Vitamin E Supplementation in Hyperinsulinism/Hyperammonemia Syndrome
Phase: NA
Conditions: Hyperinsulinism-Hyperammonemia Syndrome
Sponsor: Elizabeth A Rosenfeld

 
 
Vitamin E Supplementation in Hyperinsulinism/Hyperammonemia Syndrome 
 
NCT03797222 
 
Study Protocol Update Date: December 2, 2019 
Study Protocol IRB Approval Date: December 11, 2019 
 
Upload Date: June 22, 2022 
 
 

 
 
 
 
 
 
 
 
ii
 
Title: 
Vitamin E Supplementation in 
Hyperinsulinism/Hyperammonemia Syndrome 
 
Short Title 
Vitamin E in HI/HA 
Supplement Name: 
Vitamin E supplements 
eIRB Number 
17-014550 
Protocol Date: 
November 5, 2019 
Amendment 1 Date:  January 23, 2019, 
revised February 7, 2019 
Amendment 4 Date: June 10, 2019 
Amendment 2 Date:  February 20, 2019 
Amendment 3 Date: April 12, 2019, revised 
May 7, 2019, revised May 20, 2019 
Amendment 5 Date: November 5, 2019 
Amendment 6 Date: December 2, 2019 
 
Site Principal Investigator: Amanda M. Ackermann, MD, PhD 
Co-Investigator: Diva D. De Leon, MD, MSCE 
Co-Investigator: Elizabeth Rosenfeld, MD 
 
Children’s Hospital of Philadelphia 
3615 Civic Center Blvd., ARC 802C 
Philadelphia, PA, 19104 
Phone: 215-590-3174 
E-mail: ackermanna@email.chop.edu 
 

 
 
 
 
 
 
 
 
iii
TABLE OF CONTENTS 
Table of Contents ........................................................................................................ iii 
Abbreviations and Definitions of Terms .................................................................... vi 
Abstract ...................................................................................................................... vii 
Protocol Synopsis ..................................................................................................... viii 
TABLE 1: SCHEDULE OF STUDY PROCEDURES ................................................... XII 
1 
BACKGROUND INFORMATION AND RATIONALE .......................................... 1 
1.1 
INTRODUCTION ......................................................................................................... 1 
1.2 
NAME AND DESCRIPTION OF INVESTIGATIONAL PRODUCT OR INTERVENTION ......... 1 
1.3 
FINDINGS FROM NON-CLINICAL AND CLINICAL STUDIES ......................................... 2 
1.3.1 Non-Clinical Studies ............................................................................................ 2 
1.3.2 Clinical Studies .................................................................................................... 4 
1.4 
SELECTION OF SUPPLEMENTS AND DOSAGES ............................................................ 5 
1.5 
RELEVANT LITERATURE AND DATA ......................................................................... 6 
1.6 
COMPLIANCE STATEMENT ........................................................................................ 7 
2 
STUDY OBJECTIVES ................................................................................................. 7 
2.1 
PRIMARY OBJECTIVE ................................................................................................ 8 
2.2 
SECONDARY OBJECTIVES ......................................................................................... 8 
3 
INVESTIGATIONAL PLAN ....................................................................................... 8 
3.1 
GENERAL SCHEMA OF STUDY DESIGN ...................................................................... 8 
3.1.1 Screening Phase .................................................................................................. 8 
3.1.2 Visit 1 (may occur on the same day as Screening Visit) ...................................... 8 
3.1.3 Interim between Visit 1 and Visit 2 ...................................................................... 9 
3.1.4 Visit 2 ................................................................................................................... 9 
3.2 
ALLOCATION TO EXPERIMENTAL GROUPS AND BLINDING ....................................... 9 
3.3 
STUDY DURATION, ENROLLMENT AND NUMBER OF SITES ....................................... 9 
3.3.1 Duration of Study Participation .......................................................................... 9 
3.3.2 Total Number of Study Sites/Total Number of Subjects Projected ...................... 9 
3.4 
STUDY POPULATION ................................................................................................. 9 
3.4.1 Inclusion Criteria ................................................................................................ 9 

  
 
 
 
 
 
 
 
iv
3.4.2 Exclusion Criteria .............................................................................................. 10 
4 
STUDY PROCEDURES ............................................................................................. 11 
4.1 
SCREENING VISIT.................................................................................................... 11 
4.2 
STUDY SUPPLEMENTATION PHASE ......................................................................... 11 
4.2.1 Visit 1 (may be the same day as screening visit) ............................................... 11 
4.2.2 Interim between Visit 1 and Visit 2 .................................................................... 12 
4.2.3 Visit 2 ................................................................................................................. 12 
4.3 
CONCOMITANT MEDICATION .................................................................................. 12 
4.4 
RESCUE MEDICATION ADMINISTRATION ................................................................ 13 
4.5 
SUBJECT COMPLETION/WITHDRAWAL .................................................................... 13 
5 
STUDY EVALUATIONS AND MEASUREMENTS .............................................. 13 
5.1 
SCREENING VISIT MEASUREMENTS ........................................................................ 13 
5.1.1 Inclusion/Exclusion Criteria Review ................................................................. 13 
5.2 
VISIT 1 MEASUREMENTS ........................................................................................ 14 
5.2.1 Demographic/Medical History Review ............................................................. 14 
5.2.2 Physical Examination ........................................................................................ 14 
5.2.3 Baseline Symptom Questionnaire (Appendix 2) ................................................ 15 
5.2.4 Laboratory Testing ............................................................................................ 15 
5.3 
INTERIM EVALUATIONS .......................................................................................... 15 
5.4 
VISIT 2 MEASUREMENTS ........................................................................................ 15 
5.4.1 Physical Examination ........................................................................................ 15 
5.4.2 Laboratory Testing ............................................................................................ 15 
5.4.3 Tolerability Questionnaire (Appendix 3) ........................................................... 16 
5.4.4 Study Period Data Review ................................................................................. 16 
6 
STATISTICAL CONSIDERATIONS ....................................................................... 16 
6.1 
PRIMARY ENDPOINT ............................................................................................... 16 
6.2 
SECONDARY ENDPOINTS......................................................................................... 17 
6.3 
CONTROL OF BIAS AND CONFOUNDING .................................................................. 17 
6.4 
STATISTICAL METHODS .......................................................................................... 17 
6.5 
SAMPLE SIZE AND POWER ...................................................................................... 17 

  
 
 
 
 
 
 
 
v
7 
STUDY SUPPLEMENT ............................................................................................. 18 
7.1 
DESCRIPTION .......................................................................................................... 18 
7.1.1 Dosing ................................................................................................................ 18 
7.1.2 Supplement Accountability ................................................................................ 18 
8 
SAFETY MANAGEMENT ........................................................................................ 19 
8.1 
CLINICAL ADVERSE EVENTS .................................................................................. 19 
8.2 
ADVERSE EVENT REPORTING ................................................................................. 19 
8.3 
DEFINITION OF AN ADVERSE EVENT ....................................................................... 19 
8.4 
DEFINITION OF A SERIOUS ADVERSE EVENT (SAE) ................................................ 19 
8.4.1 Relationship of SAE to study drug or other intervention ................................... 20 
8.5 
IRB/IEC NOTIFICATION OF SAES AND OTHER UNANTICIPATED PROBLEMS .......... 20 
8.5.1 Follow-up report ................................................................................................ 21 
8.6 
MEDICAL EMERGENCIES ......................................................................................... 21 
9 
STUDY ADMINISTRATION .................................................................................... 21 
9.1 
DATA COLLECTION AND MANAGEMENT ................................................................. 21 
9.2 
CONFIDENTIALITY .................................................................................................. 21 
9.3 
REGULATORY AND ETHICAL CONSIDERATIONS ...................................................... 22 
9.3.1 Data and Safety Monitoring Plan ...................................................................... 22 
9.3.2 Risk Assessment ................................................................................................. 22 
9.3.3 Potential Benefits of Study Participation .......................................................... 22 
9.3.4 Risk-Benefit Assessment .................................................................................... 22 
9.4 
RECRUITMENT STRATEGY ...................................................................................... 22 
9.5 
INFORMED CONSENT/ASSENT AND HIPAA AUTHORIZATION ................................. 23 
9.6 
PAYMENT TO SUBJECTS/FAMILIES .......................................................................... 23 
9.6.1 Payments to the family for time and inconvenience, parking (i.e. compensation)
 
