A Pilot, Safety and Feasibility Trial of High-Dose Omega-3 Fatty Acids and High-Dose Cholecalciferol (Vitamin D) Supplementation in Type 1 Diabetes
Phase: PHASE1
Conditions: Diabetes Mellitus, Type 1, Hypoglycemia, Diabetes Mellitus, Diabetes, Autoimmune
Sponsor: Rodolfo Alejandro

POSEIDON STUDY 
 
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A Pilot, Safety and Feasibility Trial of High-Dose Omega-3 Fatty 
Acids and High-Dose Cholecalciferol Supplementation in Type 1 
Diabetes 
 
POSEIDON STUDY: PILOT STUDY OF OMEGA-3 AND VITAMIN D HIGH DOSES IN T1D  
 
 
VERSION 4.1 dated April 10th, 2023  
 
 
SPONSOR(S)  
Camillo Ricordi, M.D.  
Rodolfo Alejandro, M.D.  
 
PRINCIPAL INVESTIGATOR 
David Alejandro Baidal, M.D. 
 
CO-PRINCIPAL INVESTIGATOR  
Janine Sanchez, M.D.  
 
 
NCT03406897 
 
Diabetes Research Institute, University of Miami  
1450 NW 10th Avenue, Miami, FL 33136 
Office: 305 243-6913; Fax: 305 243-4404 
 
CONFIDENTIAL 
  
 

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PREFACE 
PILOT STUDY OF OMEGA-3 AND VITAMIN D HIGH DOSES IN T1D (POSEIDON) Protocol, A Pilot, 
Safety and Feasibility Trial of High-Dose Omega-3 Fatty Acids and High-Dose Cholecalciferol 
Supplementation in Type 1 Diabetes, describes the background, design, and organization of the study.  
The Diabetes Research Institute will maintain the protocol over the course of the study through new 
releases of the protocol, or issuance of updates either in the form of revisions of complete chapters or 
pages thereof, or in the form of supplemental protocol memoranda. 
 
 
AUTHORSHIP TEAM (in alphabetical order) 
The following individuals provided substantial input during protocol development: 
 
• 
Rodolfo Alejandro, M.D. Ψ  
• 
David Alejandro Baidal, M.D.*  
• 
Carlos Eduardo Blaschke, F.M.G.  
• 
Khemraj Hirani, Ph.D., RPh, RAC, CHRC 
• 
Della Lorraine Matheson, R.N., CDE  
• 
Shari Messinger, M.E., Ph.D.  
• 
Alberto Pugliese, M.D.  
• 
Lisa Emily Rafkin, MS, RD, CDE 
• 
Camillo Ricordi, M.D.Ψ 
• 
Luis A. Roque, Quality Assurance Auditor 
• 
Janine Sanchez, M.D. * 
• 
Juliana Maria Vera Ortiz, M.D. 
* Principal/Co- Investigator(s)  
ΨSponsor 
 
The sponsors, investigators, DRI leadership and study authors appreciate support and feedback from 
patient advocacy group such as Children with Diabetes in the creation and design of this study.  
 

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Glossary of Abbreviations 
 
AE 
 
 
Adverse event 
APC 
 
 
Antigen presenting cell 
ATG 
 
 
Anti-Thymocyte Globulin (Thymoglobulin®) 
AUC 
 
 
Area under Curve 
BB Rat  
 
Bio-Breeding Rat 
CBC 
 
 
Complete blood count 
CFR 
 
 
Code of Federal Regulations 
cGMP  
 
Current Good Manufacturing Practice 
CHO  
 
Carbohydrates 
CMV  
 
Cytomegalovirus 
CRF 
 
 
Case report form 
CRS 
 
 
Cytokine Release Syndrome 
DC 
 
 
Dendritic Cell 
DRI 
 
 
Diabetes Research Institute  
DSMB  
 
Data and Safety Monitoring Board 
EBV 
 
 
Epstein-Barr virus 
FACS  
 
Fluorescence activated cell sorting 
FDA 
 
 
US Food and Drug Administration 
FOXP3 
 
Forkhead box P3 
FWA  
 
Federal-wide Assurance 
GAD 
 
 
Glutamate decarboxylase 
GCP 
 
 
Good Clinical Practice 
GCSF  
 
Granulocyte colony-stimulating factor (GCSF/Neulasta®) 
HbA1c  
 
Hemoglobin A1c 
HBsAg  
 
Hepatitis B surface antigen 
HIV 
 
 
Human immunodeficiency virus 
ICA 
 
 
Islet cytoplasmic antibodies 
IGRA  
 
Interferon-γ release assays 
IL-2 
 
 
Interleukin 2 
IND 
 
 
Investigational New Drug 
IRB 
 
 
Institutional Review Board 
LIFT 
 
 
Long Term Investigative Follow-Up 
MMTT  
 
Mixed-meal tolerance test 
NCI-CTCAE 
 
National Cancer Institute Common Terminology Criteria for Adverse Events 
NOD  
 
Nonobese diabetic 
OHRP  
 
Office for Human Research Protections 
PBMC  
 
Peripheral Blood Mononuclear Cell 
PCR 
 
 
Polymerase chain reaction 
PI 
 
 
Principal Investigator 
QA 
 
 
Quality Assurance 
RMSE  
 
Residual Mean Square Error 
SAE 
 
 
Serious adverse event 
SC 
 
 
Subcutaneous  
SOE 
 
 
Schedule of events 
SOP 
 
 
Standard operating procedure 
T1D 
 
 
Type 1 diabetes, previously known as insulin-dependent diabetes mellitus 
Tregs  
 
Regulatory T cells 
ZnT8  
 
Zinc Transporter 8 

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TABLE OF CONTENTS 
 
1. 
INTRODUCTION ........................................................................................................................... 7 
1.1. 
Study Overview ......................................................................................................................... 7 
2. 
BACKGROUND AND RATIONALE FOR THE PROPOSED TRIAL ...................................... 12 
2.1. 
Type 1 diabetes........................................................................................................................ 12 
2.2. 
Omega-3 Fatty Acids .............................................................................................................. 13 
2.3. 
Omega-3 Fatty Acids and Inflammation ................................................................................. 14 
2.4. 
Omega-3 Polyunsaturated Fatty Acid Intake and T1D ........................................................... 14 
2.5. 
Vitamin D in T1D ................................................................................................................... 16 
3. 
STUDY RATIONALE .................................................................................................................. 17 
4. 
STUDY DESIGN........................................................................................................................... 18 
4.1. 
Overview ................................................................................................................................. 18 
4.2. 
Objectives ................................................................................................................................ 19 
4.2.1. 
Primary Objective(s) ........................................................................................................ 19 
4.2.2. 
Secondary Objective(s) .................................................................................................... 19 
4.3. 
Investigational Plan ................................................................................................................. 19 
4.4. 
Summary of Inclusion and Exclusion Criteria ........................................................................ 20 
4.4.1. 
Inclusion Criteria ............................................................................................................. 20 
4.4.2. 
Exclusion Criteria ............................................................................................................ 20 
4.5. 
Informed Consent .................................................................................................................... 22 
4.6. 
Description of Treatment Groups ............................................................................................ 22 
4.7. 
Treatment Assignment ............................................................................................................ 22 
4.8. 
Study Assessments .................................................................................................................. 23 
4.9. 
Quality Assurance ................................................................... Error! Bookmark not defined. 
4.10. Post-treatment Follow-up ........................................................................................................ 23 
5. 
PATIENT MANAGEMENT ......................................................................................................... 23 
5.1. 
Screening ................................................................................................................................. 23 
5.2. 
Randomization ........................................................................................................................ 24 
5.3. 
Intensive Diabetes Management ............................................................................................. 24 
5.4. 
Administration of Study Drugs ............................................................................................... 24 
5.4.1. 
Omega-3 Fatty Acids ....................................................................................................... 25 
5.4.2. 
Vitamin D......................................................................................................................... 27 
5.4.3. 
Assessment of Participant Compliance with Study Agent .............................................. 29 
5.4.4. 
Prior and Concomitant Treatments .................................................................................. 30 
5.4.5. 
Removal of Subjects from Therapy or Subject Withdrawal or Early Termination ......... 30 
5.4.6. 
Discontinuation of study drug(s) in an individual patient ............................................... 30 

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5.4.7. 
Re-Entry into Study Treatment ........................................................................................ 31 
5.4.8. 
Risk to the Subjects .......................................................................................................... 31 
6. 
STUDY ASSESSMENTS ............................................................................................................. 32 
6.1. 
General Assessments ............................................................................................................... 32 
6.2. 
Enrollment ............................................................................................................................... 33 
6.3. 
Screening (visit -1) and Baseline visits ................................................................................... 33 
6.4. 
Diabetes Management ............................................................................................................. 33 
6.5. 
Laboratory Assessments .......................................................................................................... 34 
6.6. 
Mechanistic Outcome Assessments ........................................................................................ 34 
6.7. 
Metabolic Outcome Assessments............................................................................................ 34 
6.8. 
Visit Windows ......................................................................................................................... 35 
6.9. 
Unscheduled visits................................................................................................................... 35 
6.10. Sample retention ...................................................................................................................... 35 
7. 
PARTICIPANT SAFETY .............................................................. Error! Bookmark not defined. 
7.1. 
Staggering Administration ...................................................................................................... 36 
7.2. 
Intensive T1D Management .................................................................................................... 36 
7.3. 
Safety Monitoring ................................................................................................................... 36 
7.4. 
Ad-hoc DSMB Reviews .......................................................................................................... 37 
7.5. 
Temporary Suspension of Enrollment/Drug(s) Dosing or Both ............................................. 37 
7.6. 
Stopping rules .......................................................................................................................... 37 
7.7. 
Inclusion and exclusion criteria............................................... Error! Bookmark not defined. 
7.8. 
Potential Benefits .................................................................................................................... 38 
7.9. 
Pregnancy ................................................................................................................................ 38 
7.10. Protecting Against or Minimizing Potential Treatment Risks ................................................ 38 
7.11. Supplement - Drug Interaction ................................................................................................ 39 
8. 
ADVERSE EVENT REPORTING AND SAFETY MONITORING ........................................... 40 
8.1. 
Adverse Event Definition ........................................................................................................ 40 
8.1.1. 
Adverse Event (AE) ......................................................................................................... 40 
8.1.2. 
Serious Adverse Event ..................................................................................................... 41 
8.1.3. 
Unexpected Adverse Event .............................................................................................. 41 
8.1.4. 
Grading Event Severity .................................................................................................... 41 
8.1.5. 
Evaluation of Adverse events .......................................................................................... 41 
8.1.6. 
Adverse Event Reporting and Monitoring ....................................................................... 42 
8.1.7. 
Recording Adverse Events ............................................................................................... 43 
8.1.8. 
Adverse Events Requiring Expedited Reporting ............................................................. 44 
8.1.9. 
Special Reporting Situations ............................................................................................ 44 
8.1.10. 
Laboratory Test Result Abnormalities ......................................................................... 44 

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8.1.11. 
Pregnancy ..................................................................................................................... 44 
8.1.12. 
Overdose....................................................................................................................... 45 
8.1.13. 
Other Safety Considerations......................................................................................... 45 
9. 
DATA QUALITY ASSURANCE ................................................................................................. 45 
9.1. 
Data Collection, Monitoring, and Transfer ............................................................................. 45 
9.2. 
Quality Control and Quality Assurance .................................................................................. 46 
10. 
STATISTICAL CONSIDERATIONS AND ANALYSIS PLAN .............................................. 46 
10.1. Overview ................................................................................................................................. 46 
10.2. Endpoints/Outcomes ............................................................................................................... 46 
10.3. Measures to Minimize Bias ..................................................................................................... 46 
10.4. Analysis Plan ........................................................................................................................... 46 
10.4.1. 
Analysis Populations .................................................................................................... 46 
10.4.2. 
Primary Analysis of Primary Endpoint(s)/Outcome(s) ................................................ 47 
10.4.3. 
Supportive Analyses of the Primary Endpoint(s)/Outcome(s) ..................................... 47 
10.4.4. 
Descriptive Analyses .................................................................................................... 47 
10.5. Interim Analyses ..................................................................................................................... 48 
10.5.1. 
Interim Analysis of Efficacy Data ................................................................................ 48 
10.5.2. 
Interim Analysis of Safety Data ................................................................................... 48 
10.5.3. 
Futility Analysis ........................................................................................................... 48 
10.6. Statistical Hypotheses ............................................................................................................. 49 
10.7. Sample Size Considerations .................................................................................................... 49 
11. 
ETHICAL CONSIDERATIONS & COMPLIANCE WITH GOOD CLINICAL PRACTICE . 49 
11.1. Statement of Compliance ........................................................................................................ 50 
11.2. Informed Consent .................................................................................................................... 50 
11.3. Study Subject Confidentiality ................................................................................................. 50 
12. 
STUDY ADMINISTRATION.................................................................................................... 51 
12.1. Groups and Committees .......................................................................................................... 51 
12.1.1. 
Protocol Development Committee ............................................................................... 51 
12.1.2. 
Clinical Site .................................................................................................................. 51 
12.1.3. 
Medical Monitor for the Clinical Protocol ................................................................... 51 
12.1.4. 
Clinical Site Monitoring ............................................................................................... 51 
12.1.5. 
Data and Safety Monitoring Board (DSMB) ............................................................... 52 
12.1.6. 
DRI Office of Regulatory Support ............................................................................... 52 
12.1.7. 
Human Subjects Research Office & Institutional Review Boards (HSRO & IRB) .... 52 
12.1.8. 
Clinical Research Centers ............................................................................................ 52 
12.1.9. 
Investigational Drug Services (Research Pharmacy) ................................................... 52 
12.1.10. 
Core Laboratories ......................................................................................................... 53 
12.2. Preservation of the Integrity of the Study ............................................................................... 53 
12.3. Participant Reimbursement and Compensation ...................................................................... 53 
13. 
STUDY TIMELINE ................................................................................................................... 53 

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REFERENCES ...................................................................................................................................... 65 
 
1. INTRODUCTION 
1.1. 
Study Overview 
 
Title 
A Pilot, Safety and Feasibility Trial of High-Dose Omega-3 Fatty Acids and High-
Dose Cholecalciferol Supplementation in Type 1 Diabetes 
 
 
Acronym  
POSEIDON study: Pilot study of omega-3 and vitamin D high doses in T1D  
 
Conducted By 
Diabetes Research Institute 
 
Study Phase 
Phase I/IIa 
 
Accrual Objective 
56 subjects over three years 
 
Study Design  
The study is a two-arm, open label, randomized trial.  All groups will receive 
standard intensive diabetes treatment with insulin and dietary management. 
There will be four groups of participants in this study 
 
Group 
Subject population  
I 
Fourteen (14) adults (18-65 years) of established T1D of more 
than 6 months duration (>180 days) and up to 10 years  
II 
Fourteen (14) adults (18-65 years) of new-onset T1D diagnosed 
within last 6 months (≤ 180 days) 
III 
Fourteen (14)children (6-17 years) of established T1D of more 
than 6 months duration (>180 days) and up to 10 years  
IV 
Fourteen (14) children (6-17 years) of new-onset T1D diagnosed 
within last 6 months (≤ 180 days) 
 
Participants in each group will be randomly assigned in a 1:1 ratio to receive 
either one year of high dose Omega-3 fatty acids and Vitamin D combination 
(Arm A) or Vitamin D alone (Arm B).  Both arms will receive Vitamin D 
supplementation.  
 
Table 1. This table represents supplements used in the study, their doses, 
routes of administrations and description of two arms 
 
Drugs with 
route of 
administration 
 
Dose 
Arm A 
Arm B 
Ultra-refined 
omega-3 
EPA/DHA 
concentrate in 
Liquid or 
Based on the body 
weight of the 
participant 
 
Initial Dose: 
Three times 
daily orally 
 

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Capsule 
formulations 
 
(ZoneLabs® 
OmegaRx®2 
Fish Oil – 
Liquid; 
containing 
3375 mg 
EPA+DHA 
[2250 mg EPA 
and 1125 mg 
DHA] in 5 mL 
 
 
 
Of note, each 
1 mL of 
OmegaRx®2 
contains 675 
mg of 
EPA+DHA. 
 
150 mg of EPA+DHA 
per Kg body weight. 
 
Maintaince Dose: 
Dose will be adjusted 
to maintain a target 
AA/EPA ratio of 1.5-
3.0. AA/EPA ratios will 
be monitored monthly 
until achieving target 
levels and as per 
protocol or as clinically 
indicated thereafter. 
 
It 
is 
150 
mg 
of 
combined 
EPA 
and 
DHA 
per 
kg 
body 
weight to be given 
either as a single or 
divided 
into 
smaller 
doses through the day. 
 
Vitamin-D 
25OH 
(Cholecalcifero
l) 
Based on the Vitamin 
D level and age of the 
participant  
 
For levels 30 -39 
ng/ml:   
 
Age ≤10 years give 
500 IU daily  
 
Age >10 years give 
1000 IU daily 
 
For levels 20 – 29 
ng/ml 
 
50,000 IU weekly for 2 
months and then: 
 
Age ≤10 years give 
1000 IU daily 
 
Age >10 years give 
2000 IU daily 
 
For levels < 20 ng/ml  
 
Daily orally  
 
Daily orally  

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50,000 IU weekly for 6 
months and then: 
 
Age ≤10 years, give 
1000 IU daily   
 
Age >10 years, give 
2000 IU daily 
 
Dose will be adjusted 
and monitored every 
four months until 
achieving target levels 
and as clinically 
indicated thereafter. 
Treatment 
Description 
Subjects will be randomized to one of the two arms. The treatment arm A 
includes Omega-3 Fatty Acids and Vitamin D. The subjects randomized in Arm 
B (control group) will receive only Vitamin D supplement. The treatment 
assignment will be conducted in a open label fashion.  
 
Study Duration 
All subjects will be treated and followed for one year, and will be followed for up 
to one additional year beyond the period of treatment.  Enrollment is expected 
to occur over three years. 
 
