Pivotal Prospective Clinical Trial to Demonstrate the Efficacy and Safety of AEYE-DS Software Device for Automated Diabetic Retinopathy Detection From Digital Fundoscopic Images
Phase: NA
Conditions: Diabetes Mellitus, Diabetic Retinopathy
Sponsor: AEYE Health Inc

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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PIVOTAL PROSPECTIVE CLINICAL TRIAL TO DEMONSTRATE 
THE EFFICACY AND SAFETY OF AEYE-DS SOFTWARE DEVICE 
FOR AUTOMATED DIABETIC RETINOPATHY DETECTION FROM 
DIGITAL FUNDUSCOPIC IMAGES  
 
Product: AEYE-DS 
 
Protocol Number: AEYE-DS-001 
Development Phase: Clinical Validation 
Sponsor:  
AEYE HEALTH INC. 
 
 
 
 
 
Protocol Approval Date: 
Current protocol Version No.3.4 November 24, 2021 
 
 
 
 
This clinical study will be conducted in accordance with current Good Clinical Practice (GCP) as 
directed by the provisions of the International Conference on Harmonization (ICH); Israel Ministry 
of Health; United States (US) Code of Federal Regulations (CFR) and the Sponsor’s Standard 
Operating Procedures (SOPs). 
This document contains confidential and proprietary information (including confidential 
commercial information pursuant to 21CFR§20.61) and is a confidential communication of AEYE 
Health INC (the “Sponsor”). The recipient agrees that no information contained herein may be 
published or disclosed without written approval from the Sponsor. 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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PROTOCOL SYNOPSIS ................................................................................................................ 4 
1 
INTRODUCTION ................................................................................................................ 9 
2 
DEVICE DESCRIPTION .................................................................................................. 10 
2.1 
General Device Description ...................................................................................... 10 
2.2 
Device Modules ........................................................................................................ 11 
2.2.1 
...................................................................................... 11 
2.2.2 
 ................................................................................... 11 
2.2.3 
 ................................................................................ 11 
2.2.4 
 ............................................................... 12 
2.2.5 
 .......................................................................................... 12 
3 
INTENDED USE AND INDICATIONS FOR USE ........................................................ 12 
4 
PREVIOUS CLINICAL STUDIES WITH THE INVESTIGATIONAL DEVICE ..... 13 
5 
RATIONALE FOR STUDY ..................................................................................... 14 
6 
STUDY OBJECTIVES ...................................................................................................... 15 
6.1 
Primary Objective ..................................................................................................... 15 
6.2 
Powered Secondary Objective .................................................................................. 15 
6.3 
Secondary Objectives ................................................................................................ 15 
7 
SELECTION OF STUDY POPULATION ...................................................................... 16 
7.1 
Inclusion Criteria:...................................................................................................... 16 
7.2 
Exclusion Criteria...................................................................................................... 17 
7.3 
Informed Consent ...................................................................................................... 17 
8 
STUDY DESIGN ................................................................................................................ 18 
9 
STUDY PROCEDURES: ................................................................................................... 20 
9.1 
Screening ................................................................................................................... 20 
9.2 
Data Acquisition (by Novice Operator): ................................................................... 20 
9.2.1 
Non-Mydriatic Fundoscopy Imaging ......................................................... 21 
9.2.2 
AEYE-DS System Operation ..................................................................... 21 
9.3 
Data Acquisition (by a Professional Ophthalmic Photographer) .............................. 22 
9.3.1 
Mydriatic Fundoscopy Imaging (Dilated Wide Field Stereo Color) .......... 22 
9.3.2 
Lens Photography for Media Opacity Assessment ..................................... 22 
9.3.3 
Macular Optical Coherence Tomography (OCT) Imaging ......................... 22 
9.4 
Determining Reference Standard / Ground Truth ..................................................... 23 
9.5 
Data Analysis ............................................................................................................ 24 
9.6 
Clinical Assessment .................................................................................................. 26 
9.7 
Flow Chart ................................................................................................................. 28 
 
28 
9.8 
Non-Mydriatic Fundoscopy Imaging with a Handheld Device ................................. 29 
9.9 
Human Factors Testing ............................................................................................. 29 
9.10 
Repeatability & Reproducibility Testing .................................................................. 30 
10 
PATIENT COMPLETION / WITHDRAWAL ......................................................... 31 
10.1 
Completion ................................................................................................................ 31 
10.2 
Withdrawal from the study ........................................................................................ 31 
10.3 
Analyzable Subjects .................................................................................................. 31 
11 
STATISTICAL CONSIDERATIONS .............................................................................. 32 
11.1 
Study Design and Objectives .................................................................................... 32 
11.2 
Study Endpoints ........................................................................................................ 32 
11.2.1 
Primary Efficacy Endpoints ....................................................................... 32 
11.2.2 
Powered Secondary Efficacy Endpoint ...................................................... 32 

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11.2.3 
Secondary Efficacy Endpoints ................................................................... 32 
11.3 
Study Hypotheses ...................................................................................................... 32 
11.4 
Sample Size ............................................................................................................... 33 
11.5 
Interim Analysis ........................................................................................................ 34 
11.5.1 
Procedure .................................................................................................... 34 
11.5.2 
Blinding ...................................................................................................... 34 
11.5.3 
Decision Rules ............................................................................................ 35 
11.5.4 
References .................................................................................................. 36 
11.6 
Data Sets Analyzed ................................................................................................... 36 
11.6.1 
Full Analysis Set (FA) ................................................................................ 36 
11.6.2 
Per Protocol (PP) Analysis Set ................................................................... 36 
11.6.3 
Safety Analysis Set (SA) ............................................................................ 36 
11.6.4 
Statistical Analysis of Analysis Sets .......................................................... 36 
11.7 
Randomization .......................................................................................................... 36 
11.8 
Blinding ..................................................................................................................... 36 
11.9 
Statistical Analysis .................................................................................................... 37 
11.9.1 
General Considerations .............................................................................. 37 
11.9.2 
Significance level and handling of Type I error ......................................... 37 
11.9.3 
Demographic and Baseline Characteristics ................................................ 37 
11.9.4 
Disposition of Subjects ............................................................................... 37 
11.9.5 
Analysis of Primary Endpoint .................................................................... 38 
11.9.6 
Analysis of Secondary Endpoints ............................................................... 39 
11.9.7 
Safety Assessments and Adverse Events .................................................... 40 
11.9.8 
Poolability .................................................................................................. 40 
11.9.9 
Handling of Missing Data .......................................................................... 40 
12 
RISK / BENEFIT ANALYSIS ................................................................................. 41 
12.1 
Risks .......................................................................................................................... 41 
12.2 
Benefits ..................................................................................................................... 41 
13 
ETHICAL ASPECTS ................................................................................................ 42 
13.1 
Study-Specific Design Considerations ...................................................................... 42 
13.2 
Regulatory Ethics Compliance .................................................................................. 42 
13.2.1 
Investigators Responsibilities ..................................................................... 42 
13.2.2 
Independent Ethics Committee or Institutional Review Board .................. 42 
13.2.3 
Informed Consent ....................................................................................... 43 
13.2.4 
Privacy of Personal Data ............................................................................ 43 
14 
ADMINISTRATIVE REQUIREMENTS ................................................................. 44 
14.1 
Protocol Modifications .............................................................................................. 44 
14.2 
Regulatory Documentation ....................................................................................... 44 
14.3 
Regulatory Approval/Notification ............................................................................. 44 
14.3.1 
Required Pre-study Documentation ........................................................... 44 
14.3.2 
Patient Identification Register and Patient Screening Log ......................... 45 
14.3.3 
Case Report Form Completion ................................................................... 45 
14.3.4 
Record Retention ........................................................................................ 45 
14.3.5 Use of Information and Publication ........................................................... 45 
15 
REFERENCES ................................................................................................................... 47 
APPENDIX A – TIME & EVENTS SCHEDULE 

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PROTOCOL SYNOPSIS 
Protocol Number 
AEYE-DS-001 
Protocol Title 
Pivotal prospective clinical trial to demonstrate the efficacy and 
safety of the AEYE-DS software device for automated diabetic 
retinopathy detection from digital funduscopic images. 
Location(s) 
5-10 primary care clinics in the US  
Phase of Development 
Clinical Validation, Pivotal Study 
Study Population 
Adult patients who are clinically diagnosed with diabetes and who 
have undergone a digital funduscopic examination, and have not been 
previously diagnosed with diabetic retinopathy. 
Study Objectives 
To evaluate the performance and usability of the AEYE-DS software 
for the detection of more than mild diabetic retinopathy (mtmDR) on 
digital funduscopic images from patients with known diabetes 
obtained from a Topcon NW400 fundoscopy device (an FDA cleared 
desktop camera). 
Number of Patients: 
A minimum of 350 and a maximum of up to 774 subjects will be 
enrolled in the Pivotal Study. 
 
