note 30 minute measurements are key 
secondary endpoints]: 
Heart rate and BP  
Urine pregnancy test (females of childbearing potential) 
Concomitant medications  
Pupil diameter (photopic and mesopic) 
Distance-corrected near VA (photopic and mesopic), BCDVA (photopic and mesopic), 
DCIVA (photopic). All VA measurements will be made binocularly except for 0.5 hours 
(DCNVA photopic) and 8 hours (DCNVA photopic, BCDVA photopic and mesopic), 
which will be measured binocularly followed by OD and OS measurements 
Biomicroscopy 
Intraocular pressure 
Ophthalmoscopy 
Ocular tolerability 
Subject questionnaire 
Adverse events 
Collection of used/unused Treatment 1 and Treatment 2 and subject diaries  
8.2.6. 
Unscheduled Visits 
An unscheduled visit may be any visit to the Investigator other than the specific visits requested 
in the protocol as possibly required for the subject’s ophthalmic condition. The Investigator will 
perform all procedures necessary to evaluate the subject at these visits and record any AEs in the 
eCRF. 
As noted in Section 5.9.4, every possible effort should be made by Investigators to assure that 
subjects who discontinue early from the study have a telephone follow-up that includes 
assessments of AEs and concomitant medications. 


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8.3.  Visit Variation 
Visit 3 may occur 1 to 3 days after Visit 2; Visit 4 may occur 8 to 15 days after completion of 
Visit 1; and Visit 5 may occur 22 to 36 days after Visit 1.  
 
9. ASSESSMENT OF SAFETY 
9.1. Specification of Safety Parameters 
The primary safety measures are subject-evaluated ocular tolerability and AEs. Other safety 
measures include biomicroscopy, ophthalmoscopy, BCDVA, IOP, subject questionnaire, and 
systemic safety (as measured by HR and BP). Urine pregnancy tests for females of childbearing 
potential will also be conducted. 
The assessment of safety will be evaluated by: 
• Ocular tolerability measured on a 4 point scale 
• Biomicroscopy of the anterior segment including evaluation of lids, lashes, cornea, 
conjunctiva, iris, lens, and anterior chamber. Following topical fluorescein 
administration, the integrity of the cornea should be evaluated by fluorescein staining 
• Ophthalmoscopy (direct or indirect) without dilation to evaluate the vitreous and 
posterior pole 
• Best-corrected distance VA 
• Intraocular pressure performed with a Tono-Pen or Goldmann Applanation tonometry 
• Subject questionnaire 
• Heart rate and BP (as per the site’s normal equipment and procedures) 
• Adverse events 
9.2. Assessing, Recording, and Analyzing Safety Parameters 
The timing for recording safety parameters may be found in Table 1 and Table 2. 
9.3. Adverse Events and Serious Adverse Events 
All AEs and SAEs that occur following consent and until the final study visit should be collected 
and recorded on the AE or SAE eCRF page. Only TEAEs will be summarized (Section 9.3.1). 
All AEs/adverse reactions occurring during the study (ie, once the subject has signed the ICF) 
must be documented, regardless of the assumption of causal relationship, on the respective 
eCRF. All TEAEs/treatment-emergent adverse reactions must be documented from the time the 
subject receives the study medication until the subject’s participation in the study has been 
completed. If a subject has ongoing AEs/adverse reactions at the time of study completion or 
discontinuation from the study, the ongoing AEs/adverse reactions must be followed up and 
provided appropriate medical care until the signs and symptoms have remitted or stabilized or 
until medical assessments have returned to acceptable or pre-study limits. 
Documentation of AEs/adverse reactions includes start date and end date, severity, relationship 
to study medications, action(s) taken, seriousness, and outcome.  


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9.3.1. 
 Adverse Event Definitions 
The following definitions of terms apply to this section: 
Adverse event. An AE is any untoward medical occurrence associated with the use of a study 
medication in humans, whether or not it is considered drug related. An AE can therefore be any 
unfavorable and unintended sign (including abnormal laboratory findings), symptom, or disease 
temporally associated with the use of the study medication, whether or not related to the study 
medication. Study medication includes the investigational drug under evaluation and the 
comparator product or vehicle placebo that is given or administered during any phase of the 
study.  
Medical conditions/diseases present before starting the investigational treatment are only 
considered AEs if they worsen after starting the investigational treatment. Abnormal test results 
constitute AEs only if they induce clinical signs or symptoms, are considered clinically 
significant, or require therapy. 
The occurrence of AEs should be sought by open-ended questioning of the subject at each visit 
during the study. At each clinic assessment/visit, study personnel should ask the following 
question: “Have you had any problems since your last assessment/visit?” Adverse events also 
may be detected when they are volunteered by the subject during or between visits or through 
study assessments. 
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 Ocuphire, its occurrence places the subject at immediate risk of death. It does not 
include an AE or suspected adverse reaction that, had it occurred in a more severe form, might 
have caused death.  
Serious adverse event or serious suspected adverse reaction. An AE or suspected adverse 
reaction is considered ‘‘serious’’ if, in the view of either the Investigator or Ocuphire, it results 
in any of the following outcomes:  
• Death 
• Life-threatening AE 
• Inpatient hospitalization or prolongation of existing hospitalization 
• Persistent or significant incapacity or substantial disruption of the ability to conduct 
normal life functions  
• Congenital anomaly/birth defect  
• Other medically important serious event 
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.  
Treatment on an outpatient emergency basis that does not result in hospital admission, or a 
hospitalization that is elective or is a preplanned treatment for a pre-existing condition that is 


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unrelated to the medication under study and has not worsened since the start of the study is not 
considered an SAE. 
Suspected adverse reaction means any AE for which there is a reasonable possibility that the 
drug caused the AE. For the purposes of Investigational New Drug safety reporting, “reasonable 
possibility” means there is evidence to 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.  
Unexpected adverse event or unexpected suspected adverse reaction. An AE or suspected 
adverse reaction is considered “unexpected” if it is not listed in the IB or is not listed at the 
specificity or severity that has been observed. “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. 
The study medication relationship for each AE/adverse reaction should be determined by the 
Investigator using these explanations: 
• Not related 
• Unlikely related 
• Possibly related 
• Probably related 
• Definitely related 
• Unknown 
Unless the relationship is considered to be “Not related” or “Unlikely related” and there is any 
valid reason, even if undetermined, for suspecting a possible cause-and-effect relationship 
between the study medication and the occurrence of the AE, then the AE should be considered 
“related.”  
If the relationship between the AE/SAE and the investigational product is determined by 
Ocuphire to be anything other than “Not related” or “Unlikely related,” the event will be 
considered to be related to the investigational product for the purposes of expedited regulatory 
reporting. 
Severity of an AE is defined as a qualitative assessment of the level of discomfort of an AE as is 
determined by the Investigator or reported to him/her by the subject. The assessment of severity 
is made irrespective of study medication relationship or seriousness of the event and should be 
evaluated according to the following scale: 
• 1 = Mild: Present, but not distressing, and no disruption of normal daily activity 
• 2 = Moderate: Discomfort sufficient to reduce or affect normal daily activity 
• 3 = Severe: Incapacitating, with inability to work or perform normal daily activity 
A change in severity for a reported AE will require an end date for the previous severity and a 
new start and end date for the new severity. For example, a change in severity may go from mild 


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to severe or from severe to moderate. In either case, the start or end times/dates should be 
recorded.  
The term “severe” is used to describe the intensity of an event/reaction; the event/reaction itself 
may be of relatively minor medical significance (such as a severe headache). This is not the same 
as a “serious” AE, which is based on a subject/event outcome or action criteria usually associated 
with events that pose a threat to the subject’s life or vital functions. “Seriousness” (NOT 
severity) serves as a guide for defining regulatory reporting obligations. 
Should a pregnancy occur, it must be reported and recorded on the pregnancy form and emailed 
to VEGA2_Safety@Oculoscr.com, as well as in the eCRF. Pregnancy in itself is not regarded as 
an AE unless there is a suspicion that an investigational product may have interfered with the 
effectiveness of a contraceptive medication. 
The outcome of all pregnancies (spontaneous miscarriage, elective termination, normal birth, or 
congenital abnormality) must be followed up and documented even if the subject was 
discontinued from the study. 
Action taken in response to an AE is coded as: 
• Dose increased: An indication that a medication schedule was modified by addition; 
either by changing the frequency, strength, or amount 
• Dose not changed: An indication that a medication schedule was maintained 
• Dose reduced: An indication that a medication schedule was modified by subtraction, 
either by changing the frequency, strength, or amount 
• Dose interrupted: An indication that a medication schedule was modified by 
temporarily terminating a prescribed regimen of medication 
• Drug withdrawn: An indication that a medication schedule was modified through 
termination of a prescribed regimen of medication 
• Not applicable: Determination of a value is not relevant in the current context 
• Unknown: Not known, not observed, not recorded, or refused 
Additional other action taken: 
• Concomitant medication 
• Hospitalization 
Outcome of an AE is coded as: 
• Fatal: The termination of life as a result of an AE 
• Not recovered/not resolved: One of the possible results of an AE outcome that 
indicates that the event has not improved or the subject has not recuperated 
• Recovered/resolved: One of the possible results of an AE outcome that indicates that 
the event has improved or the subject has recuperated 


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• Recovered/resolved with sequelae: One of the possible results of an AE outcome 
where the subject recuperated but retained pathological conditions resulting from the 
prior disease or injury 
• Recovering/resolving: One of the possible results of an AE outcome that indicates 
that the event is improving 
• Unknown: Not known, not observed, not recorded, or refused 
In previous clinical studies of Nyxol, the most frequently reported AE was conjunctival 
hyperemia. Investigators are cautioned to use the appropriate verbatim term on the AE form to 
describe this observation:  
• Redness related to instillation that is transient (ie, is no longer present 2 hours after 
instillation) = “conjunctival erythema upon instillation” 
• Redness that is NOT transient (ie, is present 2 hours after instillation) = “conjunctival 
hyperemia” 
Expedited reporting of serious and unexpected adverse events: All SAEs (related and 
unrelated) will be recorded following subject signature of the ICF consent and until the 
completion of Visit 5. Any SAEs “suspected” to be related to the study medication and 
discovered by the Investigator at any time after the study must be reported. 
Any SAE that occurs must be reported to the CRO within 24 hours of its occurrence or within 
24 hours of learning of its occurrence. Recurrent episodes, complications, or progression of the 
initial SAE must be reported to the CRO as follow-up to the original episode within 24 hours of 
the Investigator receiving the information. Information about all SAEs will be collected and 
recorded on the SAE eCRF. All pertinent medical records and information collected during the 
treatment and follow-up of the subject should be maintained at the site with a copy emailed 
to VEGA2_Safety@Oculoscr.com. The Investigator must assess the SAE relationship and 
complete the SAE eCRF. The CRO may request additional information. Follow-up information 
(eg, discharge summary) will be retained in the subject’s chart and a copy will be emailed to 
VEGA2_Safety@Oculoscr.com. In addition, all SAEs should be recorded on the AE eCRF page 
with the serious question marked “Yes.” 
It is the Investigator’s responsibility to notify the approving IRB of any SAEs on a timely basis, 
as instructed by Ocuphire following Ocuphire’s determination of causality. All subjects who 
experience an SAE should be followed clinically and undergo the appropriate diagnostic 
evaluations until stabilization or resolution of the event. Ocuphire will report all SAEs to the 
FDA on the appropriate schedule, depending if the event is drug related or not drug related, 
expected, or unexpected (based on the available information in the IB). 
Any death occurring during the study and follow-up period must be reported as an SAE. For any 
death occurring through the end of the study, regardless of the degree of relationship to study 
medication, the SAE resulting in the death must be reported to the CRO. A death occurring after 
completion of the study that is not reasonably associated with study medication administration 
does not require completion of the SAE form. 


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9.3.2. 
 Follow-Up of Subjects After Adverse Events 
If an AE/adverse reaction occurs, the Investigator will institute support and/or treatment as 
deemed appropriate. All SAEs ongoing at the time of the last visit or discontinuation from the 
study will be followed up until the AE/adverse reaction is resolved or stabilized, the subject is 
lost to follow-up, or there is other resolution to the event.  
 
10. STATISTICS 
10.1. Sample Size 
Using α = 0.05 significance and a two-tailed test, 300 total subjects (150 subjects each in the 
Nyxol and placebo arms) who are evaluable for efficacy will provide greater than 80% power to 
detect a difference of 17% between treatments in percent of subjects with ≥ 15 letters of 
improvement in photopic binocular DCNVA and with < 5 letters of loss in photopic binocular 
BCDVA from Baseline at 12 hours post-dose at Visit 2 (Stage 1 Day 8). This calculation is based 
on a 12% placebo effect and a 29% Nyxol treatment effect. Excess power will be employed for a 
hierarchical analysis of selected secondary efficacy endpoints. 
Subjects will be randomized 1:1:1:1 to Nyxol + LDP, Nyxol + LDP vehicle, Placebo + LDP, and 
Placebo + LDP vehicle respectively. As a result, subjects will be randomized 1:1 to Treatment 1 
(Nyxol or placebo). 
An additional 20 subjects will be enrolled to account for dropouts, for a total of approximately 
320 subjects to be randomized. 
A diagram outlining the study design can be found in Appendix 4. 
10.2. Analysis Populations 
Modified Intention-to-Treat Population: The mITT Population will include all randomized 
subjects who received at least 1 drop of Treatment 1. The mITT Population will be used to 
analyze the primary endpoint as well as other efficacy endpoints. 
Per Protocol Stage 1 Population: The PP Stage 1 (PPS1) Population will include all subjects in 
the mITT Population who receive 1 drop of Treatment 1 the day prior to Visit 2, have binocular 
DCNVA and BCDVA in photopic conditions at Baseline (Visit 1) and at 12 hours post-dose at 
Visit 2, and have no major protocol deviations considered to have significant impact on 
treatment outcome in Stage 1. The PPS1 Population will be used for analysis of select efficacy 
endpoints as specified in the SAP.  
Per Protocol Stage 2 Population: The PP Stage 2 (PPS2) Population will include all subjects in 
the PPS1 Population who receive 1 drop of Treatment 1 the day prior to Visit 4, receive 1 drop of 
Treatment 2 at Visit 4, have binocular DCNVA and BCDVA in photopic conditions for at least 1 
timepoint at Visit 4 after Treatment 2 dosing, and have no major protocol deviations considered 
to have significant impact on treatment outcome in Stage 2. The PPS2 Population will be used 
for analysis of select efficacy endpoints as specified in the SAP.  
All Randomized Population (ARP): The ARP will include all randomized subjects. This 
population is also known as the Intention-to-Treat (ITT) Population. The ARP will be used in 
confirmatory efficacy analyses. 


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Safety Population (SP): The SP will include all randomized subjects who have received at least 
1 drop of study treatment (Treatment 1 or Treatment 2). The SP will be used to summarize safety 
variables. 
Per Protocol Populations include PPS1 and PPS2 throughout this document. 
10.3. Statistical Methods  
10.3.1.  General Considerations 
All continuous variables will be summarized by treatment and timepoint (as applicable) using 
descriptive statistics (n, mean, median, standard deviation, minimum, and maximum). All 
categorical variables will be summarized by treatment and timepoint (as applicable) using 
frequency counts and percentages.  
All study data will be listed by treatment, subject, and timepoint (as applicable). 
All statistical tests will be performed using a significance level of 5% (two-tailed). The p-values 
for the analysis of secondary efficacy endpoints will be considered descriptive. 
10.3.2.  Demographic and Baseline Characteristics 
Demographic and Baseline characteristics, such as age, race, and sex, will be summarized by 
treatment arm using the mITT Population, PP Populations, the ARP, and the SP. These data will 
also be provided in by-subject listings. 
10.3.3.  Subject Disposition 
Subject disposition, including randomization, completion, and withdrawal from the study, will be 
summarized using the ARP. These data will also be provided in by-subject listings. 
10.3.4.  Medical History and Prior/Concomitant Medications 
Medical history will be coded using the latest version of Medical Dictionary for Regulatory 
Activities (MedDRA, Version 25 or higher) and will be summarized by treatment arm using the 
mITT Population.  
Prior medications (medications with an end date prior to the date of randomization) and 
concomitant medications (medications with a start or end date after the date of randomization) 
will be coded using World Health Organization Drug Dictionary (March 2022 or later edition 
available at study start) and will both be summarized by treatment arm using the SP. 
Medical history and prior and concomitant medications will also be provided in by-subject 
listings.  
10.3.5.  Analysis of Efficacy 
Efficacy will be assessed using the mITT and PP Populations, with subjects included in the 
treatment arm into which they were randomized. For the analysis of the primary efficacy 
endpoint, appropriate imputation techniques will be performed for missing observations if 
applicable; details will be provided in the study SAP. The primary comparison of interest is 
Stage 2 Nyxol + LDP versus placebo + LDP vehicle for the mITT Population. Confirmatory 
analyses may be performed using the ARP, with imputation performed for missing data. For the 
analysis of the secondary efficacy endpoints, only observed case data will be used. If warranted, 


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confirmatory analyses using the ARP with imputation for missing data will also be performed for 
the secondary efficacy endpoints. 
 For all efficacy endpoints, Baseline values are defined as the latest values taken prior to 
administration of Treatment 1, which will usually be the screening assessment values on Visit 1. 
All efficacy data will be summarized by treatment group, study visit, and timepoint, as 
appropriate. 
The primary efficacy endpoint is the percent of subjects with ≥ 15 letters of improvement in 
photopic binocular DCNVA and with < 5 letters of loss in photopic binocular BCDVA at 30 min 
post-LDP/vehicle comparing Nyxol + LDP to placebo + LDP vehicle at Visit 5 (Stage 2 Day 8). 
The primary efficacy endpoint will be analyzed using a logistic regression model with treatment, 
light/dark irides, and Baseline DCNVA as a covariate. The percentage of subjects in each 
treatment arm meeting the criteria, the odds ratio (OR) with 95% confidence interval (CI), and p-
value will be provided. The analysis will be performed using the mITT and PP Populations, with 
subjects included in their randomized treatment regardless of the treatment they actually 
received. A sensitivity analysis will be completed for the primary efficacy endpoint using the 
same model but excluding the light/dark irides factor. 
Secondary efficacy endpoints are indicated in Section 5.1. Each of the continuous secondary 
efficacy endpoints will be analyzed using analysis of covariance (ANCOVA) with change from 
Baseline as the dependent variable, treatment and light/dark irides as factors, and the respective 
Baseline value included as the covariate. Each ANCOVA will be performed using the mITT and 
PP Populations, with subjects included in their randomized treatment regardless of the treatment 
they actually received. The output from each ANCOVA will include the least-squares mean 
(LSM) and standard error for all treatment groups, along with the placebo-corrected LSM for 
each treatment difference, its 95% CI, and associated p-value.  
All continuous secondary endpoints derived from VA assessments, such as change in DCNVA, 
DCIVA, and BCDVA, will be analyzed using ETDRS letters correctly read.  
For each of the secondary endpoints related to percent of subjects achieving certain criteria, the 
analysis will be performed using a logistic regression model with treatment, light/dark irides, and 
the respective Baseline as a covariate. For each analysis, the percentage of subjects in each 
treatment arm meeting the criteria, the OR with 95% CI, and p-value will be provided. For these 
endpoints, the mITT and PP Populations will be used, with subjects included in their randomized 
treatment regardless of the treatment they actually received. 
In addition, each secondary efficacy endpoint will be analyzed by light/dark irides using the 
same model indicated above but without irides as a factor, as appropriate. Other subgroups, such 
as age, sex, and race, may be analyzed as well.  
Select efficacy endpoints will be included in a hierarchical analysis, which will be specified in 
the SAP. The first endpoint in the hierarchy will be the primary efficacy endpoint, followed by 
key secondary endpoints. 