23 
9.6.2 Payment to the family for transportation and lodging. ..................................... 23 
10 REFERENCES ............................................................................................................ 25 
11 APPENDIX 1 ................................................................................................................ 27 

  
 
 
 
 
 
 
 
vi
12 APPENDIX 2 ................................................................................................................ 28 
13 APPENDIX 3 ................................................................................................................ 32 
14 APPENDIX 4 ................................................................................................................ 35 
 
ABBREVIATIONS AND DEFINITIONS OF TERMS 
 
 
 
 
 
 
AE 
ATP 
CHOP 
CHPS 
CRF 
EGCG 
ECG 
GDH 
GIR 
GTP 
HEK 
HI 
HI/HA 
IU 
OPTT 
SAE 
 
Adverse event 
Adenosine triphosphate 
Children’s Hospital of Philadelphia 
Center for Human Phenomic Science 
Case report form 
Epigallocatechin gallate 
Epicatechin gallate 
Glutamate dehydrogenase 
Glucose infusion rate 
Guanosine triphosphate 
Human embryonic kidney 
Hyperinsulinism 
Hyperinsulinism/Hyperammonemia 
International units 
Oral protein tolerance test 
Serious adverse event 

 
 
 
 
 
 
 
 
 
 
vii
ABSTRACT 
Context:  Congenital hyperinsulinism (HI) is a rare disorder of pancreatic beta cell insulin 
secretion that causes persistent and severe hypoglycemia starting at birth. 
Hyperinsulinism/hyperammonemia (HI/HA) syndrome is the second most common type of 
congenital HI and is caused by activating mutations in glutamate dehydrogenase (GDH). 
Patients with HI/HA exhibit fasting hyperinsulinemic hypoglycemia, protein-induced 
hypoglycemia, hyperammonemia, seizures, and intellectual disability independent of 
hypoglycemia. These effects result from abnormal GDH activity in the beta cells, liver and 
kidney cells, neurons, and astrocytes. The only available treatment for HI/HA syndrome is 
diazoxide, which acts on the beta cells to decrease insulin secretion but has no effect on 
GDH activity itself or on other cell types. Thus, there remains a significant unmet need for 
improved therapies for this disorder. Preliminary data show that Vitamin E inhibits GDH 
activity in cell lines and improves hypoglycemia in a GDH HI mouse model. Therefore, we 
hypothesize that Vitamin E will inhibit GDH activity, as measured by plasma ammonia 
levels and parameters of the oral protein tolerance test (OPTT), in subjects with HI/HA 
syndrome. This hypothesis will be tested in a future study. The intent of the current pilot 
study is to use Vitamin E as a supplement to obtain tolerability and feasibility data, and not 
to cure, prevent, treat, mitigate, or diagnose HI. 
 
Objectives:  The primary objective of this study is to determine the tolerability of short-term 
oral Vitamin E supplementation in subjects with HI/HA syndrome. Secondary objectives are 
to assess the effects of Vitamin E supplements on plasma Vitamin E levels, on plasma 
glucose, insulin, and ammonia concentrations during fasting oral protein tolerance test 
(OPTT), and on hypoglycemia episodes in subjects with HI/HA syndrome. 
Study Design:  This open-label pilot and feasibility clinical study will use a before-and-after 
design, with assessments performed prior to and after 2 weeks of daily oral Vitamin E 
supplementation in individuals with HI/HA syndrome. 
Setting/Participants:  This single-site study will recruit 14 participants from 1 to 40 years of 
age (inclusive) with HI/HA syndrome from CHOP’s Congenital Hyperinsulinism Center. 
Study procedures will be performed in the outpatient Center for Human Phenomic Science. 
 
 

 
 
 
 
 
 
 
 
 
 
viii
PROTOCOL SYNOPSIS 
 
Study Title 
Vitamin E Supplementation in Hyperinsulinism/Hyperammonemia 
Syndrome. 
Funder 
University of Pennsylvania Orphan Disease Center, 
Pediatric Endocrine Society, 
Children’s Hospital of Philadelphia 
Clinical Phase 
Pilot Study 
Study Rationale 
Congenital hyperinsulinism (HI) is a rare genetic disorder of 
dysregulated pancreatic beta cell insulin secretion that causes 
persistent and severe hypoglycemia starting at birth, which can 
result in developmental delays, brain damage, and death if 
inadequately controlled. The second most common type of 
congenital HI is caused by dominant activating mutations in 
glutamate dehydrogenase (GDH) that result in 
hyperinsulinism/hyperammonemia (HI/HA) syndrome. In addition 
to fasting hyperinsulinemic hypoglycemia, which is seen in all 
patients with HI, patients with HI/HA also exhibit protein-induced 
hypoglycemia, hyperammonemia, generalized seizures, and 
intellectual disability independent of hypoglycemia. These effects 
result from abnormal GDH activity in the insulin-secreting beta 
cells, liver and kidney cells, neurons, and astrocytes. Currently, the 
only treatment for HI/HA syndrome is diazoxide, which acts on beta 
cells to decrease insulin secretion. However, diazoxide has no effect 
on GDH activity itself or on other cell types. Diazoxide also has 
side effects of fluid retention and excessive hair growth. Thus, there 
remains a significant unmet need for improved therapies for patients 
with HI/HA syndrome. Targeting GDH activity in all cells would be 
a more effective approach to treat HI/HA syndrome. 
Our preliminary data show that alpha-tocopherol (Vitamin E) 
effectively inhibits GDH activity in human embryonic kidney cells 
with an activating GDH mutation, and it improves hypoglycemia in 
transgenic mice with GDH HI. Based on these preclinical in vitro 
and in vivo studies, we hypothesize that alpha-tocopherol will 
inhibit GDH activity and may impact hyperinsulinemic 
hypoglycemia and hyperammonemia in subjects with HI/HA 
syndrome. This hypothesis will be tested in a future study. In this 
initial pilot study, we propose the following objectives: 
Study Objective(s) 
Primary 

  
 
 
 
 
 
 
 
ix
• To determine the tolerability of supplemental oral alpha-
tocopherol in subjects with HI/HA syndrome. 
Secondary 
• To assess the effects of Vitamin E supplements on plasma 
Vitamin E levels, on plasma glucose, insulin, and ammonia 
concentrations during fasting oral protein tolerance test (OPTT), 
and on hypoglycemic episodes in subjects with HI/HA 
syndrome. 
Test Article 
Vitamin E is a fat-soluble vitamin made exclusively by plants. 
Vitamin E supplements are available over-the-counter in liquid and 
capsule formulations. Vitamin E supplements are widely used and 
have very low risk of intoxication or side effects. 
Study Design 
 
This is an open-label pilot and feasibility clinical study to assess 
tolerability of Vitamin E supplements in subjects with HI/HA 
syndrome. The duration of this study will be two weeks, during 
which subjects take a daily oral Vitamin E supplement and will 
undergo outpatient evaluations at baseline and at study end. 
Subject Population 
key criteria for 
Inclusion and Exclusion: 
Inclusion Criteria 
1. Males or females age ≥1 year and ≤40 years. 
2. Diagnosis of HI/HA syndrome. 
3. Currently being treated with diazoxide. 
4. Females 11 years of age or menstruating must have a 
negative urine/serum pregnancy test and must use an 
acceptable method of contraception, including abstinence, a 
barrier method (diaphragm or condom), Depo-Provera, or an 
oral contraceptive, for the duration of the study. 
5. Informed consent for participants ≥18 years. 
Parental/guardian permission (informed consent) and, if 
appropriate, child assent for participants <18 years. 
Exclusion Criteria 
1. Males or females age <1 year or >40 years. 
2. Individuals who have experienced an allergic reaction to 
Vitamin E. 
3. Individuals with a known allergy to dairy, whey, or soy. 
4. On concurrent therapy with a medication known to 
adversely interact with Vitamin E (refer to Appendix 1). 
5. Known increased risk of bleeding (bleeding disorder or on 
antiplatelet or anticoagulation therapy). 
6. Vitamin E supplementation within 30 days prior to 
enrollment, including multivitamins containing Vitamin E. 