 
. 
Inclusion Criteria: 
1. 
Age≥22 
2. 
Male or female 
3. 
Documented diagnosis of diabetes mellitus, meeting the 
criteria established by the American Diabetes Association 
(ADA) and World Health Organization (WHO): 
a. Elevated Hemoglobin A1c (HbA1c) ≥ 6.5% 
(48mmol/mol), based on repeated assessments, 
performed in a laboratory using a method that is 
NGSP certified and standardized to the DCCT assay 
OR 
b. Fasting Plasma Glucose (FPG) ≥ 126 mg/dL (7.0 
mmol/L), based on repeated assessments, where 
'fasting' is defined as no caloric intake for at least 8 
hours  OR 
c. Oral Glucose Tolerance Test (OGTT) with two hour 
plasma glucose (2-hr PG) ≥200 mg/dl (11.1 mmol/L), 
using the equivalent of an oral 75 g anhydrous 
glucose dose dissolved in water OR 
d. Symptoms of hyperglycemia or hyperglycemic crisis 
with a random plasma glucose (RPG) ≥ 200 mg/dl 
(11.1 mmol/L) 

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4. 
Understand the study and volunteer to sign the informed 
consent 
Exclusion Criteria: 
1. Uncorrectable vision loss (e.g., with the use of eyeglasses), 
blurred vision, or floaters. 
2. Diagnosed with macular edema, severe non-proliferative 
retinopathy, proliferative retinopathy, radiation retinopathy, or 
retinal vein occlusion. 
3. History of laser treatment of the retina or injections into either 
eye, or any history of retinal surgery. 
4. Currently participating in another investigational eye study and 
actively receiving investigational product for DR or DME. 
5. Participant has a condition that, in the opinion of the 
investigator, would preclude participation in the study (e.g., 
unstable medical status including blood pressure or glycemic 
control, microphthalmia or previous enucleation). 
6. Participant is contraindicated for imaging by fundus imaging 
systems used in the study: 
6.1. Participant is hypersensitive to light 
6.2. Participant recently underwent photodynamic therapy (PDT) 
6.3. Participant is taking medication that causes photosensitivity 
6.4. Participant has a history of angle-closure glaucoma or 
narrow anterior chamber angles 
Investigational 
Product:  
AEYE-DS 
Duration of Study: 
3 to 5 months. Study duration will include performing the data 
acquisition alongside analysis by AEYE-DS novice operators, 
followed by additional image data acquisition by a professional 
ophthalmic photographer and reading by a panel of expert readers at 
the Reading Center and statistical analysis. 
Primary Objective 
• 
Primary Objective: 
o To assess the sensitivity of the AEYE-DS software device for 
the detection of more than mild diabetic retinopathy (mtmDR) 
on digital funduscopic images in patients with known diabetes 
undergoing screening for diabetic retinopathy.  
o 
To assess the specificity of the AEYE-DS software device for 
the detection of more than mild diabetic retinopathy (mtmDR) 
on digital funduscopic images in patients with known diabetes 
undergoing screening for diabetic retinopathy. 
Digital funduscopic images will be acquired by the TopCon NW400 
fundoscopy device based on one macula centered image obtained per 
eye. 
Powered Secondary 
Objective 
• 
Powered Secondary Objective: 
o To assess the sensitivity of the AEYE-DS software device for 
the detection of more than mild diabetic retinopathy (mtmDR) 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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on digital funduscopic images in patients with known diabetes 
undergoing screening for diabetic retinopathy.  
o 
To assess the specificity of the AEYE-DS software device for 
the detection of more than mild diabetic retinopathy (mtmDR) 
on digital funduscopic images in patients with known diabetes 
undergoing screening for diabetic retinopathy. 
Digital funduscopic images will be acquired by the TopCon NW400 
fundoscopy device based on 2 images obtained per eye, one macula 
centered and one optic-disc centered. 
Secondary Objectives 
• 
Secondary Objectives: 
o To assess the imageability of AEYE-DS, defined as the 
percentage of participants with a disease level output 
(mtmDR+ or mtmDR-) from the AEYE-DS device. 
o 
To assess the Usability of the AEYE-DS device, including 
User Manual comprehension and subsequent usability of the 
device in the hands of potential users (i.e., primary care 
clinicians). 
o 
To assess the sensitivity and specificity of the AEYE-DS 
software device for the detection of more than mild diabetic 
retinopathy (mtmDR) on digital funduscopic images obtained 
from a handheld fundoscopy device, in patients with known 
diabetes undergoing screening for diabetic retinopathy. 
Digital funduscopic images will be acquired using the 
Optomed Aurora handheld fundoscopy device based on one 
macula-centered image obtained per eye. 
 
 
Study Hypothesis 
In this study, we will test the following pairs of hypotheses on the 
primary endpoint, as well as on the powered secondary 
endpoint: 
• 
H0:  Se ≤ 82%,  
H1: Se > 82% 
• 
H0:  Sp≤ 87% 
      H1: Sp> 87% 
Where Se and Sp are the sensitivity and specificity of the mtmDR 
classification of the AEYE-DS software device. 
Sample Size 
At least 66 positive and 265 negative subjects are required to meet 
the primary and powered secondary endpoints. 
The study will follow a sample size adaptive design with one interim 
analysis planned to allow for sample size increase up to at most 774 
subjects, or completion per original sample size. 
 
 
 

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Abbreviations 
Abbreviation 
Description 
AAO 
American Association of Ophthalmologists 
AMD 
Age-Related Macular Degeneration 
CI 
Confidence Interval 
CRF 
Case Report Form 
DR 
Diabetic Retinopathy 
FDA 
US Food and Drug Administration 
GCP 
Good Clinical Practice 
HbA1c 
Hemoglobin A1c (Glycated hemoglobin is a form of 
hemoglobin that is measured primarily to identify the 
three-month average plasma glucose concentration) 
HIPAA 
Health Insurance Portability and Accountability Act 
ICF 
Informed Consent Form 
IEC 
Independent Ethics Committee 
IRB 
Institutional Review Board 
mtmDR 
more than mild Diabetic Retinopathy 
NPV 
Negative Predictive Value 
PE 
Physical Examination 
PI 
Principal Investigator 
PPV 
Positive Predictive Value 
SOP 
Standard Operating Procedure 
SW 
Software 
WHO 
World Health Organization 

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1 
INTRODUCTION 
Diabetic retinopathy is a highly specific neurovascular complication of both type 1 and type 
2 diabetes, the prevalence of which strongly correlates to both the duration of diabetes and 
level of glycemic control. Diabetic Retinopathy (DR) is the leading cause of vision loss in 
adults between ages 20-74 [Cheung et al., 2010], and ranked as one of the most common 
causes of preventable blindness [Bourne,2013]. Based on a recent global report [WHO, 
2014], there are over 422 million people worldwide with diabetes, and over 35% have signs 
of DR, a third of them with vision threatening DR. Early detection of retinopathy is an 
important part of managing care for people with diabetes, yet many patients with diabetes 
are not adequately screened for diabetic retinopathy. 
 
Retinal imaging is known to be a diagnostic tool for DR [Walker et al. 1990] as well as 
other known conditions, such as Age Related Macular Degeneration (AMD), Glaucoma, 
Papilledema, and Hypertensive Retinopathy to name a few [Wiedemann, 1997]. High-
quality fundus photographs can detect most clinically significant diabetic retinopathy. 
Interpretation of the images should be performed by a trained eye care provider. For this 
reason, the AAO recommends periodic fundus examination starting at the age of 40 [AAO, 
2014]. The 2017 revision of the American Academy of Ophthalmology’s Preferred Practice 
Pattern recommends people with no or mild DR be followed annually, whereas those with 
more than mild DR, and/or DME (abbreviated to mtmDR), are recommended to receive 
evaluation and consideration for treatment. 
 
The ability to detect the presence of DR by computer models has been demonstrated more 
than once in previous work 
 In 2018, a medical 
device, IDx-DR, indicated for use as a screening diagnostic tool for DR, received FDA 
clearance for that purpose [FDA De Novo DEN180001]. This type of device permits the 
availability of a novel artificial intelligence technology that can be used in a primary care 
doctor’s office and improve patient access to needed health care. 
 