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10.3.6.  Analysis of Safety 
Safety will be assessed using the SP, with subjects included in the treatment arm they actually 
received, regardless of their randomized treatment. Observed case data will be used; no 
imputation will be performed for missing safety data.  
For HR and BP, Baseline is defined as the Screening value. Heart rate and BP values and change 
from Baseline in the values will be summarized by treatment arm and select timepoints.  
For IOP, Baseline is defined as the Screening value. Observed values and change from Baseline 
in IOP at select timepoints will be summarized by treatment arm for the study eye and the fellow 
eye. 
For BCDVA, the number of subjects who lose ≥ 5 letters from Baseline will be summarized by 
treatment arm and select timepoints. 
Ocular tolerability values will be summarized by treatment arm at select timepoints.  
Verbatim descriptions of AEs will be coded using MedDRA. Only TEAEs (those that occur after 
the first dose of study medication or increasing in severity after initiation of study medication) 
will be summarized. Treatment-emergent AEs and SAEs will be summarized by treatment group, 
by system organ class (SOC), severity, and relationship to study medication. Deaths, withdrawal 
from study medication due to AEs, and withdrawal from the study due to AEs will each by 
summarized by treatment group. Note that in MedDRA, ocular events are coded to the SOC of 
“special senses.” Thus, using SOC in the summaries will provide a separation of ocular and non-
ocular AEs. 
All safety data will be provided in by-subject listings.  
10.4. Procedure for Accounting for Missing, Unused, or Spurious Data 
For the summarization and analysis of efficacy data, the focus will be on observed case data 
only. As appropriate, confirmatory efficacy analyses will be performed using imputation for 
missing data; details of the imputation, if performed, will be included in the study SAP. For the 
summarization of safety data, observed case data only will be used. 
10.5. Procedure for Reporting Deviations From the Statistical Plan 
Any deviations from the SAP will be described and a justification given in the final Clinical 
Study Report.  
 
11. DIRECT ACCESS TO SOURCE DATA AND DOCUMENTS 
The Investigator will permit study-related monitoring visits, audits, IRB review, and regulatory 
inspection(s) by providing direct access to source data and documents. 
 
12. QUALITY CONTROL AND QUALITY ASSURANCE 
The progress of the study will be monitored by onsite, written, and telephone communications 
between personnel at the Investigator’s site and the Medical Monitor. Should the COVID-19 
pandemic restrict monitors from traveling to a site, remote review will be conducted to the extent 


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possible, while still ensuring the study is monitored appropriately per applicable regulations and 
guidelines. The Investigator will allow Ocuphire, the Study Monitor, and the Medical Monitor to 
inspect all eCRFs, subject records (source documents), signed ICFs, records of study medication 
receipt, storage, preparation, disposition, and regulatory files related to this study.  
 
13. ETHICAL CONSIDERATIONS AND GOOD CLINICAL PRACTICE 
COMPLIANCE  
13.1. Good Clinical Practice Compliance 
The proposed study is subject to all applicable governmental rules and regulations concerning 
the conduct of clinical trials on human subjects. This includes, but is not necessarily limited to, 
the approval of IRBs, the Helsinki Declaration, US FDA law, International Council for 
Harmonisation (ICH) GCP guidelines, obtaining prospective informed consent, monitoring of the 
conduct of the study and the completeness of the eCRFs by Ocuphire or its designee(s), and 
appropriate record retention by the Investigator.  
13.2. Institutional Review Board  
This protocol, materials used to recruit subjects, and materials used to document consent must be 
approved by the IRB prior to initiation of the study. Written IRB approval must adequately 
identify the protocol and informed consent. In addition to approving the protocol, the IRB must 
also approve the subject information and consent form, as well as any advertising tools that will 
be used for the study. Copies of all approved materials, all correspondence with the IRB and 
written approval from the IRB must be made available to Ocuphire, prior to the start of subject 
enrollment into the study. 
13.3. Protocol Deviations/Violations 
The Investigator should not deviate from the requirements of this protocol without prior written 
approval of the Medical Monitor or Sponsor, except in the event of a medical emergency. 
A reportable protocol deviation is defined as nonadherence to the protocol that involves 
inclusion/exclusion criteria, affects subject safety, rights, or welfare, or has the potential to affect 
the integrity of the data. Examples of major protocol deviations include study enrollment by 
ineligible subject, loss of key data such as equipment malfunction (eg, pupillometer), and/or use 
of a prohibited medication during the study.  
All protocol deviations will be reported by entering the event in the appropriate eCRF page. 
Protocol deviations should be reported to the IRB in accordance with IRB-specific guidelines. If 
there is any question as to whether the deviation is reportable, Ocuphire or designee and the IRB 
should be contacted. 
All changes to the protocol will be made by the Sponsor or designee as an approved amendment 
to the protocol, submitted to the FDA, and approved by the IRB prior to implementation.  
Changes implemented without prior approval will be considered protocol violations. 


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13.4.  Informed Consent Requirements 
Written informed consent will be obtained from each subject. A copy of the signed and dated 
consent document will be given to each subject. The original signed and dated ICF must be 
maintained in the study files at the Investigator’s site.  
The Investigator is responsible for ensuring that no subject is subject to any study-related 
examination or activity before that subject has given informed consent. The subject must give 
written consent after the receipt of detailed information. The verbal explanation will cover all the 
elements specified in the written information provided for the subject. 
It should be emphasized that the subject is at liberty to withdraw consent to participate at any 
time, without penalty or loss of benefits to which the subject is otherwise entitled. Subjects who 
refuse to give or who withdraw written informed consent may not be included or continued in 
this study, but this will not impact on their subsequent care.  
The Investigator will inform the subject of the aims, methods, anticipated benefits, and potential 
hazards of the study, including any discomfort it may entail. The subject must be given every 
opportunity to clarify any points he/she does not understand and, if necessary, ask for more 
information. At the end of the interview, the subject may be given time to reflect if this is 
required, or if the subject requests more time. Subjects will be required to sign and date the ICF.  
A copy of the signed and dated ICF will be given to each subject. The original signed and dated 
ICF must be maintained in the study files at the Investigator’s site. Signed informed consent 
must be obtained prior to the conductance of any study procedures. 
 
14. DATA HANDLING AND RECORD KEEPING 
All 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 Entry 
Study-specific data that have been outlined in the protocol will be entered into the clinical 
database by individual(s) designated by the Investigator.  
14.2. Data Quality Control and Reporting 
Data will be verified electronically using a series of programmed edit checks that have been 
created by the Clinical Data Manager and programmed by the Clinical Data Programmer or 
designee. Data discrepancies will be brought to the attention of the clinical team and investigated 
by the clinical research associate (CRA) and Site Staff. Clinical research associates will review 
and verify all data collected in the eCRF against source documentation during scheduled 
monitoring visits. The CRA will work closely with the Site Staff to address any discrepancies 
that have been found so that proper resolutions can be made and documented in the clinical 
database. An audit trail within the system will track all changes made to the data. 
14.3. Archiving of Data 
Archived versions of the database will be saved by Ocuphire consistent with ICH GCP 
guidelines, complying with whichever of the requirements is longer. Ocuphire will notify the 


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Investigator when documents should be returned. 
14.4.  Records Retention 
The Investigator’s site and clinical laboratory will retain all records related to the study in 
compliance with ICH GCP guidelines. 
14.5. Amendments to the Protocol 
Modifications of the signed protocol are only possible by 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 by the committee. This should be fully documented. 
The Investigator must not implement any deviation from or change to the protocol, without 
discussion with, and agreement by Ocuphire and prior review and documented 
approval/favorable opinion of the amendment from the relevant ethics committee, except where 
it is necessary to eliminate an immediate hazard to study subjects, or where the change(s) 
involves only logistical or administrative aspects of the study (eg, change in monitor, change of 
telephone number). 
Protocol amendments will be submitted to the appropriate authority(ies) as required by the 
applicable regulatory requirement(s). 
 
 


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15. REFERENCES 
1. 
McDonald MB, Pitlick W, D.R. V, Meyer AR, E.D. D. Phentolamine Mesylate Treatment of 
Severe Night Vision Complaints. AAO Abstracts, PO433. 2010. 
2. 
Holladay JT, Meyer A, Pitlick B. Phentolamine Mesylate Ophthalmic Solution Once Daily 
Reduces Pupil Diameter and Improves Night Vision Disturbances. AAO Annual Meeting, PA025. 
2018. 
3. 
Pepose JS, Pitlick B, Meyer A, Jaber R, Charizanis K, Slonim C, et al. Phentolamine 
mesylate ophthalmic solution provides long lasting pupil modulation and improves visual acuity. 
Investigative Ophthalmology & Visual Science. 2020;61(7):5100-. 
4. 
Leavitt JA, Wayman LL, Hodge DO, Brubaker RF. Pupillary response to four concentrations 
of pilocarpine in normal subjects: application to testing for Adie tonic pupil. Am J Ophthalmol. 
2002;133(3):333-6. 
5. 
Edgar DF, Crabb DP, Rudnicka AR, Lawrenson JG, Guttridge NM, O'Brien CJ. Effects of 
dipivefrin and pilocarpine on pupil diameter, automated perimetry and LogMAR acuity. Graefes 
Arch Clin Exp Ophthalmol. 1999;237(2):117-24. 
6. 
AOA. American Optometric Association - Adult Vision: 41 to 60 Years of Age. 
7. 
McDonnell PJ, Lee P, Spritzer K, Lindblad AS, Hays RD. Associations of presbyopia with 
vision-targeted health-related quality of life. Arch Ophthalmol. 2003;121(11):1577-81. 
8. 
Fricke TR, Tahhan N, Resnikoff S, Papas E, Burnett A, Ho SM, et al. Global Prevalence of 
Presbyopia and Vision Impairment from Uncorrected Presbyopia: Systematic Review, Meta-
analysis, and Modelling. Ophthalmology. 2018;125(10):1492-9. 
9. 
Waring GO, Price FW, Wirta D, et al, Safety and Efficacy of AGN-190584 in Individuals 
with Presbyopia, The GEMINI 1 Phase 3 Randomized Clinical Trial, JAMA Ophthalmol. 
2022;140(4):363-71. 
10.  
Tucker J, Charman WN. The depth-of-focus of the human eye for Snellen letters. Am J 
Optom Physiol Opt. 1975;52(1):3-21. 
11. 
Dexl AK, Seyeddain O, Riha W, Hohensinn M, Hitzl W, Grabner G. Reading performance 
after implantation of a small-aperture corneal inlay for the surgical correction of presbyopia: Two-
year follow-up. J Cataract Refract Surg. 2011;37(3):525-31. 
12. 
Xu R., Thibos L., Bradley A., Effect of Target Luminance on Optimum Pupil Diameter for 
Presbyopic Eyes. Optometry and Vision Science. 2016;91(11):1409-19. 
13. 
Hogan TS, McDaniel DD, Bartlett JD, Hart KK, Paggiarino DA. Dose-response study of 
dapiprazole HCl in the reversal of mydriasis induced by 2.5% phenylephrine. J Ocul Pharmacol 
Ther. 1997;13(4):297-302. 


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16.    SUMMARY OF CHANGES 
Bold text shows additions; strikethrough text shows deletions. For changes that affect multiple 
sections of the protocol, the change is listed once at the first instance in the table below, and each 
subsequent protocol section incorporating that change is also listed at that point. Administrative and 
minor editing changes that do not affect the content or conduct of the protocol have been made; 
these are not listed. 
16.1. Protocol OPI-NYXP-301 Amendment 1 
Protocol OPI-NYXP-301 Amendment 1, issued 31 October 2022, makes the following changes to 
the original protocol dated 08 July 2022: 
Table 5 
Protocol OPI-NYXP-301 Amendment 1 Summary of Changes 
Section/Location  
Description of Change 
Rationale for Change 
Synopsis: Inclusion 
criteria (p.7) 
 
6.1.Subject Inclusion 
Criteria (p.37) 
Inclusion criterion #8: 
Photopic PD of ≥ 3 mm in either eye. 
To clarify that photopic PD 
must be ≥ 3 mm in at least 
one eye (both eyes not 
required) 
Synopsis: Exclusion 
criteria (p.8) 
 
6.2.Subject Exclusion 
Criteria (p.38) 
Exclusion criterion #15: 
Contact lens wear on the day of any study visit and 
contact lenses must be removed for home dosing and for 
at least 10 minutes following dosing. 
To clarify that contact lens 
wearers must remove 
contact lenses prior to 
dosing and leave them out 
for at least 10 minutes after 
dosing 
Synopsis: Key 
Secondary Efficacy 
Endpoints (p.10) 
 
 
The key secondary endpoint to evaluate LDP adjunctive to 
Nyxol in Stage 2 is the percent of subjects with ≥ 15 letters 
of improvement in photopic binocular DCNVA and with < 
5 letters of loss in photopic binocular BCDVA at 30 min 
post-LDP/vehicle comparing LDP as adjunctive therapy to 
Nyxol to placebo, Nyxol alone, and LDP alone at Visit 5 
(Stage 2 Day 8). 
To clarify that the key 
secondary endpoint is to be 
evaluated at Visit 5 (Stage 
2 Day 8) 


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Section/Location  
Description of Change 
Rationale for Change 
5.4.1 Study 
Medication: Table 4 
(p.32-33) 
Investigational product 
name/formulation 
Low-Dose Pilocarpine 
Hydrochloride Ophthalmic 
Solution 0.4% (LDP) 
Aqueous isotonic solution, 
compounded by Pine 
Pharmaceuticals LLC and 
released by Iuvo BioScience 
LLC  
Concentration active 
0.4% 
Drug used in 
compounding the 
investigational 
product 
PILOCARPINE 
HYDROCHLORIDE 4% - 
pilocarpine hydrochloride 
solution/ drops  
NDC 61314-0206-15 
Manufacturer drug 
product used for 
compounding 
Sandoz, Inc. 
Low-Dose Pilocarpine investigational drug is a sterile 
solution manufactured under current Good Manufacturing 
Practice (cGMP) conditions. In addition to the active drug 
substance, the formulation also comprises the following 
inactive excipients: benzalkonium chloride NF, 
hypromellose 2910 USP, sodium citrate USP, boric acid 
NF, sodium chloride USP, and USP purified water. The pH 
is adjusted with sodium hydroxide NF (10%) and 
hydrochloric acid NF (37%) to achieve a target pH of 3.5 to 
5.5.Pilocarpine hydrochloride 0.4% investigational drug 
is compounded in pharmacy, starting with pilocarpine 
hydrochloride 4% produced by Sandoz, Inc. In addition 
to the active drug, the formulation also comprises the 
following inactive excipients: benzalkonium chloride 
NF, hypromellose 2910 USP, sodium citrate USP, boric 
acid NF, sodium chloride USP and sterile water for 
injection. The pH is adjusted with sodium hydroxide NF 
(10%) and hydrochloric acid NF (37%) to achieve a 
target pH of 3.5 to 5.5. 
To describe the 
formulation of the drug 
product (manufactured by 
Sandoz, Inc.) that will be 
compounded by Pine 
Pharmaceuticals LLC and 
released by Iuvo 
BioScience LLC 
5.1 Primary and 
Secondary Endpoints 
(p.26) 
In photopic lighting conditions, the distance and near 
illuminated charts will be at a luminance level of 
approximately 85 to 160 cd/m2 (85 to 160 nits or 267 to 
502 lux). In mesopic conditions, the distance and near 
illuminated charts will be at a luminance level of 
approximately 3 cd/m2 (3 nits or 9.42 lux). 
To provide the equivalence 
luminance levels used in 
lux 


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Appendix 3: 
Adrenergic and 
Cholinergic Drugs 
(p.63-64) 
Alpha-1-agonists 
Metaraminol 
Methoxamine 
Midodrine 
Amidephrine 
Alpha-2-agonists 
Norepinephrine 
Lofexidine 
Medetomidine 
Undetermined alpha agonists 
Ergotamine 
Etilefrine 
Indanidine 
Mephentermine 
Metaraminol 
Methoxamine 
Non-selective alpha-antagonists 
Trazodone 
Phentolamine 
Alpha-1-antagonists  
Silodosin 
Fuzosin 
Alpha-2-antagonists 
Raulscine 
Mirtazapine 
Direct-acting acetylcholine receptor agonists 
Choline esters 
Acetylcholine 
Bethanechol 
Carbachol 
Methacholine 
Plant alkaloids 
Arecoline 
Nicotine 
Muscarine 
Pilocarpine 
Indirect-acting acetylcholine receptor agonists 
Reversible cholinesterase inhibitors 
Donepezil 
Edrophonium 
Neostigmine 
Physostigmine 
Pyridostigmine 
Rivastigmine 
To make this list of 
adrenergic and cholinergic 
drugs more comprehensive 


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Tacrine 
Caffeine 
Huperzine A 
Irreversible cholinesterase inhibitors 
Echothiophate 
Isoflurophate 
Malathion 
Antimuscarinic agents 
Antipsychotics (clozapine, quetiapine) 
Atropine 
Benztropine 
Biperiden 
Chlorpheniramine 
Certain SSRIs (Paroxetine) 
Dicyclomine (Dicycloverine) 
Dimenhydrinate 
Diphenhydramine 
Doxepin 
Doxylamine 
Flavoxate 
Glycopyrrolate 
Glycopyrronium 
Hyoscyamine 
Ipratropium 
Orphenadrine 
Oxitropium 
Oxybutynin 
Promethazine 
Propantheline bromide 
Scopolamine 
Solifenacin 
Tolterodine 
Tiotropium 
Tricyclic antidepressants 
Trihexyphenidyl 
Tropicamide 
Umeclidinium 
Antinicotinic agents 
Bupropion  
Dextromethorphan 
Doxacurium 
Hexamethonium 
Mecamylamine 
Tubocurarine 


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16.2. Protocol OPI-NYXP-301 Amendment 2 
Protocol OPI-NYXP-301 Amendment 2, issued 15 November 2022, makes the following changes to 
the Protocol OPI-NYXP-301 Amendment 1 dated 31 October 2022: 
Table 6 
Protocol OPI-NYXP-301 Amendment 2 Summary of Changes 
Section/Location  
Description of Change 
Rationale for Change 
Synopsis: Duration 
of Study (p.3) 
 
5.5. Expected 
Duration of 
Subject 
Participation (p.35) 
22 to 2936 days 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
Synopsis: Design 
(p.4) 
 
 
Subjects will be randomized 1:1:1:1 to treatments for both 
stages at Visit 1 (Screening/Baseline). Note that Visit 1 
(Screening/Baseline) assessments are recommended to be 
performed on the same day; however, in instances in which 
subjects are unable to complete the full day of assessments 
for Visit 1, subjects may be scheduled to return for the 
completion of Visit 1 within 1 week of Screening. If Visit 1 is 
split over 2 separate days, all visit assessments up to and 
including randomization must occur on the first day, and 
then the subject should return to complete their post-
randomization assessments on the second day.    
 
In Stage 1, the safety and efficacy of Nyxol as a single agent 
will be evaluated following 1 day and 7 days of daily dosing, in 
which Treatment 1 (Nyxol or placebo) is dosed daily in the 
evenings near bedtime starting with the day after completion 
of Visit 1, except for Visit 1 [Stage 1 Day 1], when the subject 
is dosed during the day at the study site, and the evening before 
Visit 2, when Treatment 1 is dosed as close to 12 hours prior to 
Time 0 min at Visit 2. 
 