Objective 
Conduct a clinical intervention study to assess, in participants with established 
T1D (>6 months and up to 10 years after diagnosis) or with new onset T1D (≤6 
months from diagnosis): 
Primary outcome: Stimulated (90 minute sample of a MMTT) C-peptide at the 1 
year visit greater or equal to baseline level. 
Secondary objectives: Stimulated C-peptide area under the curve (AUC) during 
a 4-hour MMTT greater or equal to baseline level, at the 1 year visit. 
Reduction in HbA1c at the one year visit compared to baseline. 
Reduction in insulin requirement at the 1 year visit compared to baseline. 
Incidence of adverse events (AE) comparable to general diabetes population. 
Inclusion Criteria 
 
 
 
 
 
 
 
 
 
 
 
 
Patients must meet all of the following criteria to be eligible to participate in this 
study: 
• 
Subject must be able to understand and provide informed consent. 
• 
Males and females, 6-17 years for children and 18-65 years of age for 
adult group. 
• 
For new onset T1D - from onset to 6 months (180 days) post diagnosis 
at the time of randomization. 
• 
For established T1D – At least 6 months and up to 10 years from the 
diagnosis at the time of randomization.  

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Exclusion Criteria 
• 
Affected by T1D, according to ADA standard criteria, and confirmed by 
positivity of at least one T1D-associated autoantibody, to GAD65, IA-2, 
ZnT8, or insulin autoantibody (if patient has been treated with insulin for 
less than 2 weeks). 
• 
T1D must be treated with insulin (except if participant is in Honeymoon 
period/phase).  
• 
Stimulated C-peptide peak level, at the baseline 1 visit MMTT, ≥ 0.2 
ng/ml. 
• 
Female subjects of childbearing potential must have a negative 
pregnancy test upon study entry. 
• 
Adequate venous access to support study required blood draws. 
 
Patients must not meet any of the following criteria to be eligible to participate: 
• 
Inability or unwillingness of a participant to give written informed consent 
or comply with study protocol. 
• 
BMI>30 kg/m2. 
• 
Contra-indications to any of the drugs used in the trial (as per package 
insert, e.g., knowledge of hypersensitivity to drugs or its excipients). 
• 
Uncompensated heart failure, fluid overload, myocardial infarction or 
liver disease or severe impairment of a vital organ within the last 6 weeks 
before enrollment. 
• 
Any of the following laboratory findings indicating hemoglobin <10.0 
g/dL; leukocytes <3,000/μL; neutrophils <1,500/μL; lymphocytes 
<800/μL; platelets <100,000/μL. 
• 
Any sign or diagnosis of significant chronic active infection (e.g., 
hepatitis, tuberculosis, EBV, or CMV), or screening laboratory evidence 
consistent with a significant chronic active infection (such as positive for 
HIV, IGRA test for TB, or hepatitis B-C).  
• 
Ongoing acute infections, e.g., acute respiratory tract, urinary tract, or 
gastrointestinal tract infections. 
• 
Ongoing or anticipated use of diabetes medications other than insulin. 
• 
Current or ongoing use of non-insulin pharmaceuticals that affect 
glycemic control within prior 7 days of screening. 
• 
Recent recipient of any licensed or investigational live attenuated 
vaccine(s) within 6 weeks of randomization. 
• 
Use of investigational drugs within 3 months of participation. 
• 
Concomitant 
therapy 
with 
immunosuppressive 
drugs, 
immunomodulators, or cytotoxic agents, or previous therapy less than 3 
months from randomization. 

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• 
History or diagnosis of malignancy, with the exception of a history of 
localized basal or squamous cell carcinoma.  
• 
History of gastroparesis or other severe gastrointestinal disease.. 
• 
Presence of an allograft. 
• 
AST, ALT or Alkaline Phosphatase >2 times upper limit of normal or total 
bilirubin >1.5 times upper limit of normal.     
• 
History of mental illness deemed to be clinically unstable or any situation 
that, in the opinion of the investigator, would interfere with the 
participant’s ability to comply with study requirements. 
• 
History of illicit drug or alcohol abuse. 
• 
Pregnancy or ongoing breastfeeding for women; unwillingness or 
inability of both females and males of childbearing age to use a reliable 
and effective form of contraception, for the entire duration of the study. 
• 
Past or current medical problems or findings from physical examination 
or laboratory testing that are not listed above, which, in the opinion of 
the investigator, may pose additional risks from participation in the study, 
may interfere with the participant’s ability to comply with study 
requirements or that may impact the quality or interpretation of the data 
obtained from the study. 
 

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2. BACKGROUND AND RATIONALE FOR THE PROPOSED TRIAL 
 
2.1. Type 1 diabetes 
Type 1 diabetes (T1D) is a chronic autoimmune disease leading to the selective destruction of insulin 
producing beta cells within the pancreatic islets. Autoreactive T cells and abnormalities in immune 
regulation, especially in regulatory T cells (Treg cells), play a key role in disease development 
(Pugliese, 2014). In the U.S. alone, approximately 1.3 million people, especially children and 
adolescents, suffer from T1D; the disease incidence is increasing in many countries (Cizza et al, 2012; 
Forlenza & Rewers, 2011; Pettitt et al, 2013; Vehik & Dabelea, 2011). Chronic autoimmune T cell 
responses against pancreatic ß-cells are considered the primary cause of T1D, leading to loss of ß-cell 
mass and insulin secretion and in turn life-long dependence on insulin injections (Coppieters et al, 
2012; Roep & Peakman, 2012). The disease severely impacts quality of life and confers risk for both 
acute and chronic complications linked to significant morbidity and mortality, such as end stage renal 
disease, blindness, cardiovascular disease, diabetic ketoacidosis and hypoglycemia (Cengiz et al, 
2013; Nathan, 2014). The economic burden caused by T1D amounts to approximately $14.4 billion in 
medical costs and lost income (Tao et al, 2010).  
The discovery of insulin in 1922 dramatically changed the prognosis of T1D. It was the first effective 
therapy, which converted a fatal condition into a chronic disease. However, the prolonged life 
expectancy allows time for the development of long-term complications, e.g. retinopathy, neuropathy, 
nephropathy and vasculopathy, which are largely related to the imperfect degree of metabolic control 
afforded by insulin therapy. The “Diabetes Control and Complications Trial” showed those intensive 
insulin regimens that achieve tight blood glucose control reduced the risk of complications in T1D 
(Nathan, 2014). Yet this level of tight control is difficult to achieve for many patients, despite many 
advances in insulin therapy and blood glucose monitoring, and it bears significant risk of hypoglycemia 
(Steffes et al, 2003; Zinman, 1998). 
T1D requires lifelong therapy with insulin injections, multiple times daily. Despite much progress in 
insulin preparations, insulin therapy is inadequate in most patients (Hirsch & Skyler, 2000; Pouwer & 
Hermanns, 2009). Insulin dosing largely remains an empiric decision and is prone to error, which in 
turn can lead to severe, life-threatening hyperglycemia or hypoglycemia. Blood glucose monitoring is 
required to adjust insulin doses. Even patients who employ frequent finger-stick blood glucose 
monitoring fail to maintain strict glycemic control within the ADA-specified optimal range 70% of the 
time (Bode et al, 2005). The use of wearable Continuous Glucose Monitoring (CGM) devices and insulin 
pumps represent a step forward, yet they fail to restore normal glucose metabolism and may still be 
hyperglycemic almost 30% of the time (Bode et al, 2005). Patients using CGM may be hypoglycemic 
approximately 8% of each day, with a propensity for hypoglycemic events to occur at night (Bode et al, 
2005), when hypoglycemia can more likely go unnoticed and can be therefore more dangerous.  In 
some cases, severe hypoglycemic events, i.e., those requiring the aid of another person to administer 
carbohydrates, glucagon, or other resuscitative assistance, can lead to seizures, unconsciousness, 
coma, and death. Frequent hypoglycemia lead to hypoglycemia unawareness, which increases their 
risk of severe hypoglycemia and serious sequelae by at least 6-fold (Seaquist et al, 2013). Moreover, 
severe and multiple hypoglycemic episodes are associated with increased risk of death lead to death 
(Weinstock et al, 2013). 

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Transplantation can reverse diabetes by replacing the lost beta cell mass. Both transplantation of whole 
pancreas and transplantation of purified pancreatic islets are possible and can lead to reversal of the 
diabetic state (Bruni et al, 2014; Mineo et al, 2009; Watson, 2015; White et al, 2009). Both procedures 
have improved over time, but many more whole pancreas transplants have been performed compared 
to islet transplants. Although it requires invasive surgery and has a higher risk of perioperative mortality 
and morbidity, pancreas transplantation is correlated with longer graft survival and function than islet 
transplantation. Pancreas transplantation is particularly beneficial for patients with T1D and end stage 
renal disease (ESRD), in whom transplantation of pancreas and kidney restores both insulin secretion 
and renal function for several years. However, in all forms of transplantation, patients are required to 
be treated with chronic immunosuppression to prevent rejection, which is associated with well-known 
side effects and health risks that also limit applicability of transplantation to younger patients, 
representing the predominant portion of newly diagnosed patients. Moreover, our own studies have 
shown that approximately 5-6% of pancreas transplant recipients may develop recurrence of type 1 
diabetes in the transplanted pancreas, despite immunosuppression that prevents rejection (Burke et al, 
2011; Vendrame et al, 2010). 
There is much hope that stem cells based approaches may provide new sources of beta cells, and yet 
it will be critical to define immunomodulation protocols that can control islet autoimmunity and prevent 
rejection of transplanted cells, recurrence of disease, and as well help patients preserve insulin 
secretion from the residual beta cell mass at diagnosis (Orlando et al, 2014). Thus, there is a need to 
develop new therapies to interdict islet autoimmunity, to prevent, treat and cure T1D. Even a partial 
level of insulin secretion affords protection from chronic complications (Steffes et al, 2003), 
hypoglycemia and diabetic ketoacidosis (Ludvigsson, 2013), which can all lead to death (Realsen et al, 
2012; Rosenbloom, 2010; Weinstock et al, 2013).  
 
2.2. Omega-3 Fatty Acids  
The human body can make most of the types of fats it needs from other fats or raw materials. That isn’t the 
case for omega-3 fatty acids (also called omega-3 fats and n-3 fats). These are essential fats—the body 
can’t make them but must get them from food. Foods high in Omega-3 include fish, vegetable oils, nuts 
(especially walnuts), flax seeds, flaxseed oil, and leafy vegetables. 
The omega-3 fats are special for many reasons. They are an integral part of cell membranes throughout 
the body and affect the function of the cell receptors in these membranes. They provide the starting point 
for making hormones that regulate blood clotting, contraction and relaxation of artery walls, and 
inflammation. They also bind to receptors in cells that regulate genetic function. Likely due to these effects, 
omega-3 fats have been shown to help prevent heart disease and stroke, may help control lupus, eczema, 
and rheumatoid arthritis, and may play protective roles in cancer and other conditions. 
Omega-3 fats are a key family of polyunsaturated fats. There are three main omega-3s: Eicosapentaenoic 
acid (EPA) and docosahexaenoic acid (DHA) come mainly from fish, so they are sometimes called marine 
omega-3s. 
Alpha-linolenic acid (ALA), the most common omega-3 fatty acid in most Western diets, is found in 
vegetable oils and nuts (especially walnuts), flax seeds and flaxseed oil, leafy vegetables, and some animal 
fat, especially in grass-fed animals. The human body generally uses ALA for energy, and conversion into 
EPA and DHA is very limited. 
 

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Cell  membranes  require  unsaturated  fatty  acids  to  maintain  structure  and  function.    Desaturase 
enzymes  present  in  mammals  add  a  double  bond  to  form  a  monounsaturated  fatty  acid,  oleic  
acid(18:1  n9),  from  stearate.    Animals,  unlike  plants,  can  insert  additional  double  bonds  to  create  
two classes  of  polyunsaturated  fatty  acids  (PUFA),  n-6 and n3. Plants  convert  oleic  acid  to  linoleic  
acid (LA)  (18:2  n-6)  and  desaturation  of  oleic  produces  α-linolenic  (ALA)  (18:3  n-3).  Marine  algae  
also  produce  EPA  (20:5  n-3)  and  DHA  (22:6  n-3)  that  are  eventually  transferred  through  the  food  
chain to  fish  whose  oils  are  high  in  ω-3  fatty  acids.  Mammals  cannot  synthesize  LA  (the  
predominant  PUFA  in  Western  diets)  and  ALA;  therefore  these  fatty  acids  are  “essential”  and  must  
be  ingested. Mammals,  unlike  plants,  cannot  interconvert  n9,  n6  and  n3  fatty  acids,  but  do  further  
convert  LA  to  other n-6 fatty  acids,  (e.g.,  arachidonic  acid  (20:4  n-6))  and  ALA  to  n-3  EPA  and  
DHA.     
 
2.3. Omega-3 Fatty Acids and Inflammation  
Omega-3  fatty  acids  reduce  inflammatory  cytokines  and  inflammatory  prostaglandins,  both  of  
which may  be  related  to  the  initial  inflammation  in  islet  cells. The  mechanism  of  omega-3  fatty  
acids reducing  inflammation  has  been  shown  secondary  to  limiting  proinflammatory  cytokines  
and  the activity  of  the  PGS2  enzyme (Litherland et al., 1999) . Therefore,  it  was  proposed  that  
the  increased  PGS2  expression and  cytokine  production  observed  in  children  at  high-risk  for  
developing  T1D  may  be  reduced through  omega-3  fatty  acid  supplementation  (Litherland et al., 
2003).    Arachidonic  acid  (AA),  DHA,  and eicosapentaenoic  acid  (EPA)  inhibit  PGS2  with  
similar  potency,  which  is  greater  than  that  seen  with other  fatty  acid  analogues.   
Endres  et  al  (1989)  found  that  the  syntheses  of  interleukin-1  (IL-1alpha  and  IL-1  beta)  and  
of  tumor necrosis  factor  (TNF),  cytokines  with  potent  inflammatory  activities,  were  reduced  by  
dietary  supplementation  with  n-3  fatty  acids.    The  anti-inflammatory  effect  of  these  n-3  fatty  
acids  may  be mediated  in  part  by  their  inhibitory  effect  on  the  production  of  lL-1  and  TNF.   
 
2.4. Omega-3 Polyunsaturated Fatty Acid Intake and T1D  
 
The  incidence  of  T1D  is  increasing,  particularly  in  very  young  children.    We  hypothesize  that  
the lack  of  omega-3  fatty  acids  in  the  diet  has  contributed  to  this  increase.  These are 
essential fats—the body can’t make them but must get them from food.  Just  100  years  ago, 
consumption  of  n-6  to  n-3  fatty  acids  in  the  diet  was  at  a  1:1  ratio.    In  1999,  the  ratio  was  
reported  to be  approximately  30:1 (Simopoulos  AP.  1999) .  Because  of  the  warnings  to  
eliminate  fish  during  pregnancy,  the  ratio  is likely  even  greater  during  pregnancy.    This  results  
in  increased  production  of  inflammatory prostaglandins  and  cytokines  during  this  critical  time  of  
fetal  development.    This  trend  towards  substitution  of  the  inflammation  producing  n-6  fatty  
acids  at  the  expense  of  the  anti-inflammatory  n-3  fatty  acids  may  be  contributing  to  the  
increase  in  the  number  of  cases  of  T1D  diagnosed  each  year.  
  
More  direct  evidence  also  suggests  a  relationship  between  omega-3  fatty  acids  and  T1D.    
Decsi and  colleagues  demonstrated  that  40  children  with  T1D  had  significantly  lower  DHA  
levels  and  a lower  n-3:  n-6  ratio  than  the  control  children,  which  may  partially  explain  the  
altered  prostanoid  and inflammatory  cytokine  production.  They  conclude  that  the  reduced  
availability  of  long-chain polyunsaturates  in  children  with  T1D  suggests  the  need  for  enhanced  
dietary  supply (Decsi  et al., 2002).    
 
Observations  have  been  made  that  children  who  have  received  omega-3  fatty  acid  
supplementation have  a  lower  risk  of  T1D.    Children  born  to  women  from  Norwegian  fishing  
villages  had  a significantly  decreased  risk  of  getting  diabetes  compared  with  the  children  of  

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women  who  lived  in cities  away  from  the  coast  (Joner  G.  1992).    Using  a  case-control  
design,  Stene  et  al  found  that  cod  liver  oil, either  taken  by  the  mother  during  pregnancy  or  
by  the  child  during  the  first  year  of  life,  was associated  with  a  decreased  risk  of  developing  
T1D (Stene et al., 2000).    They  concluded  that  either  the  vitamin  D or  the  eicosapentaenoic  
acid  (EPA)  or  the  docosahexaenoic  acid  (DHA)  or  all  three  cod  liver  oil components  have  
protective  effects  against  developing  T1D.    In  a  larger  and  more  recent  study, Stene  and  
colleagues  in  Norway  found  that  supplementation  with  cod  liver  oil  during  the  first  year  of life  
was  associated  with  a  significantly  lower  risk  of  T1D  before  age  15,  odds  ratio  (OR)  of  0.7  
(p<0.001)  (Stene  and Joner, 2003).    This  is  consistent  with  the  hypothesis  that  omega-3  fatty  
acids  (or  vitamin  D)  have  a protective  effect.    It  now  appears  that  the  protective  effect  may  
occur  during  pregnancy  or  during  infancy  or  both. 
 