AEYE has also developed an AI software device (AEYE-DS) to be used as a diagnostic 
tool to assist primary care clinicians in screening for diabetic retinopathy using digital 
funduscopic images. The AEYE-DS device also automatically detects more than mild 
diabetic retinopathy (mtmDR) in adults (22 years of age or older) diagnosed with diabetes 
who have not been previously diagnosed with diabetic retinopathy. 
 
The development of artificial intelligence medical devices carries with it responsibilities as 
it sets precedent for use of AI applications in the medical field, especially, when used for 
diagnosis in a primary care setting, with less specialized medical expertise.  
 
Consequently, the goal of this pivotal clinical study is to demonstrate the safety and efficacy 
of the AEYE-DS diagnostic screening tool for automatic detection of more than mild 
Diabetic Retinopathy (mtmDR) from digital funduscopic images.  
 

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2.2 
Device Modules 
 
2.2.1 
 
 
 
 
 
 
 
. 
 
2.2.2 
 
 
 
 
 
  
2.2.3 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
  

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The system outputs one of three possible options:  
 
1) Insufficient image(s) quality – low quality images can be retaken immediately, while the 
patient is still at the camera.  
2) The patient is negative for referable mtmDR (mtmDR-).  
3) The patient is positive for mtmDR (mtmDR+). 
 
2.2.4 
 
 
 
 
 
 
 
 
 
2.2.5 
 
 
 
3 
INTENDED USE AND INDICATIONS FOR USE 
The AEYE-DS is indicated for use by health care providers to automatically detect more 
than mild diabetic retinopathy (mtmDR) in adults (22 years of age or older) diagnosed with 
diabetes who have not been previously diagnosed with diabetic retinopathy. The AEYE-
DS is indicated for use with the Topcon NW400. 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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4 
PREVIOUS CLINICAL STUDIES WITH THE INVESTIGATIONAL 
DEVICE 
 
 
 
 
 
 
 
 
 
 
  
 
 
 
 
 
  
 
  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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5 
RATIONALE FOR STUDY 
Based on a recent global report [WHO, 2014], there are over 422 million people worldwide 
with diabetes, and over 35% have signs of DR, a third of them with vision threatening DR. 
With the prevalence of diabetes expected to increase 50% over the next 10 years, screening 
rates, detection, and treatment must be addressed in order to prevent a significant increase 
in unnecessary blindness across the United States. Early detection of retinopathy through 
periodic screening is an important part of managing care for people with diabetes, yet 
screening rates are low, with only 50-60% of diabetic patients properly evaluated. An 
automated DR screening device would allow for immediate screening results at the point 
of care and standardization of results. 
 
 
 
 
 
  The current proposed study will 
validate the sensitivity and specificity of the AEYE-DS output by a novice operator 
compared to the reference standard, diagnostic determination by an independent, blinded 
panel of expert ophthalmologist at the reading center. 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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6 
STUDY OBJECTIVES 
6.1 
Primary Objective 
The primary study objective is to evaluate the performance of the AEYE-DS software for 
the detection of more than mild diabetic retinopathy (mtmDR) on digital funduscopic 
images from patients with known diabetes (acquired by the TopCon NW400 fundoscopy 
device based on one macula centered image obtained per eye ). 
 
6.2 
Powered Secondary Objective 
The powered secondary objective of the study is as follows: 
To evaluate the performance of the AEYE-DS software for the detection of more than mild 
diabetic retinopathy (mtmDR) on digital funduscopic images from patients with known 
diabetes (acquired by the TopCon NW400 fundoscopy device based on 2 images obtained 
per eye, one macula centered and one optic disc centered). 
 
6.3 
Secondary Objectives 
The secondary objectives of the study are as follows: 
• To assess the imageability of AEYE-DS, defined as the percentage of participants with 
a disease level output (mtmDR+ or MTMDR-) from the AEYE-DS device. 
• To assess the Usability of the AEYE-DS device, including User Manual comprehension 
and subsequent usability of the device in the hands of potential users (i.e., primary care 
clinicians). 
• To assess the sensitivity and specificity of the AEYE-DS software device for the 
detection of more than mild diabetic retinopathy (mtmDR) on digital funduscopic 
images obtained from a handheld fundoscopy device, in patients with known diabetes 
(acquired by the Optomed Aurora handheld fundoscopy device based on one macula-
centered image obtained per eye). 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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7 
SELECTION OF STUDY POPULATION 
The goal of the selection criteria is to identify adult patients who have undergone digital 
funduscopic imaging with clinical evidence of diabetes and are screened to rule out diabetic 
retinopathy. The study population will consist of adult patients (at least 22 years of age) 
with previously diagnosed diabetes, who are participating in a routine retinal screening test 
for diabetic retinopathy (DR) in primary care clinics, and have digital funduscopic images 
of sufficient quality from a specific funduscopic camera, i.e., the Topcon Model NW400. 
The patients referred to the screening program must be without prior diagnoses of DR. 
"Diabetic” patients are defined in the Inclusion Criteria (Section 7.1 below). 
 
The AEYE-DS device will be used in the study by clinicians who represent the actual end-
users. These will be clinical personnel working in primary care facilities, who perform 
various screening activities for patients. The users will upload the de-identified study 
images and simulate the generation of a clinical decision based on the AEYE-DS algorithm 
results. 
 
Evaluation of the study participant inclusion/exclusion criteria will be based on the review 
of clinical medical records and/or documented patient interviews conducted by the 
investigator or under the direction of the investigator in the application of the study 
inclusion and exclusion criteria, as summarized below. 
 
To be enrolled in the study, patients must meet ALL of the inclusion criteria and NONE of 
the exclusion criteria designated below. 
 
7.1 
Inclusion Criteria: 
1. Age ≥22 
2. Male or female 
3. Documented diagnosis of diabetes mellitus, meeting the criteria established by the 
American Diabetes Association (ADA) and World Health Organization (WHO): 
a. Elevated Hemoglobin A1c (HbA1c) ≥ 6.5% (48mmol/mol), based on repeated 
assessments, performed in a laboratory using a method that is NGSP certified and 
standardized to the DCCT assay OR 
b. Fasting Plasma Glucose (FPG) ≥ 126 mg/dL (7.0 mmol/L), based on repeated 
assessments, where 'fasting' is defined as no caloric intake for at least 8 hours  OR 
c. Oral Glucose Tolerance Test (OGTT) with two hour plasma glucose (2-hr PG) ≥200 
mg/dl (11.1 mmol/L), using the equivalent of an oral 75 g anhydrous glucose dose 
dissolved in water OR 
d. Symptoms of hyperglycemia or hyperglycemic crisis with a random plasma glucose 
(RPG) ≥ 200 mg/dl (11.1 mmol/L) 
4. Understand the study and volunteer to sign the informed consent 
 

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7.2 
Exclusion Criteria 
1. Uncorrectable vision loss (e.g., with the use of eyeglasses), blurred vision, or floaters. 
2. Diagnosed with macular edema, severe non-proliferative retinopathy, proliferative 
retinopathy, radiation retinopathy, or retinal vein occlusion. 
3. History of laser treatment of the retina or injections into either eye, or any history of 
retinal surgery. 
4. Currently participating in another investigational eye study and actively receiving 
investigational product for DR or DME. 
5. Participant has a condition that, in the opinion of the investigator, would preclude 
participation in the study (e.g., unstable medical status including blood pressure or 
glycemic control, microphthalmia or previous enucleation). 
6. Participant is contraindicated for imaging by fundus imaging systems used in the study: 
a. Participant is hypersensitive to light 
b. Participant recently underwent photodynamic therapy (PDT) 
c. Participant is taking medication that causes photosensitivity 
d. Participant has a history of angle-closure glaucoma or narrow anterior chamber 
angles 
 
7.3 
Informed Consent 
Written informed consent will be obtained from each study patient prior to enrollment into the 
study.  A written informed consent (
) 
must be signed and dated by the patient (or legally authorized representative, if appropriate), and 
the investigator.  Patients will be given a copy of the signed informed consent document.  The 
signed informed consent will be retained with the study records at the site. It is the responsibility of 
the Investigator to assure that informed consent is obtained from each patient in accordance with 
GCP guidelines. Subjects may withdraw their consent to participate in the study at any time without 
prejudice. The investigator may withdraw a subject if, in his clinical judgement, it is in the best 
interest of the subject or if the subject cannot comply with the protocol.  

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8 
STUDY DESIGN 
The proposed study is designed to compare AEYE-DS diagnostic results obtained from 
fundoscopy images for the detection of more than mild diabetic retinopathy (mtmDR), to 
the blinded diagnostic results from expert ophthalmologists used as the reference standard. 
 
The study is a prospective, multi-center, single-arm study. 
 