During Stage 1, subjects will dose with Treatment 1 (Nyxol or 
placebo [ie, Nyxol vehicle]) in the evenings near bedtime 
starting with the day after completion of Visit 1, except for 
Visit 1 [Stage 1 Day 1], when the subject is dosed during the 
day at the study site, and the evening before Visit 2, when 
Treatment 1 is dosed as close to 12 hours prior to Time 0 min at 
Visit 2. 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
Synopsis: Design 
(p.5) 
 
5. Study Design 
(p.24) 
The study eye and fellow eye will both be evaluated at all 
assessments. The baseline values are determined during 
Screening assessments at Visit 1. 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 


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Section/Location  
Description of Change 
Rationale for Change 
Synopsis: Design 
(p.5) 
 
At Visit 1 (Screening/Baseline), eligible subjects will be 
randomized, and the first dose of Treatment 1 will be 
administered (note: if Visit 1 is split into 2 separate days, then 
dosing occurs on the second day). Efficacy and safety 
measurements are made before dosing with Treatment 1 (Time 0 
min) and at multiple post-dose timepoints from 30 min to 8 
hours. Prior to discharge onfollowing completion of Visit 1 
(Stage 1 Day 1), Treatment 1 study medication for Stages 1 and 
2 will be dispensed for evening dosing by the subject before 
bedtime, to begin starting the night of Stage 1 Day 2. 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
Synopsis: Study 
Drug Dispensing 
and Treatment 
Adherence (p.6) 
 
7.1. Treatment 
Adherence (p.40) 
AtFollowing the completion of Visit 1 (Stage 1 Day 1), 
subjects will be dispensed Treatment 1 study drug for dosing at 
home in the evenings and a Treatment 1 subject diary to 
document dosing.  
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
3.5. Study 
Population (p.22) 
At Visit 1 (Screening/Baseline), eligible subjects will be 
randomized, and the first dose of Treatment 1 will be 
administered (note: if Visit 1 is split into 2 separate days, then 
dosing occurs on the second day). Efficacy and safety 
measurements are made before dosing with Treatment 1 (Time 0 
min) and at multiple post-dose timepoints from 30 min to 8 
hours. If Visit 1 is split over 2 separate days, all Screening 
assessments up to and including randomization must occur 
on the first day, and then the subject should return to 
complete their post-randomization assessments in the 
following order on the second day (ie, confirm concomitant 
medications, subject questionnaire, in-office Treatment 1 
dosing, ocular tolerability, followed by all other assessments 
at time points 0.5, 1, 3, 5, and 8 hours post-treatment). Prior 
to discharge onfollowing completion of Visit 1 (Stage 1 Day 1), 
Treatment 1 study medication for Stages 1 and 2 will be 
dispensed for evening dosing by the subject before bedtime, to 
begin startingthe night of Stage 1 Day 2. 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
5. Study Design 
(p.24) 
Baseline for the study is defined as pre-treatment up to and 
including randomization (prior to Time 0 min) at Visit 1 
(Screening/Baseline). 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
Table 1: Schedule 
of Events for Stage 
1 (p.28) 
 
Table 2: Schedule 
of Events for Stage 
2 (p.30) 
Moved “Subject questionnaire” row up to occur right before in-
office Treatment 1 dosing 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 


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Section/Location  
Description of Change 
Rationale for Change 
Table 1: Schedule 
of Events for Stage 
1 (p.28-29) 
[a] If the subject meets all the inclusion criteria and none of the 
exclusion criteria, this Screening Visit becomes the Baseline 
Visit. Visit 1 (Screening/Baseline) assessments are 
recommended to be performed on the same day; however, in 
instances in which subjects are unable to complete the full 
day of assessments for Visit 1, subjects may be scheduled to 
return for the completion of Visit 1 within 1 week of 
Screening. If Visit 1 is split over 2 separate days, all 
Screening assessments up to and including randomization 
must occur on the first day, and then the subject should 
return to complete their post-randomization assessments on 
the second day. *Post-randomization assessments to be 
performed on the second day (ie, confirm concomitant 
medications, subject questionnaire, in-office Treatment 1 
dosing, ocular tolerability, followed by all other assessments 
at time points 0.5, 1, 3, 5, and 8 hours post-treatment).    
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
Table 1: Schedule 
of Events for Stage 
1 (p.29) 
[j] Treatment 1: At Visit 1 (either first or second day of Visit 
1, see * above), one drop of Nyxol or placebo (ie, Nyxol 
vehicle) will be dosed at the study site after the completion of 0 
hour (Time 0 min) ocular assessments. 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
5.5. Expected 
Duration of 
Subject 
Participation (p.35) 
 
8.2.1. 
Screening/Baseline 
Visit 1 (Stage 1 
Day 1) (p.41-42) 
Note that Visit 1 (Screening/Baseline) assessments are 
recommended to be performed on the same day; however, in 
instances in which subjects are unable to complete the full 
day of assessments for Visit 1, subjects may be scheduled to 
return for the completion of Visit 1 within 1 week of 
Screening. If Visit 1 is split over 2 separate days, all 
Screening assessments up to and including randomization 
must occur on the first day, and then the subject should 
return to complete their post-randomization assessments on 
the second day.    
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
8.2.1. 
Screening/Baseline 
Visit 1 (Stage 1 
Day 1) (p.42) 
Efficacy and safety measurements are made before dosing with 
Treatment 1 (Time 0 min) and at multiple post-dose timepoints 
from 30 min to 8 hours. If Visit 1 is split over 2 separate days, 
all Screening assessments up to and including 
randomization must occur on the first day, and then the 
subject should return to complete their post-randomization 
assessments in the following order on the second day (ie, 
confirm concomitant medications, subject questionnaire, in-
office Treatment 1 dosing, ocular tolerability, followed by all 
other assessments at time points 0.5, 1, 3, 5, and 8 hours 
post-treatment). 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 
8.3. Visit Variation 
(p.45) 
Visit 3 may occur 1 to 3 days after Visit 2; Visit 4 may occur 8 
to 15 and days after completion of Visit 1; and Visit 5 may 
occur 22 to 2936 days after Visit 1. 
The change is to allow this 
visit to occur on 2 separate 
days so that the subject can 
return for dosing and Day 1 
post-dose measurements. The 
rationale for this change is to 
allow for more flexible 
scheduling for the subject. 


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16.3. Protocol OPI-NYXP-301 Amendment 3 
Protocol OPI-NYXP-301 Amendment 3, issued 29 September 2023, makes the following changes to 
the Protocol OPI-NYXP-301 Amendment 2, issued 15 November 2022: 
Table 7 
Protocol OPI-NYXP-301 Amendment 3 Summary of Changes 
Section/Location  
Description of Change 
Rationale for Change 
Synopsis: Study Objectives 
(p.3) 
 
4.Objectives and Purpose 
(p.23) 
The primary objective of this study is: 
• 
To evaluate the efficacy of Nyxol as a single agent to 
improve distance-corrected near visual acuity 
(DCNVA) without loss of best-corrected distance 
visual acuity (BCDVA) compared to placebo in 
subjects with presbyopia LDP as adjunctive 
therapy to Nyxol (Nyxol + LDP) to improve 
distance-corrected near visual acuity (DCNVA) 
without loss of best-corrected distance visual 
acuity (BCDVA) compared to placebo + LDP 
vehicle in subjects with presbyopia 
Additional objectives of this study are: 
• 
To evaluate the efficacy of LDP as adjunctive 
therapy to Nyxol (Nyxol + LDP) to improve 
DCNVA without loss of BCDVA compared to 
placebo, Nyxol alone, and LDP alone, in subjects 
with presbyopia 
• 
To evaluate the efficacy of Nyxol + LDP to 
improve DCNVA without loss of BCDVA 
compared to placebo + LDP (LDP alone) and 
Nyxol + LDP vehicle (Nyxol alone) in subjects 
with presbyopia 
• 
To evaluate the efficacy of Nyxol as a single agent 
to improve DCNVA without loss of BCDVA 
compared to placebo in subjects with presbyopia 
To move evaluation of 
Nyxol as a single 
agent from the primary 
objective to a 
secondary objective 
and to move 
evaluation of Nyxol + 
LDP from a secondary 
objective to the 
primary objective 
Synopsis: Design (p.6) 
 
3.5 Study Population 
(p.22) 
 
8.2.2 Visit 2 (Stage 1 Day 
8) (p.43) 
At Visit 2 (Stage 1 Day 8), efficacy and safety measurements 
will be made at 12 hours after the dose of Treatment 1 study 
medication (12-hour primary endpoint). 
Synopsis: Primary Efficacy 
Endpoint (p.10) 
 
5.1 Primary and Secondary 
Endpoints (p.25) 
 
8.1 Specification of the 
Efficacy Parameters (p.41) 
 
10.3.5 Analysis of Efficacy 
(p.52) 
The primary efficacy endpoint is the percent of subjects 
with ≥ 15 letters of improvement in photopic binocular 
DCNVA and with < 5 letters of loss in photopic binocular 
BCDVA at 30 min post-LDP/vehicle comparing Nyxol + 
LDP to placebo + LDP vehicle at Visit 5 (Stage 2 Day 8). 
The primary efficacy endpoint is the percent of subjects with 
≥ 15 letters of improvement in photopic binocular DCNVA 
and with < 5 letters of loss in photopic binocular BCDVA 
from Baseline comparing Nyxol-treated subjects to placebo-
treated subjects at 12 hours post-dose at Visit 2 (Stage 1 Day 
8). 


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Section/Location  
Description of Change 
Rationale for Change 
Synopsis: Key Secondary 
Efficacy Endpoints (p.10) 
 
5.1 Primary and Secondary 
Endpoints (p.25) 
The key secondary endpoints are: 
• 
The percent of subjects with ≥15 letters of 
improvement in photopic binocular DCNVA and 
with < 5 letters of loss in photopic binocular 
BCDVA at 30 min post-LDP/vehicle comparing 
Nyxol + LDP to LDP alone at Visit 5 (Stage 2 
Day 8) 
• 
The percent of subjects with ≥ 15 letters of 
improvement in photopic binocular DCNVA and 
with < 5 letters of loss in photopic binocular 
BCDVA at 30 min post-LDP/vehicle comparing 
Nyxol + LDP to Nyxol alone at Visit 5 (Stage 2 
Day 8) to evaluate LDP adjunctive to Nyxol in 
Stage 2 is the percent of subjects with ≥15 letters of 
improvement in photopic binocular DCNVA and 
with < 5 letters of loss in photopic binocular 
BCDVA at 30 min post-LDP/vehicle comparing 
LDP as adjunctive therapy to Nyxol to placebo, 
Nyxol alone, and LDP alone at Visit 5 (Stage 2 Day 
8). 
These comparisons will be analyzed second, and third, and 
fourth using a hierarchical testing method. 
Synopsis: Other Secondary 
Efficacy Endpoints (p.10-
11) 
 
5.1 Primary and Secondary 
Endpoints (p.25) 
• 
Percentage of subjects with ≥ 15 letters of 
improvement in binocular photopic DCNVA and 
with < 5 letters of loss in binocular photopic 
BCDVA from Baseline (excluding primary and key 
secondary outcome timepoints) 
10.3.5 Analysis of Efficacy 
(p.51) 
The primary comparison of interest is Stage 21 Nyxol + LDP 
versusand placebo + LDP vehicle alone for the mITT 
Population. 
 
Synopsis: Other Secondary 
Efficacy Endpoints (p.11) 
 
5.1 Primary and Secondary 
Endpoints (p.26) 
• 
Subject questionnaire responses related to change 
in near vision and satisfaction with near vision 
To add subject 
questionnaire 
responses as an 
efficacy endpoint 
 
 
 


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APPENDIX 1: IRIS COLOR CHART 
Study enrollment includes both light- and dark-colored eyes. Examples of light irides (pictures A, B, 
C, D, and E) and dark irides (pictures F, G, and H) for the purposes of this study are detailed in the 
chart below.  
 
 
 
 
 
 
 
 
A = Light blue 
B = Dark blue 
C = Blue with peripupillary brown 
D = Uniform green 
E = Green with brown iris ring 
F = Central brown and peripheral green 
G = Brown with some peripheral green 
H = Brown 


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APPENDIX 2: VISUAL ACUITY CHARTS 
DCNVA will be measured using the Near Visual Acuity Chart (logarithmic Visual Acuity Chart 
2000, chart #1 and #2) in the Precision Vision Small 914 Illuminator Cabinet (light box) at 16 inches 
(~40 cm) (letters recorded, later converted to LogMAR and number of lines). 
 
Near High-Contrast Chart 
 
            
 
 
BCDVA will be measured with a Standard ETDRS illuminated 100% high-contrast chart (on wall or 
stand) at 4 m. 
 
Distance High-Contrast Chart 
 
 
 
 
 


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APPENDIX 3: ADRENERGIC AND CHOLINERGIC DRUGS 
The following drugs are examples of drugs which cannot be used within 7 days prior to Screening or 
during the study unless the drug, dose, and regimen has been consistent for the 7 days prior to 
Screening; however, tamsulosin is specifically excluded. This list is not inclusive of all drugs in 
these classes. If there is any doubt, please consult with the Medical Monitor. 
Alpha-1-agonists 
Methyl 
norepinephrine 
Naphazoline 
Oxymetazoline 
Tetrahydrozoline 
Phenylephrine 
Xylometazoline 
Metaraminol 
Methoxamine 
Midodrine 
Amidephrine 
 
Alpha-2-agonists 
Brimonidine 
Clonidine 
Guanfacine 
Guanabenz 
Guanoxabenz 
Guanethidine 
Xylazine 
Tizanidine 
Methyldopa 
Norepinephrine 
Lofexidine 
Medetomidine 
 
Undetermined 
alpha agonists 
Ergotamine 
Etilefrine 
Indanidine 
Mephentermine 
Metaraminol 
Methoxamine 
 
Non-selective alpha-
antagonists 
Phenoxybenzamine 
Tolazoline 
Labetalol 
Carvedilol 
Trazodone 
Phentolamine 
 
Alpha-1-antagonists  
Alfuzosin 
Prazosin 
Doxazosin 
Tamsulosin 
Terazosin 
Silodosin 
Fuzosin 
 
Alpha-2-antagonists 
Atipamezole 
Idazoxan 
Yohimbine 
Raulscine 
Mirtazapine  
 
Direct-acting 
acetylcholine receptor 
agonists 
Choline esters 
Acetylcholine 
Bethanechol 
Carbachol 
Methacholine 
 
Plant alkaloids 
Arecoline 
Nicotine 
Muscarine 
Pilocarpine 
 
Indirect-acting 
acetylcholine receptor 
agonists 
Reversible 
cholinesterase inhibitors 
Donepezil 
Edrophonium 
Neostigmine 
Physostigmine 
Pyridostigmine 
Rivastigmine 
Tacrine 
Caffeine 
Huperzine A 
 
Irreversible 
cholinesterase inhibitors 
Echothiophate 
Isoflurophate 
Malathion 
 
Acetylcholine receptor 
antagonists 
Scopolamine 
Gastrointestinal 
Atropine  
Belladonna 
 
Parkinsonism 
Amantadine 
Benztropine 
Biperiden 
Trihexyphenidyl 
 
Antimuscarinic 
agents 
Antipsychotics 
(clozapine, 
quetiapine) 
Atropine 
Benztropine 
Biperiden 
Chlorpheniramine 
 
Certain SSRIs 
(Paroxetine) 
Dicyclomine 
(Dicycloverine) 
Dimenhydrinate 
Diphenhydramine 
Doxepin 
Doxylamine 
Flavoxate 
Glycopyrrolate 
Glycopyrronium 
Hyoscyamine 
Ipratropium 
Orphenadrine 
Oxitropium 
Oxybutynin 
Promethazine 
Propantheline 
bromide 


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Dicycloverine 
Tolterodine 
Oxybutynin 
Ipratropium 
Mamba Toxin (MT7) 
Pirenzepine 
Telenzepine 
 
Antivertigo 
Meclizine  
Scopolamine 
Scopolamine 
Solifenacin 
Tolterodine 
Tiotropium 
 
Tricyclic 
antidepressants 
Trihexyphenidyl 
Tropicamide 
Umeclidinium 
 
Antinicotinic agents 
Bupropion  
Dextromethorphan 
Doxacurium 
Hexamethonium 
Mecamylamine 
Tubocurarine 
 
 
 


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APPENDIX 4: STUDY DESIGN 
 
                                       
                                                                                         
       
        
     
        
      
                 
               
      
      
     
    
          
        
            
                     
                    
              
                 
     
       
       
       
        
     
        
                                    
        
                               
              
       
           
              
             
                
                          
                  
                             
                
       
        
     
        
                                                      
     
     
    
    
    
    
                                                        
                                                       
          
              
             
                
                     
                        
                  
                     
                      
      
        
        
           
           
     
     
       
             
       
       
       
       
          
          
       
       
     
        


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VEGA-2 Baseline Subject Questionnaire  
9/29/2023 
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APPENDIX 5: SUBJECT QUESTIONNAIRE FOR BASELINE VISIT 
Subject Number: _____________ 
 
 
 
 
Visit Date___________ 
 
 
 
 
 
 
 
 
 
Time: ________________ 
 
VEGA-2 Subject Questionnaire – Baseline Visit  
  
As you answer the subject questionnaire, please use the following examples as a reference:  
Near vision: seeing things up close, such as a cell phone or book  
Intermediate vision: seeing things that are not handheld, but still close, such as the dashboard of a car or 
a desktop computer  
Distance vision: seeing things far away, such as road signs while driving or a TV screen  
Using/doing something to manage near vision: may include wearing reading or bifocal glasses, contact 
lenses, squinting, or adjusting font size or zooming in (when using screens)  
  
I. For questions 1-6, circle the answer most applicable to you, without something to manage your 
vision for questions 4-6. 
  
1.  
What do you use most to 
help with managing your 
near vision?  
 
 
 
 
 
 
  
2.  
For how many hours in a 
typical day do you use or 
do something to help 
manage your near vision?  
All waking 
hours  
1  
 
Working hours 
(e.g., 9-5)  
2  
  
Morning  
3  
Afternoon/  
evening  
4  
Task-
dependent  
5  
 
  
3.  
How important is clear 
near vision from the time 
you wake up?  
Not at all 
important  
1  
Somewhat 
important  
2  
Moderately 
important  
3  
Very 
important  
4  
Extremely 
important  
5  
  
4.  
How would you describe 
your near vision in normal 
or day light?  
Very poor  
1  
Poor  
2  
Fine  
3  
Good  
4  
Excellent  
5  
  
5.  
How would you describe 
your near vision in dim or 
evening light?  
Very poor  
1  
Poor  
2  
Fine  
3  
Good  
4  
Excellent  
5  
 


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VEGA-2 Baseline Subject Questionnaire  
9/29/2023 
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6.  
How difficult is it for you to 
see distance while driving
in dim or night light?  
Very 
difficult  
1  
Moderately 
difficult   
2  
Somewhat 
difficult  
3  
A little 
difficult  
4  
Not 
difficult  
5  
I don’t 
drive  
N/A  
  
Page Break  
  
II. For each of the questions 7-12, circle the number that corresponds with the frequency that is 
most applicable to you over the past week. If you have not used screens in the past week, please 
circle “N/A” for questions 9 and 10.   
  
Over the past week…  
All of the 
time  
Most of the 
time  
Some of 
the time  
Rarely  
Never  
Not 
applicable  
7.   
How frequently did you use 
some form of near vision 
correction?  
1  
2  
3  
4  
5  
  
8.   
How often did you need to 
use or do something to help 
you read small-sized text at 
a near distance (e.g., a 
menu or prescription label)  
1  
2  
3  
4  
5  
  
9.   
How often did you need to 
use or do something to help 
you read information on a 
cell phone or tablet screen 
at a near distance?  
1  
2  
3  
4  
5  
N/A  
10.   
How often did you need to 
use or do something to help 
you read information on a 
computer screen at an 
intermediate distance?  
1  
2  
3  
4  
5  
N/A  
11.   
How often did you need to 
use or do something to help 
you see things clearly from 
a distance?  
1  
2  
3  
  
4  
  
5  
  
12.   
How often was your self-
confidence negatively 
affected by the ways you 
manage your near vision?  
1  
2  
3  
4  
5  
  
Page Break  
 


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VEGA-2 Visits 2, 4, & 5 Subject Questionnaire  
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Page 74 of 76 
APPENDIX 6: SUBJECT QUESTIONNAIRE FOR VISITS 2, 4, AND 5  
Subject Number: _____________  Visit #_________  Date ___________ Time ____________ 
 
 
 
  
 
VEGA-2 Subject Questionnaire – Visits 2, 4, and 5  
  
As you answer the subject questionnaire, please use the following examples as a reference:  
Near vision: seeing things up close, such as a cell phone or book  
Intermediate vision: seeing things that are not handheld, but still close, such as the dashboard of a car 
or a desktop computer  
Distance vision: seeing things far away, such as road signs while driving or a TV screen  
Using/doing something to manage near vision: may include wearing reading or bifocal glasses, 
contact lenses, squinting, or adjusting font size or zooming in (when using screens)  
  
I. For each of the questions 1-8, circle the answer that is most applicable to your overall 
experience with your study medication.   
1.   
My study medication made my 
near vision…  
Far   
worse  
1  
Slightly 
worse  
2  
No   
change  
3  
Slightly 
better  
4  
Far   
better  
5  
 
  
2.   
How convenient was it to use 
your study medication for your 
near vision?  
Very 
inconvenient 
1  
  
Inconvenient 
2  
Neutral  
3  
Convenient 
4  
Very 
convenient  
5  
  
 
  
3.  
How frequently would you use 
the study medication if it were 
available to you?  
Never  
1  
Less than 
once a week  
2  
Once a week  
3  
2-3 times a 
week  
4  
Every   
day   
5  
 
  
4.  
How satisfied are you with 
how long the effects of your 
study medication lasted 
throughout the day?  
Very 
dissatisfied  
1  
  
Dissatisfied  
2  
Neutral  
3  
Satisfied  
4  
Very 
satisfied  
5  
  
 
 
5.  
How satisfied are you with 
your study medication to help 
you see clearly up close 
when you first wake up?  
Very 
dissatisfied  
1  
  
Dissatisfied  
2  
Neutral  
3  
Satisfied  
4  
Very 
satisfied  
5  
  
 
  


Ocuphire Pharma, Inc. 
Clinical Protocol 
OPI-NYXP-301 
VEGA-2 
 
 
 
VEGA-2 Visits 2, 4, & 5 Subject Questionnaire  
9/29/2023 
Page 75 of 76 
6.  
My study medication made my 
near vision in dim/low light …  
Far   
worse  
1  
Slightly 
worse  
2  
No   
change  
3  
Slightly 
better  
4  
Far   
better  
5  
 
  
7.  
My study medication made my 
distance vision…  
Far   
worse  
1  
Slightly 
worse  
2  
No   
change  
3  
Slightly 
better  
4  
Far   
better  
5  
 
  
8.  
My study medication made my 
distance vision while driving at 
night…   
Far worse  
1  
Slightly 
worse  
2  
No 
change  
3  
Slightly 
better  
4  
Far   
better  
5  
I do not 
drive  
N/A  
 
  
II. For question 9, please circle every answer that applies to you.    
9.  
If the study medication 
were available to me, I 
would use it for…   
Social 
settings 
(e.g., 
restaurants)  
1  
Work  
2  
Leisure 
(e.g., 
reading)  
3  
Phone/  
tablet use  
4  
Driving  
5  
Everyday 
tasks (e.g., 
cooking)  
6  
 
  
III. For each of the questions 10-13, circle the number that corresponds with the frequency that is 
most applicable to you over the past week. If you have not used screens in the past week, please 
circle “N/A” for questions 12 and 13.    
 