  
 
 
 
 
 
 
 
x
7. Severe hypoglycemia (plasma glucose <50 mg/dL on repeat 
checks using home glucose meter) more than once weekly 
within 30 days prior to enrollment. 
8. Evidence of a medical condition that might alter results or 
compromise the interpretation of results, including active 
infection, kidney failure, severe liver dysfunction, severe 
respiratory or cardiac failure. 
9. Evidence of severe hematologic abnormality including 
severe anemia and/or thrombocytopenia. 
10. Any investigational drug use within 30 days prior to 
enrollment. 
11. Pregnant or lactating females. 
12. Parents/guardians or subjects who, in the opinion of the 
Investigator, may be non-compliant with study schedules or 
procedures. 
13. Unable to provide informed consent (e.g. impaired cognition 
or judgment). 
14. Parents/guardians or subjects with limited English 
proficiency. 
Number Of Subjects 
Fourteen total, with goal of ten evaluable 
Study Duration 
Each subject’s participation will last approximately two weeks. 
The entire study is expected to last one year. 
Study Phases 
 
(1) Screening: Screen for eligibility and obtain consent 
(2) Visit 1: Baseline OPTT and plasma alpha-tocopherol level 
(3) Vitamin E oral supplementation at home for two weeks, with 
two study phone calls 
(4) Visit 2: Post-supplementation laboratory assessment, OPTT (for 
subjects without severe hypoglycemia or hypoglycemic 
symptoms during Visit 1 OPTT), plasma alpha-tocopherol level, 
and tolerability questionnaire 
Outcome Evaluations 
The primary outcome is tolerability of Vitamin E as measured by: 
• the change in fasting plasma alpha-tocopherol concentration 
before versus after alpha-tocopherol supplementation 
• responses to a subject/parent-reported tolerability questionnaire 
after Vitamin E supplementation compared to baseline (refer to 
Appendices 2 and 3) 
The secondary outcomes will assess the effects of Vitamin E 
supplementation on the following measurements: 
• delta-glucose (fasting plasma glucose - nadir plasma glucose 
during OPTT) 

  
 
 
 
 
 
 
 
xi
• fasting plasma glucose 
• nadir plasma glucose 
• fasting plasma insulin 
• peak plasma insulin 
• delta-insulin (peak plasma insulin during OPTT - fasting plasma 
insulin) 
• fasting plasma ammonia 
• delta-ammonia (plasma ammonia at 60 minutes - plasma 
ammonia at 0 minutes) 
• hypoglycemic episodes (plasma glucose <70 mg/dL) detected 
on home glucose meter 
Safety Evaluations 
Subjects will remain under the care of their primary medical team 
and continue to receive their usual clinical care during this study. 
Adverse events will be monitored by the study PI. 
Statistical And Analytic 
Plan 
In this pilot study, paired t-tests will be conducted to evaluate 
differences in the primary and secondary objectives before versus 
after Vitamin E supplementation. 
Descriptive statistics will be used to characterize subject 
demographics. 
DATA AND SAFETY 
MONITORING PLAN 
The study investigators will be responsible for data quality 
management and ongoing assessment of safety. 

 
 
 
 
 
 
 
 
 
 
xii
TABLE 1: SCHEDULE OF STUDY PROCEDURES 
 
Screening 
Visit/Visit 1* 
Telephone 
Call 1 
Telephone 
Call 2 
Visit 2 
 
Day 0 
Day 4 
(± 1 day) 
Day 8 
(± 1 day) 
Day 14 
(± 1 day) 
Informed consent 
X 
 
 
 
Review inclusion/exclusion criteria 
X 
 
 
 
Demographics/medical history review 
X 
 
 
 
Home glucose meter review 
X 
 
 
X 
Physical exam 
X 
 
 
X 
Fasting laboratory studies 
X 
 
 
X 
Oral protein tolerance test 
X 
 
 
X† 
Tolerability questionnaire 
 
X 
X 
X 
Prior/concomitant medication review 
 
 
 
X 
Adverse event monitoring 
X 
 
* Vitamin E supplements will be dispensed to parent/caregiver or subject during Visit 1. 
† Visit 2 oral protein tolerance test will not be performed for subjects who experience severe 
hypoglycemia or hypoglycemic symptoms during Visit 1 oral protein tolerance test. 
Participants will be asked to partake in a 15 minute phone interview with study staff, during 
which the tolerability questionnaire will be administered, on Day 4 (± 1 day) and Day 8 (± 1 
day), between study visits.

 
  
 
 
 
1
1 
BACKGROUND INFORMATION AND RATIONALE 
1.1 
Introduction 
Congenital hyperinsulinism (HI) is the most common cause of permanent neonatal 
hypoglycemia. It is a rare disease and occurs in approximately 1 in 40,000 individuals. This 
disease results from various mutations that affect the secretion of insulin from the beta cell 
of the pancreas. Gain of function mutations in the glutamic dehydrogenase (GDH) enzyme 
result in the second most common form of congenital HI, also known as 
hyperinsulinism/hyperammonemia (HI/HA) syndrome (1-3). 
In addition to fasting hyperinsulinemic hypoglycemia, which is observed in all patients with 
congenital HI (4), patients with HI/HA syndrome also exhibit protein-induced 
hypoglycemia, hyperammonemia, and epilepsy (1). The severity of the hypoglycemia is 
greatest in younger children, with a milder phenotype observed in adults. The only current 
mode of treatment for hypoglycemia caused by GDH HI is diazoxide, which works at the 
level of the beta cell to inhibit insulin secretion. However, the other consequences of GDH 
activity (due to gain-of-function mutations in the kidneys, liver, and brain) including 
hyperammonemia, epilepsy, and intellectual disability persist and remain untreated. 
Furthermore, diazoxide causes side effects including fluid retention, pulmonary 
hypertension, heart failure, and neutropenia (5; 6) that may be avoided by use of a GDH-
specific therapy. 
There has been an ongoing search for a multisystem therapy for HI/HA syndrome that 
targets the GDH enzyme. Recently, we identified that Vitamin E is an effective GDH 
inhibitor (see Section 1.3.1). Furthermore, in a mouse model of GDH HI, Vitamin E (alpha-
tocopherol) oral administration improved fasting hypoglycemia compared to mice who did 
not receive Vitamin E. 
We hypothesize that Vitamin E will be a feasible, targeted, multisystem treatment for HI/HA 
syndrome due to its inhibition of GDH activity in all cells. In this pilot tolerability and 
feasibility study, we will evaluate plasma alpha-tocopherol levels and adverse events, in 
addition to plasma ammonia, glucose, and insulin levels during an oral protein tolerance test 
(OPTT) as initial, short-term, measurable effects of Vitamin E supplementation, while the 
subjects remain on their usual diazoxide treatment regimen. These initial tolerability 
measures will inform future studies addressing clinical efficacy of Vitamin E alone in 
treating the hyperinsulinemic hypoglycemia, hyperammonemia, and neurological 
manifestations of HI/HA syndrome. 
 