 
 
Study cases will be sourced from different primary care clinics in the USA. Each clinic will 
have its own separate staff for the acquisition of the retinal images. Staff will include a 
novice operator who had not previously performed ocular imaging and professional 
ophthalmic photographer who will be certified by the Fundus Reading Center. Each clinic 
will screen diabetic patients to check for potential presence of DR.  
 
 
.  The study population who represent the target population for this 
procedure consists of stable, visually asymptomatic subjects with previously diagnosed 
diabetes without prior diagnoses of DR, who are participating in a routine retinal screening 
test for diabetic retinopathy (DR) in primary care clinics, and have digital funduscopic 
images of sufficient quality from a specific funduscopic camera, i.e., the Topcon Model 
NW400. Patients of both genders, all ethnicities, ≥22 years of age will be recruited to the 
study. Patients’ demographics will include a comparable and representative distribution of 
the population of the United States and its different ethnicities, including but not limited to 
White, Asian, African-American and Latino populations. Investigators will screen 
participants based on the inclusion/exclusion criteria described above. General patient 
demographics, medical history, concomitant medications, fundoscopy system used, OCT 
system used, etc., will be obtained for each study case.  
 
Both fundus images needed for the AEYE-DS and for the reading center (to establish 
ground truth) will be acquired from each eye of the patient, after the patient has signed an 
informed consent form. 
 
The novice operator will obtain two fundoscopy images from each eye of the patient, after 
the patient has signed an informed consent form. The operator will activate the AEYE-DS 
system on the fundoscopy images and obtain a result of more than mild DR (mtmDR) 
detected, more than mild DR not detected, or insufficient quality. The results for each case 
will be recorded in the eCRF. 
 
After the novice operator has generated an AEYE-DS output, but during the same visit, 
each participant will undergo additional retinal imaging captured by the professional 
ophthalmic photographer. The professional ophthalmic photographer will be masked at all 
times to the AEYE-DS output and will use a different, FDA-cleared camera system 
) to obtain dilated four widefield 
stereo color fundus images, lens photography for media opacity assessment and macular 
optical coherence tomography (OCT) imaging. These images will all be sent to an 
independent Reading Center, where the severity of retinopathy and diabetic macular edema 
(DME) will be determined according to the Early Treatment for Diabetic Retinopathy 

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Study severity (ETDRS) scale. These readings will form the reference standard (ground 
truth) for the study. Three experienced and certified readers will review the images. If at 
least two readers agree, a majority voting paradigm will be implemented. 
 
 
The readers will be masked to the 
AEYE-DS output at all times, masked to the funduscopy images when evaluating the OCT 
images and masked to the OCT readings when evaluating the fundoscopy images. Each 
participant will be categorized as mtmDR+ or mtmDR-. The worst of two eyes will be 
compared with the AEYE-DS output at the participant level. The results for each participant 
will be recorded in the eCRF.  
 
The reference standard (ground truth) results from the Reading Center will be statistically 
compared to the AEYE-DS device results. The independent co-primary outcome 
parameters of the study will be sensitivity and specificity assessed in the Intent-to-Screen 
(ITS) population for whom AEYE-DS provided a diagnostic result and the Reading Center 
provided a usable reference standard. Furthermore, the AEYE-DS positive predictive value 
and negative predictive value will be determined. The imageability of AEYE-DS in terms 
of the percentage of readable images diagnosed successfully by the device will also be 
measured. 
 
Usability of the AEYE-DS device will also be assessed including User Manual 
comprehension and usability of the device in the hands of potential users (i.e., primary care 
clinicians).  
 
Details on the timing of all study procedures are given in the Time and Events Schedule in 
Attachment B of the Protocol. 
 
 
The blinding status will be maintained until the last participant has been completed in the 
study. 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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9 
STUDY PROCEDURES: 
9.1 
Screening 
Investigators will screen participants based on the inclusion/exclusion criteria described 
above. Diabetic patients who require fundoscopy examination, meet the inclusion criteria 
and do not meet any clause of the exclusion criteria and volunteer to participate will join 
the study. A consent form will be signed by each patient. General patient demographics 
(including age, gender, race and ethnicity), diabetic history (including HbA1C, FPGL, etc.), 
ophthalmic history (including lens status, reported light sensitivity, as applicable), medical 
history, concomitant medications, fundoscopy system used, OCT system used, etc., will be 
obtained for each study case. 
 
At the start of the study, all participants with diabetes who meet inclusion and exclusion 
criteria will be enrolled sequentially. To avoid excessive enrollment in any one stratum 
(no/mild DR or mtmDR), the totals will be monitored monthly and the study population 
will be enriched to ensure sufficient numbers of subjects with mtmDR. 
 
 
. 
 
 
 
 
  
 
9.2 
Data Acquisition (by Novice Operator):  
 
The novice operator will obtain fundoscopy images from each eye of the patient, after the 
patient has signed an informed consent form. The novice operator is a staff personal from 
a primary care site, who has not previously performed ocular imaging. 
 
 
 
 
 
 
. 
 
. 
 
The novice operators will undergo a one-time standardized training program on how to 
acquire images, how to improve image quality if the AEYE-DS gives an insufficient quality 
output, and how to submit images for analysis to the AEYE-DS device. No additional 
training will be provided to the operators for the duration of the study. 
 
   

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
21 
 
 
9.2.1 Non-Mydriatic Fundoscopy Imaging 
 
Fundus imaging involves photographing the fundus (i.e., the rear of an eye). Fundoscopy 
devices are specialized cameras which consist of an intricate microscope attached to a flash 
enabled camera. The main structures that can be visualized on a fundus photo are the central 
and peripheral retina, optic disc and macula.  
The novice operator will obtain two digital fundoscopy image from each eye of the patient 
, using the non-mydriatic, TopCon NW400 ocular fundoscopy 
imaging device, according to a standardized imaging protocol with fovea/macula centered 
and optic-disc centered 45° images 
.  
 
 
 
 
 
  
 
 
 
 
 
 
 
9.2.2 AEYE-DS System Operation 
 
The operator will activate the AEYE-DS system on the fundoscopy images and obtain a 
result of more than mild DR (mtmDR) detected, more than mild DR not detected, or 
insufficient quality, 
. In case the result is ‘insufficient quality’, 
fundoscopy can be re-taken for the relevant eye(s) and fixation point(s). 
 
It is anticipated that the presence of lens opacities due to cataract will significantly increase 
the number of imaging attempts required to get sufficient quality images, as well as the 
requirement for dilation. If after three (3) imaging attempts by the operator (without 
pharmacologic dilation), the system still indicates that at least one image is of insufficient 
quality, pharmacological dilation will be performed. In this case, the participant’s pupils 
will be dilated with tropicamide 1.0% eyedrops, until the pupil diameter is at least 5mm in 
each eye or 30 minutes have passed, and all 4 fundus images (1 macular centered and 1 
disc centered for each eye) will be reacquired. Three (3) more attempts may be made to 
capture an image of sufficient quality. If the AEYE-DS system still outputs that at least one 
image is of insufficient quality, the AEYE-DS output of insufficient quality will be 
recorded as the final result. 
The results for each case will be recorded in the eCRF.  
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
22 
 
The time required to process the fundoscopy images with the AEYE-DS system until a 
result is obtained, will also be recorded. 
 
As this is an investigational device study, no clinical action will be taken based on the 
results presented by the AEYE-DS device. 
 
9.3 
Data Acquisition (by a Professional Ophthalmic Photographer) 
After the novice operator has generated an AEYE-DS output, but during the same visit, 
each participant will undergo additional retinal imaging captured by the professional 
ophthalmic photographer trained by the reading center. The professional ophthalmic 
photographer will be masked at all times to the AEYE-DS output and will use a different, 
FDA-cleared camera system (
 
 to obtain dilated four widefield stereo color fundus images, lens photography 
for media opacity assessment and macular optical coherence tomography (OCT) imaging.  
9.3.1 Mydriatic Fundoscopy Imaging (Dilated Wide Field Stereo Color) 
If the study participant is not already dilated, dilating eye drops of tropicamide 1.0% will 
be administered. Four (4) Mydriatic (dilated) fundoscopy imaging with Wide Field Stereo 
Color will be performed on each eye of the patient by the professional ophthalmic 
photographer, at the primary care center using the 
 
 
 
 
  
 
The four widefield stereo image pairs will be obtained to determine the severity of 
retinopathy according to the well-established Early Treatment for Diabetic Retinopathy 
Study severity (ETDRS) scale. 
 