Over the past week…  
All of the 
time  
Most of 
the time  
Some of 
the time  Rarely  
Never  
N/A  
10.   
How frequently did you use some form 
of near vision correction?  
1  
2  
3  
4  
5  
  
11.  
How often did you need to use or do 
something to help you read small-sized 
text at a near distance (e.g., a menu or 
prescription label)?  
1  
2  
3  
4  
5  
  
12.  
How often did you need to use or do 
something to help you read information 
on a screen (e.g., a cell phone or 
tablet) at a near distance?  
1  
2  
3  
4  
5  
N/A  
13.   
How often did you need to use or do 
something to help you read information 
on a screen at an intermediate 
distance?  
1  
2  
3  
4  
5  
N/A  
 


Ocuphire Pharma, Inc. 
Clinical Protocol 
OPI-NYXP-301 
VEGA-2 
 
 
 
VEGA-2 Visits 2, 4, & 5 Subject Questionnaire  
11/15/2022 
Page 76 of 76 
  
IV. For question 14, circle the answer that is most applicable to you.   
14.  
How likely would you be to 
use the study medication for 
near vision correction?  
Extremely 
unlikely 
1 
 
Unlikely  
2  
Possibly  
3  
Likely 
4  
Extremely likely 
5  
 
 

SAP_001.pdf: 
  
 
CONFIDENTIAL 
 
 
 
 
Ocuphire Pharma, Inc. 
 
STATISTICAL ANALYSIS PLAN 
 
 
 
Protocol Title: 
Randomized, Double-Masked, Placebo-Controlled, Multicenter, 
Phase 3 Study of the Safety and Efficacy of Nyxol 
(Phentolamine Ophthalmic Solution 0.75%) as a Single Agent 
and With Adjunctive Low-Dose Pilocarpine Hydrochloride 
Ophthalmic Solution 0.4% in Subjects With Presbyopia 
Study Number: 
OPI-NYXP-301 (VEGA-2) 
Phase: 
Phase 3 
Sponsor: 
Ocuphire Pharma, Inc. 
37000 Grand River Avenue, Suite 120 
Farmington Hills, MI 48335 USA 
 
Author: 
Summit Analytical, LLC 
8354 Northfield Blvd. 
Bldg. G Suite 3700 
Denver, CO 80238 
SAP Date: 
2023-09-29 
Status: 
FINAL V1.0 
 
 
 


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1. 
TABLE OF CONTENTS 
1. 
TABLE OF CONTENTS ................................................................................................................................2 
2. 
LIST OF ABBREVIATIONS AND DEFINITION OF TERMS ...................................................................5 
3. 
INTRODUCTION ...........................................................................................................................................7 
3.1. 
Preface ................................................................................................................................................7 
3.2. 
Purpose of Analyses ...........................................................................................................................7 
3.3. 
Summary of Statistical Analysis Changes to the Protocol .................................................................7 
4. 
STUDY OBJECTIVES AND ENDPOINTS ..................................................................................................8 
4.1. 
Study Objectives ................................................................................................................................8 
4.2. 
Study Endpoints .................................................................................................................................8 
4.2.1. 
Primary Endpoints ............................................................................................................8 
4.2.2. 
Secondary Endpoints ........................................................................................................9 
5. 
STUDY METHODS .....................................................................................................................................12 
5.1. 
General Study Design and Plan ........................................................................................................12 
5.2. 
Inclusion – Exclusion Criteria and General Study Population .........................................................18 
5.3. 
Subject Inclusion Criteria .................................................................................................................18 
5.4. 
Subject Exclusion Criteria ................................................................................................................18 
5.5. 
Randomization and Masking ............................................................................................................20 
5.6. 
Analysis Variables ............................................................................................................................20 
6. 
SAMPLE SIZE ..............................................................................................................................................22 
7. 
GENERAL CONSIDERATIONS .................................................................................................................23 
7.1. 
Analysis Populations ........................................................................................................................23 
7.1.1. 
Modified Intention-to-Treat (mITT) ..............................................................................23 
7.1.2. 
Per Protocol Stage 1 Population (PPS1) ........................................................................23 
7.1.3. 
Per Protocol Stage 2 Population (PPS2) ........................................................................23 
7.1.4. 
All Randomized Population (ARP) ................................................................................23 
7.1.5. 
Safety Population (SP) ...................................................................................................23 
7.2. 
Covariates and Subgroups ................................................................................................................24 
7.2.1. 
Planned Covariates .........................................................................................................24 
7.2.2. 
Planned Subgroups .........................................................................................................24 
7.3. 
Management of Analysis Data .........................................................................................................24 
7.3.1. 
Data Handling ................................................................................................................24 
7.3.2. 
Missing Data ..................................................................................................................24 
7.3.2.1. 
Handling of Missing Date Values .............................................................24 
7.3.2.2. 
Missing Baseline Data ..............................................................................25 
7.3.2.3. 
Imputation Methods ..................................................................................25 
7.3.3. 
Handling of Early Termination Visit Information .........................................................26 
7.3.4. 
Pooling of Investigational Sites ......................................................................................26 
7.3.5. 
Coding Conventions for Events and Medications ..........................................................26 
7.3.6. 
Analysis Software ..........................................................................................................26 
7.3.7. 
Study Data ......................................................................................................................26 
7.4. 
Planned Study Analyses ...................................................................................................................28 
7.4.1. 
Statistical Summaries: Descriptive and Inferential ........................................................28 
7.4.2. 
Interim Analyses and Data Monitoring ..........................................................................28 
7.4.3. 
Final Analysis and Publication of Study Results ...........................................................28 
7.5. 
Multiple Testing Procedures ............................................................................................................28 
7.6. 
Baseline Values ................................................................................................................................29 


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8. 
SUMMARY OF STUDY DATA ..................................................................................................................30 
8.1. 
Subject Disposition ..........................................................................................................................30 
8.2. 
Protocol Deviations ..........................................................................................................................30 
8.3. 
Demographics and Baseline Characteristics ....................................................................................30 
8.4. 
Medical History ................................................................................................................................31 
8.5. 
Prior and Concomitant Medications .................................................................................................31 
8.6. 
Treatment Exposure and Compliance ..............................................................................................32 
9. 
EFFICACY ANALYSES ..............................................................................................................................33 
9.1. 
Clinical Efficacy ...............................................................................................................................33 
9.1.1. 
Primary Efficacy Analysis and Key Secondary Analyses .............................................33 
9.1.2. 
Secondary Efficacy Analyses .........................................................................................34 
10. SAFETY ANALYSES ..................................................................................................................................36 
10.1. 
Adverse Events .................................................................................................................................36 
10.2. 
Deaths, Serious Adverse Events and Other Significant Adverse Events .........................................37 
10.2.1. 
Deaths .............................................................................................................................37 
10.2.2. 
Serious Adverse Events ..................................................................................................37 
10.2.3. 
Adverse Events Leading to Withdrawal from the Study ................................................37 
10.2.4. 
Adverse Events Leading to Discontinuation of Study Medication ................................37 
10.3. 
Subjective Ocular Tolerability .........................................................................................................37 
10.4. 
Vital Signs ........................................................................................................................................38 
10.5. 
IOP ...................................................................................................................................................38 
10.6. 
Subject Questionnaire ......................................................................................................................38 
10.7. 
Biomicroscopy and Ophthalmoscopy ..............................................................................................38 
10.8. 
Other Safety Measures .....................................................................................................................38 
11. REFERENCES ..............................................................................................................................................39 
12. APPENDICES ...............................................................................................................................................40 
12.1. 
List of Planned Tables ......................................................................................................................40 
12.2. 
List of Planned Listings ....................................................................................................................48 
12.3. 
Testing Hierarchy .............................................................................................................................49 
 
 


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LIST OF TABLES 
 
Table 1 
Schedule of Events for Stage 1 ......................................................................................14 
Table 2 
Schedule of Events for Stage 2 ......................................................................................16 
Table 3 
Snellen Equivalents to VA Assessment Letters Read ....................................................34 
 
 
LIST OF FIGURES 
 
Figure 1 
SDTM, ADaM, and TFL Development and Validation .................................................27 
 
 
 


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2. 
LIST OF ABBREVIATIONS AND DEFINITION OF TERMS 
Only abbreviations and terms relevant to the SAP are repeated herein. The reader is referred 
to the protocol for the complete and comprehensive list of abbreviations and definitions of 
terms for the study. 
Abbreviation/Term 
Definition 
ADaM 
Analysis Data Model 
AE 
adverse event 
ANCOVA 
analysis of covariance 
ARP 
All Randomized Population 
ATC 
Anatomical Therapeutic Chemical  
BCDVA 
best-corrected distance visual acuity  
BP 
blood pressure 
°C 
degrees Celsius 
CDISC 
Clinical Data Interchange Standards Consortium  
CI 
confidence interval 
CSR 
clinical study report 
DCIVA 
distance-corrected intermediate visual acuity 
DCNVA 
distance-corrected near visual acuity 
DLD 
dim light vision disturbances (also referred to as night vision 
disturbances or NVD) 
eCRF 
electronic Case Report Form 
EDC 
electronic data capture 
ETDRS 
Early Treatment Diabetic Retinopathy Study 
°F 
degrees Fahrenheit 
FDA 
Food and Drug Administration 
HR 
heart rate 
IOP 
intraocular pressure 
LDP 
Low-Dose (0.4%) Pilocarpine Ophthalmic Solution 
LSM 
least-squares mean 
MedDRA 
Medical Dictionary for Regulatory Activities 
mITT 
Modified Intention-to-Treat 
Nyxol 
0.75% Phentolamine Ophthalmic Solution or 1% Phentolamine 
Mesylate Ophthalmic Solution  
Nyxol + LDP 
Nyxol dosed in the evening with LDP dosed during the day 
OD 
oculus dexter (right eye) 


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Abbreviation/Term 
Definition 
OR 
odds ratio 
OU 
oculus uterque (both eyes) 
PD 
pupil diameter 
POS 
Phentolamine Ophthalmic Solution 
PP 
Per Protocol 
PPS1 
Per Protocol Stage 1 
PPS2 
Per Protocol Stage 2 
PT 
preferred term 
SAE 
serious adverse event 
SAP 
statistical analysis plan 
SDTM 
study data tabulation model 
SE 
standard error 
SOC 
system organ class 
SP 
Safety Population 
TEAE 
treatment-emergent adverse event 
TFL 
tables, figures, and listings 
VA 
visual acuity 
WHO-DD 
World Health Organization Drug Dictionary 


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3. 
INTRODUCTION 
3.1. 
Preface 
This document presents a statistical analysis plan (SAP) for Ocuphire Pharma, Inc. Protocol 
OPI-NYXP-301 (VEGA-2) (Randomized, Double-Masked, Placebo-Controlled, Multicenter, 
Phase 3 Study of the Safety and Efficacy of Nyxol (Phentolamine Ophthalmic Solution 
0.75%) as a Single Agent and With Adjunctive Low-Dose Pilocarpine Hydrochloride 
Ophthalmic Solution 0.4% in Subjects With Presbyopia). 
Reference materials for this statistical plan include Amendment 3 of the protocol OPI-NYXP-
301 (29SEP2023) and Case Report Forms (CRFs; Version 09DEC2022).  
The SAP described hereafter is an a priori plan. The SAP will be finalized and approved 
prior to unmasking of any study data.     
For the reasons stated here, the conduct of the study in the field is considered to be 
independent of any study outcome that might materialize upon enactment of the currently 
proposed statistical plan.  
3.2. 
Purpose of Analyses 
The VEGA-2 study is a randomized, placebo-controlled, double-masked study of the safety 
and efficacy of Nyxol (POS 0.75%) in combination with low-dose Pilocarpine (LDP) 0.4% 
in subjects with presbyopia. The Sponsor intends to use this Phase 3 pivotal study to 
evaluate Nyxol as a single agent and Nyxol + LDP for the indication of “treatment of 
presbyopia.” 
Post-hoc exploratory analyses not identified in this SAP may be performed to further 
examine the study data. These analyses will be clearly identified, where appropriate, in the 
final clinical study report (CSR). Additional analyses not prospectively identified in this SAP 
may also be completed for publications, or regulatory or funding inquiries. These analyses, if 
performed, may not be reported in the CSR but will be fully detailed in the document 
containing the additional analyses. 
3.3. 
Summary of Statistical Analysis Changes to the Protocol 
The analyses described in this analysis plan are consistent with the analyses described in the 
study protocol. 


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4. 
STUDY OBJECTIVES AND ENDPOINTS 
Study objectives and endpoints defined in the protocol include safety and efficacy endpoints. 
Objectives and pre-specified endpoints are as follows: 
4.1. 
Study Objectives 
The objectives of this study are as follows: 
Primary objective 
• To evaluate the efficacy of LDP as adjunctive therapy to Nyxol (Nyxol + LDP) to 
improve DCNVA without loss of BCDVA compared to placebo + LDP vehicle in 
subjects with presbyopia 
 
Additional objectives 
• To evaluate the efficacy of Nyxol + LDP to improve DCNVA without loss of 
BCDVA compared to placebo + LDP (LDP alone) and Nyxol + LDP vehicle (Nyxol 
alone) in subjects with presbyopia  
• To evaluate the efficacy of Nyxol as a single agent to improve DCNVA without loss 
of BCDVA compared to placebo in subjects with presbyopia  
• To evaluate additional efficacy measures, such as visual acuity (VA) and pupil 
diameter (PD), across multiple timepoints and visits for Nyxol as a single agent and 
Nyxol + LDP  
• To evaluate the effect of iris color, Baseline PD, and Baseline DCNVA on efficacy of 
Nyxol as a single agent and Nyxol + LDP 
• To evaluate the ocular and systemic safety of Nyxol as a single agent and Nyxol + 
LDP  
• To evaluate subject-reported outcomes of Nyxol as a single agent and Nyxol + LDP 
4.2. 
Study Endpoints 
4.2.1. 
Primary Endpoints 
The primary efficacy endpoint is the percent of subjects with ≥ 15 letters of improvement in 
photopic binocular DCNVA and with < 5 letters of loss in photopic binocular BCDVA from 
Baseline at 30 min post-LDP/vehicle comparing subjects treated with Nyxol + LDP to 
subjects treated with placebo + LDP vehicle at Visit 5 (Stage 2 Day 8).  
This comparison will be analyzed first using a hierarchical testing method.  


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4.2.2. 
Secondary Endpoints 
Efficacy: 
The first key secondary endpoint is the percent of subjects with ≥ 15 letters of improvement 
in photopic binocular DCNVA and with < 5 letters of loss in photopic binocular BCDVA at 
30 min post-LDP/vehicle comparing Nyxol + LDP to LDP alone at Visit 5 (Stage 2 Day 8).  
The second key secondary endpoint is the percent of subjects with ≥ 15 letters of 
improvement in photopic binocular DCNVA and with < 5 letters of loss in photopic 
binocular BCDVA at 30 min post-LDP/vehicle comparing Nyxol + LDP to Nyxol alone at 
Visit 5 (Stage 2 Day 8). 
These comparisons will be analyzed second and third using a hierarchical testing method. 
Other secondary endpoints measured at various timepoints and visits by treatment arms in 
Stage 1 and Stage 2 include: 
• Percentage of subjects with ≥ 15 letters of improvement in binocular photopic 
DCNVA and with < 5 letters of loss in binocular photopic BCDVA from Baseline 
(excluding primary and key secondary outcome timepoints) 
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of improvement in photopic 
DCNVA from Baseline (monocular and binocular) 
• Change in photopic DCNVA from Baseline (monocular and binocular) 
• Percent of subjects with photopic DCNVA ≤ 20/15, ≤ 20/20, ≤ 20/25, ≤ 20/32, ≤ 
20/40, ≤ 20/50, ≤ 20/63, ≤ 20/80, ≤ 20/100, ≤ 20/125, and ≤ 20/160  
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of improvement in photopic 
binocular distance-corrected intermediate visual acuity (DCIVA) from Baseline  
• Change in photopic binocular DCIVA from Baseline  
• Percentage of subjects with loss or improvement in photopic and mesopic 
BCDVA from Baseline (monocular and binocular) 
• Change in photopic and mesopic BCDVA from Baseline (monocular and 
binocular) 
• Percentage of subjects with ≥ 5, ≥ 10, and ≥ 15 letters of improvement in mesopic 
binocular DCNVA from Baseline  
• Change in mesopic binocular DCNVA from Baseline  
• Change and percent change in photopic PD from Baseline  
• Percentage of subjects with photopic PD of < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm  
• Percentage of subjects with photopic PD of 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 2.5 
to < 3.0 mm, and 3.0 to < 3.5 mm  


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• Change and percent change in mesopic PD from Baseline  
• Percentage of subjects with mesopic PD of < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm  
• Percentage of subjects with mesopic PD of 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 2.5 
to < 3.0 mm, and 3.0 to < 3.5 mm  
• Percentage of subjects with Baseline photopic DCNVA ≤ 20/63, ≤ 20/80, ≤ 
20/100, ≤ 20/125, and ≤ 20/160, with a ≥ 10-letter and ≥ 15-letter improvement in 
photopic DCNVA  
• Percentage of subjects with Baseline photopic DCNVA from 20/50 to 20/63, 
20/50 to 20/80, 20/50 to 20/100, 20/50 to 20/125, and 20/50 to 20/160, with a ≥ 
10-letter and ≥ 15-letter improvement in photopic DCNVA  
• Subject questionnaire responses related to change in near vision and satisfaction 
with near vision. 
Measurements: 
• Best-corrected distance VA measured in photopic conditions using a high-contrast 
Standard Early Treatment Diabetic Retinopathy Study (ETDRS) illuminated chart 
(on wall or stand) at 4 m  
• Distance-corrected intermediate VA measured in photopic conditions using a 
high-contrast Near Visual Acuity Chart in the Precision Vision Small 914 
Illuminator Cabinet (light box) at 26 inches (66 cm) 
• Distance-corrected near VA measured in photopic and mesopic conditions using a 
high-contrast Near Visual Acuity Chart in the Precision Vision Small 914 
Illuminator Cabinet (light box) at 16 inches (~40 cm) 
• Pupil diameter measured with a NeurOptics pupillometer  
All of the efficacy endpoints will be analyzed overall and by light/dark irides at all 
timepoints. Subjects will also be analyzed by binocular, study eye, and fellow eye, where 
applicable. 
Safety: 
Safety measures are AEs, ocular tolerability, intraocular pressure (IOP), BCDVA, 
biomicroscopy, ophthalmoscopy, heart rate (HR) and blood pressure (BP), and subject 
questionnaire. Urine pregnancy tests for females of childbearing potential will also be 
conducted. 
 