T1D is an autoimmune disease that is characterized by the destruction of insulin-producing beta cells 
in the pancreatic islets. Although it is not yet known what initiates the autoimmune process, it is likely 
that both genetic background and environmental factors contribute to the disease process. Dietary 
factors have been implicated in the etiology of type 1 diabetes as well as in initiating the autoimmune 
process that leads to clinical disease. A case-control study from Norway (Stene and Joner, 2003) 
reported that children with diabetes were less likely to have been given cod liver oil during infancy than 
children without diabetes. Given that cod liver oil contains both vitamin D and the marine omega-3 fatty 
acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), it was not clear whether the 
protective factor in cod liver oil was the vitamin D, the marine fatty acids, or both.  
Although two studies reported that children with diabetes were less likely to have taken vitamin D 
supplements in infancy than children without diabetes (EURODIAB Substudy 2 Study Group. Vitamin 
D supplement in early childhood and risk for type I insulin-dependent diabetes mellitus, and Hypponen 
et al 2001), similar investigations focusing on the intake of marine omega-3 fatty acids have not been 
conducted to resolve this important question. The clinical phase of type 1 diabetes, where 
hyperglycemia manifests, is preceded by an asymptomatic period that varies in duration, ranging from 
a few months to several years, in which autoantibodies to the beta cells and their antigens are 
detectable in the blood. Persistent positivity of these autoantibodies confers a high risk of subsequent 
development of type 1 diabetes in relatives of those individuals with diabetes and in the general 
population. Because autoantibodies appear before clinical diabetes, examination of risk factors for the 
appearance of these autoantibodies would yield important clues regarding the early pathogenic events 
leading to autoimmunity, and perhaps the pathogenesis of type 1 diabetes itself.  
Studies suggest that macrophage infiltration and inflammatory cytokine production are early events in 
the pathogenesis of type 1 diabetes (Chase et al., 2004; Jansen et al., 1994; Green and Flavell, 1994; 
Dahlen et al., 1998). Therefore, identifying factors that either promote or block the impact of these early 
pathogenic inflammatory events may be key to promoting or inhibiting the development of type 1 
diabetes. Several studies have demonstrated a strong effect of omega-3 fatty acids on inflammatory 
responses in animals and humans (Endres et al., 1989; Calder, 2001; De Caterina et al., 2004). A 
relative deficiency of omega-3 fatty acids, a characteristic of many Western diets, may predispose to 
heightened inflammatory reactions and thus increase the risk for autoimmune diseases, such as type 
1 diabetes. Alpha-linolenic acid (ALA) is the principal omega-3 fatty acid in Western diets and is found 
in the green leaves of plants, and also in selected seeds, nuts, and legumes (eg, flax, canola, walnuts, 
and soy). Alpha-linolenic acid may serve in a limited capacity as a precursor for EPA and DHA, 2 
omega-3 fatty acids that are primarily obtained from fish. Linoleic acid is the most abundant omega-6 

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fatty acid in the diet and is found primarily in nut, seed, and vegetable oils. Arachidonic acid is an 
omega-6 fatty acid that can be derived from linoleic acid and is also found in meat and poultry. Because 
ALA and linoleic acid compete for key enzymes involved in fatty acid metabolism and conversion to 
either pro-inflammatory or anti-inflammatory eicosanoids, it is important to examine the effects of 
omega-3 and omega-6 fatty acid intakes together.  
 
2.5. Vitamin D in T1D 
As  noted  above,  the  observation  that  cod  liver  oil  was  associated  with  lower  risk  of  T1D  
suggests the  possibility  of  a  synergistic  effect  of  both  of  the  components  of  cod  liver  oil  (DHA  
and  vitamin  D) (Stene and Joner, 2003).  A  large  retrospective  cohort  study  demonstrated  an  
association  between  vitamin  D  supplementation  in  the  first  year  of  life  and  a  lower  risk  of  T1D  
[24].    This  association  was  also demonstrated  in  a  prospective  study  in  which  infants  received  
vitamin  D  supplementation (Stene et al., 2000; Stene and Joner, 2003).    The  mechanism  may  be  
related  to  the  immunosuppressive  properties  of  Vitamin  D  or  possibly  a direct  effect  on  islet  
cell  insulin  secretion.    Vitamin  D  receptor  (VDR)  gene  polymorphisms  have  recently  been  
associated  with  susceptibility  to  T1D  in  humans (Alician et al.,2009; Lowe et al., 2007).      
Interestingly,  Stene  et  al (2000) found  an  indication  for  a  protective  effect  with  certain  
polymorphisms,  but  this  was  strongest  for children  who  took  cod  liver  oil  during  the  first  year  
of  life .   
Serum 25-hydroxyvitamin D (25-OHD) concentrations are largely determined by environmental factors, 
mainly through vitamin D intake and ultraviolet exposure. The sun is the primary source of vitamin D, 
which is synthesized endogenously in skin to produce cholecalciferol (vitamin D3), although a small 
proportion (<20%) of vitamin D comes through diet from a limited range of foods (in the form of 
ergocalciferol [vitamin D2] and vitamin D3) (Holick 2006). The main marker of vitamin D status is the 
metabolite 25-OHD, which is synthesized in the liver (Chiu et al., 2004; Norman et al., 1980).   
A relationship between type 1 diabetes mellitus and vitamin D deficiency has been reported (Luong  et 
al., 2005; Mathieu et al., 2005). The prevalence of vitamin D deficiency in patients with type 1 diabetes 
was 15% to 90.6% (Greer and Rogers, 2007; Svoren et al., 2009; Bener et  al., 2009).  There is 
evidence that vitamin D is important in the prevention of islet cell death and might be useful in improving 
the survival of islet cell grafts, and it improves the production of insulin. Low vitamin D levels were 
shown to have a negative effect on beta-cell function (Chiu et al., 2004; Norman et al., 1980).  Regular 
doses of vitamin D early in life have been shown to reduce the risk of developing type 1 diabetes (Luong  
et al., 2005). Vitamin D treatment has also been shown to improve glycemic control and insulin 
sensitivity in people with type 1 and type 2 diabetes and in normal individuals. Increasing vitamin D 
levels from 25 to 75 nmol/L results in a 60% improvement in insulin sensitivity. (Chiu et al., 2004; 
Norman et al., 1980).  Borissova et al., 2003; Gedik and Akalin 1986; Schwalfenberg 2008). These 
effects have been mainly attributed to the immunomodulatory actions of vitamin D (Luong  et al., 2005).   
Geographic location seems to play a part in the development of type I diabetes with risk increasing as 
farther from the equator. The location on Earth contributes to the amount of vitamin D thebody 
produces. This observation is suggestive, but not proof, that vitamin D affects the development of type 
1 diabetes. Finland, a country that is at latitude far from the tropics, has the highest incidence of type 1 
diabetes in the world. In the late 1960s, it was common to supplement an infant’s diet with 2,000 IU 

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vitamin D. When researchers compared the development of type 1 diabetes in people 30 years later 
based on whether their mothers recalled giving them vitamin D during the first year after birth, they 
found an 80 percent reduction in the risk of type 1 diabetes later in life. Calcitriol makes insulin-secreting 
beta cells resistant to immune system attack. The active vitamin appears to reduce the production of 
cytokines, substances that kill beta cells. 
Currently, project NCT03046927 conducted at the University of Massachusetts, Worcester is studying 
the role of vitamin D supplementation on the honeymoon phase of type 1 diabetes in children who are 
on standardized insulin treatment. The investigation is based on hypothesis that early intervention with 
vitamin D lead to significant changes in the approach to the early phase of type 1 diabetes with a strong 
emphasis on prolonging the honeymoon phase by using vitamin D and maintaining these patients on 
a standardized insulin regimen. The study rationale also indicates role of vitamin D in reducing the long-
term complications of type 1 diabetes. 
 
3. STUDY RATIONALE  
The anti-inflammatory properties of omega-3 long-chain polyunsaturated fatty acids (LCPUFA) 
[eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] have been extensively reported in 
the literature. Derivatives from omega-3 LCPUFA have been associated with marked anti-
inflammatory effects whereas arachidonic acid (AA), an omega-6 LCPUFA, is the precursor of 
eicosanoids associated with pro-inflammatory properties. Thus, a high AA/EPA ratio would suggest 
that the balance is shifted towards a pro-inflammatory state. Treatment with omega-3 fatty acids 
aimed at lowering the AA/EPA ratio may then prove to be beneficial particularly in diseases 
associated with a high inflammatory state. Bioactive metabolites such as resolvins and protectins are 
derived from omega-3 fatty acids and are postulated to mediate the anti-inflammatory properties of 
omega-3 fatty acids. These mediators act by several mechanisms including reduction of neutrophil 
infiltration, attenuation of TNF stimulated NF-ĸB activation, inhibition of interleukin-1β (IL-1β), 
reduction in tumor necrosis factor (TNF) and interferon-ɣ secretion, inhibition of T-cell migration and 
regulation of T-cell apoptosis. Notably, IL-1β and TNF have both been implicated in beta-cell death 
and IFN signatures as well as T cell infiltration of the pancreatic islets have been associated with type 
1 diabetes (T1D). 
 
A number of studies suggest that omega-3 fatty acids may be beneficial in T1D: 
• 
Bellenger et al. reported that transgenic fat-1 mice, a model of endogenous omega-3 LCPUFA 
synthesis, were protected against streptozotocin-induced diabetes. This protection was 
associated with a reduction in pro-inflammatory cytokines with a concomitant increase in anti-
inflammatory cytokines.   
• 
The Diabetes Autoimmunity Study in the Young (DAISY) follows children at high risk for T1D 
development; in this study, dietary intake of omega-3 fatty acids was associated with a 
reduced risk of developing islet autoimmunity, especially of developing multiple 
autoantibodies, which are a strong risk factor for future disease. There was no association 
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• 
The Nutritional Intervention to Prevent (NIP) Type 1 Diabetes Study was a pilot study which 
evaluated the effect of DHA supplementation on inflammatory cytokine levels in infants at high 
genetic risk for T1D. Although DHA supplementation of infant diets was found to be safe, 
inflammatory cytokine production was not consistently reduced. However, the study was not 
designed to adjust dietary intake of omega-3 fatty acids to target specific AA/EPA ratios and it 
is unclear whether reaching specific ratios may translate into improved clinical outcomes. 
However, in a pilot study of T1D patients with established disease (>4 years duration) omega-3 fatty 
acids failed to improve glycemic control after 4 weeks of therapy. This may suggest that anti-
inflammatory therapies may need to be introduced early in the course of T1D in order to have an 
impact on prevention of beta-cell loss. In addition, considering that T1D is a chronic autoimmune and 
inflammatory disease, anti-inflammatory strategies may need to be administered over a prolonged 
period of time in order to observe a clinical benefit. 
The anti-inflammatory properties of vitamin D are well established. In particular, it has been shown 
that Vitamin D prevents both insulitis and type 1 diabetes mellitus in mouse models of T1D and 
retrospective studies have shown apparent beneficial effects of vitamin D supplementation in early life 
on successive lifetime risk of T1DM. Vitamin D has been linked to several immunomodulatory effects. 
Studies in new onset T1D have shown that treatment with cholecalciferol (vitamin D3) results in both 
an increase in the percentage and the suppressive capacity of regulatory T-cells. A recent study in 
adults with vitamin D deficiency showed that daily treatment with 4,000 IU of vitamin D3 significantly 
reduced CD4+ T-cell activation compared to treatment with 400 units of vitamin D3. However, the role 
of vitamin D in preservation of beta-cell function in new onset T1D is controversial. Two studies 
showed no significant effect of therapy with calcitriol (1,25(OH)2D3)  whereas one study using 2,000 
IU of cholecalciferol daily showed a higher stimulated C-peptide at 18 months compared to placebo.  
Further studies evaluating the role of vitamin D supplementation in prevention of beta-cell loss at T1D 
onset are warranted.  
Considering the safety profile and anti-inflammatory properties of high dose omega-3 fatty acids 
combined with Vitamin D supplementation makes this therapy a possible candidate for T1D 
intervention trials. 
 
4. STUDY DESIGN 
4.1. 
Overview 
Based on the findings presented in this document, we propose to test the safety and efficacy of a 
regimen that combines Omega-3 Fatty Acids and Vitamin D in a design that considers timing and 
duration of administration in relation to their effects and predicted synergies. These agents may 
promote sustained immune regulation, reduce inflammation, and provide support for the residual beta 
cell mass. This integrated therapeutic regimen addresses major pathogenic mechanisms in T1D and 
thus represents a rational and well supported approach to preserve insulin secretion in T1D. This 
approach could halt the disease progress, preserve β-cell function and hopefully reduce dose of insulin 
required to manage T1D. Even partial beta cell function can facilitate disease management, reduce the 
likelihood of severe hypoglycemia and is associated with reduced risk of chronic complications. We 
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established T1D and residual stimulated C-peptide secretion will be safe and may preserve insulin 
secretion; 
The safety will be measured by the AE profile. In addition, potential decreases in hypoglycemic events 
will be evaluated as this may be expected to be one of the clinical indications where combination 
therapy is likely to exert a clinical benefit. Other measures include the ability of combined supplement 
therapy to increase insulin production as assessed by serum C-peptide measurements, changes in 
exogenous insulin dose, and measurements of glycemic control.  
The study endpoints vary between the two arms and include safety, tolerability, and efficacy.  Safety 
endpoints include AEs over one year of treatment and an additional one year of follow-up (total two 
years). Efficacy endpoints include the ability of combining supplement products to produce insulin, 
assessed by serum C-peptide measurements following a Mixed Meal Tolerance Test (MMTT), changes 
in recorded exogenous insulin dose and other measures of glycemic control. 
4.2. 
Objectives 
4.2.1. Primary Objective(s) 
• 
Stimulated (90 minute sample of a MMTT) C-peptide at the 1 year visit greater or equal to 
baseline level. 
4.2.2. Secondary Objective(s) 
• 
Stimulated C-peptide area under the curve (AUC) during a 4-hour MMTT greater or equal to 
baseline level, at the 1 year visit. 
• 
Reduction in HbA1c at the one year visit compared to baseline. 
• 
Reduction in insulin requirement at the 1 year visit compared to baseline. 
• 
Incidence of adverse events (AE) comparable to general diabetes population.  
 
4.3. 
Investigational Plan 
This is a randomized, open-label, parallel-group, pilot phase I/IIa clinical study. Approximately 56 
subjects will be randomly assigned to receive either Omega-3 Fatty Acids and Vitamin D combination 
or Vitamin D alone. All subjects will be given Vitamin D supplement. Patients to be included in this study 
are those diagnosed with new onset T1D (diagnosed within the last 180 days) or with established T1D 
(more than 6 months). 
Participation for each subject will last approximately 2 years. Potential participants will have a screening 
visit to review their overall health, measure diabetes T1D-associated autoantibodies and perform a 
MMTT, which will also serve as the baseline MMTT.  Eligible subjects will be invited to participate in 
the intervention trial for one year, and then followed for a second year to monitor safety and whether 
any potential therapeutic benefit, whether metabolic, endocrine, and/or immunological, persists after 
the therapy has been stopped. 
We will evaluate beta cell function by examining C-peptide responses during a MMTT. 
 

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4.4. 
Summary of Inclusion and Exclusion Criteria 
This study can fulfill its objectives only if appropriate patients are enrolled. The following eligibility 
criteria are designed to select patients for whom protocol treatment is considered appropriate. All 
relevant medical and non-medical conditions will be taken into consideration when deciding whether 
this protocol is suitable for a particular participant.  
We will enroll T1D patients of both genders, 6-65 years -old, to examine the Omega-3 Fatty Acids and 
Vitamin D supplements covering as much of the age spectrum of the disease in both established as 
well as new onset T1D patients. There will be no restriction on race and ethnicity. However, T1D is 
more prevalent in populations of European descent, and enrollment will likely reflect disease prevalence 
in the Miami area, where the population is diverse and enriched in ethnicities from Latin America. 
Subjects who fulfill the following eligibility criteria will be recruited regardless of their race or gender. 
 
4.4.1. Inclusion Criteria 
Patients must meet all of the following criteria to be eligible to participate in this study: 
1. Subjects or their parents if under 18 years old must be able to understand and provide 
informed consent. 
2. Males and females, 6-65 years of age. 
3. For new onset T1D subjects, ≤180 days from T1D diagnosis at the time of randomization with 
a MMTT stimulated C-peptide peak level ≥0.2 ng/ml prior to randomization. 
4. For established T1D subjects, >180 days and ≤10 years of T1D duration at the time of 
randomization and MMTT stimulated C-peptide peak level ≥0.2 ng/ml prior to randomization.  
5. Affected by T1D, according to ADA standard criteria, and confirmed by positivity of at least 
one T1D-associated autoantibody, to GAD65, IA-2, ZnT8, or insulin autoantibodies (if patient 
has been treated with insulin for less than 2 weeks).  
6. Female subjects of childbearing potential must have a negative pregnancy test upon study 
entry.  
7. Adequate venous access to support study required blood draws. 
8. T1D must be treated with insulin (except if participant is in Honeymoon period/phase).  
 
 
4.4.2. Exclusion Criteria 
Potential participants must not meet any of the following exclusion criteria: 
1. Inability or unwillingness of a participant or their parents to give written informed consent or 
comply with study protocol. 
2. BMI>30 Kg/m2. 

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3. Contra-indications to Omega-3 Fatty Acids and/or Vitamin-D (e.g., knowledge of 
hypersensitivity to drugs or its excipients, allergies with fish or shellfish etc.). 
4. Uncompensated heart failure, fluid overload, myocardial infarction or liver disease or severe 
impairment of a vital organ within the last 6 weeks before enrollment. 
5. Any sign or diagnosis of significant chronic active infection (e.g., hepatitis, tuberculosis, EBV, 
or CMV), or screening laboratory evidence consistent with a significant chronic active infection 
(such as positive for HIV, IGRA test for TB, or hepatitis B-C).  
6. Ongoing acute infections, e.g., acute respiratory tract urinary tract, or gastrointestinal tract 
infections. 
7. Subjects on weight altering medications, such as Orlistat. 
8. Subjects with eating disorders 
9. Ongoing or anticipated use of diabetes medications other than insulin. 
10. Current or ongoing use of non-insulin pharmaceuticals that affect glycemic control within prior 
7 days of screening. 
11. People who chronically take drugs that affect bleeding time, such as anticoagulants (“blood 
thinners”) or nonsteroidal anti-inflammatory drugs (NSAIDs), will not qualify to enroll in the 
study.  
12. Recent recipient of any licensed or investigational live attenuated vaccine(s) within 6 weeks of 
randomization. 
13. Use of investigational drugs within 4 months of participation. 
14. Concomitant therapy with immunosuppressive drugs, immunomodulators, or cytotoxic agents, 
or previous therapy less than 3 months from randomization. 
15. History or diagnosis of malignancy.  
16. History of gastroparesis or other severe gastrointestinal disease. 
17. History or diagnosis of malignancy with the exception of a history of localized basal or 
squamouos cell carcinoma. There is conflicting evidence about whether omega-3 fatty acids 
found in seafood and fish oil might increase the risk of prostate cancer. Until additional 
research on the association of omega-3 consumption and prostate cancer risk is conducted, 
subjects with family history of prostate cancer in a first-degree relative will be excluded from 
the study.  
18. Presence of an allograft. 
19. AST, ALT or Alkaline Phosphatase >2 times upper limit of normal or total bilirubin >1.5 times 
upper limit of normal. 
20. History of a mental illness deemed to be clinically unstable or any situation that, in the opinion 
of the investigator, would interfere with the participant’s ability to comply with study 
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21. History of illicit drug or alcohol abuse. 
22. Pregnancy or ongoing breastfeeding for women; unwillingness or inability of both females and 
males of childbearing age to use a reliable and effective form of contraception, for the entire 
duration of the study. 
23. Past or current medical problems, or findings from physical examination, or laboratory testing, 
that are not listed above which, in the opinion of the investigator, may pose additional risks from 
participation in the study, may interfere with the participant’s ability to comply with study 
requirements or that may impact the quality or interpretation of the data obtained.  
24. All patients who have coagulation, bleeding, or blood disorders will be excluded due to the 
effect of high dose of Omega 3 Fatty Acids on coagulation and bleeding process.   
 