9.3.2 Lens Photography for Media Opacity Assessment  
In the presence of media opacity, the image quality of fundoscopy devices is less 
satisfactory and may lead to ungradable images. For this reason, the above mentioned 
mydriatic funduscopy imaging capturing the red reflex (
 
) will be performed to evaluate lens status 
given that cataract and other media opacities are more prevalent in the diabetic population. 
Anterior segment photography for media opacity assessment will be performed according 
to the Age Related Eye Disease Study 2: AREDS2 Report 7 (Domalpally et al 2013). 
 
9.3.3 Macular Optical Coherence Tomography (OCT) Imaging 
Macular Optical Coherence Tomography (OCT) imaging will also be performed on each 
eye of the patient by the professional ophthalmic photographer, at the primary care center 
using the using the 
 to produce 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
23 
 
a cube scan containing at least 121 B scans. A major limitation of many current DR 
screening programs based on fundus photography is the inability to accurately identify 
DME on 2-dimensional fundus photographs. 
 
 
 
 
 
 
 
 
 
 
Macular OCT images will be obtained to detect the presence of center involved DME 
according to the Diabetic Retinopathy Clinical Research Network (DRCR) grading 
paradigm.  
 
9.4 
Determining Reference Standard / Ground Truth 
The four mydriatic widefield stereo color fundus images, lens photography for media 
opacity assessment and macular optical coherence tomography (OCT) imaging obtained 
with the FDA-cleared camera system (
 
 will all be sent to an independent Reading 
Center, where the severity of retinopathy and diabetic macular edema (DME) will be 
determined according to the well-established Early Treatment for Diabetic Retinopathy 
Study severity (ETDRS) scale.  
 
These readings will form the reference standard (ground truth) for the study. 
 
 
 
 
 
 
 All certified readers will be senior, board-certified 
ophthalmologists with expertise in the field of retinal diseases with at least 10 years of 
clinical experience and certified by the Reading Center.  The readers will be masked to the 
AEYE-DS output at all times, masked to the funduscopy images when evaluating the OCT 
images and masked to the OCT readings when evaluating the fundoscopy images.  
 
The Reading Center diagnostic grading protocol will consist of the following:  
The four mydriatic widefield stereo color fundoscopy image pairs will be read by two 
readers (or three readers in case of disagreement using a majority voting paradigm) 
according to the ETDRS scale.  
 
The macular OCT images will be read by the same readers for the presence of center 
involved DME according to the Diabetic Retinopathy Clinical Research (DRCR) network 
grading paradigm.  
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
24 
 
Because DME can be identified on the basis of retinal thickening on stereoscopic 
fundoscopy images, as well as on the basis of retinal thickening on OCT, performance using 
both definitions will be analyzed.  
 
Stereoscopic fundus-based Clinically Significant DME (CSDME) will be identified if there 
is either retinal thickening or adjacent hard exudates < 600μm from the foveal center, or a 
zone of retinal thickening > 1 disc area, part of which is less than 1 disc diameter from the 
foveal center, according to the Reading Center, in any eye.  
 
OCT based center involved DME will be identified if a participant had central subfield (a 
1.0mm circle centered on the fovea) thickness that is >300μm according to the Reading 
Center, in any eye. 
 
Each participant will be categorized as mtmDR+ (i.e., ETDRS level 35 or higher and /or 
DME present), or mtmDR- (ETDRS level 10-20 and DME absent). The worst of two eyes 
will be compared with the AEYE-DS output at the participant level.  
 
In summary, the definition of mtmDR+ is defined as: 
• ETDRS level ≥ 35 (determined from fundoscopy images) and/or 
• CSDME (determined from fundoscopy images)  
and multimodal mtmDR+ is defined as: 
• ETDRS level ≥ 35 (determined from fundus photographs); and/or  
• CSDME (determined from fundoscopy images); and/or  
• Center-involved DME (determined from OCT). 
 
The reader may also classify the patient’s images as “insufficient quality" for reading. 
Lens status will be assessed using the following grading: 
• Phakic with opacities or Cannot Grade; or  
• Pseudophakic or no opacities 
 
Phakic and pseudophakic are terms used to describe the status of an individual’s lens.  
Phakic refers to a person with an intact natural lens while a pseudophakic individual had a 
lens extracted and an IOL placed.  
 
The readers will be allowed to manipulate the images to grade subtle lesions. In the case 
that incidental findings unrelated to DR (e.g., optic nerve pathology, drusen, pigmented 
choroidal lesions) are identified by the readers, these will be recorded and the sites will be 
notified along with the clinical assessment, as described in section 9.6.    
 
The results for each participant will be recorded in the eCRF. 
 
9.5 
Data Analysis 
The Reading Center will enter the ETDRS result and indication of detection of macular 
edema in the EDC within 4 weeks of each participant's completion of the study protocol. A 
study participant is considered to have completed the study protocol upon completion of 
the AEYE-DS imaging protocol (either resulting in sufficient or insufficient quality 
images) and the OCT and mydriatic four wide field fundscopy imaging, according to 
section 10.1.  

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
25 
 
 
All Reading Center ETDRS results will be converted to an International Clinical Diabetic 
Retinopathy (ICDR) level based on the American Academy American Academy of 
Ophthalmology’s (AAO) International Clinical Diabetic Retinopathy (ICDR) Disease 
Severity Scale (see table below). ICDR results will also indicate whether the Reading 
Center detected macular edema. 
  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
The following table described how the ETDES severity levels of the worst eye will be 
mapped and combined with fundus based DME to the dichotomous categories mtmDR- 
(non or mild NPDR and no DME) and mtmDR+ (more than mild DR or DME).  
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
26 
 
 
All blinded datasets will be locked after eCRF data monitoring is completed. The datasets 
will be merged and unblinded and the diagnostic results will be compared and analyzed. 
Performance for mtmDR detection will be calculated as device alone (AEYE-DS) vs. 
ground truth (Expert Reader committee decision) according to the data analysis plan 
described in Section 11 – Statistical Section. 
 
9.6 
Clinical Assessment 
The converted ICDR results and corresponding American Academy of Ophthalmology 
management recommendations will be provided to the study investigators, who are 
responsible for appropriately managing study subjects. To support referral determinations 
based on the ICDR results provided to them, the study investigators should review the 
Management Recommendations for Patients with Diabetes as contained in the AAO’s 
Preferred Practice Pattern (AAO PPP) Recommendations for Diabetic Retinopathy (See 
Table 
6 
at 
http://www.aao.org/preferred-practice-pattern/diabetic-retinopathy-ppp-
updated-2016). 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
27 
 
 
 
In correspondence about Reading Center results, clinicians will also be provided with the 
following notes regarding the PPP: 
 
• The AAO PPP recommends a 12-month follow-up for patients with normal, minimal 
non proliferative diabetic retinopathy (NPDR) or mild NPDR when there is no 
presence of macular edema (ME). 
• The AAO PPP recommends that it may be appropriate for patients with moderate 
NPDR to receive follow-up examination by an eye care professional in an interval less 
than 12 months. 
• Based on the AAO PPP, those patients with severe NPDR, Non-high risk PDR, or High 
Risk PDR – as well as those with macular edema – may be indicated for treatment based 
on evaluation by an eye care professional. These subjects should be referred. 
 
On the basis of Reading Center ICDR results and AAO PPP Recommendations, physicians 
will be able to provide appropriate eye care referrals for study participants. 
 
Furthermore, in the case that incidental findings unrelated to DR (e.g., optic nerve 
pathology, drusen, pigmented choroidal lesions) are identified by the readers, these will be 
recorded and the sites will be notified of these findings, along with the clinical assessment. 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
28 
 
9.7 
Flow Chart 
 
Data Collection by Novice Operator (at Primary Care Sites) 
 
Identify subjects according to eligibility criteria & sign Informed Consent 
 
Collect data for each subject, including the following: 
 
                Subject Data              
      Fundoscopy Imaging            AEYE-DS Operation 
(demographics, procedural, etc.)     (using TopCon NW400)          (AEYE-DS Diagnosis) 
 
 
 
 
 
   
 
 
 
 
Data Collection by Professional Ophthalmic Photographer (at Primary Care Sites) 
 
Collect data for each subject, including the following: 
 
              OCT Imaging     
 
             Fundoscopy Imaging                 Lens Photography 
                                                         (Mydriatic 4 WF Stereo Color)              
        
                 
          
     
       
                   
                
    
 
 
 
  
             READING      CENTER 
 
      DME Detection                      Severity of Retinopathy                   Lens Status  
   (according to DRCR grading)         (according to ETDRS scale) 
 
 
 
 
 
 
                        DATA        ANALYSIS 
 
ICDR Diagnosis: mtmDR+, mtmDR- or “insufficient quality" 
 
 
AEYE-DS Diagnosis: mtmDR+, mtmDR- or “insufficient quality" 
 