1.2 
Name and Description of Investigational Product or Intervention 
Vitamin E is an essential nutrient only synthesized by plants. Over-the-counter Vitamin E 
supplements are commonly used for the anti-oxidant properties and have very few side 
effects. In this study, Vitamin E will be administered orally in either liquid (50 IU/mL, 
aqueous) or capsule (200 IU) formulation, depending on subject age and preference. 
 

 
  
 
 
 
2
1.3 
Findings from Non-Clinical and Clinical Studies 
1.3.1 Non-Clinical Studies 
Previous studies from our group identified the green tea extracts epigallocatechin gallate 
(EGCG) and epicatechin gallate (ECG) (Figure 1) as potent allosteric inhibitors of wild-
type and mutant GDH activity in vitro (7) (Figure 2B), as well as in transgenic mice (8). 
However, these compounds have very poor intestinal absorption, low bioavailability, and are 
rapidly metabolized, making them unsuitable therapeutic candidates. To identify other 
potential candidates, our group conducted several in vitro chemical screens (9) and found 
several small molecules that effectively inhibited wild-type and mutant GDH activity in 
vitro with low toxicity profiles (unpublished data). Some of these lead compounds have a 
similar structure to alpha-tocopherol, the bioavailable form of Vitamin E (Figure 1). 
Therefore, we hypothesize that Vitamin E inhibits mutant GDH activity.  
 
We first evaluated alpha-tocopherol’s effect on GDH activity in vitro. In this established 
model, human embryonic kidney (HEK293T) cells were transduced with lentivirus to over-
express either wild-type GDH or a common mutant form of GDH, H454Y, in which the 
454th amino acid is changed from histidine to tyrosine. This mutation has been described in 
many patients with HI/HA syndrome (3) and has been shown to prevent enzymatic 
inhibition by guanosine triphosphate (GTP) in cells isolated from these patients, as well as in 

 
  
 
 
 
3
cell lines over-expressing this GDH mutant (Figure 2A). We found that alpha-tocopherol 
effectively inhibited activity of wild-type and H454Y mutant GDH in these human cells 
(Figure 2C), with 50% inhibitory concentration (IC50) of 4.1 and 3.1 μM, respectively. 
Alpha-tocopherol also effectively inhibited the activity of other GDH mutants previously 
identified in patients with HI/HA syndrome, with IC50 <10 μM (Figure 3A), including 
mutations that affect the GTP binding site (Figure 3B) and those that do not (Figure 3C). 
We also evaluated alpha-tocopherol’s effect in transgenic mice expressing the H454Y 
mutant GDH in beta cells, which causes fasting hypoglycemia (8) similar to patients with 
HI/HA syndrome. We treated these transgenic mice with oral alpha-tocopherol for 2 days 
and then assessed fasting hypoglycemia, which was significantly improved in alpha-
tocopherol-treated transgenic mice (nadir plasma glucose 86 ± 4 mg/dL) versus vehicle-
treated transgenic mice (nadir plasma glucose 67 ± 5 mg/dL, P <0.01, Figure 4). These 
preclinical in vitro and in vivo studies suggest that alpha-tocopherol is a promising treatment 
for HI/HA syndrome via direct GDH inhibition. 
 

 
  
 
 
 
4
1.3.2 Clinical Studies 
1.3.2.1 Human Vitamin E Plasma Concentrations 
Vitamin E is a fat-soluble antioxidant that protects cell membranes from oxidative damage. 
In vitro studies of Vitamin E have also demonstrated a role in immune and endothelial cell 
function as well as inhibition of platelet aggregation (10).  Prior national surveys have found 
that approximately 90% of children and adults in the United States have inadequate Vitamin 
E dietary intake (11) and Vitamin E insufficiency based on plasma alpha-tocopherol 
concentration <30 µM, while only 1% are truly deficient based on plasma concentration <12 
µM (12). Vitamin E deficiency typically causes neurological deficits, which can be 
ameliorated by Vitamin E supplementation because alpha-tocopherol crosses the blood-brain 
barrier (13). However, clinically apparent Vitamin E deficiency is rare and has not been 
reported to occur in healthy individuals.  Based upon results of the 2005-2006 National 
Health and Nutrition Examination Survey, the mean serum Vitamin E concentration in 
individuals age 6 and older was 25.3 µM (14). 
1.3.2.2 Pharmacokinetic Data 
Pharmacokinetic studies in adults administered 800 mg all racemic alpha-tocopherol found a 
mean absorption time of 3 hours and mean elimination half-life of 73 hours (15).  Maximum 
plasma levels occurred at 12-13 hours.  Pharmacokinetic studies in healthy children are 
lacking.  Serum levels achieved after varying doses of Vitamin E administration in pediatric 
studies are detailed in Section 1.3.2.4. 
1.3.2.3 Clinical Studies in Adults 
Adults supplemented with 400 or 800 IU/day for 4 weeks demonstrated increased plasma 
alpha-tocopherol concentrations from 26.8±2.6 to 41.8±4.3 and from 25.2±2.4 to 53.5±3.86 
µM, respectively (16). A meta-analysis of randomized studies in adults showed an 
association between alpha-tocopherol doses >400 IU/day and all-cause mortality, which was 
not observed with lower doses (17). Furthermore, a prospective randomized placebo-
controlled study of 14,641 men >50 years old given 400 IU Vitamin E every other day for 
10 years showed increased risk of hemorrhagic stroke (HR, 1.74 [95% CI, 1.04-2.91]; 
P=.04) (18). In vitro and rodent studies indicate that alpha-tocopherol inhibits platelet 
aggregation (19). Although higher plasma alpha-tocopherol levels were associated with 
increased risk of bleeding in patients on anticoagulation therapy (20), healthy adults 
supplemented with alpha-tocopherol 800 IU/day for 2 weeks had no differences in platelet 
aggregation, coagulation profile, or bleeding time (21). No other specific side effects or risks 
have been associated with Vitamin E supplementation. 
1.3.2.4 Clinical Studies in Children 
Most children’s multivitamins contain 18-30 IU of Vitamin E. Higher supplementation 
doses are commonly used for children and adults with malabsorption disorders, including 
cystic fibrosis. Vitamin E oral supplementation in children has been evaluated in multiple 
studies, including: 
• a retrospective study of 232 children ages 0-17 years with cystic fibrosis chronically 
supplemented with age-based doses of alpha-tocopherol ranging from approximately 
47.5-137.2 mg daily (equivalent to approximately 70-200 IU daily), median serum alpha 
-tocopherol levels ranged between 18 and 25 mmol/l (775 and 1079 mg/dL) (22) 

 
  
 
 