 
 
4.5. 
Informed Consent 
The process of assuring that individuals are making an informed decision about participating in this 
study includes both verbal and written communication.  The consent form will be reviewed with 
participants and the participant will be given time to review the written consent form and ask questions.  
The participant will be given a copy of their signed consent forms.  
Participants will also be informed about the common adverse effects of study drugs and it will be 
recommended that they stop eating once they feel satiety to decrease the risk of nausea and vomiting 
during the treatment period. Study subjects will receive standard medical care from their 
endocrinologist, but the study team will closely monitor the patients to assess adverse events. At these 
phone or email contacts and at the study visits, the frequency of hypoglycemia will be documented. 
Hypoglycemia will be assessed following consensus definition (Agiostratidou, 2017). Three levels of 
hypoglycemia will be recorded. Level 1: glucose < 70 mg/dl (3.9 mmol/l) and glucose ≥ 54 mg/dl (3.0 
mmol/l). Level 2: glucose < 54 mg/dl (3.0 mmol/l) and Level 3: a severe event characterized by altered 
mental and/or physical status requiring assistance. 
 
4.6. 
Description of Treatment Groups 
This open label protocol will enroll a total of 56 participants who will be randomly assigned (1:1) to the 
following groups: 
• 
28 participants will be assigned to receive Omega-3 Fatty Acids and Vitamin D combination 
following randomization, and then will be followed for an additional year.  
• 
28 participants will receive only Vitamin D.  
 
4.7. 
Treatment Assignment  

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Eligible participants who have provided written informed consent will be randomized to one of the 
treatment groups.  The participant, the clinical investigator and clinical personnel will be known to the 
treatment assignment.   
4.8. 
Study Assessments 
During the course of the study, participants will frequently undergo assessments of their insulin 
production, immunologic status, overall health and well-being and diabetes care (see schedule of 
assessments in Appendix 1).  The participant’s insulin production will be assessed by C-peptide 
measurements during mixed meal glucose tolerance tests (MMTT) conducted as per the study 
schedule. The MMTT will consist of a standard liquid meal (Boost Plus®, Nestlé HealthScience) at a 
dose of 6 mL/Kg to a maximum of 360 mL. The participant’s diabetes control will be evaluated by 
measuring glycosylated hemoglobin (HbA1c) every four months. 
Remaining samples will be stored in the Diabetes Research Institute for future analysis.  Samples will 
be stored only with the participant’s permission.  Participants who decline consent for storage of 
remaining samples will still be eligible to participate in this study. 
 
 
4.9. 
Post-treatment Follow-up 
All subjects will be treated with supplement combination or vitamin D alone over a one year period. 
Subjects will also be asked to undergo additional follow-up post-treatment for up to one year with a visit 
every 4 months.  
 
5. PATIENT MANAGEMENT 
 
5.1. 
Recruitment and Screening 
Information about the clinical trial will be made available to potential participants as follows: a) flyers at 
the Diabetes Research Institute and the Mailman Center for Child Development, University of Miami, 
b) on-line at www.Clinicaltrials.gov and Diabetes Research Institute Foundation webiste 
www.diabetesresearch.org, c) social media: Diabetes Research Institute Foundation Facebook and 
twitter accounts and YouTube chanel, d) by press release(s) from the Diabetes Research Institute, 
University of Miami.  
We will also provide information about the clinical trial to physicians within the Adult and Pediatric 
Divisions of Endocrinology, Diabetes and Metabolism at the University of Miami. We expect this 
approach to result in widespread awareness of this clinical trial and lead to successful recruitment 
numbers.   
 
After informed consent is obtained, subjects will undergo assessments to determine if they meet 
eligibility criteria.    Documentation of the subjects understanding of the risks and benefits of the study 
will also be discussed during the informed consent process.  

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5.2. 
Randomization 
Individuals complying with all inclusion and exclusion criteria and consenting to study participation will 
be randomized at the clinical site. A blinded statistician will create color and number coded envelopes 
using a stratified blocked randomization (new diagnosis vs established, child vs adult). Each envelope 
will contain a card with the corresponding treatment group assignment. Envelopes will be kept in a file 
cabinet in a secured room at the Diabetes Research Institute. The study PI will perform the 
randomization of  all newly enrolled subjects in a 1:1 ratio to each of the two arms by selecting the 
corresponding color and number coded envelope to ensure these factors are equally distributed across 
the two arms of the intervention.  
For each subject, the Subject Identificaton Number (SIN), treatment and participant’s name will be 
entered into the study log.   
 
5.3. 
Intensive Diabetes Management 
During the study period, all participants will receive “intensive” management of their diabetes.  The goal 
of the treatment will be to keep the HbA1c levels within the currently recommended American Diabetes 
Association target range in the absence of level 3 hypoglycemia or diabetic ketoacidosis.  The primary 
responsibility for diabetes management will be the treating or referring diabetes care provider, but the 
research study team will provide close additional support through regular interaction at study visits.  
Subjects will not be permitted to use non-insulin pharmaceuticals for glycemic control. 
Glucose levels should be checked frequently using either a continuous glucose monitor (CGM) or 
manually via study participant’s glucometer. In addition, subject will complete a 7-day Blood Sugar 
record and Hypoglycemia log at the screening visit (+/- 7 days) and at each study visit (+/- 7 days) 
starting at visit 2. Subjects using a CGM can use glucose values from the CGM to complete the 7-day 
blood sugar record. If subjects are using an insulin pump, pump data will be downloaded to document 
insulin use. Subjects on multiple daily injections will record their insulin use on the 7-day blood glucose 
record unless they are using a smart insulin pen with capability to record insulin dosing in which case 
data will be downloaded from the device and serve as the insulin dosing documentation. If patient is 
using a CGM and pump, downloaded data from pump and CGM will be used instead of the 7-day blood 
sugar log. Data obtained from participants’ diabetes management devices  and blood sugar log will 
serve as source documentation of glucose monitoring and insulin use.The study team will be evaluating 
the HbA1c data and provide additional guidance to the participants as needed to bring diabetes control 
within goals.  Any episodes of level 3 hypoglycemia will prompt review by the study physician and site 
medical monitor with recommendations, if any, for changes in diabetes management conveyed to the 
participants in conjunction with the principal investigator(s).  
 
5.4. 
Administration of Study Drugs 
Due to current status of COVID-19 pandemic, clinical research activites have been affected and are 
being conducted as recommended by institutional guidelines to minimize potential exposure to SARS-
CoV-2 for both research subjects and clinical research study team. Study investigational product(s) 

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may be mailed to participants as determined by PI. For international patients, they may be able to obtain 
over the counter vitamin D3 and/or omega 3 fish oil locally (in their country of residence) if the 
investigational product cannot be mailed overseas (at the discretion of the PI). 
In addition, the study team will try to conduct as much as possible “remote” study visits (e.g telephone 
communication, video conferencing, obtaining study test results from Quest Diagnostics Lab) to obtain 
required SOE information. 
5.4.1. Omega-3 Fatty Acids  
Omega-3 fatty acids (omega-3s) are a group of polyunsaturated fatty acids that are important for a 
number of functions in the body. Some types of omega-3s are found in foods such as fatty fish and 
shellfish. Another type is found in some vegetable oils. Omega-3s are also available as dietary 
supplements. This fact sheet provides basic information about omega-3s—with a focus on dietary 
supplements, summarizes scientific research on effectiveness and safety, and suggests sources for 
additional information. 
Administration of high-dose omega 3 fatty acids (18g of marine lipid concentrate) for 6 weeks to 
healthy volunteers reduced the AA/EPA ratio from 20.9±2.2 to 2.4±0.2 (nearly 90%) and resulted in a 
decrease in IL-1β, IL-1α, and TNF and was well tolerated [1].  A recent study in patients with stable 
coronary artery disease showed that long-term treatment (1 year) with high-dose omega 3 fatty acids 
(3360 mg EPA and DHA daily) was associated with an increase in specialized pro-resolving lipid 
mediators and enhancement of macrophage phagocytosis of blood clots [2]. These data suggest that 
high-doses of omega 3 fatty acids are required to mitigate cytokine responses and may be critical to 
generate resolvins and protectins, which have been postulated as the molecular basis for the anti-
inflammatory properties of omega-3 fatty acids.  
 
Further, high-doses of omega 3 have been associated with improved clinical outcomes. In children 
with ADHD, a study evaluating an 8-week treatment course with high-dose omega 3 fatty acids (250-
325 mg EPA and DHA/Kg/day) aimed at achieving an AA/EPA ratio between 1.5 and 3.0 resulted in 
significant improvements in behavior with the AA/EPA ratio correlating with the global severity of 
illness scores [3, 4].  In addition, two separate case reports in children with new onset type 1 diabetes 
have documented treatment with high-dose of omega 3 fatty acids (55-110 mg EPA and DHA/Kg/day) 
for over 1 year and showed improvement in glycemic control and preservation of C-peptide [5, 6]. 
Notably, these high doses of omega 3 fatty acids were well tolerated.  
 
To appropriately gauge the dose of omega 3 fatty acids we are proposing the use of the AA/EPA ratio 
to guide dosing. As noted above, AA/EPA ratios in the 1.5 to 3.0 range have been studied in healthy 
volunteers and in children with ADHD have shown an association with clinical outcomes and noted to 
be safe. Further, such low ratios can be achieved by diet alone as documented in the Japanese 
population where the average fish consumption has been reported to be one serving of 85 g (3 oz; 
900 mg EPA and DHA) per day and 90% of individuals eat fish at least once a week [7].  
 

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In view of the chronic nature of the immunopathogenesis of type 1 diabetes it is then not unexpected 
that therapeutic interventions started at disease onset to preserve beta cell function have usually 
been administered for a duration of 6 month to 2 years [8-10]. It is important, however, to highlight 
that one of the main factors responsible for determining the duration of therapy is the safety profile of 
the proposed intervention.   
 
Data from TrialNet intervention trials has allowed to study the decline in C-peptide 2 years following 
type 1 diabetes onset which shows a biphasic profile with a steeper decline during the first year [11] 
suggesting that processes involved with beta-cell loss may be more aggressive during the first year of 
diagnosis. Thus, therapies aimed to preserve beta-cell function in new onset disease may likely need 
to be administered over a prolonged period of time and for at least 1 year following disease onset.  
 
In established disease, duration of therapy is also dependent on the mechanism of the intervention 
and the safety profile. Since inflammation affecting glycemic control may be a chronic state in patients 
with type 1 diabetes and in view of the reportedly safety profile of high-dose omega 3 fatty acids, we 
have proposed treating patients with established type 1 diabetes for a duration of 1 year. 
 
5.4.1.1. 
Ultra-refined omega-3 EPA/DHA concentrate (ZoneLabs® OmegaRx®2 Fish Oil) 
Administration 
Participants will receive ultra-refined omega-3 EPA/DHA concentrate (ZoneLabs® OmegaRx®2 Fish 
Oil – Liquid; containing 3375 mg EPA+DHA [2250 mg EPA and 1125 mg DHA] in 5 mL).  
Research subjects will be started at a dose of 150 mg of EPA+DHA per Kg body weight.  
For example: 
- 
The dose for a 22.5 Kg child will be: 
150 mg x 22.5 Kg = 3375 mg of EPA+DHA fish oil corresponding to 5 mL of Omega Rx ®2 
Fish Oil Liquid. 
- 
The dose for a 30 Kg child will be:  
150 mg x 30 Kg = 4500 mg of EPA+DHA fish oil corresponding to 6.7 mL of Omega Rx ®2 
Fish Oil Liquid.  
Participants who are unable to tolerate the liquid formulation will be switched to the OmegaRx®2 
capsule formulation  at the same dose as described above. 
It is 150 mg of combined EPA and DHA per kg body weight to be given either as a single or divided 
into smaller doses through the day. 
 
5.4.1.2. 
Modification or Discontinuation of omega-3 EPA/DHA concentrate  
Dose will be adjusted to maintain a target AA/EPA ratio of 1.5-3.0 as follows: 
- 
If AA/EPA ratio is ≥ 7, double the dose and recheck ratio in one month  
- 
If AA/EPA ratio is 4-6, increase dose by 50% and recheck ratio in one month 
- 
If AA/EPA ratio is at target, maintain dose as prescribed s 
- 
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AA/EPA ratios will be monitored monthly until achieving target levels and as per protocol or as clinically 
indicated thereafter. 
 
 
5.4.1.3. 
Discontinuation of omega-3 EPA/DHA concentrate  
Subjects would be continuously evaluated for any untoward effects of Omega-3 fatty acid supplements 
or negative side effects. Typically side effects consist of minor gastrointestinal symptoms, such as 
belching, indigestion, or diarrhea, stomach pain or discomfort, burping, heartburn, vomiting, 
constipation, nausea, and change in the sense of taste. Joint pain may be a side effect as well.  
Patients with coagulation, bleeding, or blood disorders will be excluded. We will monitor clotting times 
of enrolled subjects by measuring the prothrombin time (PT/PT-INR) during all subject visits (except for 
visit 1), in addition to CBC with differentials (see schedule of assessment). 
 
Symptomatic management of the side effects will be evaluated by the study physician and should the 
investigator believes that the study treatment is no longer in the best interest of the participant, subjects 
would be discontinued from the treatment. Similarly, if subject begins taking medications that affect 
clotting chronically, subjects would be discontinued and appropriately noted in the study record. 
Participants who prematurely discontinue study treatment will remain in the study and undergo all 
efficacy and safety assessments. 
 
5.4.2. Vitamin D  
Vitamin D is a nutrient found in some foods that is needed for health and to maintain strong bones. It 
does so by helping the body absorb calcium (one of bone's main building blocks) from food and 
supplements. People who get too little vitamin D may develop soft, thin, and brittle bones, a condition 
known as rickets in children and osteomalacia in adults. 
Vitamin D is important to the body in many other ways as well. Muscles need it to move, for example, 
nerves need it to carry messages between the brain and every body part, and the immune system 
needs vitamin D to fight off invading bacteria and viruses. Together with calcium, vitamin D also helps 
protect older adults from osteoporosis. Vitamin D is found in cells throughout the body.  
 
5.4.2.1. 
Vitamin D Administration 
Vitamin D supplementation 
1. For levels of Vitamin D 25OH of 30 -39 ng/ml:   
a. age ≤10 years give 500 IU daily  
b. age >10 years give 1000 IU daily 
2. For levels of Vitamin D 25OH of 20 – 29 ng/ml give 50,000 IU weekly for 2 months and then: 
a. age ≤10 years give 1000 IU daily 
b. age >10 years give 2000 IU daily 
3. For levels of Vitamin D 25OH of < 20 ng/ml give 50,000 IU weekly for 4 months and then: 
a. age ≤10 years give 1000 IU daily   

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b. age >10 years 2000 IU daily 
  
Repeat vitamin D 25OH levels 1 week after the last 50,000 IU dose: 
• 
For children whose levels are not above 30 ng/ml on repeat testing, repeat treatment as 
above. 
• 
For children whose levels are 30 -39 ng/ml on repeat testing: 
o if age ≤10 years increase vitamin D supplementation to 1000 IU daily  
o if age is >10 years give 2000 IU daily 
 
Thereafter, monitor vitamin D 25OH levels every 4 months (or more frequently at the discretion of the 
investigator) and adjust Vitamin D treatment to maintain levels > 40 ng/ml. 
 
Study participants with or without a history of vitamin D deficiency who are on vitamin D replacement 
will continue their current dose if 25OH vitamin D levels checked at screening are in line with study 
targets. This may defer from the doses indicated above which apply to participants who are not on 
vitamin D replacement.  Doses will then be adjusted at the discretion of the Principal Investigator in 
order to maintain 25OH vitamin D levels >40 ng/mL.  
 
The Dietary Reference Intake Committee to Review Dietary Reference Intakes for Vitamin D and 
Calcium from Institutie of Medicine, recommends following upper intake levels for Vitamin D (see 
page 403-477)(https://www.ncbi.nlm.nih.gov/books/NBK56070/pdf/Bookshelf_NBK56070.pdf ) 
Table Representing Tolerable Upper Intake Levels (ULs) for Vitamin D  
Age 
Male 
Female 
Pregnancy 
Lactation 
0–6 months 
1,000 IU 
(25 mcg) 
1,000 IU 
(25 mcg) 
  
  
7–12 months 
1,500 IU 
(38 mcg) 
1,500 IU 
(38 mcg) 
  
  
1–3 years 
2,500 IU 
(63 mcg) 
2,500 IU 
(63 mcg) 
  
  
4–8 years 
3,000 IU 
(75 mcg) 
3,000 IU 
(75 mcg) 
  
  
≥9 years 
4,000 IU 
(100 mcg) 
4,000 IU 
(100 mcg) 
4,000 IU 
(100 mcg) 
4,000 IU 
(100 mcg) 
 
Microcrystalline cellulose is the only excipient in Biotech Pharmacal Vitamin D3 capsule with following 
quantities: 
 
• 
D3-5000 IU capsule contains 89 mg of microcrystalline cellulose 
• 
D3-1000 IU capsule contains 92.8 mg of microcrystalline cellulose 
• 
D3 50,000 IU capsule contains 115 mg of microcrystalline cellulose 
 

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Type of microcrystalline cellulose is Vivapur Type 101, it is pure microcrystalline cellulose that meets 
the requirements for Ph. Eur, NF, and JP monographs for Microcrystalline Cellulose and it complies 
with E460 (i) monograph and FCC as well.  
 