Data Monitoring, Data Lock & Unblinding 
 
Statistical Analysis of Primary Endpoints: Sensitivity and Specificity 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
29 
 
 
 
 
 
 
 
 
 
;  
 
 
 
 
 
 
9.8 
Non-Mydriatic Fundoscopy Imaging with a Handheld Device 
In selected sites, additional fundus imaging using an FDA cleared, handheld fundoscopy 
device may also be performed and processed using the AEYE-DS device. The handheld 
fundoscopy device is a specialized camera which consists of an intricate microscope 
attached to a flash enabled camera, only much smaller in size. Various handheld devices 
may be used for the purpose of this sub-study. The type of handheld device used will be 
recorded on the CRF.  
The novice operator will obtain two digital fundoscopy images (one from each eye of the 
patient)
, using the non-mydriatic, handheld fundoscopy imaging 
device, according to a standardized imaging protocol. Images will be fovea/macula 
centered  
 
 
 
 
.  
The novice operator will activate the AEYE-DS system on the fundoscopy images obtained 
from the handheld device and obtain a result of more than mild DR (mtmDR) detected, 
more than mild DR not detected, or insufficient quality, in the same manner described in 
section 9.2.2 above. The 'software upload client' will be modified for the data collection (as 
only one image per eye is submitted for analysis) and installed at the applicable 
participating sites. 
 
 
9.9 
Human Factors Testing 
The human factors testing will be performed in accordance with the FDA Guidance 
Applying Human Factors and Usability Engineering to Medical Devices, dated February 
3, 2016, as well as IEC 62366 Medical devices – Application of usability engineering to 
medical devices.  
 
Human factors testing will include preliminary, iterative evaluation of the workflow, device 
training and labeling materials and device use, followed by interviews with probing 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
30 
 
questions during feasibility and simulated studies. The standardized workflow, training 
program, and operator materials developed by this process will be implemented and tested 
as part of this pivotal study as described below. 
 
Human factors testing will be performed as part of the clinical study to demonstrate that 
the instructions for use and training are sufficient for users to operate AEYE-DS, as 
intended.  The human factors testing will validate the setting in which the device is intended 
to be used. A list of critical tasks associated with the use of the device will be provided to 
the novice operators including uploading the fundoscopy images, operating the AEYE-DS 
device and obtaining the diagnostic results from the device.  
 
A post-use questionnaire will be administered to the novice users for feedback, and 
recorded and documented as part of the eCRFs. A similar observer questionnaire will be 
used to capture and record any confusion or user errors and documented as part of the 
eCRFs, as well. 
 
The human factors testing to be conducted as part of the AEYE-DS pivotal study will 
validate the fundoscopy use and imaging protocol and the standardized labeling materials 
which enable the capture of at least two medical grade image by previously untrained 
fundoscopy operators for at least 80% of the subjects imaged who will complete the full 
imaging protocol on a fundoscopy device intended for use with AEYE-DS device.  
 
 
9.10 Repeatability & Reproducibility Testing 
Adequate validation will be performed and controls implemented to characterize and ensure 
consistency (repeatability and reproducibility) of the diagnostic output. The reproducibility 
(variability between different devices) and repeatability (variability between results from 
the same operator) analysis will be performed in a separate validation sub-study, under a 
separate sub-study protocol, but on subjects who will have participated in the pivotal study. 
Half of the participants will have been diagnosed as mtmDR- based on the original Expert 
Reader grading and half of the participants will have been diagnosed as having mtmDR+. 
 
Each participant in the sub-study will have completed the entire AEYE-DS imaging 
protocol 10 times, imaged by three different novice operators on two different Topcon 
NW400 fundoscopy devices. 
 
 
 
 
  
 
The binary output of AEYE-DS will be used to assess repeatability and reproducibility and 
compute the percent complete agreement of the AEYE-DS outputs across repeats, novice 
operators and fundoscopy devices. Full details of the study design and analysis will be 
provided in the Repeatability and Reproducibility Study Protocol. 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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10 PATIENT COMPLETION / WITHDRAWAL  
 
10.1 
Completion 
A subject will be considered to have completed the study if the required, fundoscopy 
imaging and AEYE-DS operation are completed by the novice operator as described in 
section 9.2 and the mydriatic fundoscopy imaging, OCT imaging and lens photography has 
been completed by the professional ophthalmology photographer as described in section 
9.3 and all imaging files have been received at the Reading Center.  
 
10.2 
Withdrawal from the study  
 
A subject will be withdrawn from the study for any of the following reasons: 
• Withdrawal of consent  
• Subject is not compliant with requirements of the study, including inclusion and 
exclusion criteria. 
• The investigator believes that for safety reasons (e.g. an adverse event) it is in the 
best interest of the patient to stop the study. 
• The study is prematurely stopped or halted (e.g. clinical halt) 
 
10.3 
Analyzable Subjects  
A subject will be fully analyzable if all of the following conditions are met: 
1. Subject is eligible to participate in the study according to the study eligibility criteria 
specified in section 7.0.  
2. FPRC Reading: 
o Subject completed the professional ophthalmic photography imaging protocol 
and met all minimal requirements according to section 9.3. 
o Subject received a positive or negative diagnosis by the FPRC.  
3. AEYE-DS Imaging Protocol 
o Subject completed the AEYE-DS imaging protocol and met all minimal 
requirements according to section 9.2.  
o Subject received a positive or negative diagnosis by the AEYE-DS System. 
Subjects who are non-analyzable due to "insufficient quality" or "missing output" by the 
AEYE-DS imaging, will be considered in the study imageability analysis as well as 
sensitivity analyses as described in the statistical section below. 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
32 
 
11 STATISTICAL CONSIDERATIONS 
11.1 Study Design and Objectives 
The study is a blinded, single arm cohort study, designed to evaluate the performance of 
the AEYE-DS software device for the detection of more than mild diabetic retinopathy 
(mtmDR) on digital funduscopic images in patients with known diabetes undergoing 
screening for diabetic retinopathy. 
 
 
. 
 
11.2 Study Endpoints 
11.2.1 Primary Efficacy Endpoints 
The primary efficacy endpoints are the sensitivity and specificity of the AEYE-DS device 
to detect mtmDR on digital funduscopic images (acquired by the TopCon NW400 
fundoscopy device based on 1 macula centered image obtained per eye). The ground truth 
will be determined by a panel of expert ophthalmologist readers at the Reader Center. 
 
11.2.2 Powered Secondary Efficacy Endpoint 
Powered secondary efficacy endpoint includes: 
• Sensitivity and specificity of the AEYE-DS device to detect mtmDR on digital 
funduscopic images (acquired by the TopCon NW400 fundoscopy device based on 
2 images obtained per eye, one macula centered and one optic-disc centered). 
 
 
11.2.3 Secondary Efficacy Endpoints 
Secondary efficacy endpoints include: 
• The percentage of cases with a disease level output (mtmDR+ or mtmDR-) from 
the AEYE-DS device. 
• Device Usability 
• Sensitivity and specificity of the AEYE-DS device to detect mtmDR on digital 
funduscopic images (
 
 
 
 
 
11.3 Study Hypotheses 
In this study, we will test the following pairs of hypotheses on the primary endpoint, as 
well as on the powered secondary endpoint: 
• H0: Se ≤82%,  
H1: Se > 82% 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
33 
 
• H0: Sp≤87%, 
      H1: Sp >87% 
 
Where Se and Sp are the sensitivity and specificity of the mtmDR classification of the 
AEYE-DS software device. 
 
 
 
 
 
11.4  Sample Size 
The two null hypotheses will be evaluated independently, therefore sample sizes will be 
calculated for both and the maximum sample size will be chosen. A sample size is 
calculated such that the lower limit of the one-sided 97.5% exact binomial confidence 
interval for sensitivity and specificity are greater than 82% and 87% respectively with over 
90% power, using the SAS POWER procedure. 
 
 
 
 
 
  
 
 
 
 
 
 
 
 
 
  
 
 
 
  
  
 
 
  
 
 
 
  
 
 
   
 
 
 
 
 
 
 
 
 
 
 
  
 
 
 
  
 
 
 
  
 
 
 
  
 
 
   
 
 
 
 
  
 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
34 
 
 
 
 
 
 
.  
 
 
 
 
 
 
 
 
 
.   
 
 
 
 
 
. 
 
 
 
 
 
 
 
 
 
 
 
 
. 
 