Measurements:                                                                                                                                                      
• Biomicroscopy, ophthalmoscopy, and HR and BP measured in accordance with 
the site’s standard practice  
• Ocular tolerability measured on a 4-point scale (0-3) 


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• Intraocular pressure measured with a Tono-Pen or Goldmann Applanation 
tonometry 
• Subject questionnaire completed by the subject 


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5. 
STUDY METHODS 
5.1. 
General Study Design and Plan 
This is a randomized, double-masked, placebo-controlled, multicenter, Phase 3 study in 
approximately 320 subjects with presbyopia. The study is performed in 2 stages, as described 
below. 
Subjects are randomized 1:1:1:1 to treatments for both stages at Visit 1 
(Screening/Baseline). Note that Visit 1 (Screening/Baseline) assessments are recommended 
to be performed on the same day; however, in instances in which subjects are unable to 
complete the full day of assessments for Visit 1, subjects may be scheduled to return for the 
completion of Visit 1 within 1 week of Screening. If Visit 1 is split over 2 separate days, all 
visit assessments up to and including randomization must occur on the first day, and then the 
subject should return to complete their post-randomization assessments on the second day. 
Stage 1 consists of 2 treatment groups (Nyxol or placebo [ie, Nyxol vehicle]), with 
approximately 160 subjects in each group. Stage 2 consists of 4 treatment groups (Nyxol + 
LDP, Nyxol + LDP vehicle, placebo + LDP, and placebo + LDP vehicle), with 
approximately 80 subjects per treatment group. Treatment assignments for Stage 1 and Stage 
2 are assigned at randomization prior to Stage 1. 
Randomization is stratified by light/dark irides. The number of subjects in each irides type 
should be as close as possible to 1:1, with a minimum of 40% within each stratum. The 
subjects are recruited from approximately 30 investigational sites in the US. 
Stage 1: Dosing of Treatment 1 
Individuals who are potential subjects are identified by the study center to schedule the 
Screening Visit. If a subject successfully completes their Screening Assessments, then Visit 1 
becomes the Baseline Visit.  
Once a subject arrives at the study center, a member of the Site Staff interviews the 
individual as to their qualifications for participation in the study, and if the subject wishes to 
continue, the informed consent form (ICF) is signed, and a subject number is assigned.  
If all eligibility criteria are met, the subject is randomized into the study.  
At Visit 1 (Stage 1 Day 1), eligible subjects were randomized, and the first dose of Treatment 
1 was administered at the study site at Time 0 min (note: if Visit 1 is split into 2 separate 
days, then dosing occurs on the second day). Efficacy and safety measurements were made 
before dosing with Treatment 1 (Time 0 min) and at multiple post-dose timepoints from 30 
min to 8 hours.  
In Stage 1, the safety and efficacy of Nyxol as a single agent was evaluated after 1 day and 7 
days of daily dosing.  
Following Visit 2 (Stage 1 Day 8), subjects discontinued dosing Treatment 1 for 7 to 14 days 
(Washout Period). During the Washout Period, subjects returned for Visit 3 (Stage 1 Day 10 
± 1 day) to assess the resolution of drug treatment effects.  


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During Stage 1, Treatment 1 is dosed daily in the evenings near bedtime starting with the day 
after completion of Visit 1, except for Visit 1 [Stage 1 Day 1], when the subject is dosed 
during the day at the study site, and the evening before Visit 2, when Treatment 1 is dosed as 
close to 12 hours prior to Time 0 min at Visit 2.  
Analysis will consist of comparison across the 2 arms: 
• Nyxol 
• Placebo 
Stage 2: Dosing of Treatments 1 and 2 
Visit 4 (Stage 2 Day 1) is the start of Stage 2. In Stage 2, the efficacy and safety of Nyxol + 
LDP was evaluated following 1 day and 7 days of dosing, in which Treatment 1 (Nyxol or 
placebo) is dosed daily in the evenings near bedtime, except the evening before Visits 4 and 
5, when Treatment 1 is dosed as close to 12 hours prior to the scheduled time for those visits. 
Treatment 2 (LDP or LDP vehicle) is dosed daily in the mornings. Treatment 2 will be dosed 
at the study site at the beginning of Visits 4 and 5 (Stage 2 Days 1 and 8, respectively).  
Analysis consists of comparison across the 4 arms:  
• Nyxol + LDP 
• Nyxol + LDP vehicle  
• Placebo + LDP 
• Placebo + LDP vehicle  
Other design considerations: 
Study medications (Treatments 1 and 2) were each dosed as a single drop in each eye, with 
Treatment 1 dosed in the evenings near bedtime and Treatment 2 dosed in the mornings. 
Baseline for the study is defined as pre-treatment (prior to Time 0 min) at Visit 1 
(Screening/Baseline).  
During Visit 2 (Stage 1 Day 8), Time 0 min is defined as the timing of the first efficacy 
assessments at the visit, approximately 12 hours after dosing with Nyxol the previous night 
(12 hour post-dose timepoint). During Visits 4 and 5 (Stage 2 Days 1 and 8, respectively), 
Time 0 min is defined as the time of Treatment 2 administration.  
The study eye is defined as the eligible eye with worse Baseline DCNVA at Visit 1 
(Screening/Baseline). In the case where both eyes have the same Baseline DCNVA, then the 
eye with the better BCDVA will be the study eye. If both DCNVA and BCDVA are 
equivalent between eyes, the study eye will be the right eye (OD). The study eye and fellow 
eye will both be evaluated at all assessments. 
The schedule for assessments and timing of events is presented in Table 1. 


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Table 1 
Schedule of Events for Stage 1 
  
Screening/Baseline Visit 1 [a] 
Visit 2[b] 
 
Visit 3[b] 
Day  
Stage 1 Day 1  
Stage 1 Day 8 
Washout Period 
7 – 14 days 
Stage 1 Day 10 
(± 1 day) 
Time (hr)[c]  
0 
0.5  
1  
3 
5 
8  
0 
0.5 
1 
3 
5 
8 
 
 
Informed consent   
X  
  
  
  
  
  
 
  
  
  
  
  
  
 
Subject ID # assigned   
X  
  
  
  
  
  
  
  
  
  
  
  
  
 
Medical/Ophthalmic history   
X  
  
  
  
  
  
 
  
  
  
  
  
  
 
Demographics    
X  
  
  
  
  
  
  
  
  
  
  
  
 
 
Prior/Concomitant medications  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
X 
Urine pregnancy test[d]  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
HR/BP  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
Manifest refraction and near add  
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
PD (photopic)[e]  
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
DCNVA (photopic)[f]  
X[g] 
 
X 
X 
X 
X 
X[g] 
X 
X 
X 
X 
X[g] 
 
X 
DCIVA (photopic) [f] 
X 
 
 
X 
X 
 
X 
 
X 
X 
 
X 
 
 
BCDVA (photopic)[f]  
X[g] 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X[g] 
 
X 
PD (mesopic)[e]  
X 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
DCNVA (mesopic)[f]  
X 
 
X 
X 
X 
X 
X 
 
 
X 
X 
X 
 
 
BCDVA (mesopic)[f]  
X[g] 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X[g] 
 
X  
Biomicroscopy  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
Corneal fluorescein staining and 
TBUT 
X 
 
 
 
 
 
 
 
 
 
 
 
 
  
IOP[h]  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
Ophthalmoscopy[i]  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
AEs  
X---------------------------------------------------------------------------------------------------------------------------X 
Randomization[a]  
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
In-office Treatment 1: Nyxol or 
placebo[j]  
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
Ocular tolerability[k]  
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
Subject questionnaire[l] 
X 
 
 
 
 
 
 
 
 
X 
 
 
 
 
Treatment 1: Nyxol or placebo and 
Subject diary dispensing/review [j]  
 
 
 
 
 
X 
 
 
 
 
 
X 
 
 
 


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AE, adverse event; BCDVA, best-corrected distance visual acuity; BP, blood pressure; DCNVA, distance-corrected near visual acuity; HR, heart rate; ID, identification; IOP, 
intraocular pressure; OD, right eye; OS, left eye; PD, pupil diameter; VA, visual acuity. 
[a] If the subject meets all the inclusion criteria and none of the exclusion criteria, this Screening Visit becomes the Baseline Visit.    
[b] Time of visits will be scheduled relative to the prior evening’s dose of Treatment 1. Visit 2 will be scheduled such that the 0 hour (Time 0 min) assessments on Visit 2 will 
occur approximately 12 hours (± 1 hour) after the prior evening’s dose of Treatment 1. Visit 3 occurs at Stage 1 Day 10 (±1 day) during the Washout Period. 
[c] An additional ± 5 min is permitted for measurements at 0.5 and 1 hour; an additional ± 10 min is permitted at 3, 5, and 8 hours for Visits 1 and 2.   
[d] Urine pregnancy test is for females of childbearing potential only.   
[e] PD performed with NeurOptics VIP-300 pupillometer.    
[f] VA performed binocularly, except where specifically noted [g]. 
[g] VA performed monocularly (OD, OS) following binocular assessment. 
[h] IOP performed with Tono-Pen or Goldmann applanation tonometry.   
[i] Direct or indirect ophthalmoscopy performed without dilation. Use of 90 D lens (indirect) is allowed.   
[j] Treatment 1: At Visit 1, one drop of Nyxol or placebo (ie, Nyxol vehicle) will be dosed at the study site after the completion of 0 hour (Time 0 min) ocular assessments. 
Subject will dose with Treatment 1 each evening near bedtime, starting the evening of Stage 1 Day 2 through the evening prior to Visit 2 (Stage 1 Day 8), except the evening 
before Visit 2, when Treatment 1 is dosed as close to 12 hours prior to Time 0 min for that visit. Subjects will discontinue dosing during the 7- to 14-day Washout Period. 
[k] Ocular tolerability performed using a 4-point scale; it will be assessed at the beginning of each specified timepoint after dosing at Visits 1 and 2. 
[l] Subject questionnaire (Protocol Appendix 5 and Appendix 6) will be completed by the subject at specified visits and timepoints.  
 
 
 


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Table 2 
Schedule of Events for Stage 2 
 
Visit 4[a] 
Visit 5 [a] 
  
Stage 2 Day 1 (Day 15 to 22) 
Stage 2 Day 8 (7 days after Stage 2 Day 1) 
Time (hr)[b]  12 hr post 
Treatment 1  
0  
0.5  
1  
3 
5  
8  
12 hr post 
Treatment 1  
0  
0.5  
1  
3 
5  
8  
Prior/Concomitant medications  
X 
 
 
 
 
 
X 
 
 
 
 
 
Urine pregnancy test[c]  
X 
  
 
 
 
 
X 
 
 
 
 
 
HR/BP  
 
 
 
 
 
 
 
 
 
 
 
 
X 
PD (photopic)[d]  
X 
  
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X 
DCNVA (photopic)[e]  
X 
 
X 
X 
X 
X 
X 
X 
 
X[f] 
X 
X 
X 
X[f] 
DCIVA (photopic)[e]  
X 
 
 
X 
X 
X 
X 
 
 
X 
X  
X  
BCDVA (photopic)[e]  
X 
 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X[f] 
PD (mesopic)[d]  
X 
 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X 
DCNVA (mesopic)[e]  
X 
 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X  
BCDVA (mesopic)[e]  
X 
  
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X[f] 
Biomicroscopy  
 
 
 
 
 
 
 
 
 
 
 
 
X 
IOP[g]  
 
 
 
 
 
 
 
 
 
 
 
 
X 
Ophthalmoscopy[h]  
 
 
 
 
 
 
 
 
 
 
 
 
X 
AEs  
           X------------------------------------------------------------------------------------------------------------------X 
In-office Treatment 2: LDP or LDP vehicle[i] 
 
X  
 
 
 
 
X  
 
 
 
Ocular tolerability[j]  
 
X  
 
 
 
 
X  
 
 
 
Subject questionnaire[k] 
 
 
 
 
X 
 
 
 
 
 
 
X 
 
 
Treatment 1: Nyxol or placebo and Subject 
diary dispensed / reviewed [l] 
  
 
 
 
 
 
X  
 
 
 
 
 
 
X 
Treatment 2: LDP or LDP vehicle and Subject 
diary dispensed / collected [i] 
 
 
 
 
 
 
X 
 
 
 
 
 
 
X 
AE, adverse event; BCDVA, best-corrected distance visual acuity; BP, blood pressure; DCNVA, distance-corrected near visual acuity; HR, heart rate; ID, identification; IOP, 
intraocular pressure; LDP, Low-Dose Pilocarpine; OD, right eye; OS, left eye; PD, pupil diameter; VA, visual acuity. 
[a] Time of visits will be scheduled relative to the prior evening’s dose of Treatment 1. Visits 4 and 5 will be scheduled such that the 12 hour post-dose assessments on Visits 
4 and 5 will occur approximately 12 hours (± 1 hour) after the prior evening’s dose of Treatment 1.  
[b] An additional ± 5 min is permitted for measurements at 0.5 and 1 hour; an additional ± 10 min is permitted at 3, 5, and 8 hours for Visits 4 and 5. 
[c] Urine pregnancy test is for females of childbearing potential only. 
[d] PD performed with NeurOptics VIP-300 pupillometer. 
[e] VA performed binocularly, except where specifically noted [f]. 


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[f] VA performed monocularly, following binocular assessment. 
[g] IOP performed with Tono-Pen or Goldmann applanation tonometry.   
[h] Direct or indirect ophthalmoscopy performed without dilation. Use of 90 D lens (indirect) is allowed.  
[i] Treatment 2: One drop of LDP or LDP vehicle in each eye dosed daily in the mornings beginning on the day after Visit 4 and daily through the day prior to Visit 5. At 
Visits 4 and 5, subject will be dosed with Treatment 2 at the study site. Subject will bring Treatment 2 kit and diary to Visits 4 and 5 for review and accountability. 
[j] Ocular tolerability performed using a 4-point scale; it will be assessed at the beginning of each specified timepoint after dosing at Visits 4 and 5.  
[k] Subject questionnaire (Protocol Appendix 5 and Appendix 6) will be completed by the subject at specified visits and timepoints.  
[l] Treatment 1: One drop of Nyxol or placebo in each eye dosed daily in the evenings near bedtime, except the evening before Visits 4 and 5, when Treatment 1 is dosed as 
close to 12 hours prior to the scheduled time for those visits. Subjects will bring their Treatment 1 kit and diary to Visits 4 and 5 for review and accountability. 
 
 


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5.2. 
Inclusion – Exclusion Criteria and General Study Population 
5.3. 
Subject Inclusion Criteria 
1. Males or females ≥ 40 and ≤ 64 years of age. 
2. Able to comply with all protocol-mandated procedures independently and to attend all 
scheduled office visits. 
3. Able and willing to give signed informed consent. 
4. Able to self-administer study medication throughout the study period. 
5. Best-corrected distance VA of 0.1 LogMAR (20/25 Snellen equivalent) or better in 
each eye in photopic conditions. 
6. Distance-corrected near VA of 0.4 LogMAR (20/50 Snellen equivalent) or worse but 
not > 0.7 LogMAR (20/100 Snellen equivalent) in photopic conditions in each eye and 
binocularly. 
7. For subjects who depend on reading glasses or bifocals, binocular best-corrected near 
VA is 0.1 LogMAR (20/25 Snellen equivalent) or better. 
8. Photopic PD of ≥ 3 mm. 
5.4. 
Subject Exclusion Criteria 
Subjects excluded from the study will be individuals with the following characteristics: 
Ophthalmic (in either eye): 
1. Use of any topical prescription (including Vuity) or over-the-counter (OTC) 
ophthalmic medications of any kind within 7 days of Screening until study completion, 
with the exception of lid scrubs with OTC products (eg, OCuSOFT® lid scrub, 
SteriLid®, baby shampoo, etc.) and artificial tears as specified in Exclusion #2 below. 
2. Use of any OTC artificial tears (preserved or unpreserved) during Visit days or 15 min 
before or after instillation of Treatment 1 or Treatment 2. 
3. Current use of any dry eye product such as topical ophthalmic therapy for dry eye (eg, 
generic cyclosporine, Restasis, Xiidra, Cequa, and Eysuvis) or intranasal dry eye 
product (eg, Tyrvaya) or other devices.  
4. Tear break-up time of < 5 seconds or corneal fluorescein staining Grade ≥ 2 in the 
inferior zone or Grade ≥ 1 in the central zone using the National Eye Institute scale.  
5. Clinically significant ocular disease (eg, cataract, glaucoma, corneal edema, uveitis, 
retinal degeneration, loss of visual field, or any macular pathology) that might 
interfere with the study as deemed by the Investigator. 


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6. Recent or current evidence of ocular infection or inflammation in either eye (such as 
current evidence of clinically significant blepharitis, conjunctivitis, keratitis, etc.). 
Subjects must be symptom free for at least 7 days. 
7. Any history of herpes simplex or herpes zoster keratitis. 
8. Known allergy, hypersensitivity, or contraindication to any component of the 
phentolamine, pilocarpine, or vehicle formulations.  
9. Prior participation in a study involving the use of Nyxol for the treatment of 
presbyopia. 
10. History of cauterization of the punctum or punctal plug (silicone or collagen) insertion 
or removal. 
11. Ocular trauma within 6 months prior to Screening.  
12. Ocular surgery or any ocular laser treatment within 6 months prior to Screening. 
13. Subjects with surgical monovision, multifocal or extended depth of focus IOLs are 
excluded (monofocal IOLs are accepted if in place > 6 months prior to Screening). 
14. History of any traumatic (surgical or nonsurgical) or nontraumatic condition affecting 
the pupil or iris (eg, irregularly shaped pupil, neurogenic pupil disorder, iris atrophy, 
iridotomy, iridectomy, iritis, etc.). 
15. Unwilling or unable to discontinue use of contact lenses at Screening until study 
completion. 
Systemic: 
16. Known hypersensitivity or contraindication to alpha- and/or beta-adrenoceptor 
antagonists (eg, chronic obstructive pulmonary disease or bronchial asthma; 
abnormally low BP or HR; second- or third-degree heart blockage or congestive heart 
failure). 
17. Known hypersensitivity or contraindication to any systemic cholinergic 
parasympathomimetic agent. 
18. Clinically significant systemic disease (eg, uncontrolled diabetes, myasthenia gravis, 
cancer, hepatic, renal, endocrine, or cardiovascular disorders) that might interfere with 
the study as deemed by the judgment of the Investigator. 
19. Initiation of treatment with, or any changes to, the current dosage, drug, or regimen of 
any systemic adrenergic or cholinergic drugs within 7 days prior to Screening or 
during the study; however, Flomax (tamsulosin) is specifically excluded. 
20. Participation in any investigational study within 30 days prior to Screening. 
21. Females of childbearing potential who are pregnant, nursing, planning a pregnancy, or 
not using a medically acceptable form of birth control. Acceptable methods include the 
use of at least one of the following: intrauterine device (IUD), hormonal (oral, 
injection, patch, implant, ring), barrier with spermicide (condom, diaphragm), or 
abstinence. A female is considered to be of childbearing potential unless she is 1 year 