 
5
• a randomized double-blind placebo-controlled study in which 120 children ages 5-17 
years with moderate persistent asthma were supplemented with alpha-tocopherol 50 mg 
daily (equivalent to 75 IU daily) for 8 weeks, pre-treatment serum vitamin E levels were 
4.1±1.41 μg/dL and 4.5±1.8 μg/dL in treated and untreated groups, respectively, with 
post-treatment levels of 8.2±2.4 μg/dL and 4.7±0.9 μg/dL (p=0.012), in the treated and 
untreated groups, respectively (23) 
• a randomized placebo-controlled study in which 49 children ages 7-15 years with Down 
syndrome were supplemented with alpha-tocopherol 400 IU daily for 4 months (24) 
• a randomized double-blind placebo-controlled study in which 76 female children ages 5-
12 years with pyelonephritis were supplemented with alpha-tocopherol 100 IU daily for 
2 weeks (25) 
• an open-label study of 9 children ages 8-15 years with type 1 diabetes mellitus were 
supplemented with weight-based doses of alpha-tocopherol (100 IU daily if <30 kg, 200 
IU daily if 30-60 kg, 300 IU daily if >60 kg) (26) 
• an open-label study of 18 children and adults ages 0.6-42 years with glycogen storage 
disease 1b (GSD1b) supplemented with Vitamin E either 600 mg (800 IU) daily if pre-
pubertal or 900 mg (1,350 IU) daily if post-pubertal for 1 year (27) 
These studies reported no adverse events in subjects receiving Vitamin E supplements. It is 
reassuring that Vitamin E has been given to children with disorders of glucose regulation 
(hyperglycemia in type 1 diabetes, and hypoglycemia in GSD1b). In summary, alpha-
tocopherol supplementation is generally considered safe in children and adults without 
increased bleeding risk. Based upon a literature review, no clinical studies of Vitamin E 
supplementation in subjects with HI/HA syndrome have yet been conducted. There are no 
other ongoing studies at CHOP targeting this patient population. 
1.4 
Selection of Supplements and Dosages 
Oral Vitamin E supplements to be used in this study 
will be provided by the CHOP Pharmacy, the 
formulations of which are on the CHOP formulary 
for inpatient and outpatient administration. Age-
based oral Vitamin E supplement dose ranges 
established by the Food and Nutrition Board at the 
Institute of Medicine (Table 2) were used to select 
the age-based doses that will be used in this study 
(Table 3). Liquid and capsule formulations were 
chosen to allow for age-based dosing and subject 
preference. Study duration of 2 weeks was 
determined based on the elimination half-life of 
Vitamin E of 73 hours; five half-lives will allow for 
evaluation at steady-state plasma levels of the study 
supplement. 
 
 
Age 
RDA (IU) 
UL (IU) 
0-6 months 
6 
nd 
7-12 months 
7.5 
nd 
1-3 years 
9 
300 
4-8 years 
10.4 
450 
9-13 years 
16.4 
900 
14-18 years 
22.4 
1,200 
19+ years 
22.4 
1,500 
Table 2. Recommended Dietary 
Allowance 
(RDA) 
and 
Tolerable 
Upper Intake Level (UL) of d-alpha-
tocopherol for different age groups. 

 
  
 
 
 
6
 
1.5 
Relevant Literature and Data 
HI/HA syndrome is the second most common cause of congenital HI and the most common 
cause of diazoxide-responsive congenital HI (1; 2). Patients with HI/HA syndrome have 
several clinical characteristics: 
Fasting hyperinsulinemic hypoglycemia in patients with HI/HA syndrome is relatively 
mild compared to other forms of congenital HI. Average fasting tolerance without 
hypoglycemia in patients with HI/HA syndrome is approximately 12 hours and improves 
with age (3). Diazoxide treatment effectively prevents significant fasting hypoglycemia in 
HI/HA syndrome (1). 
Protein-induced hyperinsulinemic hypoglycemia is acute and profound in patients with 
HI/HA syndrome, with decrease of plasma glucose concentration by 9-62 mg/dL (mean 23 
mg/dL) within 3 hours of an oral protein load (1.5 g/kg body weight), without carbohydrate 
intake (3). This decline in plasma glucose results in significant hypoglycemia in most 
patients with HI/HA syndrome, compared to controls who only experience a decrease in 
plasma glucose of 4 mg/dL. Patients with HI/HA syndrome can avoid protein-induced 
hypoglycemia by consuming carbohydrates with protein-containing meals. Diazoxide 
treatment is moderately effective in preventing significant protein-induced hypoglycemia. 
Hyperammonemia with plasma ammonia levels typically 2-5 times the upper limit of 
normal (35 μmol/L) occurs in patients with HI/HA syndrome (3). However, there is no clear 
correlation between plasma ammonia and plasma glucose levels, likely because the primary 
source of elevated plasma ammonia is the kidney, rather than the beta cell (28). Diazoxide 
treatment does not affect plasma ammonia levels. 
Central nervous system (CNS) effects, including seizures and developmental delays, are 
seen in patients with HI/HA syndrome, although the underlying mechanisms are not 
understood (1). While hyperinsulinemic hypoglycemia likely contributes to some degree of 
Age 
Daily Dose 
(mg) Active 
Form 
Equivalent 
Daily Dose (IU) 
Natural Form 
Equivalent 
Daily Dose (IU) 
Synthetic Form 
Liquid Dose 
(50 IU/mL) 
Natural 
Liquid Dose 
(50 IU/ mL) 
Synthetic 
Capsule Dose 
(200 IU/capsule) 
Natural 
Capsule Dose  
(200 IU/capsule) 
Synthetic 
1-3 years  67 
 100 
 148.7 IU 
– round to 150 
IU 
2 mL 
 3 mL 
-- 
 -- 
4-8 years  134 
 200 
 297 IU 
– round to 300 
IU 
4 mL 
 6 mL 
-- 
 -- 
9-17 years  201 
 300 
 446.2 IU 
– round to 450 
IU 
6 mL 
 9 mL 
-- 
 -- 
18+ years  268 
 400 
 594.9 IU 
– round to 600 
IU 
8 mL 
 12 mL 
2 capsules 
 3 capsules 
Table 3.  Alpha-tocopherol daily doses to be used in this study.  1 mg of alpha-tocopherol active form is 
equivalent to 1.49 IU of natural form (d-alpha-tocopherol) or 2.22 IU of synthetic form (dl-alpha-tocopherol). 

 
  
 
 
 
7
neurodevelopmental deficits, as is seen in other forms of congenital HI, seizures 
(particularly absence seizures) are observed in patients with HI/HA syndrome even when 
plasma glucose is normal. In comparison to patients with other genetic causes of 
hyperammonemia, such as urea cycle defects, CNS effects in patients with HI/HA syndrome 
are relatively mild. It is likely that these outcomes are consequences of GDH hyperactivity 
in the CNS, rather than solely due to hyperammonemia. However, the direct effects of GDH 
hyperactivity in the CNS are not understood. 
The only current treatment available for HI/HA syndrome is diazoxide, which opens the 
ATP-gated potassium (KATP) channels on beta cells to reduce insulin secretion. Diazoxide 
effectively prevents fasting and protein-induced hypoglycemia, but it has no effect on 
hyperammonemia, seizures, or developmental delays (1). Furthermore, diazoxide causes 
significant side effects, including fluid retention, pulmonary hypertension, heart failure, and 
neutropenia (5; 6). Thus, an alternative therapy for HI/HA syndrome, directly targeting 
GDH in all cell types to treat the disease’s widespread pathophysiology, is warranted. 
Glutamate Dehydrogenase (GDH) Activating Mutations Cause HI/HA Syndrome 
GDH is a mitochondrial matrix enzyme that is highly expressed in kidney, liver, brain, and 
beta cells (1). It is responsible for metabolizing glutamate into alpha-ketoglutarate and 
ammonia. In beta cells, elevated alpha-ketoglutarate leads to increased ATP production via 
the citric acid cycle, which stimulates insulin secretion. GDH activity is normally 
suppressed by binding of guanosine triphosphate (GTP), which alters conformation of the 
catalytic domain and prevents conversion of glutamate into alpha-ketoglutarate and 
ammonia (1). Dominant gain-of-function missense mutations in GDH interfere with GTP 
binding, impairing GTP-mediated inhibition and thus increasing GDH catalytic activity, 
leading to excess amino acid catabolism and ammonia production, which results in HI/HA 
syndrome. 
1.6 
Compliance Statement 
This study will be conducted in full accordance of all applicable Children’s Hospital of 
Philadelphia Research Policies and Procedures and all applicable federal and state laws and 
regulations including 45 CFR 46 and HIPPA. All episodes of noncompliance will be 
documented. 
The investigators will perform the study in accordance with this protocol, will obtain 
consent and assent, and will report unanticipated problems involving risks to subjects or 
others in accordance with the Children’s Hospital of Philadelphia Institutional Review 
Board Policies and Procedures and all federal requirements. Collection, recording, and 
reporting of data will be accurate and will ensure the privacy, health, and welfare of research 
subjects during and after the study. 
 