5.4.2.2. 
Modification or Discontinuation of Vitamin D  
 
Vitamin D is likely safe when taken by mouth in recommended amounts. Most people do not commonly 
experience side effects with vitamin D, unless too much is taken. Some side effects of excess vitamin 
D include weakness, fatigue, sleepiness, headache, loss of appetite, dry mouth, metallic taste, nausea, 
and vomiting. 
Taking vitamin D for long periods of time in doses higher than 4000 units daily is possibly unsafe and 
may cause excessively high levels of calcium in the blood. However, much higher doses are often 
needed for the short-term treatment of vitamin D deficiency.  
Vitamin D in very high doses can make the intestines absorb too much calcium which can cause high 
levels of calcium in the blood (hypercalcemia). This can cause side effects including nausea, vomiting, 
constipation, poor appetite, weakness, weight loss, confusion and disorientation.  Hypercalcemia can 
also lead to kidney stones, damage to the kidneys, and cause calcium deposits in heart and lungs. We 
will monitor blood levels of vitamin D to minimize the risk of these side-effects. 
 
5.4.2.3. 
Contraindications for Vitamin D Supplement  
There are conditions where Vitamin D treatment require special precautions and these subjects will not 
be included in this study:  
Kidney disease: Vitamin D may increase calcium levels and increase the risk of arteriosclerosis” in 
people with serious kidney disease. This must be balanced with the need to prevent renal 
osteodystrophy, a bone disease that occurs when the kidneys fail to maintain the proper levels of 
calcium and phosphorus in the blood.  
Sarcoidosis: Vitamin D may increase calcium levels in people with sarcoidosis. This could lead to 
kidney stones and other problems..  
Histoplasmosis: Vitamin D may increase calcium levels in people with histoplasmosis. This could lead 
to kidney stones and other problems.. 
Over-active parathyroid gland (hyperparathyroidism): Vitamin D may increase calcium levels in 
people with hyperparathyroidism.. 
Lymphoma: Vitamin D may increase calcium levels in people with lymphoma. This could lead to kidney 
stones and other problems.. 
Tuberculosis: Vitamin D might increase calcium levels in people with tuberculosis. This might result in 
complications such as kidney stones.  
 
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Patients will be closely monitored to assess compliance of study drug administration. Each participant 
will return the empty bottles to the Coordinator to assess compliance, which in turn will be returned to 
the Pharmacy for appropriate on-site destruction and for documentation purposes. 
 
5.4.4. Prior and Concomitant Treatments 
The use of concomitant medications will be assessed at each study visit and recorded on an appropriate 
source document and CRF. Participants are allowed to use preparations of insulin as advised by the 
investigator or the referring physician. 
The following medications are prohibited during participation in this study: 
• 
Agents that influence insulin sensitivity or secretion (e.g. pramlintide, sulfonylureas, metformin, 
diphenylhydantoin, thiazide, or other potassium-depleting diuretics, beta-adrenergic blockers, 
niacin etc). 
If participants receive, or if the investigator believes that participants must receive, any of the above 
medications, the case must be immediately discussed with the medical monitor. The use of prohibited 
medications must be documented on the source document and CRF, and a protocol deviation must be 
requested. A decision regarding continuation of the participant in the trial will be made by the study PI, 
and the medical monitor. 
 
5.4.5. Removal of Subjects from Therapy or Subject Withdrawal or Early Termination 
• 
Withdrawal of consent - Participants who withdraw consent will be asked to complete all the 
assessments listed for the end of treatment follow-up  
• 
Failure to return - Participants who do not return for visits and who do not respond to repeated 
attempts by the site staff to have them return will be considered lost to follow-up.   
• 
Investigator judgment - A severe or serious AE occurs, which, based on the medical judgment 
of the investigator, prevents completion of participation in the study. 
 
5.4.6. Discontinuation of study drug(s) in an individual patient 
The dosing and administration of investigational medication according to study specification will be 
discontinued for an individual participant if any of the following criteria is met: 
• 
Subjects who have 3 severe hypoglycemic reactions on separate days (requiring assistance 
from another individual)  
• 
Subjects who have nausea or vomiting that precludes adherence to diet will discontinue the 
use of the study drug. 
• 
Subjects who lose more than 10 kg in weight from baseline. 

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• 
Any grade 3 or higher AE occurs that, based on the medical judgment of the investigator, 
the medical monitor, and the clinical trial physician, prevents completion of the course of 
study treatment. 
• 
Any unexpected, treatment-related SAE resulting in permanent treatment discontinuation 
and not related to glycemic events. 
• 
The investigator determines that it is in the participant’s best interest to discontinue 
treatment. 
• 
The participant, or participant’s legal representative, requests that treatment be halted. 
• 
The participant becomes pregnant. 
Further care will be provided according to the judgment and practice of the investigator. The participant 
will be asked to remain in the study and participate in follow-up. If study treatment is discontinued, the 
medical monitor will be notified. Participants who prematurely terminate from the study will not be 
replaced. 
 
5.4.7. Re-Entry into Study Treatment 
In some circumstances, a participant may temporarily discontinue the study medication and/or not 
return to the study clinic for follow-up visits. If the participant decides to return for study treatment and/or 
follow-up assessments at a later date, he or she will be allowed and encouraged to do so. 
The study will be conducted according to the intent-to-treat principle.  This means that once randomized 
into the study, a participant will be expected to undergo all scheduled follow-up assessments and will 
remain in the assigned treatment group for purposes of statistical analysis regardless of the actual 
course of treatment administered.  Withdrawal from treatment does not automatically entail withdrawal 
from the study.  Withdrawal from the study will only occur if the participant dies or withdraws consent.  
Subjects who withdraw consent are classified as inactive but may again become active upon re-entry 
into the study, if they so choose.  
 
5.4.8. Risk to the Subjects  
The risks of these over the counter drugs is summarize below: 
• 
Omega-3 Fatty Acids may cause gastrointestinal side- effects including stomach pain or 
discomfort, burping, heartburn, vomiting, constipation, diarrhea, nausea, and change in the 
sense of taste. Joint pain may be a side effect as well.  
• 
Vitamin D in very high doses can make the intestines absorb too much calcium which can cause 
high levels of calcium in the blood (hypercalcemia). This can cause side effects including 
nausea, vomiting, constipation, poor appetite, weakness, weight loss, confusion and 
disorientation.  Hypercalcemia can also lead to kidney stones, damage to the kidneys, and 
cause calcium deposits in heart and lungs. We will monitor blood levels to minimize these risks. 

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Allergic reaction: As with any drug, it is possible that subjects could experience an allergic reaction to 
OMEGA-3, or Vitamin D. Such allergic reactions include: itching, skin rash, sudden drop in blood 
pressure, loss of consciousness and/or associated with seizures, including the possibility of death. 
Blood draw risks: Drawing blood may cause temporary pain from the needle stick, bruising or swelling 
at the site, and rarely, infection or fainting.  
Hypoglycemia:  These medications could lower blood sugar too much (hypoglycemia). This could make 
subjects feel tired, dizzy, sweaty, and/or nauseated. Also, it could cause heart to feel as if it is racing. 
There may also be other effects. Untreated hypoglycemia could cause convulsions, loss of 
consciousness, and can lead to death. The clinical research team will provide added training and 
education to subjects to better manage hypoglycemia and other diabetes related complications.  
Unknown Risks: The experimental drug may have side effects that no one knows about yet. The 
researchers will let you know if they learn anything that might make you change your mind about 
participating in the study. 
Females of child bearing potential must have a negative pregnancy test prior to enrolling in the study 
and will be required to use at least two FDA approved birth control during the study.  At every study 
visit the sexual activity of female participants of reproductive age will be re-assessed.  Pregnancy 
testing will be conducted in females in  reproductive age at 4, 8,12 and 16 months study visit. If a 
subject who was previously sexually inactive becomes sexually active, she will be counseled about the 
need to use a reliable forms of birth control.    Subjects will be requested to avoid pregnancy for 6 
months following the last study administration and instructed to use birth control. If subject becomes 
pregnant, study intervention will be discontinued and subject withdrawn from study. 
 
6. STUDY ASSESSMENTS 
Due to current status of COVID-19 pandemic, clinical research activites have been affected and are 
being conducted as recommended by institutional guidelines to minimize potential exposure to SARS-
CoV-2 for both research subjects and clinical research study team. Study investigational product(s) 
may be mailed to participants as determined by PI. For international patients, they may be able to obtain 
over the counter vitamin D3 and/or omega 3 fish oil locally (in their country of residence) if the 
investigational product cannot be mailed overseas (at the discretion of the PI). 
In addition, the study team will try to conduct as much as possible “remote” study visits (e.g telephone 
communication, video conferencing, obtaining study test results from Quest Diagnostics Lab) to obtain 
required SOE information. 
 
6.1. 
General Assessments 
Patients visits will occur for all subjects at baseline (day 0), and every four months thereafter (month 4, 
8, and 12) and consist of treatment adherence and outcome assessments.  Subjects will be provided 
with supply of study drugs to take home and administer orally.  Subjects will be educated and trained 
for the drug administration and study personnel will evaluate drug returns and treatment adherence. 
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one year, all subjects will continue with assessments up to an additional one year (months 16, 20, and 
24). 
The time each assessment will be performed relative to dosing is shown in the Schedule of Study 
Activities (Appendix 1). 
 
General assessments include:  
• 
Medical history: To determine if there are any clinically significant diseases or medical 
procedures other than the disease under study.  
• 
Physical examination: Includes any body system with clinical signs or reported symptoms of 
adverse events and weight loss.  
• 
Concomitant medications 
• 
Adverse events 
 
6.2. 
Enrollment 
The research study will be explained in lay terms to each potential research participant. The potential 
participants will sign an informed consent before undergoing any study procedures.  The consent form 
will be reviewed with participants and the participants will be given time to review the written consent 
form and ask questions.  The participant will be given a copy of their signed consent form.   
 
6.3. 
Screening (visit 0) and Baseline (visit 1) visits  
Patients will be asked to sign consent for screening to determine eligibility. Patients will receive a 
physical exam, medical history will be taken, and basic physical parameters will be recorded (height, 
body weight, heart rate, blood pressure). Blood will be drawn for laboratory assessments (see section 
6.5) for evaluation of inclusion and exclusion criteria.  
A patient that meets the inclusion/exclusion entry criteria will be randomized to one of the two treatment 
arms. Blood will be drawn for baseline visit (visit1) assessments. The patient will receive the first 
treatment dose within 48 hr from the baseline visit. 
 
6.4. 
Diabetes Management  
During the study period, all participants will receive “intensive” management of their diabetes.  The 
patients will continue to see their primary endocrinologist during the study period. All participants will 
be expected to take a sufficient number of daily insulin injections to meet the glycemic targets.  In 
general, the expectation is that all participants will receive at least three injections of insulin daily, 
including short- and long-acting insulin preparations, or will utilize continuous subcutaneous insulin 
infusion (CSII insulin pump).  
Glucose levels should be checked either via continuous glucose monitor or manually at least 4 times 
daily via the study participant’s glucometer. Records of glucose measurements will be kept as source 
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required to record the daily amount of insulin they have used at each study visit (+/- 7 days) using the 
7-day Blood Sugar and Hypoglycemia log provided.  Insulin pump data or smart insulin pen data will 
be used for documentation of insulin use in lieu of the 7-day blood sugar log for those participants using 
these devices.  
Upon review of these records, the investigator may make adjustments in the insulin regimen, refer a 
participant to a registered dietician or if necessary take other approaches that will help to maintain or 
improve a participant’s glucose control.  
 
6.5. 
Laboratory Assessments 
The following laboratory assessments will be performed during the study: 
• 
Chemistries (sodium, potassium, chloride, CO2, glucose, BUN, creatinine) 
• 
Liver function tests (ALT, AST, alkaline phosphatase, total protein, albumin, total and direct bilirubin) 
• 
Hematology (complete blood count with differential,  platelets, Prothrombin time [PT/PT-INR]) 
• 
Pregnancy test as appropriate 
• 
Autoantibodies (IA2, GAD65, ZnT8, insulin autoantibodies are induced by insulin injections and 
thus are not useful at this stage) 
• 
Samples for serology (i.e. Hepatitis B, Hepatitis C, EBV, CMV and Human Immunodeficiency Virus) 
and HbA1c 
• 
4-hours Mixed Meal Tolerance Test (MMTT) 
• 
Samples for 25OH vitamin D levels and AA/EPA ratio 
 
 
6.6. 
Mechanistic Outcome Assessments 
The DRI will perform immune assays to further understand mechanisms that may be underlying the 
Type 1 disease process and response to therapy.  For this purpose, samples will be obtained and 
stored for future analysis. 
Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles 
6.7. 
Metabolic Outcome Assessments 
Metabolic assessments will consist of: 
• 
Glucose records and reports of hypoglycemia  
• 
Insulin dose 
• 
HbA1c 
• 
Clarke Score 

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• 
Mixed meal tolerance test (MMTT)  
 
6.8. 
Visit Windows 
The initial study drug administration must begin within 180 days from the day of diagnosis.  All 
subsequent treatment visits and follow up visits in Appendix 1 must occur within the time limits specified 
below: 
 
The table below depicts summary of visit windows. 
 
Visit Windows 
Visit 0 
At least 2 weeks before visit 1  
Visit 1 
New onset group (randomization ≤180 days from  
T1D diagnosis) or established T1D (>180 days and 
<10 years from T1D diagnosis)  
Visit 2-4 (Days 120, 240, 360) 
± 14 days 
Follow-up between year 1 and 2 
Every four months  
 
± 14 days 
 
During the course of the study, participants who complete one year treatment will receive combination 
therapy of Omega-3 Fatty Acids and Vitamin D or Vitamin D alone. In addition, participants will 
frequently undergo assessments of their insulin production, immunological status, and overall health.  
The participant’s insulin production will be measured by a series of MMTTs conducted regularly during 
the study.  The participant’s diabetes control will be evaluated by measuring HbA1c every four months 
and clinical records including insulin types, doses, administration times and blood glucose records. 
 
6.9. 
Unscheduled visits 
Patients will be instructed to contact the study coordinator for any health concern, side effects 
potentially associated with the study, or other concern that may affect their participation and require 
an unscheduled visit. 
 
 
 
6.10. Sample retention 
Specimens for future mechanistic studies will be obtained throughout the study. Residual specimens 
may be used by the investigators for studies of immunologic mechanisms involved in T1D.   
Specimens collected in this trial may be used to reevaluate biologic responses as new research tools 
become available.  These  specimens will be encoded with specific study ID and stored at the Diabetes 
Research Institute of University of Miami.   

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6.11. Staggering Administration 
Please note that the enrollment of patients in this trial is staggered across individuals. Initially, a total 
of 6 subjects will be randomized in each of the following groups:   
 
Group 
Subject population  
I 
Fourteen (14) adults (18-65 years) of established T1D of more 
than 6 months duration (>180 days) and up to 10 years 
II 
Fourteen (14) adults (18-65 years) of new-onset T1D diagnosed 
within last 6 months (≤ 180 days) 
III 
Fourteen (14) children (6-17 years) of established T1D of more 
than 6 months duration (>180 days) and up to 10 years 
IV 
Fourteen (14) children (6-17 years) of new-onset T1D diagnosed 
within last 6 months (≤ 180 days) 
 
All 24 subjects will be monitored for acute and subacute adverse events for 90 days and a formal DSMB 
(Data and Safety Monitoring Board) will review the data and safety prior to treating additional subjects. 
This will limit the number of subjects who might be exposed to any unanticipated safety risk.  
 
6.12. Intensive T1D Management  
During the study period, all participants will receive “intensive” management of their diabetes.  The goal 
of the treatment will be to keep the HbA1c levels within the currently recommended American Diabetes 
Association age-specific target range in the absence of significant or level 3 hypoglycemia or diabetic 
ketoacidosis.  The primary responsibility for diabetes management will be the treating or referring 
diabetes care provider, but the research study team will provide close additional support through regular 
interaction at study visits.  Subjects will not be permitted to use non-insulin pharmaceuticals for glycemic 
control. Glucose levels should be checked frequently and records of the glucose levels communicated 
to the study team at study visits.   
 
6.13. Safety Monitoring  
The members of the Data Safety Monitoring Board (DSMB) are independent and serve in an individual 
capacity to provide their expertise and recommendations. They will review the 90 day data of the first 
6 subjects, and then periodically review and evaluate the accumulated study data for participant safety, 
study conduct and progress, and, when appropriate, efficacy, and make recommendations to study 
investigators concerning the continuation, modification, or termination of the trial. The DSMB members 
will comprise of experts in the field of T1D immunotherapy and will consider study-specific data as well 
as relevant background knowledge about the disease, test agents, and patient population under study.  
The DSMB is responsible for defining its own deliberative processes, including event triggers that would 
call for an unscheduled review, stopping guidelines, and voting procedures prior to initiating any data 
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6.14. Ad-hoc DSMB Reviews  
In addition to the pre-scheduled data reviews and planned safety monitoring, the DSMB may be called 
upon for ad hoc reviews. The DSMB will review any event that potentially impacts safety at the request 
of the study investigator.  In addition, the following events will trigger an ad-hoc comprehensive DSMB 
Safety Review:  
• 
Any death that occurs in the study.  
• 
The occurrence of a Grade 3 or higher related and unexpected SAE in three or more of the 
study participants who have received a study treatment. 
After review of the data, the DSMB will make recommendations regarding study conduct and/or 
continuation. 
 
6.15. Temporary Suspension of Enrollment/Drug(s) Dosing or Both  
A temporary halt in enrollment will be implemented if an ad-hoc DSMB safety review is required.  
Subjects already on investigational product will continue on therapy unless the ad-hoc DSMB review is 
completed and the DSMB determines that its findings warrant cessation of study drug(s).  Subjects in 
the screening phase of the study may continue to undergo screening procedures. 
 
6.16. Stopping rules 
As stated earlier, the progress of the study will be monitored by the DSMB which will review safety data 
and make recommendations regarding continuation, termination, or modification of the study.  Based 
on a 12 month enrollment period and an additional study period of 12 months, the DSMB will formally 
review the safety data after participant number 24 reaches day 90 of the study.  The number of subjects 
who discontinue study treatment will also be included in the reports prepared for the DSMB. 
In addition, safety data will be reviewed by the DSMB when an event occurs that is of sufficient concern 
to the medical monitor and/or study investigator(s) to warrant review, or when an event occurs that 
contributes to a stopping rule listed below.  
• 
Patient stopping rules: Subjects who have 3 severe hypoglycemic reactions (requiring 
assistance from another individual) on separate days or who have nausea or vomiting that 
precludes adherence to diet will discontinue the use of the study drug. Subjects who lose more 
than 10 kg will also discontinue study drug. Subjects who discontinue the drug treatment will 
not be replaced.  
• 
Trial stopping rules: Interim analysis will be conducted when recommended by the DSMB to 
review safety data, including metabolic, immunological assessments and adverse events. The 
DSMB may recommend early stopping if there is sufficient evidence to conclude that the 
investigational treatment is harmful based on interim analysis of safety endpoints. 
 