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postmenopausal or 3 months post-surgical sterilization. All females of childbearing 
potential, including those with post−tubal ligation, must have a negative urine 
pregnancy test result at Visit 1 (Screening/Baseline).  
22. Resting HR outside the range of 50 to 110 beats per minute following at least a 5-min 
rest period in the sitting position at Visit 1 (Screening/Baseline). Heart rate may be 
repeated only once if outside the specified range, following another 5-min rest period 
in the sitting position. 
23. Hypertension with resting diastolic BP > 105 mmHg or systolic BP > 160 mmHg 
following at least a 5-min rest period in the sitting position at Visit 1 
(Screening/Baseline). Blood pressure may be repeated only once if outside the 
specified range, following another 5-min rest period in the sitting position. 
5.5. 
Randomization and Masking 
A randomization code for allocating subjects to each treatment group will be prepared by an 
unmasked biostatistician not connected with the study. Subjects will be then randomized 
1:1:1:1 into 1 of 4 treatment arms at Visit 1:  
• Nyxol in Stage 1; Nyxol + LDP in Stage 2 
• Nyxol in Stage 1; Nyxol + LDP vehicle in Stage 2 
• Placebo in Stage 1; Placebo + LDP in Stage 2 
• Placebo in Stage 1; Placebo + LDP vehicle in Stage 2 
Randomization will be stratified by iris color (light/dark irides). The number of subjects in 
each irides type should be as close as possible to 1:1, with a minimum of 40% within each 
stratum. 
The Treatment 1 (Nyxol/placebo) and Treatment 2 (LDP/LDP vehicle) study medications 
was masked to Investigators, Site Staff, study subjects, Ocuphire, and all other study 
personnel involved in the conduct of the study. Only in case of medical emergency or 
occurrence of SAEs will the randomization code be unmasked by the Medical Monitor and 
made available to the Investigator, Ocuphire, and/or other personnel involved in the 
monitoring or conduct of this study.  
5.6. 
Analysis Variables 
Variables to be summarized include demographics and baseline characteristics, medical 
(non-ocular) and ocular history, concomitant medications, and study drug accountability.  
Efficacy variables include: 
• Photopic and mesopic DCNVA (i.e., Near VA) 
• Photopic and mesopic BCDVA (i.e., Distance VA) 
• Photopic DCIVA (i.e., Intermediate VA) 


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• Photopic and mesopic PD 
• Subject questionnaire responses related to change in near vision and satisfaction with 
near vision 
Safety variables include: 
• AEs  
• Subjective ocular tolerability measured on a 4-point scale: 
o 0 – No discomfort 
o 1 – Mild discomfort 
o 2 – Moderate discomfort 
o 3 – Severe discomfort 
• IOP 
• BCDVA 
• Biomicroscopy  
• Ophthalmoscopy  
• Subject questionnaire 
• Vital signs (HR and BP) 
• Urine pregnancy tests for females of childbearing potential 
 


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6. 
SAMPLE SIZE 
Using α = 0.05 significance and a two-tailed test, 300 total subjects (150 subjects each in the 
Nyxol and placebo arms) who are evaluable for efficacy will provide greater than 80% power 
to detect a difference of 17% between treatments in percent of subjects with ≥ 15 letters of 
improvement in photopic binocular DCNVA and with < 5 letters of loss in photopic 
binocular BCDVA from Baseline at 12 hours post-dose at Visit 2 (Stage 1 Day 8). This 
calculation is based on a 12% placebo effect and a 29% Nyxol treatment effect. An additional 
20 subjects will be enrolled to account for dropouts. 
For the analysis of Stage 2, using α = 0.05 significance and a two-tailed test, 150 total 
subjects (75 each in the Nyxol + LDP and Placebo + LDP Vehicle arms) who are evaluable 
for efficacy will provide greater than 90% power to detect a difference of 34% between 
treatments in percent of subjects with ≥ 15 letters of improvement in photopic binocular 
DCNVA and with < 5 letters of loss in photopic binocular BCDVA from Baseline at Visit 5 
(Stage 2 Day 8), 30 minutes post-LDP/vehicle. This calculation is based on a 26% Nyxol + 
LDP Vehicle effect and a 60% Placebo + LDP treatment effect. An additional 5 subjects per 
treatment group will be enrolled to account for dropouts. Comparisons of Nyxol + LDP to 
the other treatment groups (Nyxol + LDP Vehicle and Placebo + LDP Vehicle) also provide 
approximately 90% power or more. 


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7. 
GENERAL CONSIDERATIONS 
7.1. 
Analysis Populations 
The following analysis populations will be defined for this study.  
7.1.1. 
Modified Intention-to-Treat (mITT) 
The mITT Population will include all randomized subjects who received at least 1 drop of 
Treatment 1 (Nyxol or placebo). The mITT Population will be used to analyze the primary 
endpoint as well as other efficacy endpoints, with subjects included in their randomized 
treatment regardless of the treatment they actually received. 
7.1.2. 
Per Protocol Stage 1 Population (PPS1) 
The PP Stage 1 (PPS1) Population will include all subjects in the mITT Population who 
receive 1 drop of Treatment 1 the day prior to Visit 2, have binocular DCNVA and BCDVA 
in photopic conditions at Baseline (Visit 1) and at 12 hours post-dose at Visit 2, and have no 
major protocol deviations considered to have significant impact on treatment outcome in 
Stage 1. The PPS1 Population will be used for analysis of select efficacy endpoints. 
7.1.3. 
Per Protocol Stage 2 Population (PPS2) 
The PP Stage 2 (PPS2) Population will include all subjects in the mITT Population who 
receive 1 drop of Treatment 1 the day prior to Visit 4 and Visit 5, receive 1 drop of 
Treatment 2 (LDP or LDP vehicle) at Visit 4 and Visit 5, have binocular DCNVA and 
BCDVA in photopic conditions at the 30 Minute timepoint at Visit 5 (after Treatment 2 
dosing), and have no major protocol deviations considered to have significant impact on 
treatment outcome in Stage 2. The PPS2 Population will be used for analysis of select 
efficacy endpoints. 
7.1.4. 
All Randomized Population (ARP) 
The ARP will include all randomized subjects. This population is also known as the 
Intention-to-Treat (ITT) Population. The ARP will be used in confirmatory efficacy analyses, 
with subjects included in their randomized treatment regardless of the treatment they actually 
received. 
7.1.5. 
Safety Population (SP) 
The SP will include all randomized subjects who have received at least 1 drop of study 
treatment (Treatment 1 or Treatment 2). The SP will be used to summarize safety variables, 
using the actual treatment a subject received. 


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7.2. 
Covariates and Subgroups 
7.2.1. 
Planned Covariates 
Planned covariates include baseline values for the given assessment.  
7.2.2. 
Planned Subgroups 
Subgroup analyses by irides type (light, dark) will be completed for efficacy endpoints.  
Other possible subgroups include age, sex, and race. 
7.3. 
Management of Analysis Data 
7.3.1. 
Data Handling 
Data from unscheduled visits may be used for the primary efficacy analysis as described in 
Section 7.3.2.3 below. Otherwise, unscheduled visit data will not be included in the analysis 
of efficacy or safety but will be listed.   
7.3.2. 
Missing Data 
The primary efficacy endpoint is the percent of subjects with ≥ 15 letters of improvement in 
photopic binocular DCNVA and with < 5 letters of loss in photopic binocular BCDVA from 
Baseline comparing Nyxol-treated subjects to placebo-treated subjects at 12 hours post-dose 
at Visit 2 (Stage 1 Day 8). For the analysis of the primary efficacy and key secondary 
endpoints, imputation will be performed for missing efficacy data as specified in Section 
7.3.2.3 for the analysis using the mITT, as well as either PP population, if required. 
Confirmatory analyses may be performed using the ARP, also using imputation for missing 
data.   
Otherwise, there will be no substitutions made to accommodate missing data points for 
efficacy data. All data recorded on the CRF will be included in data listings that will 
accompany the CSR. 
Safety data will be imputed in limited situations. If the severity of an AE is missing, then the 
severity will remain missing. If relationship of the AE to study drug is missing, the 
relationship will remain missing. Missing or partial dates for AEs or concomitant 
medications will be imputed as described in Section 7.3.2.1. Otherwise, all summaries of 
safety endpoints will be completed using observed cases in the SP; no imputation will be 
completed. 
7.3.2.1. 
Handling of Missing Date Values 
Partial or Missing Dates 


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The following conventions will be used to impute missing portions of dates for AEs and 
concomitant medications, if warranted. Note that the imputed values outlined here may not 
always provide the most conservative date. In those circumstances, the imputed value may be 
replaced by a date that will lead to a more conservative analysis. 
A. 
Start Dates 
1) 
If the year is unknown, then the date will not be imputed and will 
be assigned a missing value.  
2) 
If the month is unknown, then: 
i) If the year matches the first dose date year, then impute 
the month and day of the first dose date. 
ii) Otherwise, assign ‘January.’  
3) 
 If the day is unknown, then: 
i) If the month and year match the first dose date month 
and year, then impute the day of the first dose date. 
ii) Otherwise, assign the first day of the month. 
B. 
Stop Dates 
1) 
If the year is unknown, then the date will not be imputed and will 
be assigned a missing value. 
2) 
If the month is unknown, then assign ‘December.’ 
3) 
If the day is unknown, then assign the last day of the month. 
 
7.3.2.2. 
Missing Baseline Data 
Every effort will be made to ensure that accurate baseline information on the subjects is 
collected. In the event that a subject is missing baseline information that is required for the 
primary efficacy analyses, the subject will be included in the SP for assessment of safety and 
excluded from the primary analyses. Each case of missing baseline data will be evaluated for 
potential inclusion in the exploratory endpoints. All baseline data will be observed cases, 
without imputation. 
7.3.2.3. 
Imputation Methods 
Imputation for efficacy data will only be performed for the primary efficacy endpoint using 
the mITT and ARP, as well as either PP population, if required. The primary efficacy time 
point at Visit 2 (12 hours after the dose of Treatment 1 study medication) is the first 
scheduled efficacy assessment at Visit 2; if the primary efficacy time point is not collected, 
then the most recent valid PD assessment prior to Visit 2 will be used for the primary 
analysis, which will usually be the last assessment taken at Visit 1 or from an unscheduled or 
early termination visit after Visit 1 but prior to the scheduled Visit 2. 
For later time points at Visit 2, an analysis with last observation carried forward (LOCF) for 
missing data will be applied if at least one Visit 2 time point is collected. Visit 1 data or any 


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other data collected prior to the scheduled Visit 2 will not be used for imputation for any time 
points, except for 12 hours after the dose of Treatment 1 study medication.  
For Visit 5, if the 12 hours after the dose of Treatment 1 study medication is not collected, 
this timepoint will be imputed using the most recent valid assessment prior to Visit 5 and 
while exposed to Treatment 2.  
For Visit 4 and Visit 5, an LOCF analysis will be conducted if there is at least one time point 
with observed data after the administration of Treatment 2 at that visit. If a subject misses 
either Visit 4 or Visit 5, no imputation for the missing visit data will be employed, except for 
the 12 hours after the dose of Treatment 1 study medication at Visit 4 and Visit 5.   
7.3.3. 
Handling of Early Termination Visit Information 
In the event that a subject is terminated early from the study, the early termination data will 
be assigned to the closest scheduled visit and/or time point. If the closest visit or time point 
has valid data, the early termination data will be assigned to the next available visit or time 
point. 
7.3.4. 
Pooling of Investigational Sites 
The data from all study centers will be pooled together for all planned analyses. 
7.3.5. 
Coding Conventions for Events and Medications 
All AEs and medical history will be mapped to the Medical Dictionary for Regulatory 
Activities (MedDRA Version 25.1) system for reporting (preferred term and body system). 
Prior and concomitant medications will be coded using WHO-DD (World Health 
Organization Drug Dictionary) (Version Global C3, September 1, 2022). 
7.3.6. 
Analysis Software 
Data manipulation, tabulation of descriptive statistics, calculation of inferential statistics, and 
graphical representations will be performed primarily using SAS (release 9.4 or higher) for 
Windows. If the use of other software is warranted, the final CSR will detail what software 
was used and for what purposes. 
7.3.7. 
Study Data 
Study data identified in the schedules for time and events (


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Table 1 and Table 2) are collected, and source verified, on the electronic data capture (EDC) 
Fountayn Version 1.0.0.  
All study data will be formulated into regulatory-compliant data sets to provide transparency, 
traceability, and integrity of trial analysis results from the collection source. Observed study 
data will be mapped to the Clinical Data Interchange Standards Consortium (CDISC) Study 
Data Tabulation Model (SDTM) and serve as the source data from the trial. All study 
analyses will be completed using analysis data sets that are derived from the SDTM and 
follow the CDISC Analysis Data Model (ADaM) architecture.  
The methods for programming the CDISC SDTM and ADaM data sets are described in 
Figure 1. 
Figure 1 
SDTM, ADaM, and TFL Development and Validation 
 
Where: 
1. Development, validation, and maintenance of SDTM domains 
2. Development and validation of ADaM data sets, with input source the appropriate 
SDTM domains. 
3. Development and validation of tables, figures, and listings (TFL), with input data 
source the SDTM domains and analysis specific ADaM data sets. 


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7.4. 
Planned Study Analyses 
7.4.1. 
Statistical Summaries: Descriptive and Inferential 
Categories for data presentation and analysis will consist of each treatment group: Nyxol and 
placebo for Stage 1, and Nyxol + LDP, Nyxol + LDP vehicle, Placebo + LDP, and Placebo + 
LDP vehicle for Stage 2.  
All statistical tests will be two-sided and a difference resulting in a p-value of less than or 
equal to 0.05 will be considered statistically significant. The p-values for the analysis of 
secondary efficacy endpoints and safety endpoints will be considered descriptive, unless the 
endpoint is used for the hierarchical testing method. All p-values will be rounded to and 
displayed in four decimals. If a p-value less than 0.0001 occurs, it will be shown in tables as 
<0.0001. 
Descriptive summaries of variables will be provided where appropriate. For continuous 
variables, the number of non-missing values (n), mean, standard deviation, median, 
minimum, and maximum will be tabulated by treatment group. For categorical variables, the 
counts and proportions of each value will be tabulated by treatment group. Expansion of 
descriptive table categories within each treatment may occur if such elaborations are thought 
to be useful. 
All study-related data collected will be presented in listings. Study-related data not subject to 
analysis according to this plan will not appear in any tables or graphs but will be included in 
the data listings. 
7.4.2. 
Interim Analyses and Data Monitoring 
No formal interim analysis or safety monitoring committee is planned for this study.  
7.4.3. 
Final Analysis and Publication of Study Results 
The final analysis will be completed after all subjects have completed the study.   
7.5. 
Multiple Testing Procedures 
A hierarchical analysis will formally test a family of endpoints beyond the primary efficacy 
endpoint. The percent of subjects with ≥ 15 letters of improvement in photopic binocular 
DCNVA and with < 5 letters of loss in photopic binocular BCDVA will be analyzed at 30 
min post-LDP/vehicle at Visit 5 (Stage 2 Day 8 [Day 22-29]), comparing Nyxol + LDP to 
placebo + LDP vehicle, Nyxol + LDP vehicle, and placebo + LDP. These comparisons will 
be analyzed second, third, and fourth in the testing hierarchy. The full hierarchy is in Section 
12.3. 
Otherwise, there will be no adjustments for multiplicity and no formal multiple testing 
procedures are to be implemented with this analysis plan. 


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7.6. 
Baseline Values 
Baseline values are the values obtained at Visit 1, 0 Hour (Time 0 min), prior to any 
treatment administration. If the Visit 1, 0 Hour assessments are missing, any value collected 
prior to treatment administration will be treated as the baseline. 
 


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8. 
SUMMARY OF STUDY DATA 
8.1. 
Subject Disposition 
A summary of the analysis sets includes the number and percentage of subjects by Stage 1 
treatment group and overall for the following categories: subjects in the ARP, subjects in the 
SP, subjects in the mITT Population, and subjects in the PP Population. All percentages will 
be based on the number of subjects in the ARP. A similar summary will be presented by 
Stage 2 treatment group. 
End of trial information will also be summarized in these tables, including the number of 
subjects completing the study, the number of subjects who prematurely discontinued the 
study with reasons for withdrawal, the number of subjects completing the study medication 
dosing, and the number of subjects who prematurely discontinued the study medication with 
reasons for study medication discontinuation.   
A by-subject data listing of study completion information including the reason for premature 
study withdrawal, if applicable, will be presented. 
8.2. 
Protocol Deviations 
Major protocol deviations, as determined by a Sponsor blinded review of the data prior to 
database lock and unblinding of the study, may result in the removal of a subject’s data from 
the PP Population. The Sponsor or designee will be responsible for producing the final 
deviation file; this file will include a description of the protocol deviation and clearly identify 
whether this violation warrants exclusion from the PP Population. This file will be finalized 
prior to database lock. 
All protocol deviations will be presented in a by-subject data listing, with a flag to indicate if 
a deviation was considered major and/or exclusionary.  
8.3. 
Demographics and Baseline Characteristics 
Subject demographic data and baseline characteristics will be tabulated and summarized 
descriptively by Stage 1 treatment group, Stage 2 treatment group, and overall. The 
demographic data and baseline characteristics will be summarized for the mITT Population, 
PP Population, SP, and ARP. If the mITT population is equivalent to any of the other 
populations, then only the mITT version will be generated rather than repeating equivalent 
summaries. 
The demographics consist of age (year), sex, race, ethnicity, and study eye, iris type (light, 
dark). A subject’s age in years is calculated using the date of the informed consent and date 
of birth. Age will be summarized using descriptive statistics. The number and percentage of 
subjects by sex, race, ethnicity, study eye, and iris type will also be reported. Percentages will 
be based on the total number of subjects in the study population presentation. 


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The following baseline characteristics will be summarized for study eye, fellow eye, and 
binocular (if applicable), using descriptive statistics: 
• BCDVA (photopic and mesopic) 
• DCNVA (photopic and mesopic) 
• DCIVA (photopic)  
• Pupil diameter (photopic and mesopic) 
• IOP 
All demographic and baseline information will be presented in by-subject listings. 
8.4. 
Medical History  
The number and percent of subjects with individual medical histories will be summarized for 
all subjects by Stage 1 treatment group, Stage 2 treatment group, and overall. Non-ocular and 
ocular medical history will be summarized separately.  
Medical history will be coded using the MedDRA Version 25.1 The number and percentage 
of subjects with any medical history will be summarized overall and for each system organ 
class (SOC) and preferred term (PT). Percentages will be calculated based on number of 
subjects in the SP.   
Subject medical history data including specific details will be presented in by-subject 
listings. 
8.5. 
Prior and Concomitant Medications 
The number and percentages of all concomitant medications will be summarized by Stage 1 
treatment group, Anatomical Therapeutic Chemical (ATC) level 4, and PT. The total number 
of concomitant medications and the number and percentages of subjects with at least 1 
concomitant medication will be summarized by Stage 1 treatment group. A similar summary 
will be presented by Stage 2 treatment group. All summaries will be performed using the SP.   
A concomitant medication is defined as any medication taken on or after the day of first 
exposure to study drug. 
Prior medications are defined as any medication that has a start and stop date prior to the day 
of first exposure to any study drug, collected from up to 30 days prior to Screening. The total 
number of prior medications and the number and percentages of subjects with at least 1 prior 
medication will be summarized by Stage 1 treatment group, Stage 2 treatment group, and 
overall. 


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8.6. 
Treatment Exposure and Compliance 
Exposure to Treatment 1 for the evening of Stage 1 Day 2 through the evening prior to Visit 
2 (Stage 1 Day 8) will be determined for each subject in the SP by counting the total number 
of days that Treatment 1 was taken, according to the subject diary. Compliance will be 
determined by calculating the percent of days exposed to the number of days that the subject 
should have taken Treatment 1 during the same period. Treatment 1 exposure and 
compliance will be tabulated and summarized descriptively by Stage 1 treatment group and 
overall. 
For Stage 2, exposure to Treatment 1 will be assessed between Visits 4 and 5, when subjects 
are expected to dose daily in the evenings, using the subject diary. Compliance will be 
determined by calculating the percent of days exposed to the number of days that the subject 
should have taken Treatment 1 during the same period. Separately, exposure to Treatment 2 
will be assessed between Visits 4 and 5, when subjects are expected to dose daily in the 
mornings, also via the subject diary. Compliance for Treatment 2 will be calculated similarly 
to Treatment 1. Treatment 1 and Treatment 2 exposure and compliance will be tabulated and 
summarized descriptively by Stage 2 treatment group and overall.  
In-office treatment administration data and diary data for both Stage 1 and Stage 2 will be 
presented in by-subject listings.  