2 
STUDY OBJECTIVES 
The purpose of this pilot clinical study is to determine the feasibility and tolerability of 
alpha-tocopherol supplementation in subjects with HI/HA syndrome. 

 
  
 
 
 
8
2.1 
Primary Objective 
To determine the tolerability of supplemental oral alpha-tocopherol in subjects with HI/HA 
syndrome as measured by responses to a subject/parent-reported tolerability questionnaire 
compared to baseline and the change in fasting plasma alpha-tocopherol concentration 
before versus after alpha-tocopherol supplementation. 
2.2 
Secondary Objectives 
To assess the effects of Vitamin E supplements on plasma Vitamin E levels, on plasma 
glucose, insulin, and ammonia concentrations during fasting oral protein tolerance test 
(OPTT), and on hypoglycemic episodes in subjects with HI/HA syndrome. 
 
3 
INVESTIGATIONAL PLAN 
3.1 
General Schema of Study Design 
This is a pilot and feasibility before-and-after study. Enrolled subjects will undergo lab 
draws at CHOP’s outpatient Center for Human Phenomic Science (CHPS) prior to and after 
two weeks of daily oral Vitamin E supplementation. Fasting oral protein tolerance test 
(OPTT) will be performed prior to Vitamin E supplementation for all subjects and will be 
repeated after Vitamin E supplementation for subjects who do not who experience severe 
hypoglycemia, defined as in section 4.4, or severe hypoglycemic symptoms, including 
seizure or altered mental status, during the initial oral protein tolerance test. For each OPTT, 
the subjects will fast for 12 hours overnight at home, then will have fasting plasma glucose, 
insulin, alpha-tocopherol, and ammonia concentrations measured. Then the subjects will be 
given a weight-based oral protein drink, and subsequent plasma glucose, insulin, and 
ammonia concentrations will be measured. Subjects will continue diazoxide and other usual 
medications throughout the study period. 
3.1.1 Screening Phase 
• Subjects with HI/HA syndrome currently treated with diazoxide will be recruited 
from the Congenital Hyperinsulinism Center at the Children’s Hospital of 
Philadelphia (CHOP). 
• Informed consent and assent (for subjects ≥7 years of age) will be obtained. 
3.1.2 Visit 1 (may occur on the same day as Screening Visit) 
• Subject demographic and medical history will be reviewed, including prior and 
concomitant medications and supplements, and recent hypoglycemic events. 
• A physical exam will be performed. 
• Complete blood count with platelet count, fasting plasma glucose, insulin, alpha-
tocopherol, and ammonia concentrations will be measured. 
• Oral protein tolerance test will be performed with plasma glucose, insulin, and 
ammonia concentration measurements. 
• Subjects/caregivers will be given the Vitamin E supplements, with instructions for 
administration over the next two weeks. 

 
  
 
 
 
9
• Subject/parent will complete the baseline symptom questionnaire (Appendix 2). 
3.1.3 Interim between Visit 1 and Visit 2 
• 
Subjects will self-administer daily oral Vitamin E supplementation with a fat-
containing meal at home. 
• 
Phone interview with study staff on Day 4 (± 1 day) and Day 8 (± 1 day) to ensure 
study compliance, inquire about adverse events, and administer a verbal 
tolerability questionnaire. Study staff will use the script provided in Appendix 4, 
which includes administration of the tolerability questionnaire (Appendix 3). 
3.1.4 Visit 2 
• Recent medical history, hypoglycemic events, and concomitant medications and 
supplements will be reviewed. 
• A physical exam will be performed. 
• Fasting plasma glucose, insulin, alpha-tocopherol, and ammonia concentrations will 
be measured. 
• Oral protein tolerance test will be performed with plasma glucose, insulin, and 
ammonia concentration measurements for subjects who do not experience severe 
hypoglycemia or hypoglycemic symptoms during Visit 1 OPTT. 
• Subject/parent will complete the Vitamin E tolerability questionnaire (Appendix 3). 
• Adverse events will be recorded. 
3.2 
Allocation to Experimental Groups and Blinding 
All recruited subjects will be exposed to Vitamin E supplementation. Post-supplementation 
data will be compared to baseline data. There will be no blinding. 
3.3 
Study Duration, Enrollment and Number of Sites 
3.3.1 Duration of Study Participation 
The study duration per subject will be up to 15 days. Based upon the half-life of Vitamin E 
(approximately 73 hours), steady-state plasma levels are expected to be reached and 
maximal biological effect to occur within this timeframe. 
3.3.2 Total Number of Study Sites/Total Number of Subjects Projected 
The study will be conducted at the Children’s Hospital of Philadelphia.  Recruitment will 
stop when approximately 14 subjects are enrolled. It is expected that approximately 14 
subjects will be enrolled to produce 10 evaluable subjects. 
3.4 
Study Population 
3.4.1 Inclusion Criteria 
1) Males or females age ≥1 year and ≤40 years. 
2) Diagnosis of HI/HA syndrome. 
3) Currently being treated with diazoxide. 

 
  
 
 
 
10
4) Females 11 years of age or menstruating must have a negative urine/serum pregnancy 
test and must use an acceptable method of contraception, including abstinence, a barrier 
method (diaphragm or condom), Depo-Provera, or an oral contraceptive, for the duration 
of the study. 
5) Informed consent for participants ≥18 years. Parental/guardian permission (informed 
consent) and, if appropriate, child assent for participants <18 years. 
3.4.2 Exclusion Criteria 
1) Males or females age <1 year or >40 years. 
2) Individuals who have experienced an allergic reaction to Vitamin E. 
3) Individuals with a known allergy to dairy, whey, or soy. 
4) On concurrent therapy with a medication known to adversely interact with Vitamin E 
(refer to Appendix 1). 
5) Known increased risk of bleeding (bleeding disorder or on antiplatelet or anticoagulation 
therapy). 
6) Vitamin E supplementation within 30 days prior to enrollment, including multivitamins 
containing Vitamin E. 
7) Severe hypoglycemia (plasma glucose <50 mg/dL on repeat checks using home glucose 
meter) more than once weekly within 30 days prior to enrollment. 
8) Evidence of a medical condition that might alter results or compromise the interpretation 
of results, including active infection, kidney failure, severe liver dysfunction, severe 
respiratory or cardiac failure. 
9) Evidence of severe hematologic abnormality including severe anemia and/or 
thrombocytopenia. 
10) Any investigational drug use within 30 days prior to enrollment. 
11) Pregnant or lactating females. 
12) Parents/guardians or subjects who, in the opinion of the Investigator, may be non-
compliant with study schedules or procedures. 
13) Unable to provide informed consent (e.g. impaired cognition or judgment). 
14) Parents/guardians or subjects with limited English proficiency. 
Subjects that do not meet all of the enrollment criteria may not be enrolled.  Subjects who 
have been screened and are later determined to be eligible can participate without having to 
be rescreened for a period of one year. The study staff will obtain verbal confirmation that 
there have been no health events that would make them ineligible if the interval between 
screening and participation is longer than 3 months. Any violations of these criteria must be 
reported in accordance with IRB Policies and Procedures. 
 