 
 
 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
36 
 
, 
 
  
 
 
 
 
 
 
 
 
 
 
  
 
  
 
 
 
 
 
 
  
 
  
 
 
 
 
 
 
 
 
 
 
 
 
11.7 Randomization 
This is a single arm, non-randomized study, no randomization will be performed. 
11.8 Blinding 
The clinician who will assess the DR score with the AEYE-DS will be blinded to the 
committee decision. 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
38 
 
 
11.9.5 Analysis of Primary Endpoint 
A 2x2 table of the results of all subjects who had valid results on both AEYE-DS and the 
Reader Center comparing the positive/negative results obtained from the AEYE-DS versus 
the reference values will be presented.  
 
The primary efficacy analysis will present the estimation of sensitivity and specificity of 
the of the AEYE-DS device to detect mtmDR on digital funduscopic images (acquired by 
the TopCon NW400 fundoscopy device based on one macula centered image obtained per 
eye) together with their lower limit of their respective one-sided exact 97.5% confidence 
intervals. If the lower limits of the one-sided 97.5% confidence intervals of the sensitivity 
and specificity are greater than or equal to 82% and 87% respectively the null-hypotheses 
will be rejected. If both null hypotheses are rejected in favor of the alternative hypotheses 
the study will be deemed successful, and the performance goals met. 
 
The positive and negative predictive values (PPV and NPV respectively) of the diagnosis 
together with their respective exact 95% confidence intervals will be presented as well. 
 
Analyses will be performed on both ground truth modalities, fundus-only and multi-
modality (using fundoscopy and OCT). 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
The percent of participants requiring pharmacologic dilation and not requiring dilation to 
obtain an AEYE-DS disease level output will be calculated. 
 
Sensitivity and specificity, analyses will be carried out on the combined cohorts. 
 
 
 
 
 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
39 
 
 
 
 
 
 
 
 
11.9.6 Analysis of Secondary Endpoints 
The powered secondary efficacy analysis will present the estimation of sensitivity and 
specificity of the AEYE-DS device to detect mtmDR on digital funduscopic images 
(acquired by the TopCon NW400 fundoscopy device based on 2 images obtained per eye, 
one macula centered and one optic disc centered) together with the lower limit of their 
respective one-sided exact 97.5% confidence intervals. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
  
 
 
 
.  
 
 
 
 
 
 
 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
40 
 
 
11.9.7 Safety Assessments and Adverse Events 
 
 
 
 
  
  
11.9.8  Poolability 
 
  
 
  
 
 
 
 
 
11.9.9  Handling of Missing Data 
Subjects with missing data will be compared to those with complete data, with respect to 
demographic and perhaps other baseline characteristics.  
 
As a sensitivity analysis, the primary and powered secondary endpoints will be analyzed 
considering a non-analyzable result of the AEYE-DS as “negative” for the calculation of 
sensitivity, and “positive” for the calculation of specificity. 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
41 
 
12 RISK / BENEFIT ANALYSIS 
12.1 
Risks 
The risks to patients resulting from potential device hazards have been analyzed using the 
Risk Management Standard - ISO 14971. The different types of hazards were identified 
and evaluated using risk assessment numerical parameters. Applicable controls for the risks 
were analyzed. 
 
 
 
 
 
 
   
 
12.2 
Benefits 
Diabetic Retinopathy (DR) is the leading cause of vision loss in adults between ages 20-74 
[Cheung et al., 2010], and ranked as one of the most common causes of preventable 
blindness [Bourne, 2013]. Based on a recent global report [WHO, 2014], there are over 422 
million people worldwide with diabetes, and over 35% have signs of DR, a third of them 
with vision threatening DR. With the prevalence of diabetes expected to increase 50% over 
the next 10 years, screening rates, detection, and treatment must be addressed in order to 
prevent a significant increase in unnecessary blindness across the United States.  
 
Early detection of retinopathy is an important part of managing care for people with 
diabetes, yet many patients with diabetes are not adequately screened for diabetic 
retinopathy. A 1990 program launched by the American Academy of Ophthalmology 
(AAO), known as Diabetes 2000, was designed to increase nationwide DR screening rates. 
However, after 20 years of implementation, screening rates continue to stagnate, with only 
50-60% of diabetic patients properly evaluated.  
 
A DR screening solution at the point of primary care is expected to help address the unmet 
need, lack of access, and racial and ethnic disparity issues that have persisted for decades. 
An automated screening device would allow for immediate screening results at the point of 
care and standardization of results. Ophthalmic key opinion leaders state that an automated 
diabetic retinopathy screening device in the hands of primary care healthcare providers 
would increase screening rates and detect more treatable vision threatening DR. 
 
The current proposed study will validate the sensitivity and specificity of the AEYE-DS 
output by a novice operator compared to the diagnostic determination by an independent, 
blinded panel of expert ophthalmologist at the reading center. 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
42 
 
13 ETHICAL ASPECTS 
 
13.1 
Study-Specific Design Considerations 
Subjects/cases will be carefully screened using the study eligibility criteria prior to 
enrollment in the study.  
 
13.2 
Regulatory Ethics Compliance 
 
13.2.1 
Investigators Responsibilities 
The investigator is responsible for ensuring that the clinical study is performed in 
accordance with the protocol, current ICH guidelines on Good Clinical Practice (GCP), and 
applicable regulatory requirements. 
GCP is an international ethical and scientific quality standard for designing, conducting, 
recording, and reporting studies that involve the participation of human subjects. 
Compliance with this standard provides public assurance that the rights, safety, and well-
being of study subjects are protected, consistent with the principles that originated in the 
Declaration of Helsinki, and that the clinical study data are credible. 
 
13.2.2 
Independent Ethics Committee or Institutional Review Board  
Before the start of the study, the investigator will provide the IEC/IRB with current and 
complete copies of the following documents: 
• Final protocol and if applicable, amendments 
• Informed consent form  
• Investigator's Brochure (or equivalent information) and amendments 
• Investigator's curriculum vitae or equivalent information (unless not required, as 
documented by IEC/IRB) 
• Information regarding funding, name of the sponsor, institutional affiliations, and 
other potential conflicts of interest 
• Any other documents that the IEC/IRB requests to fulfill its obligation 
This study will be undertaken only after IEC/IRB has given full approval of the final 
protocol, amendments (if any), and informed consent and after the sponsor has received a 
copy of this approval. This approval letter must be dated and must clearly identify the 
documents being approved. 
During the study the investigator will send the following documents to the IEC/IRB for 
their review and approval, where appropriate: 
• Protocol amendments 
• Investigator's Brochure amendments or new edition(s) 
• Summaries of the status of the study (at least annually or at intervals stipulated in 
guidelines of the IEC/IRB) 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
43 
 
• Reports of any serious adverse events, if applicable 
• Deviations from or changes to the protocol  
• Notification if new investigator is responsible for the study at the site 
• Any other requirements of the IEC/IRB 
At least once a year the IEC/IRB will be asked to review and re-approve this clinical study. 
This request and approval should be documented in writing. 
At the end of the study, the investigator (or sponsor where required) will notify the IEC/IRB 
about the study completion. 
 
13.2.3 
Informed Consent 
Each subject must give written consent according to local requirements after the nature of 
the study has been fully explained. The consent form must be signed before performance 
of any study-related activity. The consent form used must be approved by both the sponsor 
and by the reviewing IEC/IRB. The informed consent should be in accordance with 
principles set forth in the Declaration of Helsinki, current ICH and GCP guidelines, 
applicable regulatory requirements, and sponsor policy. 
Before entry into the study, the investigator or an authorized member of the investigational 
staff must explain to potential subjects the aims, methods, reasonably anticipated benefits, 
and potential hazards of the study, and any discomfort it may entail. Subjects will be 
informed that their participation is voluntary and that they may withdraw consent to 
participate at any time. They will be informed that choosing not to participate will not affect 
the care the subject will receive for the treatment for his/her disease. Subjects will be told 
that alternative treatments are available if they refuse to take part and that such refusal will 
not prejudice future treatment. Finally they will be told that their records may be accessed 
by health authorities without violating the confidentiality of the subject, to the extent 
permitted by the applicable law(s) or regulations. By signing the informed consent form 
the subject is authorizing such access. 
The subject will be given sufficient time to read the informed consent form and the 
opportunity to ask questions. After this explanation and before entry into the study, consent 
should be appropriately recorded by name of the subject, subject's signature and date of 
signature. After having obtained the consent, a copy of the informed consent form must be 
given to the subject. If the subject or legally acceptable representative is unable to read or 
write, an impartial witness should be present for the entire informed consent process (which 
includes reading and explaining all written explanations) and should personally date and 
sign the inform consent form after the oral consent of the subject or legally acceptable 
representative is obtained. 
 