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6.17. Potential Benefits 
The risks of this study in detailed are presented in the informed consent form. This study will examine 
whether combination supplement therapy intervention will preserve beta cell function, but there is no 
guarantee that this will occur.  
This is a pilot feasibility trial with the prospect of direct benefit to the individual subjects for their 
participation in the study. The potential direct benefit to research subjects is that administration of 
specific combination supplement therapy or vitamin D alone could stabilize or improve ß-cell function, 
thus facilitating attainment of improved glycemic control. All subjects will benefit from being closely 
involved with an experienced team of diabetes experts, with the likely benefit that their glycemic control 
will be better than would be otherwise obtained. Thus, these potential benefits include the recognized 
benefits of being in a clinical study, including close monitoring and additional resources available to 
maintain tight glycemic control offered to all subjects, regardless of group assignment.  These additional 
resources include frequent in-person and other contact with study associate diabetes educator 
throughout the duration of the study.  Further, the intervention has the prospect of direct benefit to a 
given subject and is likely to yield general knowledge about T1D which is of importance for the 
understanding and amelioration of T1D. The study procedures, while greater than minimal risk, offers 
the possibility of benefit due to the close monitoring for all participants.   
If effective in preserving or increasing ß-cell function, combination therapy could be an important new 
treatment to change the course of T1D, thus benefiting the individual research participant and 
potentially society at large. Moreover, the proposed studies will produce new information about the 
immune system and its regulation, in the context of T1D and in relation to therapy with these over the 
counter agents that would be relevant to other autoimmune diseases. 
 
6.18. Pregnancy 
Pregnant and lactating women will not be included in the study.  Sexually active females must have a 
negative pregnancy test prior to enrolling in the study and will be required to use at least two FDA 
approved birth control during the study.  At every study visit the sexual activity of female participants of 
reproductive age will be re-assessed.  If a subject who was previously sexually inactive becomes 
sexually active, she will be counseled about the need to use a reliable forms of birth control.    Subjects 
will be requested to avoid pregnancy for 6 months following the last study administration and instructed 
to use birth control.  
 
6.19. Protecting Against or Minimizing Potential Treatment Risks 
Subjects will not be enrolled who have other active serious medical problems.  Frequent monitoring of 
patients with history, physical examination, and laboratory studies will allow for early identification of 
adverse events.  All participants will be required to have adequate hemoglobin to allow frequent 
venipuncture.  The supplement drugs will be dispensed to subjects free of cost, and subjects will be 
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Subjects will be counseled about the potential risk for gastrointestinal side effects and the need to report 
any change in health status between or at the time of visits.  Directed questioning about concurrent 
illness will occur before each treatment.   
 
6.20. Supplement - Drug Interaction  
Vitamin D 
Like most dietary supplements, vitamin D may interact or interfere with other medicines or 
supplements.. Here are several examples: 
- 
Corticosteroid medicines to reduce inflammation impair how the body handles vitamin D, which 
leads to lower calcium absorption and loss of bone over time. 
- 
Both the weight-loss drug orlistat (brand names Xenical® and Alli®) and the cholesterol-
lowering drug cholestyramine (brand names Questran®, LoCholest®, and Prevalite®) can 
reduce the absorption of vitamin D and other fat-soluble vitamins (A, E, and K). 
- 
Both phenobarbital and phenytoin (brand name Dilantin®), used to prevent and control epileptic 
seizures, increase the breakdown of vitamin D and reduce calcium absorption. 
 
Omega-3 Fatty Acids  
Omega-3 Fatty Acids can affect medications that influence blood clotting phenomenon, some of which 
includes:  
• 
Antiplatelet Medication: Anagrelide (Agrylin®), aspirin(any brand, all doses), cilostazol 
(Pletal®), 
clopidogrel 
(Plavix®), 
dipyradamole 
(Persantine®), 
dipyridamole/aspirin 
(Aggrenox®), enteric-coated aspirin (Ecotrin®), ticlopidine (Ticlid®) 
• 
Anticoagulant Medication: Anisindione (Miradon®), Arixtra, enoxaparin (Lovenox®) injection, 
Fragmin, heparin injection, Pradaxa, pentosan polysulfate (Elmiron®), warfarin (Coumadin®), 
Xerelto  
• 
Nonsteroidal Anti-Inflammatory Drugs: Celebrex, diclofenac (Voltaren®, Cataflam®), diflunisal 
(Dolobid®), etodolac (Lodine®), fenoprofen (Nalfon®), flurbiprogen (Ansaid®), ibuprofen 
(Motrin®, Advil®, Nuprin®, Rufen®), indomethacin (Indocin®), ketoprofen (Orudis®, Actron®), 
ketorolac (Toradol®), meclofenamate (Meclomen®), meloxican (Mobic®), nabumeton 
(Relafen®), naproxen (Naprosyn®, Naprelan®, Aleve®), oxaprozin (Daypro®), piroxicam 
(Feldene®), salsalate (Salflex®, Disalcid®), sulindac (Clinoril®), sulfinpyrazone tolmetin 
(Tolectin®), trilisate (salicylate combination)  
• 
Herbs/Vitamins: Ajoene birch bark, cayenne, Chinese black tree fungus, cumin, evening 
primrose oil, feverfew, garlic, ginger, ginkgo biloba, ginseng, grape seed extract, milk thistle, 
onion extract, St. John’s wort, tumeric, vitamins C and E.  
The study team will evaluate the list of all medications and its interaction with Vitamin D or Omega-3 
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if dietary supplements might interact or interfere with their prescription or over-the-counter medicines, 
or if the medicines might interfere with how patients body absorbs, uses, or breaks down nutrients. 
7. ADVERSE EVENT REPORTING AND SAFETY MONITORING 
7.1. 
Adverse Event Definition 
7.1.1. Adverse Event (AE) 
In this clinical study, an adverse event is any occurrence or worsening of an undesirable or unintended 
sign, symptom or disease whether or not associated with the treatment and study procedures.   
Throughout the study, the investigator must record all adverse events on source documents.  Events 
not related to hypo or hyperglycemia that are Grade 2 or greater per the NCI CTCAE (see Grading 
Event Severity below) will be reviewed by study physician and medical monitor on the appropriate 
adverse event form.  The investigator will treat participants with adverse events appropriately and 
observe them at suitable intervals until the events resolve or stabilize. 
Adverse events may be discovered through: 
• 
observation of the participant; 
• 
questioning the participant; 
• 
unsolicited complaint by the participant 
In questioning the participant the questioning should be conducted in an objective manner. 
 
7.1.1.1. 
Disease specific AE 
For the purposes of this study, hypoglycemia events will be recorded as follows: 
• 
Level 1: glucose < 70 mg/dl (3.9 mmol/l) and glucose ≥ 54 mg/dl (3.0 mmol/l). and  
• 
Level 2: glucose < 54 mg/dl (3.0 mmol/l) 
• 
Level 3: a severe event characterized by altered mental and/or physical status requiring 
assistance. 
 
• 
.  Level 2 and Level 3 hypoglycemic events must be reported as adverse events on the case 
report forms. All episodes of Level 3  hypoglycemia will be reported as SAEs to the DSMB. 
 
Hypoglycemia grading according to CTCAE version 4.0 
Adverse Event 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Grade 5 
Hypoglycemia 
<LLN-
54mg/dL 
<54 -40 mg/dL 
<40-30 mg/dL 
<30 mg/dL; life 
threatening 
consequences; seizures 
Death 
 
.  

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Hyperglycemia will not be collected as an adverse event, unless it results in an inpatient 
hospitalization. 
 
7.1.2. Serious Adverse Event 
For this trial, an adverse event associated with the treatment or study procedure that suggests a 
significant hazard, contraindication, side effect or precaution (as described below) is to be reported as 
a serious adverse event (SAE).  A serious adverse event (experience) or reaction is any untoward 
medical occurrence that: 
• 
results in death, 
• 
is life-threatening, 
• 
requires inpatient hospitalization or prolongation of existing hospitalization, 
• 
results in persistent or significant disability/incapacity, or 
• 
is a congenital anomaly/birth defect. 
Important medical events that may not result in death, be life threatening, or require hospitalization may 
be considered serious adverse events when, based upon appropriate medical judgment, they may 
jeopardize the patient and may require medical or surgical intervention to prevent one of the outcomes 
listed above. 
 
7.1.3. Unexpected Adverse Event 
An adverse event is considered unexpected when the nature (specificity) or severity of the event is not 
consistent with the risks described in the protocol or informed consent document for a particular 
protocol. 
7.1.4. Grading Event Severity 
This study will adopt the National Cancer Institute (NCI) Common Technology Criteria for Adverse 
Events (CTCAE) and/or study-specific criteria for classification to describe the severity of adverse 
events with the exception of hyper and hypoglycemia. For this study, a reportable hypoglycemic event 
is defined as Level 3 (a severe event characterized by altered mental and/or physical status requiring 
assistance) and hyperglycemic event is one resulting in Diabetes Keto Acidosis (DKA).  
 
7.1.5. Evaluation of Adverse events  
A non-serious adverse event is an AE not classified as serious. Both serious and non-serious AEs 
should be graded with respect to severity on the following 3 point scale and reported, in detail, on the 
appropriate CRF page: 
 

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Mild 
Discomfort noticed, but no disruption of normal daily activities; event usually requires 
no intervention. 
Moderate 
Discomfort sufficient to reduce or affect normal daily activities; even may require 
intervention. 
Severe 
Incapacitating, with inability to perform normal daily activities; event usually requires 
treatment or other intervention. Subject may not be able to continue in the study. 
 
The Investigator should evaluate the relationship of each AE to the study treatment regimen, using the 
following criteria: 
Unrelated: 
Another cause of the AE is more plausible; a clinically plausible temporal 
sequence is inconsistent with the onset of the AE and administration of the study 
treatment regimen; or a causal relationship is considered biologically impossible. 
Possibly related: 
There is a clinically plausible time sequence between onset of the AE and 
administration of the study treatment regimen, but the AE could also be 
attributed to concurrent or underlying disease, or the use of other drugs or 
procedures. “Possibly Related” should be used when the study treatment 
regimen is one of several biologically plausible AE causes. 
Definitely Related: The AE is clearly related to use of the study treatment regimen. 
 
 
7.1.6. Adverse Event Reporting and Monitoring 
Study personnel will assess adverse events and the use of concomitant medications throughout the 
study. They will be graded as to severity according to common toxicity criteria or study-specific criteria 
and the investigator will make a determination as to the relation to therapy. Events will be assessed 
and reported in accordance with the ICH Guideline for Good Clinical Practice and per the guidance of 
the DHHS Office for Human Research Protections (OHRP). The adverse event case report for the 
protocol must be documented for all adverse events (AE) of Grade 2 or greater severity regardless of 
relationship to therapy. For reporting serious adverse events (SAE), the MedWatch Form should also 
be completed and sent to medical monitor within 24 hours of when the site was notified of the event. 
This will be reviewed by the study physicians and Medical Monitor, and the Data and Safety Monitoring 
Board (DSMB) as appropriate. Deaths must be reported immediately. Event outcome and other follow-
up information regarding the treatment and resolution of the event will be obtained and reported when 
available, if not known at the time the event is reported. The follow-up information should contain 
sufficient detail to allow for a complete medical assessment of the case and an independent 
determination of possible causality. 
Adverse events will be assessed by the Medical Monitor. The DSMB will conduct regular safety reviews 
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assignment. Serious adverse events as well as adverse events leading to treatment discontinuation will 
be reviewed by the DSMB. 
Fatal or life-threatening events thought to be caused by study drug(s) should be reported to the Medical 
Monitor, or CRA immediately, as follows: 
Position 
Name 
Telephone 
Number 
Pager Number 
Medical 
Monitor 
George W. Burke,III M.D., FACS 
Professor of Surgery  
Director, Division of Kidney and 
Pancreas Transplantation, 1801 
NW 9th Floor, Miami, FL 33136 
Email: GBurke@med.miami.edu   
(305) 355-5111   
(305) 355-5134 
CRA 
Clinical Research Operations & 
Regulatory Support (CRORS), 
Miller Office of Research, 
University of Miami, Miller School 
of Medicine, Dominion Towers 
4th floor, 1400 N.W. 10th 
Avenue; Suite 401, Miami, FL 
33136 
(305) 243-0133   
(305) 243-5392 
 
7.1.7. Recording Adverse Events 
To improve the quality and precision of AE data, Investigators should observe the following 
guidelines: 
• 
Whenever possible, use recognized medical terms when recording AEs on the AE page of the 
CRF. Do not use colloquialisms, jargon, or abbreviations.  
• 
If known, record the diagnosis (ie, disease or syndrome) rather than component signs and 
symptoms on ae pages of the CRF (eg, record “congestive heart failure” rather than 
“dyspnea”, “rales”, and “cyanosis”). However, signs and symptoms that are considered 
unrelated to an encountered syndrome or disease should be recorded as individual AEs on 
the CRF page. For example, if congestive heart failure and severe headache are observed at 
the same time, each event should be recorded as an individual AE. 
• 
Adverse events occurring secondary to other events (ie, sequelae) should be identified by the 
primary cause. A primary ae, if clearly identifiable, generally represents the most accurate 
clinical term to record on ae pages of the CFR. If a primary SAE is recorded on an AE CRF 
page, events occurring secondary to the primary event should be described in the narrative 
description of the event. 
• 
Laboratory abnormalities that are defined as clinically significant are to be considered AEs 
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7.1.8. Adverse Events Requiring Expedited Reporting 
Serious AEs require expedited reporting to the Sponsor or designee, regardless of the relationship of 
the event to the study treatment regimen. Refer to the previous section for the definition of a serious 
adverse event. 
Investigators should report all SAEs to the Sponsor or designee within 24 hours of the observation or 
learning of the event. For initial SAE reports, the Investigator should record all case details that can be 
garnered on the SAE form and the AE CRF page. 
The completed SAE form and Cover Sheet should be faxed within 24 hours to the medical monitor at 
(305) 355-5134 or via email GBurke@med.miami.edu. Relevant follow-up information is to be 
submitted to the Sponsor or its designee as soon as it becomes available. 
 
7.1.9. Special Reporting Situations 
Death: Death is an outcome of an event. The event that resulted in death should be recorded and 
reported on the SAE form and the AE CRF page. 
Hospitalization for Surgical or Diagnostic Procedures: The illness leading to the surgical or diagnostic 
procedure is to be recorded as the SAE, not the procedure itself. The procedure is to be captured in 
the case narrative as part of the action taken in response to the illness. 
 
7.1.10. Laboratory Test Result Abnormalities 
The following laboratory test result abnormalities should be captured on the non-serious AE CRF page 
or SAE Report Form (paper or electronic) as appropriate: 
• 
Any laboratory test result that is clinically significant or meets the definition of an SAE  
• 
Any laboratory test result abnormality that required the subject to have study drug discontinued 
or interrupted 
• 
Any laboratory test result abnormality that required the subject to receive specific corrective 
therapy. 
It is expected that wherever possible, the clinical rather than laboratory term would be used by the 
reporting investigator (e.g. anemia versus low hemoglobin value). 
7.1.11. Pregnancy 
If, following initiation of the study product(s), it is subsequently discovered that a study subject is 
pregnant or may have been pregnant at the time of investigational product exposure, including up to 
12 weeks after last product administration, the study product(s) will be permanently discontinued in an 
appropriate manner (e.g. dose tapering if necessary for subject safety).  

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They must be reported to the medical monitor within 24 hours by fax to (305) 355-5134 with a 
confirmatory telephone call to (305) 355-5111 and in accordance with SAE reporting procedures 
described above. 
Any pregnancy that occurs in a female partner of a male study participant should be reported to the 
medical monitor.  
. 
 
7.1.12. Overdose 
All occurrences of overdose must be reported as SAEs (see SAE reporting section for details). 
An overdose is defined as the accidental or intentional administration of any dose of a product that is 
considered both excessive and medically important. All occurrences of overdose must be reported as 
an SAE. 
 
7.1.13. Other Safety Considerations 
Any significant worsening noted during interim or final physical examinations, electrocardiogram, x-ray 
filming, any other potential safety assessment required or not required by protocol should also be 
recorded as a non-serious or serious AE, as appropriate, and reported accordingly.  
 
8. DATA QUALITY ASSURANCE 
Accurate, consistent, and reliable data will be ensured through the use of standard practices and 
procedures. These are described in the following sections. 
 
8.1. 
Data Collection, Monitoring, and Transfer 
Adverse events will be collected from the initiation of study drug and be followed to resolution or until 
30 days after the participant completes participation, whichever comes first. 
Adverse events may be collected as follows: 
• 
Observing the participant. 
• 
Questioning the participant in an objective manner. 
• 
Receiving an unsolicited complaint from the participant. 
An abnormal value or result from a clinical or laboratory can also indicate an adverse event.  If this is 
the case, then the evaluation that produced the value or result should be repeated until the value or 
result returns to normal or can be explained and the participant’s safety is not at risk.  If an abnormal 
value or result is determined by the investigator to be clinically significant, it must be indicated as such 
on the appropriate laboratory evaluation form(s), and must also be reported as an adverse event on 
the adverse event form. Abnormal vital sign measurements or laboratory finding  deemed not clinically 
significant by the site investigator must be documented as such and are not required to be listed in the 

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adverse event source documents or adverse event case report form. The DRI will perform data 
collection, monitoring, and transfer tasks. A Clinical Research Monitor designated by Sponsor will be 
responsible for monitoring the clinical study in accordance with current federal and relevant foreign 
regulations. The Monitor will periodically collect the CRFs and brought them to Sponsor. Data on CRFs 
will be double-entered into the database system and verified. 
 
8.2. 
Quality Control and Quality Assurance 
 The database will be verified against the CRFs as outlined in the monitoring plan.  
9. STATISTICAL CONSIDERATIONS AND ANALYSIS PLAN 
 
9.1. 
Overview 
This study will test the hypothesis that combination therapy, administered for one year to patients with 
either new onset or established T1D with residual stimulated C-peptide, will retain C-peptide secretion 
compared to therapy with vitamin D alone. 
 