 
  
 
 
 
11
4 
STUDY PROCEDURES 
All subjects will undergo an oral protein tolerance test (OPTT) at Visit 1. Subjects who do 
not experience severe hypoglycemia, defined as in section 4.4, or severe hypoglycemic 
symptoms, including seizure or altered mental status, during the Visit 1 OPTT will also 
undergo OPTT at Visit 2. OPTT is a standard test that patients with HI/HA undergo 
annually to evaluate efficacy of diazoxide treatment. 
4.1 
Screening Visit 
• Review of inclusion/exclusion criteria 
• Informed consent, assent 
• Physical exam 
• Medical record review 
• Recent/current medication and supplement review 
• Home glucose meter review 
• Urine pregnancy test for females who are menstruating or ≥11 years old 
4.2 
Study Supplementation Phase 
The supplementation period will span approximately 2 weeks (14 days ± 1 day) from Visit 1 
to Visit 2. 
4.2.1 Visit 1 (may be the same day as screening visit) 
• Demographic/medical history review 
• Physical exam 
• Prior/concomitant medication and supplement review 
• Home glucose meter review 
• Subject/parent completion of the baseline symptom questionnaire (Appendix 2). 
• Placement of peripheral intravenous (IV) line 
• Fasting baseline plasma glucose, insulin, alpha-tocopherol, and ammonia 
concentrations, and complete blood count with platelet count drawn from IV line.  
Ammonia sample will be drawn from free-flowing IV line without tourniquet and 
placed immediately on ice. 
• Oral protein tolerance test: 
o Enteral intake of Beneprotein (1.5 g/kg body weight protein, maximum 60 
grams protein = 1.75g/kg body weight protein powder, maximum 70 grams 
powder) mixed with carbohydrate-free Crystal Light in 6 ounces of water. 
o Plasma glucose and insulin concentrations obtained from IV line will be 
measured at 15, 30, 45, 60, 90, 120, 150, and 180 minutes. 
o Plasma ammonia will also be measured at 60 minutes from free-flowing IV 
line without tourniquet, and sample will be placed immediately on ice. 
o Plasma glucose measurements will be repeated as needed (if <70 mg/dL). 

 
  
 
 
 
12
• Dispensing of Vitamin E oral supplement with instructions for administration over 
the next two weeks and instructions to not take any other Vitamin E-containing 
supplements, including multivitamins containing Vitamin E, during the study period. 
4.2.2 Interim between Visit 1 and Visit 2 
• 
Daily Vitamin E oral supplementation taken with a fat-containing meal 
• 
Phone interview with study staff on Day 4 (± 1 day) and Day 8 (± 1 day) to ensure 
study compliance, inquire about adverse events, and administer a verbal 
tolerability questionnaire. Study staff will use the script provided in Appendix 4, 
which includes administration of the tolerability questionnaire. 
4.2.3 Visit 2 
• Physical exam 
• Prior/concomitant medication and supplement review 
• Home glucose meter review 
• Placement of peripheral intravenous (IV) line 
• Fasting baseline plasma glucose, insulin, alpha-tocopherol, and ammonia 
concentrations drawn from IV line.  Ammonia sample will be drawn from free-
flowing IV line without tourniquet and placed immediately on ice. 
• Oral protein tolerance test for subjects who do not experience severe hypoglycemia 
or hypoglycemic symptoms during Visit 1 oral protein tolerance test: 
o Enteral intake of Beneprotein (1.5 g/kg body weight protein, maximum 60 
grams protein = 1.75g/kg body weight protein powder, maximum 70 grams 
powder) mixed with carbohydrate-free Crystal Light in 6 ounces of water. 
o Plasma glucose and insulin concentrations obtained from IV line will be 
measured at 15, 30, 45, 60, 90, 120, 150, and 180 minutes. 
o Plasma ammonia will also be measured at 60 minutes from free-flowing IV 
line without tourniquet, and sample will be placed immediately on ice. 
o Plasma glucose measurements will be repeated as needed (if <70 mg/dL). 
 
• Subject/parent will complete the Vitamin E tolerability questionnaire (Appendix 3). 
• Adverse event monitoring. 
• Collection of any unused Vitamin E supplement. 
4.3 
Concomitant Medication 
All prior and concomitant medications used to treat hypoglycemia within 30 days prior to 
the screening visit and through the end of the study will be recorded. All medications and 
supplements regardless of indication will be recorded during the study period.  The dates of 
administration, dosage, and reason for use will be included. If a subject is taking a vitamin E 
supplement at the time of screening and then discontinues the supplement for 30 days, s/he 
can be eligible at that point for inclusion in the study. Subjects will be instructed to continue 
diazoxide per previously-prescribed dose throughout the study period. If hypoglycemia 

 
  
 
 
 
13
(plasma glucose <70 mg/dl measured on twice repeated point-of-care glucose meter) is 
noted during Visit 1 or Visit 2, then the Investigator may recommend the individual consult 
with his/her physician regarding increasing the diazoxide dose.  No medical management 
will be directed nor prescribed solely as means for study participation. If a subject later 
achieves better glucose control, s/he can be eligible at that point for inclusion in the study. 
4.4 
Rescue Medication Administration 
Hypoglycemia will be treated per usual clinical regimen used by CHOP’s Congenital 
Hyperinsulinism Center. During Visit 1 and Visit 2, if the subject has severe hypoglycemia, 
defined as plasma glucose <50 mg/dL measured on twice repeated point-of-care glucose 
meter, then after that time point blood sample is drawn, the subject will be given enteral 
fluids containing 15 grams carbohydrates, and the oral protein tolerance test will end. 
Plasma glucose will subsequently be measured with point-of-care glucose meter 20 minutes 
later to verify that plasma glucose has increased to >70 mg/dL. If plasma glucose is <70 
mg/dL on repeat checks, then the subject will be given a second enteral dose of 15 grams 
carbohydrates, and plasma glucose will be measured 20 minutes later. If plasma glucose 
remains <70 mg/dL after two doses of enteral carbohydrates, or if subject is unable to 
tolerate enteral fluids, then 10% dextrose 2 mL/kg bolus will be administered via peripheral 
IV as needed until plasma glucose is ≥70 mg/dL. 
4.5 
Subject Completion/Withdrawal 
Subjects who are found to have severe thrombocytopenia, defined as a platelet count 
<100,000 per microliter, during Visit 1 will be withdrawn from the study. Subjects may 
withdraw from the study at will at any time without prejudice to their care.  They may also 
be discontinued from the study at the discretion of the Investigator for lack of adherence to 
study or visit schedules. The Investigator may also withdraw subjects who violate the study 
plan, or to protect the subject for reasons of safety or for administrative reasons. It will be 
documented whether or not each subject completes the clinical study. If the Investigator 
becomes aware of any serious, related adverse events after the subject completes or 
withdraws from the study, they will be recorded in the source documents and on the case 
report forms. 
Subjects who were enrolled in this study prior to Amendment 4 being approved and whom 
were withdrawn due to severe hypoglycemia during the Visit 1 OPTT may be re-enrolled in 
this study to complete Vitamin E supplementation at home for two weeks and Visit 2 
procedures (with the exception of the repeat OPTT at Visit 2).  This will allow for 
tolerability of the supplement to still be assessed in individuals for whom a second OPTT is 
not warranted. 
 