13.2.4 
Privacy of Personal Data 
The collection and processing of personal data from patients enrolled in this study will be 
limited to those data that are necessary to investigate the efficacy, safety, quality, and utility 
of the investigational product(s) used in this study. This data must be collected and 
processed with adequate precautions to ensure confidentiality and compliance with 
applicable data privacy protection laws and regulations. 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
44 
 
14 ADMINISTRATIVE REQUIREMENTS 
 
14.1 
Protocol Modifications 
 
The investigator will not modify this protocol without a formal amendment. Protocol 
amendments must not be implemented without prior IEC/IRB approval, or when the 
relevant competent authority has raised any grounds for non-acceptance, except when 
necessary to eliminate immediate hazards to the patients in which case the amendment must 
be promptly submitted to the IEC/IRB and relevant competent authority. When the 
change(s) involves only logistic or administrative aspects of the study, the IRB (and IEC 
where required) only needs to be notified. 
The investigator or other physician in attendance will contact the appropriate sponsor 
representative by fax or telephone regarding any situations requiring a departure from the 
protocol. If possible, contact will be made before implementing any departure from the 
protocol. In all cases contact with the sponsor must be made as soon as possible in order to 
discuss the situation and agree on an appropriate course of action. The data recorded in the 
CRF and source document will reflect any departure from the protocol, and the source 
documents will describe this departure and the circumstances requiring it. 
 
14.2 Regulatory Documentation 
 
14.3 Regulatory Approval/Notification 
This protocol and any amendment(s) must be submitted to the appropriate regulatory 
authorities in each respective country, if applicable. A study may not be initiated at a site 
until all local regulatory requirements are met.  
 
14.3.1 
Required Pre-study Documentation  
The following documents must be available and maintained during the study: 
• Approved Study Protocol and amendment(s) 
• A copy of the dated and signed written IEC/IRB approval of the protocol and any 
amendments. This approval must clearly identify the specific protocol by title and 
number and must be signed by the chairman or authorized designee. 
• An approved informed consent form 
• Regulatory authority approval or notification, if applicable 
• Documentation of investigator qualifications (e.g., curriculum vitae) 
• Completed financial disclosure form  
• Signed and dated clinical trial agreement, which includes the financial agreements 
• Other documentation required by local regulations 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
45 
 
14.3.2 
Patient Identification Register and Patient Screening Log 
The investigator agrees to complete a subject/case identification register to permit easy 
identification of each subject/case during and after the study.  
The subject/case identification register will be treated as confidential. To ensure patient 
confidentiality, no copy will be made. All reports and communications relating to the study 
will identify subject/case by initials and assigned number only. 
The investigator will also complete a subject/case enrollment log, which reports all 
subject/case who were determined eligible for inclusion in the study. 
 
14.3.3 
Case Report Form Completion  
All data relating to the study will be recorded on source documents and then entered into 
an EDC with electronic CRFs (eCRFs). Data will be collected in English. The source 
documents and eCRFs are to be completed at the time of the data collection. Every effort 
should be made to ensure that all measures are recorded on the source documents and 
eCRFs. The investigator must verify that all data entries on the source documents and in 
the eCRFs are accurate and correct. 
 
14.3.4 
Record Retention  
In compliance with the ICH/GCP guidelines, the investigator/institution will maintain all 
source documents and eCRFs that support the data collected from each patient, as well as 
all study documents as specified in ICH/GCP Section 8, Essential, Documents for the 
Conduct of a Clinical Trial, and all study documents as specified by the applicable 
regulatory requirement(s). The investigator/institution will take measures to prevent 
accidental or premature destruction of these documents. 
 
 
 
 
 
 
 
If the responsible investigator retires, relocates, or for other reasons withdraws from the 
responsibility of keeping the study records, custody must be transferred to a person who 
will accept the responsibility. The sponsor must be notified in writing of the name and 
address of the new custodian. Under no circumstance shall the investigator relocate or 
dispose of any study documents before having obtained written approval from the sponsor. 
 
 
 
14.3.5 
Use of Information and Publication 
 
 
 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
47 
 
15 REFERENCES 
1. Cheung N, Mitchell P, Wong TY. Diabetic retinopathy. Lancet. 2010;376(9735):124–36. doi: 
10.1016/S0140-6736(09)62124-3. 
 
2. Bourne RR, Stevens GA, White RA, Smith JL, Flaxman SR, Price H, et al. Causes of vision loss 
worldwide, 1990–2010: a systematic analysis. Lancet Glob Health. 2013;1(6):e339–49. doi: 
10.1016/S2214-109X(13)70113-X.  
 
3. Yau JW, Rogers SL, Kawasaki R, Lamoureux EL, Kowalski JW, Bek T, et al. Global prevalence 
and major risk factors of diabetic retinopathy. Diabetes Care. 2012;35(3):556–64. doi: 
10.2337/dc11-1909. 
 
4. Walker HK, Hall WD, Hurst JW, “Clinical Methods: The History, Physical, and Laboratory 
Examinations. 3rd edition.”Boston: Butterworths; 1990. 
 
5. FDA Application: DEN180001 Trade/Device Name: IDx-DR Regulation Number: 21 CFR 
886.1100, 2018 
 
6. 
Michael 
Larsen; Jannik 
Godt; Nicolai 
Larsen; Henrik 
Lund-Andersen; Anne 
KatrinSjølie; Elisabet Agardh; Helle Kalm; Michael Grunkin; David R. Owens. “Automated 
Detection of Fundus Photographic Red Lesions in Diabetic Retinopathy”, Investigative 
Ophthalmology & Visual Science February 2003, Vol.44, 761-766. doi:10.1167/iovs.02-0418 
 
7. Oliver Faust, Rajendra Acharya U., E. Y. K. Ng, Kwan-Hoong Ng, Jasjit S. Suri, ““Algorithms 
for the Automated Detection of Diabetic Retinopathy Using Digital Fundus Images: A Review”, 
Journal of Medical Systems, February 2012, Volume 36, Issue 1, pp 145–157  
 
8. Diabetes Mellitus – epidemiology. 2. Diabetes Mellitus – prevention and control. 3. Diabetes, 
Gestational. 4. Chronic Disease. 5. Public Health. I. World Health Organization. ISBN 978 92 4 
156525 7 (NLM classification: WK 810) 
 
9. Wiedemann P., Kohen L., “Macular and Retinal Diseases”, Karger Publishing, 1997 
 
10. American Association of Ophthalmologists (AAO): Article https://www.aao.org/eye-
health/tips-prevention/screening 
 
11. Trans Am OphthalmolSoc 2015;113:T9[1-18]. ©2015 by the American Ophthalmological 
Society. 
 
12. Early Treatment Diabetic Retinopathy Study Research Group. Grading diabetic retinopathy 
from stereoscopic color fundus photographs: an extension of the modified Airlie House 
classification. ETDRS report number 10. Ophthalmology 1999;98 (5Suppl):786-806.  
 
13. Domalpally A, Danis RP, Chew EY, et al. Evaluation of optimized digital fundus reflex 
photographs for lens opacities in the Age-Related Eye Disease Study 2: AREDS2 Report 7. 
Invest Ophthalmol Vis Sci. 2013;54:5989–5994.). 
 

Protocol AEYE-DS-001                                                                                Ver. 3.4 24 November 2021 
 
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APPENDIX A – TIME AND EVENTS SCHEDULE 
 
Study Procedure 
Primary Care Site 
(by Novice Operator) 
Primary Care Site 
(by Professional 
Ophthalmic 
Photographer) 
Reading 
Center 
CRO 
Informed Consent 
√ 
 
 
 
Eligibility Criteria  
√ 
 
 
 
Patient Demographics 
√ 
 
 
 
Diabetic History 
√ 
 
 
 
Ophthalmic History 
√ 
 
 
 
Medical History 
√ 
 
 
 
Concomitant Medications 
√ 
 
 
 
Procedural Information 
√ 
 
 
 
Fundoscopy Images 
Acquisition with TopCon 
NW400 
√ 
 
 
 
Fundoscopy Images 
Acquisition with handheld 
device (Optomed Aurora) 
√ 
 
 
 
AEYE Operation & mtmDR 
Diagnosis with TopCon 
NW400 images 
√ 
 
 
 
AEYE Operation & mtmDR 
Diagnosis with handheld 
fundoscopy images 
√ 
 
 
 
Mydriatic Fundoscopy 
Images Acquisition 
 
√ 
 
 
Macular Optical Coherence 
Tomography (OCT) Imaging 
 
√ 
 
 
Lens Photography 
 
√ 
 
 
ETDRS Results 
 
 
√ 
 
Detection of Macular Edema 
 
 
√ 
 
Lens Status Determination 
 
 
√ 
 
ICDR Results 
 
 
 
√