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9. 
EFFICACY ANALYSES 
Unless otherwise noted, efficacy will be assessed using the mITT and PP populations, with 
subjects included in their randomized treatment regardless of the treatment they actually 
received. For the analysis of the primary efficacy endpoint, imputation will be performed for 
missing data as described in Section 7.3.2.3. Confirmatory analysis of the primary efficacy 
endpoint may be performed using the ARP, if different from the mITT, also using imputation 
for missing data.  
All efficacy assessment data, regardless of whether they are included in the analysis, will be 
presented in by-subject listings.  
9.1. 
Clinical Efficacy   
All efficacy data will be summarized by treatment group for the Baseline assessment; for 
Visit 1 (Stage 1 Day 1) post-treatment time points (0.5 hours, 1 hour, 3 hours, 5 hours, and 8 
hours); Visit 2 (Stage 1 Day 8) time points (12 hour post-Treatment 1 dose, 12.5 hours, 13 
hour, 15 hours, 17 hours, and 20 hours); Visit 3 (Stage 1 Day 10); Visit 4 (Stage 2 Day 1) 
time points (12 hour post-Treatment 1 dose, 0.5 hours, 1 hour, 3 hours, 5 hours, and 8 hours 
post-Treatment 2); and Visit 5 (Stage 2 Day 8) time points (12 hour post-Treatment 1 dose, 
0.5 hours, 1 hour, 3 hours, 5 hours, and 8 hours post-Treatment 2). 
9.1.1. 
Primary Efficacy Analysis and Key Secondary Analyses 
The primary efficacy endpoint is the percent of subjects with ≥ 15 letters of improvement in 
photopic binocular DCNVA and with < 5 letters of loss in photopic binocular BCDVA on 
Visit 2 at the 12 hour post-dose timepoint for Nyxol compared to Placebo. If the analysis 
using the PP Population shows a positive effect for Nyxol at the 0.05 level of significance, 
the primary endpoint will be considered met. The primary efficacy endpoint and key 
secondary endpoints will be analyzed using a logistic regression model with treatment and 
light/dark irides as factors and the baseline DCNVA as a covariate. The percentage of 
subjects in each treatment group meeting the criteria, the odds ratio (OR) with 95% CI, and 
p-value will be provided. Example SAS code is as follows: 
proc logistic data=efficacydata;  
class trtname irides;  
model outcome = trtname irides base/cl rl firth; 
run; 
 
By including irides type as a factor in the primary efficacy analysis model, the model 
efficiency as well as a change in the treatment effect will be increased. Including this factor 
in the model will also make the results more generalizable to other studies in which the 
sample characteristics may differ from the current study [2].  


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In addition, the primary efficacy endpoint will be analyzed by light/dark irides using the 
same model indicated above but without irides as a factor. For this subgroup analyses, 
observed case data only will be used; that is, missing values will not be imputed.   
9.1.2. 
Secondary Efficacy Analyses  
Secondary efficacy endpoints are indicated in Section 4.2.2. Secondary efficacy endpoints 
will be analyzed by study eye, fellow eye, and binocular (if available) using the the PPS1 and 
PPS2 populations, and the mITT population for selected endpoints (see the list of planned 
tables, Section 12.1, for details).  
All continuous secondary endpoints derived from VA assessments, such as change in 
DCNVA, DCIVA, and BCDVA, will be analyzed using Early Treatment Diabetic 
Retinopathy Study (ETDRS) letters correctly read. Any secondary endpoints using Snellen 
measurements will be converted from letters read as follows: 
Table 3 
Snellen Equivalents to VA Assessment Letters Read 
DCNVA/DCIVA letters 
Snellen 
BCDVA letters 
≥75 
20/15 
≥60 
70 - 74 
20/20 
55 - 59 
65 - 69 
20/25 
50 - 54 
60 - 64 
20/32 
45 - 49 
55 - 59 
20/40 
40 - 44 
50 - 54 
20/50 
35 - 39 
45 - 49 
20/63 
30 - 34 
40 - 44 
20/80 
25 - 29 
35 - 39 
20/100 
20 - 24 
30 – 34 
20/125 
15 - 19 
25 - 29 
20/160 
10 - 14 
<25 
20/200 
<10 
 
Each of the continuous secondary efficacy endpoints will be analyzed using analysis of 
covariance (ANCOVA), with observed value as the dependent variable, treatment, and 
light/dark irides as factors, and the respective baseline value included as the covariate. 
Another ANCOVA will be applied with change from baseline as the dependent variable 
using the same factors and covariates. 
The output from each ANCOVA will include the LSM and SE for all treatment groups, along 
with each LSM, the 95% CI and associated p-value for Nyxol vs. Placebo for Stage 1 time 
points, and Nyxol + LDP vs. Nyxol + LDP vehicle, Placebo + LDP, and Placebo + LDP 
vehicle for Stage 2 time points. Continuous pupil diameter endpoints will be assessed using 
the study eye, fellow eye, and best eye, which is defined as the eye with the greatest 
reduction in diameter. 
For each of the secondary endpoints related to percent of subjects achieving certain criteria, 
the analysis will be performed using a logistic regression model with treatment and light/dark 
irides as factors, and the respective baseline as a covariate. For each analysis, the percentage 


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of subjects in each treatment group meeting the criteria will be presented, and the OR with 
95% CI and p-value will be provided for Nyxol vs. Placebo for Stage 1 time points, and 
Nyxol + LDP vs. Nyxol + LDP vehicle, Placebo + LDP, and Placebo + LDP vehicle for 
Stage 2 time points. Categorical pupil diameter endpoints will be assessed using the study 
eye, fellow eye, and either eye, which is defined as achieving a given criterion in either eye. 
In addition, each secondary efficacy endpoint will be analyzed by light/dark irides using the 
same model indicated above but without irides as a factor.  
Finally, select questions from the Subject Questionnaire will be analyzed.  The results for 
Question 1 (change in near vision) and Question 5 (satisfaction with near vision) will be 
analyzed at Visit 2 (Stage 1 Day 8) and Visit 5 (Stage 2 Day 8) using an ANCOVA model 
with numeric observed response as the dependent variable (1=Far worse to 5=Far better for 
Question 1, and 1=Very dissatisfied to 5=Very satisfied for Question 5) and treatment as a 
factor. Only observed subject questionnaire data will be used; no imputation will be 
performed for missing data. 


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10. 
SAFETY ANALYSES 
All safety analyses will be conducted using the SP unless specified otherwise below. All 
safety analyses will be completed using the actual treatment a subject received. Observed 
case data will be used; no imputation will be performed for missing safety data except for the 
limited situations described in Section 7.3.2. 
All safety data will be presented in by-subject listings. Unscheduled assessments will not be 
summarized but will be included in the listings.  
10.1. 
Adverse Events 
AEs will be coded using MedDRA, Version 25.1.   
Treatment-emergent adverse events (TEAEs) are defined as any AE that begins or worsens 
after initiation of the investigational product and through the subject’s last study visit (study 
completion or early termination). 
If the onset of an AE is on or after the date of first dose of study medication or is increasing 
in severity after first dose of study medication, then the AE will be considered treatment 
emergent. 
Only TEAEs will be summarized; all AEs (TEAE, non-TEAE) will be included in a by-
subject listing.  
TEAEs that start during Stage 1 will only be counted in the Stage 1 tables using the 
Treatment 1 that was actually taken, regardless of the TEAE stop date. TEAEs that start 
during Stage 2 will be counted in both Stage 1 and Stage 2 tables. 
The number and percent of subjects with any TEAEs will be summarized by SOC and PT by 
treatment group and overall. At each level of tabulation (e.g., at the PT level), subjects will 
be counted only once if they had more than one such event reported during the AE collection 
period.  
 
Note that in MedDRA, ocular events are coded to the SOC of “Eye Disorders”. Thus, using 
SOC in the summaries will provide a separation of ocular and non-ocular adverse events. 
The following summary tables will be presented for TEAE data:  
• 
Overall summary of TEAEs (completed for both the SP and mITT population) 
• 
Summary table of TEAEs by SOC and PT  
• 
Summary table of TEAEs by SOC, PT, and by greatest relationship level to study 
drug (not related, unlikely related, possibly related, probably related, definitely 
related, or unknown) 
• 
Summary table of TEAEs by SOC, PT, and maximum severity (mild, moderate, 
severe) 


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• 
Summary table of serious TEAEs by SOC and PT  
• 
Summary table of TEAEs leading to withdrawal from the study by SOC and PT 
 
• 
Summary table of TEAEs leading to study medication discontinuation by SOC and 
PT 
 
10.2. 
Deaths, Serious Adverse Events and Other Significant Adverse Events 
10.2.1. 
Deaths 
The AE listing will include all AEs, including deaths, regardless of causality; one of the 
columns in the listing will specify whether the AE was fatal.     
10.2.2. 
Serious Adverse Events 
The AE listing will include all AEs, including SAEs; one of the columns in the listing will 
specify whether the AE was an SAE.     
10.2.3. 
Adverse Events Leading to Withdrawal from the Study 
The AE listing will include all AEs, including AEs leading to withdrawal from the study; one 
of the columns in the listing will specify whether the AE led to withdrawal from the study.   
10.2.4. 
Adverse Events Leading to Discontinuation of Study Medication 
The AE listing will include all AEs, including AEs leading to discontinuation of study 
medication; one of the columns in the listing will specify whether the AE led to 
discontinuation of study medication.   
10.3. 
Subjective Ocular Tolerability 
Results from the subjective ocular tolerability assessment, measured on a 4-point scale, will 
be summarized descriptively using counts and percentages for each treatment group at the 0 
hour time point for each visit. Additionally, the categories “No Discomfort” and “Mild 
Discomfort” will be pooled into a single category and summarized descriptively, as will the 
categories “Moderate Discomfort” and “Severe Discomfort”.  
Treatments will be compared for the two pooled categories using a Fisher’s exact test for 
Nyxol vs. Placebo for Stage 1 time points, and Nyxol + LDP vs. Nyxol + LDP vehicle, 
Placebo + LDP, and Placebo + LDP vehicle for Stage 2 time points. Separate summaries will 
be created for the study eye and the fellow eye.  


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10.4. 
Vital Signs 
Descriptive statistics of observed values will be presented for vital sign data at baseline, Visit 
2 (Stage 1 Day 8) at the 12 hour post-dose time point, and Visit 5 (Stage 2 Day 8) at the 8 
hour time point by treatment group. Changes from baseline to each time point will be 
presented.  
10.5. 
IOP 
Observed values and change from baseline in IOP at baseline, Visit 2 (Stage 1 Day 8) at the 
12 hour post-dose time point, and Visit 5 (Stage 2 Day 8) at the 8 hour time point will be 
summarized for the study eye and the fellow eye by treatment group. Treatments will be 
compared using the same ANCOVA model proposed for the continuous secondary efficacy 
endpoints. 
10.6. 
Subject Questionnaire 
Results from the subject questionnaire at baseline, and the 3 hour time point for Visit 2 
(Stage 1 Day 8), Visit 4 (Stage 2 Day 1) and Visit 5 (Stage 2 Day 8) will be summarized by 
treatment group.  
10.7. 
Biomicroscopy and Ophthalmoscopy 
Results from biomicroscopic and ophthalmoscopic examinations at baseline, Visit 2 (Stage 1 
Day 8) at the 12 hour post-dose time point, and Visit 5 (Stage 2 Day 8) at the 8 hour time 
point will be summarized by treatment group for the study eye and the fellow eye by 
treatment group. 
10.8. 
Other Safety Measures 
The percent of subjects who lose ≥ 5 letters from Baseline in photopic BCDVA will be 
summarized within the summary of secondary efficacy endpoint Percentage of subjects with 
loss or improvement in photopic and mesopic BCDVA from baseline. Treatments will be 
compared using the same logistic regression model proposed for the secondary efficacy 
endpoints. 
Urine pregnancy tests for females of childbearing potential will be presented in by-subject 
listings. Dry eye tests (tear break-up time and cornea fluorescein staining), performed only at 
baseline, will also be listed. 
 
 


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11. 
REFERENCES 
[1] ICH E9 Expert Working Group. Statistical Principles for Clinical Trials: ICH 
Harmonized Tripartite Guideline, September 1998 
[2] Hauck WM, Anderson S, and Marcus SM, Should We Adjust for Covariates in Nonlinear 
Regression Analyses of Randomized Trials? Controlled Clin Trials 1998;19:249–256 
 
 


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12. 
APPENDICES 
12.1. 
List of Planned Tables 
The list of planned tables includes all of the main tables to be presented for the study. Cells 
that are highlighted in yellow are what we are considering for the top-line report.  
Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
14.1.1 
Subject Disposition  
X 
 
 
 
 
14.1.2.1.1 - 5 Demographics and Baseline Characteristics 
X 
X 
X 
X 
X 
14.1.3.1 
Non-ocular Medical History  
 
X 
 
 
 
14.1.3.2 
Ocular Medical History  
 
X 
 
 
 
14.1.4.1 
Prior Medications  
 
X 
 
 
 
14.1.4.2.1 
Concomitant Medications – Stage 1 
 
X 
 
 
 
14.1.4.2.2 
Concomitant Medications – Stage 2 
 
X 
 
 
 
14.1.5.1 
Study Treatment Exposure and Compliance – 
Stage 1 
 
X 
 
 
 
14.1.5.2 
Study Treatment Exposure and Compliance – 
Stage 2 
 
X 
 
 
 
14.2.1.1.1/2 
Percent of Subjects with ≥ 15 Letters of 
Improvement in Photopic DCNVA and < 5 
Letters of Loss in Photopic Binocular 
BCDVA by Time Point – Stage 1 
 
 
X 
X 
 
14.2.1.2.1/2 
Percent of Subjects with ≥ 15 Letters of 
Improvement in Photopic DCNVA and < 5 
Letters of Loss in Photopic Binocular 
BCDVA by Time Point – Stage 2 
 
 
X 
 
X 
14.2.1.3.1/2 
Percent of Subjects with ≥ 15 Letters of 
Improvement in Photopic DCNVA and < 5 
Letters of Loss in Photopic Binocular 
BCDVA by Time Point and Irides Type – 
Stage 1 
 
 
X 
X 
 
14.2.1.4.1/2 
Percent of Subjects with ≥ 15 Letters of 
Improvement in Photopic DCNVA and < 5 
Letters of Loss in Photopic Binocular 
BCDVA by Time Point and Irides Type – 
Stage 2 
 
 
X 
 
X 
14.2.2.1.1/2 
Percent of Subjects with Improvement in 
Photopic DCNVA by Time Point – Stage 1 
 
 
X 
X 
 


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Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
14.2.2.2.1/2 
Percent of Subjects with Improvement in 
Photopic DCNVA by Time Point – Stage 2 
 
 
X 
 
X 
14.2.2.3.1/2 
Percent of Subjects with Improvement in 
Photopic DCNVA by Time Point and Irides 
Type – Stage 1 
 
 
X 
X 
 
14.2.2.4.1/2 
Percent of Subjects with Improvement in 
Photopic DCNVA by Time Point and Irides 
Type – Stage 2 
 
 
X 
 
X 
14.2.3.1.1/2 
Observed Values and Change from Baseline 
in Photopic DCNVA (Letters Read) by Time 
Point – Stage 1 
 
 
X 
X 
 
14.2.3.2.1/2 
Observed Values and Change from Baseline 
in Photopic DCNVA (Letters Read) by Time 
Point – Stage 2 
 
 
X 
 
X 
14.2.3.3.1/2 
Observed Values and Change from Baseline 
in Photopic DCNVA (Letters Read) by Time 
Point and Irides Type – Stage 1 
 
 
X 
X 
 
14.2.3.4.1/2 
Observed Values and Change from Baseline 
in Photopic DCNVA (Letters Read) by Time 
Point and Irides Type – Stage 2 
 
 
X 
 
X 
14.2.4.1 
Percent of subjects with photopic DCNVA ≤ 
20/15, ≤ 20/20, ≤ 20/25, ≤ 20/32, ≤ 20/40, ≤ 
20/50, ≤ 20/63, ≤ 20/80, ≤ 20/100, ≤ 20/125, 
and ≤ 20/160 by Time Point – Stage 1 
 
 
 
X 
 
14.2.4.2 
Percent of subjects with photopic DCNVA ≤ 
20/15, ≤ 20/20, ≤ 20/25, ≤ 20/32, ≤ 20/40, ≤ 
20/50, ≤ 20/63, ≤ 20/80, ≤ 20/100, ≤ 20/125, 
and ≤ 20/160 by Time Point – Stage 2 
 
 
 
 
X 
14.2.4.3 
Percent of subjects with photopic DCNVA ≤ 
20/15, ≤ 20/20, ≤ 20/25, ≤ 20/32, ≤ 20/40, ≤ 
20/50, ≤ 20/63, ≤ 20/80, ≤ 20/100, ≤ 20/125, 
and ≤ 20/160 by Time Point and Irides Type – 
Stage 1 
 
 
 
X 
 
14.2.4.4 
Percent of subjects with photopic DCNVA ≤ 
20/15, ≤ 20/20, ≤ 20/25, ≤ 20/32, ≤ 20/40, ≤ 
20/50, ≤ 20/63, ≤ 20/80, ≤ 20/100, ≤ 20/125, 
and ≤ 20/160 by Time Point and Irides Type – 
Stage 2 
 
 
 
 
X 
14.2.5.1 
Percent of Subjects with Improvement in 
Photopic DCIVA by Time Point – Stage 1 
 
 
 
X 
 


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Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
14.2.5.2 
Percent of Subjects with Improvement in 
Photopic DCIVA by Time Point – Stage 2 
 
 
 
 
X 
14.2.6.1 
Observed Values and Change from Baseline 
in Photopic DCIVA (Letters Read) by Time 
Point – Stage 1 
 
 
 
X 
 
14.2.6.2 
Observed Values and Change from Baseline 
in Photopic DCIVA (Letters Read) by Time 
Point – Stage 2 
 
 
 
 
X 
14.2.7.1 
Percent of Subjects with Improvement or 
Loss in Photopic BCDVA by Time Point – 
Stage 1 
 
 
 
X 
 
14.2.7.2 
Percent of Subjects with Improvement or 
Loss in Photopic BCDVA by Time Point – 
Stage 2 
 
 
 
 
X 
14.2.7.3 
Percent of Subjects with Improvement or 
Loss in Photopic BCDVA by Time Point and 
Irides Type – Stage 1 
 
 
 
X 
 
14.2.7.4 
Percent of Subjects with Improvement or 
Loss in Photopic BCDVA by Time Point and 
Irides Type – Stage 2 
 
 
 
 
X 
14.2.8.1 
Observed Values and Change from Baseline 
in Photopic BCDVA (Letters Read) by Time 
Point – Stage 1 
 
 
 
X 
 
14.2.8.2 
Observed Values and Change from Baseline 
in Photopic BCDVA (Letters Read) by Time 
Point – Stage 2 
 
 
 
 
X 
14.2.8.3 
Observed Values and Change from Baseline 
in Photopic BCDVA (Letters Read) by Time 
Point and Irides Type – Stage 1 
 
 
 
X 
 
14.2.8.4 
Observed Values and Change from Baseline 
in Photopic BCDVA (Letters Read) by Time 
Point and Irides Type – Stage 2 
 
 
 
 
X 
14.2.9.1 
Percent of Subjects with Improvement or 
Loss in Mesopic BCDVA by Time Point – 
Stage 1 
 
 
 
X 
 
14.2.9.2 
Percent of Subjects with Improvement or 
Loss in Mesopic BCDVA by Time Point – 
Stage 2 
 
 
 
 
X 


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Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
14.2.9.3 
Percent of Subjects with Improvement or 
Loss in Mesopic BCDVA by Time Point and 
Irides Type – Stage 1 
 
 
 
X 
 
14.2.9.4 
Percent of Subjects with Improvement or 
Loss in Mesopic BCDVA by Time Point and 
Irides Type – Stage 2 
 
 
 
 
X 
14.2.10.1 
Observed Values and Change from Baseline 
in Mesopic BCDVA (Letters Read) by Time 
Point – Stage 1 
 
 
X 
X 
 
14.2.10.2 
Observed Values and Change from Baseline 
in Mesopic BCDVA (Letters Read) by Time 
Point – Stage 2 
 
 
X 
 
X 
14.2.10.3 
Observed Values and Change from Baseline 
in Mesopic BCDVA (Letters Read) by Time 
Point and Irides Type – Stage 1 
 
 
X 
X 
 
14.2.10.4 
Observed Values and Change from Baseline 
in Mesopic BCDVA (Letters Read) by Time 
Point and Irides Type – Stage 2 
 
 
X 
 
X 
14.2.11.1 
Percent of Subjects with Improvement in 
Mesopic DCNVA by Time Point – Stage 1 
 