9.2. 
Endpoints/Outcomes 
Primary endpoint: 
• 
Stimulated (90 minute sample of a MMTT) C-peptide at the 1 year visit greater or equal to 
baseline level. 
Secondary Objective(s) 
 
• 
Stimulated C-peptide area under the curve during a 4-hour MMTT greater or equal to baseline 
level, at the 1 year visit. 
• 
Reduction in HbA1c at the one year visit compared to baseline. 
• 
Reduction in insulin requirement at the 1 year visit compared to baseline. 
• 
Incidence of adverse events (AE) comparable to general diabetes population.  
 
 
9.3. 
Measures to Minimize Bias  
Participants who completed the screening visit and are determine to be eligible for study participation 
will undergo randomization. Participants will be randomized by the study PI at the clinical site. The 
participants will be informed regarding the intervention assignment.   
 
9.4. 
Analysis Plan 
The details of the analyses will be provided in the statistical analysis plan (SAP) below. 
 
9.4.1. Analysis Populations 

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Analyses of study data will be conducted to address all objectives of the trial and other interrelationships 
among data elements of interest to the investigators and of relevance to the objectives of the study. 
Primary analysis of treatment effect will be conducted under the intention-to-treat principle of eligible 
patients whereby outcome data from all eligible patients will be included regardless of treatment 
compliance. 
9.4.2. Primary Analysis of Primary Endpoint(s)/Outcome(s) 
The metabolic primary outcome is the 90-minute stimulated C-peptide following MMTT, conducted at 
the 1 year visit. The primary statistical hypothesis to be assessed is that the proportion of patients with 
maintained or increased level of 90-minute at 12 months as compared to baseline will be greater in 
subjects receiving combination therapy than for subjects receiving vitamin D alone. 
The analysis of the primary outcome will initially involve a two sample test of proportions comparing the 
proportion of patients in the combination therapy group maintaining or increasing 90-C-peptide with 
respect to baseline with the proportation of vitamin D alone treated subjects with 90-minute C-peptide 
that is maintained or increased from baseline value. A covariate-adjusted analysis will additionally be 
performed by way of a logistic regression model, with a categorical index of treatment group adjusted 
for baseline age and time since diagnosis; this can provide us with adjustment for potential confounding 
of these factors and/or provide increased efficiency and precision by including these potentially 
significant explanatory variables. 
 
9.4.3. Supportive Analyses of the Primary Endpoint(s)/Outcome(s)  
90-min C-peptide measured at screening and , 12 months and 24 months in the different treatment 
groups will additionally be fit to a mixed linear regression model, allowing estimation and testing of 
means and differences between groups at each time point, while simultaneously accommodating the 
correlation arising from the longitudinal study design in estimates of variances used in tests of 
significance. We will also estimate and test the differential rate of change in mean 90-min C-peptide 
over time between these groups, by incorporating an interaction term between time point and treatment 
group into the models. Although this investigation is not specifically powered to test this interaction, this 
might provide us with useful insight as to the differences in the effect of combination therapy over time 
on 90-min C-peptide. 
 
9.4.4. Descriptive Analyses  
In addition to the detailed statistical analysis described above, descriptive analysis will include reporting 
and comparison baseline characteristics of the study subjects to evaluate whether any substantial 
differences exist between the vitamin D alone and combination treatment groups for key variables such 
as age, gender, BMI, HbA1c, baseline C-peptide, insulin requirements and other variables such as 
concomitant use of other medications to determine any potential bias. These will involve standard two 
sample comparisons such as T-tests for continuous variables (possibly transformed when necessary 
to preserve distributional assumptions) and Chi-square or Fisher’s exact tests for categorical variables. 
As described earlier, potentially confounding factors arising from imbalance will be included into more 
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9.5. 
Interim Analyses  
The DSMB will be convened to review safety and efficacy data.  Descriptive interim analyses will include 
distributions of endpoints, and adverse events.  Interim analyses will be conducted periodically during 
the study and will be reviewed by the DSMB for assessment of effectiveness and safety. Group-
sequential spending function with an O’Brien-Fleming boundaries may be used to protect the type I 
error probability from early and multiple testing and to assess the significance of the interim results that 
emerge during the trial.   
 
9.5.1. Interim Analysis of Efficacy Data 
Interim analysis for efficacy will be planned when half the subjects have reached their endpoint.  
However we note that this is a small trial and if we were to consider a priori planning for interim analysis 
the limited number of observations could lead to unstable estimates of efficacy.  This can be further 
discussed with the DSMB. The inflation factor needed for augmenting sample size to preserve power 
if we were to plan for interim analysis at one look with group sequential methods using standard Obrien-
Fleming boundaries is 1.02, which does not significantly affect the parameters of our design (sample 
size needs to be inflated by 1.02).  
 
9.5.2. Interim Analysis of Safety Data 
Interim analysis may be conducted as recommended by the DSMB, to review safety data, including 
metabolic, immunological assessments and adverse events. The DSMB may recommend early 
stopping if there is sufficient evidence to conclude that the investigational treatment is harmful based 
on interim analysis of safety endpoints.   
At any time, if there are significantly more adverse events in the experimental arm based on Fisher’s 
exact tests for comparison, early termination should be considered.   For example, a scenario where 9 
patients have enrolled with 6 in the combination treatment arm and 3 in the vitamin D alone arm with 5 
adverse events in the combination treatment arm and 0 in the vitamin D treatment arm would be 
considered significantly increased likelihood of adverse events in the combination treatment arm and 
would potentially result in trial termination.   The same holds in the scenario where 15 patients have 
been enrolled with 10 in the combination treatment arm and 5 in the vitamin D alone treatment arm, 
where the combination treatment group experiences 9 adverse events while the vitamin D group 
experiences only 1.  Similar scenarios exist throughout the trial and at any time if the exact test 
comparing groups under the null that the likelihood of adverse events is equal between groups exceeds 
a significance threshold of 0.05, this would warrant early termination. 
 
9.5.3. Futility Analysis 
Futility is unlikely to be shown during a one year treatment period, but the DSMB can decide to terminate 
the study should futility be shown. We do not intend to stop the study for futility as we intend to obtain 
an estimate and 95% confidence interval for the effect of treatment between groups at the 1 year mark 

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using the full information offered by our design.  Additionally, not stopping for futility incorporates the 
consideration that the C-peptide AUC curves of combination therapy and vitamin D alone groups may 
separate later on follow-up, as AUC values decline further in the vitamin D alone group. Essentially all 
type 1 diabetes trials have been allowed to reach their one-year endpoint, even those that showed no 
efficacy. 
 
9.6. 
Statistical Hypotheses 
 
Primary hypothesis: 
• 
Null: Patients receiving combination supplement therapy and patients in the Vitamin-D alone 
group have comparable proportions with maintained or increased C-peptide production, 
assessed by the measurement of the stimulated 90-minute C-peptide at one year of follow up. 
• 
Alternative: Patients receiving combination therapy have higher proportions with maintained or 
increased C-peptide production, assessed by the measurement of the stimulated 90-minute C-
peptide as compared to the proportion of patients with maintained or increased C-Peptide in the 
vitamin D alone group at one year of follow up. 
 
9.7. 
Sample Size Considerations  
Metabolic primary outcome 
Sample size considerations correspond to the primary hypothesis which is that the proportion of 
patients with maintained or increased 90-min C-pepetide at 12 months compared to baseline value is 
greater in the combination therapy group than in the vitamin D alone group.  Standard power 
calculations for two sample tests of proportions indicate that 12 subjects in each group are needed to 
achieve 80% power to detect differences between less than 5% in the vitamin D alone group and at 
least 50% in the combination therapy group.  Allowing for 10% dropout/attrition, sample size 
considerations include an allowance for 10% of the subjects to have missing data (one-year MMTT was 
not done or subject withdrew prior to the one-year assessment).  
Primary endpoints should be available for all enrolled subjects.  An exception will be if a death occurs 
or if the subject withdraws consent to be followed, although we expect this to be very limited. We also 
have included consideration for attrition in our design.  Should there be any missing data, missing data 
will be assessed for mechanism of missingness and to see if missing at random assumptions are valid. 
If necessary, appropriate imputation strategies can be applied for sensitivity analysis. We will explore 
the data to assess if missingness at a given time point is related to observed data at the previous time 
point. If we find that, conditional on treatment group, observed values strongly predict subsequent 
values where observed, but are not related to missingness at subsequent time points, we may regard 
the missing data as a random sample of previously observed data. 
 
10. 
ETHICAL CONSIDERATIONS & COMPLIANCE WITH GOOD CLINICAL PRACTICE 

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10.1. Statement of Compliance 
The study will be conducted in accordance with the protocol, Good Clinical Practices, the relevant ICH 
guidelines, the applicable regulatory requirements, and the ethical principles that have their origins in 
the Declaration of Helsinki. As required by United States Food and Drug Administration (FDA) Code of 
Federal Regulations (CFR) (21 CFR 56) and the Declaration of Helsinki, the study protocol, 
amendments, and Informed Consent form will be reviewed and approved, according to 21 CFR §50 
and §56, respectively, by University of Miami’s IRB. Any amendments to the protocol or consent 
materials must also be approved before they are implemented. 
 
10.2. Informed Consent 
The consent process will be conducted by qualified study personnel (the Study Coordinator and/or 
Investigator or other designee).  All participants (or their legally acceptable representative) must read, 
sign and date a consent or assent form prior to participation in the study, and/or undergoing any study-
specific procedures. 
The informed consent form must be updated or revised whenever there is new clinically significant 
information applicable to the safety of the participants  when indicated for a protocol amendment, and/or 
whenever any new information becomes available that may affect a patient’s participation in the study.   
 
10.3. Study Subject Confidentiality 
The investigational site participating in this study will maintain the highest degree of confidentiality 
permitted for the clinical and research information obtained from subjects.  As a part of the quality 
assurance and legal responsibilities of an investigation, the investigational site must permit authorized 
representatives of the Quality Assurance division of University of Miami and regulatory agencies to 
examine (and when required by applicable law, to copy) records for the purposes of quality assurance 
reviews, audits and evaluation of the study safety and progress. Unless required by the laws permitting 
copying of records, only the coded identity associated with documents or other participant data may be 
copied (obscuring any personally identifying information). Authorized representatives, as noted above, 
are bound to maintain the strict confidentiality of medical and research information that may be linked 
to identify individuals. The investigational site will normally be notified in advance of auditing visits. 
Study records with the study subject’s information for internal use at the clinical site will be secured at 
the study site during the study. At the end of the study, all records will continue to be kept in a secure 
location. There are no plans to destroy the records.  
Study subject data, which is for reporting purposes, will be stored at the Diabetes Research Institute.  
Case report forms will identify participants by the unique Identification Number.  At the end of the study, 
all study databases will be archived at the Center, and the data collection forms will be saved for long-
term storage.   
Stored samples including genetic samples could be utilized to learn more about causes of type 1 
diabetes, its complications (such as eye, nerve, and kidney damage) and other conditions for which 

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individuals with diabetes are at increased risk, and how to improve treatment.  The results of these 
future analyses, and any mechanistic studies will not be made known to the participant.  
 
11. STUDY ADMINISTRATION 
This study is investigator initiated study sponsored by the investigators from Diabetes Research 
Institute. Funding will cover the costs of administration and laboratory tests associated with this study 
during the participant’s period of follow-up and procedures related to drug administration. Study drugs 
will 
be 
free 
of 
charge 
for 
the 
participant’s 
entire 
length 
of 
treatment.  
 
Since dosage of study drugs is dependent on blood levels (i.e., 25OH vitamin D and AA/EPA ratio) and 
results may take up to 1 week to be ready following blood collection during the participants’ visits, 
participants who live far from the study site and logistically cannot come to the site to pick-up their study 
drug outside of their scheduled study visit windows, may have the study drugs shipped to them. Both 
study drugs, vitamin D and omega 3 fatty acids, are stable at room temperature and do not require 
additional special handling during shipment except for adequate packing of the omega 3 bottles to 
ensure they do not break during transportation.  
 
11.1. Groups and Committees 
11.1.1. Protocol Development Committee 
As acknowledged on the preface of this study protocol, protocol development committee met 
periodically over four months to finalize this protocol. Members of this committee bring interdisciplinary 
and translational expertise in pediatrics, immunology, transplantation and endocrinology of T1D, 
research nurse, research pharmacist and regulatory experts. Committee consists of Subject Matter 
Experts with unique and proven expertise in different immune pathways involved in T1D. 
 
11.1.2. Clinical Site 
The study will be conducted at Diabetes Research Institute and Mailman Center for Child Development. 
Principal Investigators of the participating clinical site will oversee all operations.  The clinical sites will 
conduct the trial and collect all laboratory and data collection form information for analysis. Clinical 
research team meets every Friday of the week to discuss development and issues related to all clinical 
research studies at the institute and facilitate evaluation of the trial management.  
 
11.1.3. Medical Monitor for the Clinical Protocol 
This trial includes the medical monitor independent of the study (George Burke, M.D., Professor of 
Surgery at the University of Miami Miller School of Medicine). 
 
11.1.4. Clinical Site Monitoring 

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In order to conduct this study consistent with established research principles and ICH-GCP guidelines, 
there will be site visits conducted during the study to evaluate study conduct. Clinical Research 
Operation and Regulatory Support (CRORS) is part of the Research, Research Education & Innovative 
Medicine (RIM) Department, within the Miller School of Medicine at the University of Miami. The 
CRORS is independent of the Diabetes Research Institute and will provide source data verification and 
periodic monitoring for patient enrollment, compliance with protocol procedures, completeness and 
accuracy of data entered on the case report forms (CRFs), and the occurrence and reporting of adverse 
events (AEs) and serious adverse events (SAEs). The site will be monitored by the Clinical Research 
Associates from the CRORS.  
 
11.1.5. Data and Safety Monitoring Board (DSMB) 
The members of the DSMB are independent and serve in an individual capacity to provide their 
expertise and recommendations. They will review the 90 day data of the first 24 subjects, and then 
periodically review as per the charter and evaluate the accumulated study data for participant safety, 
study conduct and progress, and, when appropriate, efficacy, and make recommendations to study 
investigators concerning the continuation, modification, or termination of the trial. The DSMB members 
are experts in the field of T1D immunotherapy and will consider study-specific data as well as relevant 
background knowledge about the disease, test agents, and patient population under study. All adverse 
events will be recorded on the adverse event forms, which will be provided to the local IRBs, per their 
reporting requirements.  The DSMB will independently evaluate whether the adverse events constitute 
grounds to discontinue the study. 
 
11.1.6. DRI Office of Regulatory Support  
This office of Quality Assurance and Regulatory Affairs provides standardized auditing functions, 
educational training, and regulatory support for clinical research at the Diabetes Research Institute. 
 
11.1.7. Human Subjects Research Office & Institutional Review Boards (HSRO & IRB) 
The HSRO provides administrative support for the University of Miami IRBs, which review proposed 
research involving human subjects to ensure the protection of those subjects and regulatory 
compliance.  
 
11.1.8. Clinical Research Centers 
Pediatric endocrinology clinics at the Mailman Center for Child Development and adult endocrinology 
clinics at Diabetes Research Institute and Clinical Translational Research Site at the University of Miami 
Miller School of Medicine provide comfortable, safe and effective environment to conduct professional, 
high-quality research involving human subjects.  
 
11.1.9. Investigational Drug Services (Research Pharmacy)  

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The Investigational Drug Service (IDS) was established to provide pharmacy support to investigators 
in managing randomized, clinical trials at the University of Miami. The IDS also has the capability to 
dispensing complex investigational therapies and as drug safety experts therein can provide 
customized value that is increasingly recognized as a mechanism for assuring safety of study patients, 
ensuring adherence to protocol guidelines, and minimizing the liability associated in conducting drugs 
or supplement trials. 
 
11.1.10. Core Laboratories 
Core laboratories participating in the study include: 
• 
Clinical research laboratories routinely providing services to the University of Miami. 
• 
The Laboratory of Dr. Alberto Pugliese at the Diabetes Research Institute, University of Miami, 
Miami, Florida, where samples from patients will be processed, divided in aliquots, frozen as 
needed for future analysis.  
 
11.2. Preservation of the Integrity of the Study 
All presentations and publications using study data must protect the main objectives of the trial.  Data 
integrity will be maintained and the investigators will publish study results and will apply accepted 
academic policies and regulations governing the publication. 
 
11.3. Participant Reimbursement and Compensation  
Participants will not be compensated for visits attended in the study. The supplement(s) during the 
course of the study will be provided at no charge to the subjects. 
  
12. 
STUDY TIMELINE 
It is anticipated that patient enrollment will occur during the first three years of the trial.  All subjects will 
be followed until one year after initial treatment, with up to one year further follow-up after the treatment 
period has ended. Study visits will have a window of ±14 days starting from visit 2. 
 
 
 
 
 
 
 
 
 
 

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APPENDIX A: Schedule of Events (SOE) 
 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial1 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
Informed Consent 
X 
 
 
 
 
 
 
 
Medical History 
X 
 
X 
X 
X 
X 
X 
X 
Physical Exam2 
X 
 
X 
X 
X 
X 
X 
X 
CBC with differential 
X 
X 
X 
X 
X 
 
 
 
Prothrombin time (PT/PT-INR) 
X 
 
X 
X 
X 
X 
X 
X 
Chemistries 
X 
 
X 
X 
X 
 
 
 
Pregnancy test (blood sample)5  
X 
 
X 
X 
X 
X 
 
 
Serum for islet autoantibodies 
X 
 
X 
X 
X 
X 
X 
X 
Serology 
X 
 
 
 
 
 
 
 
Mechanistic assessments3 
 
X 
X 
X 
X 
X 
X 
X 
Hemoglobin A1c  
X 
 
X 
X 
X 
X 
X 
X 
Stimulated C-peptide at 90 min 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
X 
 
 
 
X 
 
 
X 
Randomization  
 
X 
 
 
 
 
 
 
Diabetes Assessments (insulin adjustment 
and Hypoglycemia awareness) 
X 
X 
X 
X 
X 
X 
X 
X 
Clarke Score 
X 
 
 
 
X 
 
 
X 
Dispensing or Administration of Study Drug 
• 
Omega-3 Fatty Acids + Vitamin-D 
OR  
• 
Vitamin-D  
 
X6 
X 
X 
 
 
 
 
Vitamin D levels 4 
X 
 
X 
X 
X 
X 
X 
X 
AA/EPA Ratio7 
X 
 
X 
X 
X 
X 
X 
X 
Counting of Returned Study Drug 
 
 
X 
X 
X 
 
 
 
Subject 7-day blood sugar record and 
hypoglycemia log  
X 
 
X 
X 
X 
X 
X 
X 
Concomitant Medication Review 
X 
X 
X 
X 
X 
 
 
 
Adverse Events Assessment 
 
 
X 
X 
X 
X 
X 
X 
 
1. Screening Visit:  Screening MMTT must be at least 3 weeks after diagnosis and preferably within one 
month (37 days) of randomization for recent onset T1D. For established T1D, screening must take place 
after 180 days of diagnosis. 
2. Physical Exam: Routine exam at screening and quarterly. 