 
X 
X 
 
14.2.11.2 
Percent of Subjects with Improvement in 
Mesopic DCNVA by Time Point – Stage 2 
 
 
X 
 
X 
14.2.11.3 
Percent of Subjects with Improvement in 
Mesopic DCNVA by Time Point and Irides 
Type – Stage 1 
 
 
X 
X 
 
14.2.11.4 
Percent of Subjects with Improvement in 
Mesopic DCNVA by Time Point and Irides 
Type – Stage 2 
 
 
X 
 
X 
14.2.12.1 
Observed Values and Change from Baseline 
in Mesopic DCNVA (Letters Read) by Time 
Point – Stage 1 
 
 
 
X 
 
14.2.12.2 
Observed Values and Change from Baseline 
in Mesopic DCNVA (Letters Read) by Time 
Point – Stage 2 
 
 
 
 
X 
14.2.12.3 
Observed Values and Change from Baseline 
in Mesopic DCNVA (Letters Read) by Time 
Point and Irides Type – Stage 1 
 
 
 
X 
 


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Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
14.2.12.4 
Observed Values and Change from Baseline 
in Mesopic DCNVA (Letters Read) by Time 
Point and Irides Type – Stage 2 
 
 
 
 
X 
14.2.13.1 
Observed Values and Change from Baseline 
in Photopic Pupil Diameter by Time Point – 
Stage 1  
 
 
 
X 
 
14.2.13.2 
Observed Values and Change from Baseline 
in Photopic Pupil Diameter by Time Point – 
Stage 2 
 
 
 
 
X 
14.2.13.3 
Observed Values and Change from Baseline 
in Photopic Pupil Diameter by Time Point and 
Irides Type – Stage 1 
 
 
 
X 
 
14.2.13.4 
Observed Values and Change from Baseline 
in Photopic Pupil Diameter by Time Point and 
Irides Type – Stage 2 
 
 
 
 
X 
14.2.14.1 
Percent of Subjects with Photopic Pupil 
Diameter < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm by Time Point – Stage 1 
 
 
 
X 
 
14.2.14.2 
Percent of Subjects with Photopic Pupil 
Diameter < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm by Time Point – Stage 2 
 
 
 
 
X 
14.2.14.3 
Percent of Subjects with Photopic Pupil 
Diameter < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm by Time Point and Irides Type – Stage 1 
 
 
 
X 
 
14.2.14.4 
Percent of Subjects with Photopic Pupil 
Diameter < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm by Time Point and Irides Type – Stage 2 
 
 
 
 
X 
14.2.15.1 
Percent of Subjects with Photopic Pupil 
Diameter 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 
2.5 to < 3.0 mm, and 3.0 to < 3.5 mm by Time 
Point – Stage 1 
 
 
 
X 
 
14.2.15.2 
Percent of Subjects with Photopic Pupil 
Diameter 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 
2.5 to < 3.0 mm, and 3.0 to < 3.5 mm by Time 
Point – Stage 2 
 
 
 
 
X 
14.2.15.3 
Percent of Subjects with Photopic Pupil 
Diameter 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 
2.5 to < 3.0 mm, and 3.0 to < 3.5 mm by Time 
Point and Irides Type – Stage 1 
 
 
 
X 
 
14.2.15.4 
Percent of Subjects with Photopic Pupil 
Diameter 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 
 
 
 
 
X 


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Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
2.5 to < 3.0 mm, and 3.0 to < 3.5 mm by Time 
Point and Irides Type – Stage 2 
14.2.16.1 
Observed Values and Change from Baseline 
in Mesopic Pupil Diameter by Time Point – 
Stage 1  
 
 
 
X 
 
14.2.16.2 
Observed Values and Change from Baseline 
in Mesopic Pupil Diameter by Time Point – 
Stage 2 
 
 
 
 
X 
14.2.16.3 
Observed Values and Change from Baseline 
in Mesopic Pupil Diameter by Time Point and 
Irides Type – Stage 1 
 
 
 
X 
 
14.2.16.4 
Observed Values and Change from Baseline 
in Mesopic Pupil Diameter by Time Point and 
Irides Type – Stage 2 
 
 
 
 
X 
14.2.17.1 
Percent of Subjects with Mesopic Pupil 
Diameter < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm by Time Point – Stage 1 
 
 
 
X 
 
14.2.17.2 
Percent of Subjects with Mesopic Pupil 
Diameter < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm by Time Point – Stage 2 
 
 
 
 
X 
14.2.17.3 
Percent of Subjects with Mesopic Pupil 
Diameter < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm by Time Point and Irides Type – Stage 1 
 
 
 
X 
 
14.2.17.4 
Percent of Subjects with Mesopic Pupil 
Diameter < 3.5, < 3.0, < 2.5, < 2.0, and < 1.5 
mm by Time Point and Irides Type – Stage 2 
 
 
 
 
X 
14.2.18.1 
Percent of Subjects with Mesopic Pupil 
Diameter 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 
2.5 to < 3.0 mm, and 3.0 to < 3.5 mm by Time 
Point – Stage 1 
 
 
 
X 
 
14.2.18.2 
Percent of Subjects with Mesopic Pupil 
Diameter 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 
2.5 to < 3.0 mm, and 3.0 to < 3.5 mm by Time 
Point – Stage 2 
 
 
 
 
X 
14.2.18.3 
Percent of Subjects with Mesopic Pupil 
Diameter 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 
2.5 to < 3.0 mm, and 3.0 to < 3.5 mm by Time 
Point and Irides Type – Stage 1 
 
 
 
X 
 
14.2.18.4 
Percent of Subjects with Mesopic Pupil 
Diameter 1.5 to < 2.0 mm, 2.0 to < 2.5 mm, 
 
 
 
 
X 


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Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
2.5 to < 3.0 mm, and 3.0 to < 3.5 mm by Time 
Point and Irides Type – Stage 2 
14.2.19.1 
Percent of Subjects with Photopic Baseline 
DCNVA ≤ 20/63, ≤ 20/80, ≤ 20/100, 
≤ 20/125, and ≤ 20/160, with a ≥ 10-letter and 
≥ 15-letter Improvement in Photopic 
DCNVA by Time Point – Stage 1 
 
 
 
X 
 
14.2.19.2 
Percent of Subjects with Photopic Baseline 
DCNVA ≤ 20/63, ≤ 20/80, ≤ 20/100, 
≤ 20/125, and ≤ 20/160, with a ≥ 10-letter and 
≥ 15-letter Improvement in Photopic 
DCNVA by Time Point – Stage 2 
 
 
 
 
X 
14.2.19.3 
Percent of Subjects with Photopic Baseline 
DCNVA ≤ 20/63, ≤ 20/80, ≤ 20/100, 
≤ 20/125, and ≤ 20/160, with a ≥ 10-letter and 
≥ 15-letter Improvement in Photopic 
DCNVA by Time Point and Irides Type – 
Stage 1 
 
 
 
X 
 
14.2.19.4 
Percent of Subjects with Photopic Baseline 
DCNVA ≤ 20/63, ≤ 20/80, ≤ 20/100, 
≤ 20/125, and ≤ 20/160, with a ≥ 10-letter and 
≥ 15-letter Improvement in Photopic 
DCNVA by Time Point and Irides Type – 
Stage 2 
 
 
 
 
X 
14.2.20.1 
Percent of Subjects with Photopic Baseline 
DCNVA from 20/50 to 20/63, 20/50 to 20/80, 
20/50 to 20/100, 20/50 to 20/125, and 20/50 
to 20/160, with a ≥ 10-letter and ≥ 15-letter 
Improvement in Photopic DCNVA by Time 
Point – Stage 1 
 
 
 
X 
 
14.2.20.2 
Percent of Subjects with Photopic Baseline 
DCNVA from 20/50 to 20/63, 20/50 to 20/80, 
20/50 to 20/100, 20/50 to 20/125, and 20/50 
to 20/160, with a ≥ 10-letter and ≥ 15-letter 
Improvement in Photopic DCNVA by Time 
Point – Stage 2 
 
 
 
 
X 
14.2.20.3 
Percent of Subjects with Photopic Baseline 
DCNVA from 20/50 to 20/63, 20/50 to 20/80, 
20/50 to 20/100, 20/50 to 20/125, and 20/50 
to 20/160, with a ≥ 10-letter and ≥ 15-letter 
Improvement in Photopic DCNVA by Time 
Point and Irides Type – Stage 1 
 
 
 
X 
 


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Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
14.2.20.4 
Percent of Subjects with Photopic Baseline 
DCNVA from 20/50 to 20/63, 20/50 to 20/80, 
20/50 to 20/100, 20/50 to 20/125, and 20/50 
to 20/160, with a ≥ 10-letter and ≥ 15-letter 
Improvement in Photopic DCNVA by Time 
Point and Irides Type – Stage 2 
 
 
 
 
X 
14.2.21.1 
Analysis of Observed Values in Subject 
Questionnaire – Stage 1 
 
 
X 
 
 
14.2.21.2 
Analysis of Observed Values in Subject 
Questionnaire – Stage 2 
 
 
X 
 
 
14.3.1.1.1 
Overall Summary of Treatment Emergent 
Adverse Events (TEAE) – Stage 1 
 
X 
 
 
 
14.3.1.1.2 
Overall Summary of Treatment Emergent 
Adverse Events (TEAE) – Stage 2 
 
X 
 
 
 
14.3.1.2.1 
Treatment-Emergent Adverse Events (TEAE) 
by System Organ Class and Preferred Term – 
Stage 1 
 
X 
 
 
 
14.3.1.2.2 
Treatment-Emergent Adverse Events (TEAE) 
by System Organ Class and Preferred Term – 
Stage 2 
 
X 
 
 
 
14.3.1.3.1 
Treatment-Emergent Adverse Events (TEAE) 
by System Organ Class, Preferred Term, and 
Severity – Stage 1 
 
X 
 
 
 
14.3.1.3.2 
Treatment-Emergent Adverse Events (TEAE) 
by System Organ Class, Preferred Term, and 
Severity – Stage 2 
 
X 
 
 
 
14.3.1.4.1 
Treatment-Emergent Adverse Events (TEAE) 
by System Organ Class, Preferred Term, and 
Relationship to Study Treatment – Stage 1 
 
X 
 
 
 
14.3.1.4.2 
Treatment-Emergent Adverse Events (TEAE) 
by System Organ Class, Preferred Term, and 
Relationship to Study Treatment – Stage 2 
 
X 
 
 
 
14.3.1.5.1 
Serious Treatment-Emergent Adverse Events 
(TEAE) by System Organ Class and Preferred 
Term– Stage 1 
 
X 
 
 
 
14.3.1.5.2 
Serious Treatment-Emergent Adverse Events 
(TEAE) by System Organ Class and Preferred 
Term – Stage 2 
 
X 
 
 
 


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Table 
Description of Table 
ARP Safety 
mITT 
PPS1 
PPS2 
14.3.1.6.1 
Treatment-Emergent Adverse Events (TEAE) 
Leading to Withdrawal From the Study by 
System Organ Class and Preferred Term – 
Stage 1 
 
X 
 
 
 
14.3.1.6.2 
Treatment-Emergent Adverse Events (TEAE) 
Leading to Withdrawal From the Study by 
System Organ Class and Preferred Term – 
Stage 2 
 
X 
 
 
 
14.3.1.7.1 
Treatment-Emergent Adverse Events (TEAE) 
Leading to Study Medication Discontinuation 
by System Organ Class and Preferred Term – 
Stage 1 
 
X 
 
 
 
14.3.1.7.2 
Treatment-Emergent Adverse Events (TEAE) 
Leading to Study Medication Discontinuation 
by System Organ Class and Preferred Term – 
Stage 2 
 
X 
 
 
 
14.3.3.1 
Subjective Ocular Tolerability – Stage 1 
 
X 
 
 
 
14.3.3.2 
Subjective Ocular Tolerability – Stage 2 
 
X 
 
 
 
14.3.4.1 
Vital Signs – Stage 1 
 
X 
 
 
 
14.3.4.2 
Vital Signs – Stage 2 
 
X 
 
 
 
14.3.5.1 
Intraocular Pressure (IOP) – Stage 1 
 
X 
 
 
 
14.3.5.2 
Intraocular Pressure (IOP) – Stage 2 
 
X 
 
 
 
14.3.6.1 
Subject Questionnaire – Stage 1 
 
X 
 
 
 
14.3.6.2 
Subject Questionnaire – Stage 2 
 
X 
 
 
 
14.3.7.1 
Biomicroscopy – Stage 1 
 
X 
 
 
 
14.3.7.2 
Biomicroscopy – Stage 2 
 
X 
 
 
 
14.3.8.1 
Ophthalmoscopy – Stage 1 
 
X 
 
 
 
14.3.8.2 
Ophthalmoscopy – Stage 2 
 
X 
 
 
 
 
12.2. 
List of Planned Listings 
Listing 
Description of Listing 
16.2.1.1 
Subject Disposition 
16.2.1.2 
Eligibility and Randomization 
16.2.2 
Treatment Assignment and Analysis Populations 
16.2.3 
Protocol Deviations 


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Listing 
Description of Listing 
16.2.4.1 
Demographics 
16.2.4.2 
General Medical History (Non-ocular) 
16.2.4.3 
Ocular Medical History 
16.2.4.4 
Manifest Refraction and Near Add 
16.2.4.5 
Prior Medications 
16.2.4.6 
Concomitant Medications 
16.2.5.1 
Study Treatment 1 Administration and Dispensation 
16.2.5.2 
Study Treatment 1 Timing and Washout 
16.2.5.3 
Study Treatment 1 Administration and Dispensation 
16.2.5.4 
Study Treatment 1 and Treatment 2 Diary 
16.2.5.5 
Study Treatment Exposure and Compliance 
16.2.6.1 
Distance Corrected Near Visual Acuity (DCNVA) 
16.2.6.2 
Best Corrected Distance Visual Acuity (BCDVA) 
16.2.6.3 
Distance Corrected Intermediate Visual Acuity (DCIVA) 
16.2.6.4 
Pupil Diameter 
16.2.7 
Adverse Events 
16.2.8 
Subjective Ocular Tolerability 
16.2.9 
Vital Signs 
16.2.10 
Intraocular Pressure (IOP) Tonometry 
16.2.11 
Subject Questionnaire 
16.2.12 
Biomicroscopy Examination 
16.2.13 
Ophthalmoscopic Examination 
16.2.14 
Corneal Fluorescein Staining 
16.2.15 
Tear Break-up Time 
16.2.16 
Urine Pregnancy Test 
 
12.3. 
Testing Hierarchy 
1. Percentage of subjects gaining ≥15 letters binocular photopic DCNVA without a loss 
of >5 letters DCNVA at Stage 2 Day 8 0.5 Hrs post-dose Nyxol + LDP vs. Placebo + 
LDP Vehicle 
2. Percentage of subjects gaining ≥15 letters binocular photopic DCNVA without a loss 
of >5 letters DCNVA at Stage 2 Day 8 0.5 Hrs post-dose Nyxol + LDP vs. LDP alone 
3. Percentage of subjects gaining ≥15 letters binocular photopic DCNVA without a loss 
of >5 letters DCNVA at Stage 2 Day 8 0.5 Hrs post-dose Nyxol + LDP vs. Nyxol 
alone 


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4. Percentage of subjects gaining ≥15 letters binocular photopic DCNVA without a loss 
of >5 letters DCNVA at Stage 2 Day 1 0.5 Hrs post-dose Nyxol + LDP vs. Placebo + 
LDP Vehicle 
5. Mean change from BL in photopic Study eye PD at Stage 2 Day 8 0.5 Hrs post-dose 
Nyxol + LDP vs. Placebo + LDP Vehicle 
6. Mean change from BL in photopic Study eye PD at Stage 2 Day 1 0.5 Hrs post-dose 
Nyxol + LDP vs. Placebo + LDP Vehicle 
7. Mean change from BL in photopic Study eye PD at Stage 2 Day 8 8 Hrs post-dose 
Nyxol + LDP vs. Placebo + LDP Vehicle 
8. Mean change from BL in binocular photopic DCNVA at Stage 2 Day 8 0.5 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 
9. Mean change from BL in binocular photopic DCNVA at Stage 2 Day 1 0.5 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 
10. Percentage of subjects gaining ≥15 letters binocular photopic DCNVA without a loss 
of >5 letters DCNVA at Stage 1 Day 8 12 Hr post-dose Nyxol vs. Placebo 
11. Percentage of subjects gaining ≥15 letters binocular photopic DCNVA without a loss 
of >5 letters DCNVA at Stage 1 Day 1 8 Hr post-dose Nyxol vs. Placebo 
12. Mean change from BL in photopic Study eye PD at Stage 1 Day 8 12 Hrs post-dose 
Nyxol vs. Placebo 
13. Mean change from BL in photopic Study eye PD at Stage 1 Day 1 8 Hrs post-dose 
Nyxol vs. Placebo 
14. Mean change from BL in photopic Study eye PD at Stage 1 Day 1 8 Hrs post-dose 
Nyxol vs. Placebo 
15. Mean change from BL in binocular photopic DCNVA at Stage 1 Day 8 12 Hrs post-
dose Nyxol vs. Placebo 
16. Mean change from BL in binocular photopic DCNVA at Stage 1 Day 1 8 Hrs post-
dose Nyxol vs. Placebo 
17. Mean change from BL in binocular photopic DCNVA at Stage 2 Day 8 8 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 
18. Mean change from BL in mesopic Study eye PD at Stage 2 Day 8 0.5 Hrs post-dose 
Nyxol + LDP vs. Placebo + LDP Vehicle 
19. Mean change from BL in mesopic Study eye PD at Stage 1 Day 8 12 Hrs post-dose 
Nyxol vs. Placebo 
20. Mean change from BL in mesopic Study eye PD at Stage 2 Day 1 0.5 Hrs post-dose 
Nyxol + LDP vs. Placebo + LDP Vehicle 
21. Mean change from BL in mesopic Study eye PD at Stage 1 Day 1 8 Hrs post-dose 
Nyxol vs. Placebo 
22. Mean change from BL in mesopic Study eye PD at Stage 2 Day 8 8 Hrs post-dose 
Nyxol + LDP vs. Placebo + LDP Vehicle 
23. Mean change from BL in binocular photopic DCIVA at Stage 2 Day 8 0.5 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 


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24. Percentage of subjects gaining ≥15 letters binocular photopic DCNVA without a loss 
of >5 letters DCNVA at Stage 2 Day 8 8 Hrs post-dose Nyxol + LDP vs. Placebo + 
LDP Vehicle 
25. Mean change from BL in binocular photopic DCIVA at Stage 2 Day 1 0.5 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 
26. Mean change from BL in binocular photopic DCIVA at Stage 2 Day 8 8 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 
27. Mean change from BL in binocular mesopic DCNVA at Stage 2 Day 8 0.5 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 
28. Mean change from BL in binocular mesopic DCNVA at Stage 2 Day 1 0.5 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 
29. Mean change from BL in binocular mesopic DCNVA at Stage 2 Day 8 8 Hrs post-
dose Nyxol + LDP vs. Placebo + LDP Vehicle 
30. Mean change in near vision (question 1) Stage 2 Day 8 Nyxol + LDP vs. Placebo + 
LDP Vehicle 
31. Mean satisfaction with near vision (question 5) Stage 2 Day 8 Nyxol + LDP vs. 
Placebo + LDP Vehicle 
32. Percentage of subjects gaining ≥15 letters binocular photopic DCNVA without a loss 
of >5 letters DCNVA at Stage 1 Day 8 20 Hr post-dose Nyxol vs. Placebo 
33. Mean change from BL in binocular photopic DCIVA at Stage 1 Day 1 8 Hrs post-
dose Nyxol vs. Placebo 
34. Mean change from BL in binocular mesopic DCNVA at Stage 1 Day 8 12 Hrs post-
dose Nyxol vs. Placebo 
35. Mean change from BL in binocular mesopic DCNVA at Stage 1 Day 1 8 Hrs post-
dose Nyxol vs. Placebo 
36. Mean change from BL in binocular photopic DCIVA at Stage 1 Day 8 12 Hrs post-
dose Nyxol vs. Placebo 
37. Mean change in near vision (question 1) Stage 1 Day 8 Nyxol vs. Placebo 
38. Mean satisfaction with near vision (question 5) Stage 1 Day 8 Nyxol vs. Placebo