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3. Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles.  The schedule for these assessments may vary as appropriate.  
At no time will the blood draw volume exceed what is allowable according to the subject’s body weight. 
4. Vitamin D levels will be monitored every 4 months or more frequently at the discretion of the investigator 
in order to adjust dose to achieve target levels.  
5. Pregnancy testing will be conducted in females in  reproductive age at 4, 8,12 and 16 months study visit. 
6. Study drug(s) will be administered within 48hr from baseline visit (visit 1).    
7. Dose will be adjusted to maintain a target AA/EPA ratio of 1.5-3.0. AA/EPA ratios will be monitored 
monthly until achieving target levels and as per protocol or as clinically indicated thereafter. 
 
F1-F3.  Follow-Up after 12 Months: Visits will be conducted approximately every 4 months (16, 20, and 24 
months). 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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APPENDIX B: Blood Sample Collection Amounts by Age Group and Weight 
 
 Ages ≥18 y/o (Adults) 
Maximum Draw Allowed Per 8 Week Period: 10.5 mL/kg or 550 mL 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
CBC with differential 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
 
 
 
Prothrombin time (PT/PT-INR) 
1.8 mL 
 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
Chemistries 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
 
 
 
Pregnancy test ¥ 
¥ 
 
¥ 
¥ 
¥ 
3.5 mL 
 
 
Serum for islet autoantibodies 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
Serology 
3.5 mL 
 
 
 
 
 
 
 
Mechanistic assessments§ 
 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
Hemoglobin A1c  
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Simulated C-peptide at 90 min* 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
38.5 
mL 
 
 
 
38.5 
mL 
 
 
 
Vitamin D levels  
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
AA/EPA Ratio 
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Total Blood collection per visit 
60.3 
mL 
23 mL 
39.3 
mL 
39.3 
mL 
77.8 
mL 
37.3 
mL 
33.8 
mL 
33.8 
mL 
 
¥ Pregnancy test for visits -1; 120; 240, and 360 will be performed using same serum sample for 
chemistry. For visit 480 a 3.5 mL SST tube (serum sample) will be collected to perform the test in the 
absence of a chemistry sample requirement  
§ Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles. The schedule for these assessments may vary as appropriate. 
At no time will the blood draw volume exceed what is allowable according to the subject’s body 
weight. 
Mechanistic Samples (total volume: 21 ml) as follows: 
Pro-inflammatory cytokines: 3.5 ml 
Phenotyping: 2 mL  
RNA: 2.5 mL 
DNA: 2.5  mL 
Resolvins: 2 mL 
LPS activation T-cell stimulation assay: 3 mL 
Plasma: 2 mL 
Serum: 3.5 mL 
 

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*Stimulated c-peptide at 90 min is a time point of the MMTT. No extra sample collection needed. 
 
 
 Ages 6 to <18 y/o 
 Weight at screening visit (Kg): 27 and greater 
Maximum Draw Allowed Per Day (5mL/Kg): 135 mL/kg 
Maximum Draw Allowed Per 8 Week Period (9.5 mL/Kg): 257 mL 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
CBC with differential 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
 
 
 
Prothrombin time (PT/PT-INR) 
1.8 mL 
 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
Chemistries 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
 
 
 
Pregnancy test ¥ 
¥ 
 
¥ 
¥ 
¥ 
3.5 mL 
 
 
Serum for islet autoantibodies 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
Serology 
3.5 mL 
 
 
 
 
 
 
 
Mechanistic assessments§ 
 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
Hemoglobin A1c  
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Simulated C-peptide at 90 min* 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
38.5 
mL 
 
 
 
38.5 
mL 
 
 
 
Vitamin D levels  
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
AA/EPA Ratio 
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Total Blood collection per visit 
60.3 
mL 
23 mL 
39.3 
mL 
39.3 
mL 
77.8 
mL 
37.3 
mL 
33.8 
mL 
33.8 
mL 
 
¥ Pregnancy test for visits -1; 120; 240, and 360 will be performed using same serum sample for 
chemistry. For visit 480 a 3.5 mL SST tube (serum sample) will be collected to perform the test in the 
absence of a chemistry sample requirement  
§ Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles. The schedule for these assessments may vary as appropriate. At 
no time will the blood draw volume exceed what is allowable according to the subject’s body weight. 
Mechanistic Samples (total volume: 21 ml) as follows: 
Pro-inflammatory cytokines: 3.5 ml 
Phenotyping: 2 mL  
RNA: 2.5 mL 
DNA: 2.5  mL 
Resolvins: 2 mL 
LPS activation T-cell stimulation assay: 3 mL 
Plasma: 2 mL 
Serum: 3.5 mL 

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*Stimulated c-peptide at 90 min is a time point of the MMTT. No extra sample collection needed. 
 
 
 
 Ages 6 to <18 y/o 
 Weight at screening visit (Kg): 25 to < 27 
Maximum Draw Allowed Per Day (5mL/Kg): 125 mL/kg 
Maximum Draw Allowed Per 8 Week Period (9.5 mL/Kg): 238 mL 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
CBC with differential 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
 
 
 
Prothrombin time (PT/PT-INR) 
1.8 mL 
 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
Chemistries 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
 
 
 
Pregnancy test ¥ 
¥ 
 
¥ 
¥ 
¥ 
3.5 mL 
 
 
Serum for islet autoantibodies 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
Serology 
3.5 mL 
 
 
 
 
 
 
 
Mechanistic assessments§ 
 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
Hemoglobin A1c  
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Simulated C-peptide at 90 min* 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
38.5 
mL 
 
 
 
38.5 
mL 
 
 
 
Vitamin D levels  
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
AA/EPA Ratio 
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Total Blood collection per visit 
60.3 
mL 
23 mL 
39.3 
mL 
39.3 
mL 
77.8 
mL 
37.3 
mL 
33.8 
mL 
33.8 
mL 
 
¥ Pregnancy test for visits -1; 120; 240, and 360 will be performed using same serum sample for 
chemistry. For visit 480 a 3.5 mL SST tube (serum sample) will be collected to perform the test in the 
absence of a chemistry sample requirement  
§ Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles. The schedule for these assessments may vary as appropriate. At 
no time will the blood draw volume exceed what is allowable according to the subject’s body weight. 
Mechanistic Samples (total volume: 21 ml) as follows: 
Pro-inflammatory cytokines: 3.5 ml 
Phenotyping: 2 mL  
RNA: 2.5 mL 
DNA: 2.5  mL 
Resolvins: 2 mL 
LPS activation T-cell stimulation assay: 3 mL 

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Plasma: 2 mL 
Serum: 3.5 mL 
 
*Stimulated c-peptide at 90 min is a time point of the MMTT. No extra sample collection needed. 
 
 
 
 Ages 6 to <18 y/o 
 Weight at screening visit (Kg): 23 to < 25 
Maximum Draw Allowed Per Day (5mL/Kg): 115 mL/kg 
Maximum Draw Allowed Per 8 Week Period (9.5 mL/Kg): 219 mL 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
CBC with differential 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
 
 
 
Prothrombin time (PT/PT-INR) 
1.8 mL 
 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
Chemistries 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
 
 
 
Pregnancy test ¥ 
¥ 
 
¥ 
¥ 
¥ 
3.5 mL 
 
 
Serum for islet autoantibodies 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
Serology 
3.5 mL 
 
 
 
 
 
 
 
Mechanistic assessments§ 
 
22 21 
mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
Hemoglobin A1c  
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Simulated C-peptide at 90 min* 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
38.5 
mL 
 
 
 
38.5 
mL 
 
 
 
Vitamin D levels  
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
AA/EPA Ratio 
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Total Blood collection per visit 
60.3 
mL 
23 mL 
39.3 
mL 
39.3 
mL 
77.8 
mL 
37.3 
mL 
33.8 
mL 
33.8 
mL 
 
¥ Pregnancy test for visits -1; 120; 240, and 360 will be performed using same serum sample for 
chemistry. For visit 480 a 3.5 mL SST tube (serum sample) will be collected to perform the test in the 
absence of a chemistry sample requirement  
§ Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles. The schedule for these assessments may vary as appropriate. At 
no time will the blood draw volume exceed what is allowable according to the subject’s body weight. 
Mechanistic Samples (total volume: 21 ml) as follows: 
Pro-inflammatory cytokines: 3.5 ml 
Phenotyping: 2 mL  
RNA: 2.5 mL 
DNA: 2.5  mL 

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Resolvins: 2 mL 
LPS activation T-cell stimulation assay: 3 mL 
Plasma: 2 mL 
Serum: 3.5 mL 
 
 
*Stimulated c-peptide at 90 min is a time point of the MMTT. No extra sample collection needed. 
 
 
 
 Ages 6 to <18 y/o 
 Weight at screening visit (Kg): 21 to < 23 
Maximum Draw Allowed Per Day (5mL/Kg): 105 mL/kg 
Maximum Draw Allowed Per 8 Week Period (9.5 mL/Kg): 200 mL 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
CBC with differential 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
 
 
 
Prothrombin time (PT/PT-INR) 
1.8 mL 
 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
Chemistries 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
 
 
 
Pregnancy test ¥ 
¥ 
 
¥ 
¥ 
¥ 
3.5 mL 
 
 
Serum for islet autoantibodies 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
Serology 
3.5 mL 
 
 
 
 
 
 
 
Mechanistic assessments§ 
 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
Hemoglobin A1c  
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Simulated C-peptide at 90 min* 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
38.5 
mL 
 
 
 
38.5 
mL 
 
 
 
Vitamin D levels  
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
AA/EPA Ratio 
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Total Blood collection per visit 
60.3 
mL 
23 mL 
39.3 
mL 
39.3 
mL 
77.8 
mL 
37.3 
mL 
33.8 
mL 
33.8 
mL 
 
¥ Pregnancy test for visits -1; 120; 240, and 360 will be performed using same serum sample for 
chemistry. For visit 480 a 3.5 mL SST tube (serum sample) will be collected to perform the test in the 
absence of a chemistry sample requirement  
§ Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles. The schedule for these assessments may vary as appropriate. At 
no time will the blood draw volume exceed what is allowable according to the subject’s body weight. 
Mechanistic Samples (total volume: 21 ml) as follows: 
Pro-inflammatory cytokines: 3.5 ml 

POSEIDON STUDY 
 
Version 4.1 
 
61 
 
 
Phenotyping: 2 mL  
RNA: 2.5 mL 
DNA: 2.5  mL 
Resolvins: 2 mL 
LPS activation T-cell stimulation assay: 3 mL 
Plasma: 2 mL 
Serum: 3.5 mL 
 
*Stimulated c-peptide at 90 min is a time point of the MMTT. No extra sample collection needed. 
 
 
 
 Ages 6 to <18 y/o 
 Weight at screening visit (Kg): 19 to < 21 
Maximum Draw Allowed Per Day (5mL/Kg): 95 mL/kg 
Maximum Draw Allowed Per 8 Week Period (9.5 mL/Kg): 181 mL 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
CBC with differential 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
 
 
 
Prothrombin time (PT/PT-INR) 
1.8 mL 
 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
Chemistries 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
 
 
 
Pregnancy test ¥ 
¥ 
 
¥ 
¥ 
¥ 
3.5 mL 
 
 
Serum for islet autoantibodies 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
Serology 
3.5 mL 
 
 
 
 
 
 
 
Mechanistic assessments§ 
 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
Hemoglobin A1c  
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Simulated C-peptide at 90 min* 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
38.5 
mL 
 
 
 
38.5 
mL 
 
 
 
Vitamin D levels  
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
AA/EPA Ratio 
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Total Blood collection per visit 
60.3 
mL 
23 mL 
39.3 
mL 
39.3 
mL 
77.8 
mL 
37.3 
mL 
33.8 
mL 
33.8 
mL 
 
¥ Pregnancy test for visits -1; 120; 240, and 360 will be performed using same serum sample for 
chemistry. For visit 480 a 3.5 mL SST tube (serum sample) will be collected to perform the test in the 
absence of a chemistry sample requirement  
§ Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles. The schedule for these assessments may vary as appropriate. At 
no time will the blood draw volume exceed what is allowable according to the subject’s body weight. 

POSEIDON STUDY 
 
Version 4.1 
 
62 
 
 
Mechanistic Samples (total volume: 21 ml) as follows: 
Pro-inflammatory cytokines: 3.5 ml 
Phenotyping: 2 mL  
RNA: 2.5 mL 
DNA: 2.5  mL 
Resolvins: 2 mL 
LPS activation T-cell stimulation assay: 3 mL 
Plasma: 2 mL 
Serum: 3.5 mL 
 
*Stimulated c-peptide at 90 min is a time point of the MMTT. No extra sample collection needed. 
 
 
 
 Ages 6 to <18 y/o 
 Weight at screening visit (Kg): 17 to < 19 
Maximum Draw Allowed Per Day (5mL/Kg): 85 mL/kg 
Maximum Draw Allowed Per 8 Week Period (9.5 mL/Kg): 162 mL 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
CBC with differential 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
 
 
 
Prothrombin time (PT/PT-INR) 
1.8 mL 
 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
Chemistries 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
 
 
 
Pregnancy test ¥ 
¥ 
 
¥ 
¥ 
¥ 
3.5 mL 
 
 
Serum for islet autoantibodies 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
Serology 
3.5 mL 
 
 
 
 
 
 
 
Mechanistic assessments§ 
 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
Hemoglobin A1c  
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Simulated C-peptide at 90 min* 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
38.5 
mL 
 
 
 
38.5 
mL 
 
 
 
Vitamin D levels  
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
AA/EPA Ratio 
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Total Blood collection per visit 
60.3 
mL 
23 mL 
39.3 
mL 
39.3 
mL 
77.8 
mL 
37.3 
mL 
33.8 
mL 
33.8 
mL 
 
¥ Pregnancy test for visits -1; 120; 240, and 360 will be performed using same serum sample for 
chemistry. For visit 480 a 3.5 mL SST tube (serum sample) will be collected to perform the test in the 
absence of a chemistry sample requirement  

POSEIDON STUDY 
 
Version 4.1 
 
63 
 
 
§ Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles. The schedule for these assessments may vary as appropriate. At 
no time will the blood draw volume exceed what is allowable according to the subject’s body weight. 
Mechanistic Samples (total volume: 21 ml) as follows: 
Pro-inflammatory cytokines: 3.5 ml 
Phenotyping: 2 mL  
RNA: 2.5 mL 
DNA: 2.5  mL 
Resolvins: 2 mL 
LPS activation T-cell stimulation assay: 3 mL 
Plasma: 2 mL 
Serum: 3.5 mL 
 
 
*Stimulated c-peptide at 90 min is a time point of the MMTT. No extra sample collection needed. 
 
 
 
 Ages 6 to <18 y/o 
 Weight at screening visit (Kg): 16 to < 17 
Maximum Draw Allowed Per Day (5mL/Kg): 80 mL/kg 
Maximum Draw Allowed Per 8 Week Period (9.5 mL/Kg): 152 mL 
Visit number 
0 
1 
2 
3 
4 
 
5 
F1 
6 
F2 
7 
F3 
Days of the trial 
-1 
1 
120 
240 
360 
480 
600 
720 
Months  
1 
1 
4 
8 
12 
16 
20 
24 
CBC with differential 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
 
 
 
Prothrombin time (PT/PT-INR) 
1.8 mL 
 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
1.8 mL 
Chemistries 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
 
 
 
Pregnancy test ¥ 
¥ 
 
¥ 
¥ 
¥ 
3.5 mL 
 
 
Serum for islet autoantibodies 
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
Serology 
3.5 mL 
 
 
 
 
 
 
 
Mechanistic assessments§ 
 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
21 mL 
Hemoglobin A1c  
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Simulated C-peptide at 90 min* 
X 
 
 
 
X 
 
 
X 
MMTT 4 hour  
38.5 
mL 
 
 
 
38.5 
mL 
 
 
 
Vitamin D levels  
3.5 mL 
 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
3.5 mL 
AA/EPA Ratio 
2 mL 
 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
2 mL 
Total Blood collection per visit 
60.3 
mL 
23 mL 
39.3 
mL 
39.3 
mL 
77.8 
mL 
37.3 
mL 
33.8 
mL 
33.8 
mL 
 

POSEIDON STUDY 
 
Version 4.1 
 
64 
 
 
¥ Pregnancy test for visits -1; 120; 240, and 360 will be performed using same serum sample for 
chemistry. For visit 480 a 3.5 mL SST tube (serum sample) will be collected to perform the test in the 
absence of a chemistry sample requirement  
§ Mechanistic Assessments: includes samples for RNA, plasma, serum, DNA, measures of B and T cell 
number/function and cytokine profiles. The schedule for these assessments may vary as appropriate. At 
no time will the blood draw volume exceed what is allowable according to the subject’s body weight. 
Mechanistic Samples (total volume: 21 ml) as follows: 
Pro-inflammatory cytokines: 3.5 ml 
Phenotyping: 2 mL  
RNA: 2.5 mL 
DNA: 2.5  mL 
Resolvins: 2 mL 
LPS activation T-cell stimulation assay: 3 mL 
Plasma: 2 mL 
Serum: 3.5 mL 
 
*Stimulated c-peptide at 90 min is a time point of the MMTT. No extra sample collection needed. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

POSEIDON STUDY 
 
Version 4.1 
 
65 
 
 
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Bener A, Alsaied A, Al-Ali M, Al-Kubaisi A, Basha B, Abraham A, et al. High prevalence of 
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Elajami, T.K., et al., Specialized proresolving lipid mediators in patients with coronary artery 
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Germano, M., et al., Plasma, red blood cells phospholipids and clinical evaluation after long 
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