5 
STUDY EVALUATIONS AND MEASUREMENTS 
5.1 
Screening Visit Measurements 
5.1.1 Inclusion/Exclusion Criteria Review 
• Date of birth 

 
  
 
 
 
14
• Relevant laboratory values for diagnosis of hyperinsulinism (including glucose, beta-
hydroxybutyrate, non-esterified free fatty acids, insulin, c-peptide, insulin-like 
growth factor binding protein 1, ammonia, and genetic testing results, if available) 
• Current use of diazoxide (including dosage and frequency) 
• Review of prior and concomitant medications and supplements 
• Review of allergies 
• Medical history review, including any history of bleeding disorder or current use of 
anticoagulation therapy 
• Review home glucose meter results for past 30 days 
• Urine pregnancy test for females who are menstruating or ≥11 years old 
5.2 
Visit 1 Measurements 
5.2.1 Demographic/Medical History Review 
• Medical record number 
• Sex 
• Race and ethnicity 
• Gestational age 
• Birth weight and length 
• Relevant co-morbidities 
• Molecular genetic study results, if available 
• Prior treatments for hyperinsulinism 
• Current treatments for hyperinsulinism 
• Review clinical care home glucose meter data over past 30 days 
• Episodes of seizures over the past month and year 
• Treatment for seizures 
• Recent/concomitant medications and supplements 
5.2.2 Physical Examination 
Physical exam by medical professional to collect the following measurements: 
• 
Length/Height (cm) 
• 
Weight (kg) 
• 
Vital signs: 
o heart rate [bpm] measured by automated device 
o blood pressure [mm Hg] measured by automated device in right upper arm 
while sitting 

 
  
 
 
 
15
o respiratory rate [bpm] 
• 
Skin exam (bruising) 
5.2.3 Baseline Symptom Questionnaire (Appendix 2) 
5.2.4 Laboratory Testing 
• Urine pregnancy test for females that are menstruating or ≥11 years of age 
• Point-of-care plasma glucose measurements from peripheral IV line at 0, 15, 30, 45, 
60, 90, 120, 150, and 180 minutes during OPTT 
o If plasma glucose is <70 mg/dL, then repeat point-of-care measurement will 
be obtained immediately. 
• Blood tests from peripheral IV: 
o plasma glucose at 0, 15, 30, 45, 60, 90, 120, 150, and 180 minutes during 
OPTT 
o plasma insulin at 0, 15, 30, 45, 60, 90, 120, 150, and 180 minutes during 
OPTT 
o plasma alpha-tocopherol at 0 minutes (fasting) 
o plasma ammonia at 0 and 60 minutes during OPTT – sample to be drawn 
without tourniquet and placed immediately on ice 
o complete blood count with platelet count at 0 minutes 
5.3 
Interim Evaluations 
• 
Phone interview with study staff on Day 4 (± 1 day) and Day 8 (± 1 day).  Study 
staff will use the script provided in Appendix 4 which includes administration of 
the tolerability questionnaire. 
5.4 
Visit 2 Measurements 
5.4.1 Physical Examination 
• 
Vital signs: 
o heart rate [bpm] measured by automated device 
o blood pressure [mm Hg] measured by automated device in right upper arm 
while sitting 
o respiratory rate [bpm] 
• 
Skin exam (bruising) 
5.4.2 Laboratory Testing 
For subjects completing Visit 2 OPTT 
• Point-of-care plasma glucose measurements from peripheral IV line at 0, 15, 30, 45, 
60, 90, 120, 150, and 180 minutes during OPTT 
o If plasma glucose is <70 mg/dL, then repeat point-of-care measurement will 

 
  
 
 
 
16
be obtained immediately. 
• Blood tests from peripheral IV: 
o plasma glucose at 0, 15, 30, 45, 60, 90, 120, 150, and 180 minutes during 
OPTT 
o plasma insulin at 0, 15, 30, 45, 60, 90, 120, 150, and 180 minutes during 
OPTT 
o plasma alpha-tocopherol at 0 minutes (fasting) 
o plasma ammonia at 0 and 60 minutes during OPTT – sample to be drawn 
without tourniquet and placed immediately on ice 
For subjects not completing Visit 2 OPTT 
• Point-of-care plasma glucose measurement from peripheral IV line 
• Fasting blood tests from peripheral IV: 
o plasma glucose 
o plasma insulin 
o plasma alpha-tocopherol 
o plasma ammonia – sample to be drawn without tourniquet and placed 
immediately on ice 
5.4.3 Tolerability Questionnaire (Appendix 3) 
5.4.4 Study Period Data Review 
• Concomitant medications and supplements during the study period 
• Hyperinsulinism treatment during the study period 
• Episodes of documented symptomatic and asymptomatic hypoglycemia (meter 
plasma glucose <70 mg/dL and <50 mg/dL): number during the study period 
• Review clinical care home glucose meter data during the study period 
• Episodes of seizures during the study period 
• Treatment for seizures during the study period 
• All adverse events that occur during the study period will be documented and 
reported per regulatory requirements 
 
6 
STATISTICAL CONSIDERATIONS 
6.1 
Primary Endpoint 
The primary endpoint will be tolerability as measured by responses to a subject/parent-
reported tolerability questionnaire compared to baseline responses and change in fasting 
plasma alpha-tocopherol concentration before versus after alpha-tocopherol 
supplementation. 

 
  
 
 
 
17
6.2 
Secondary Endpoints 
Secondary endpoints will include assessments of the effect of Vitamin E supplementation on 
the following measurements: 
• fasting plasma ammonia 
• delta-ammonia (plasma ammonia at 60 minutes during OPTT - plasma ammonia at 0 
minutes) 
• delta-glucose (fasting plasma glucose - nadir plasma glucose during OPTT) 
• fasting and nadir plasma glucose 
• fasting and peak plasma insulin 
• delta-insulin (peak plasma insulin during OPTT - fasting plasma insulin) 
• hypoglycemic episodes (plasma glucose <70 mg/dL) detected on clinical care home 
glucose meter 
• adverse events, including bleeding and bruising 
6.3 
Control of Bias and Confounding 
As a means to control bias and confounding variables, all patients with HI/HA syndrome 
aged 1 year to 40 years of age (inclusive) receiving ongoing treatment for recurring 
hypoglycemia followed at the Children’s Hospital of Philadelphia’s Congenital 
Hyperinsulinism Center (approximately 40 total) will be evaluated for eligibility for this 
study. Up to 14 eligible subjects whose parents consent to the study will be enrolled. Each 
subject serves as their own control. 
6.4 
Statistical Methods 
Descriptive statistics will be used to characterize demographics and outcome measures in 
subjects. As a general strategy, continuous endpoints will be summarized using the five-
number summary (mean, standard deviation, median, minimum, and maximum). Categorical 
endpoints will be summarized using frequency distributions. Paired t-tests (or if data are not 
normally distributed, Wilcoxon signed-rank tests) will be used to determine statistical 
significance of the primary and secondary outcomes, with alpha = 0.05. The number of 
subjects who experience any versus no adverse event will be tabulated, and the proportion 
will be calculated and reported, along with its corresponding 95% CI, based on all subjects 
in the study. The acceptable level of tolerability for this study is defined as if ≥75% of 
subjects find Vitamin E tolerable. 
6.5 
Sample Size and Power 
The proposed sample size of 14 subjects for this pilot clinical study is a sample of 
convenience based on the number of subjects that we can feasibly study during one year. 
The data generated from this study will be used to examine the feasibility and tolerability of 
alpha-tocopherol in subjects with HI/HA syndrome, in order to design and optimize a phase 
II/III study within this population in terms of sample size and selecting appropriate 
endpoints. 
 

 
  
 
 
 
18
7 
STUDY SUPPLEMENT 
7.1 
Description 
Vitamin E supplements are available over-the-counter in liquid and oral formulations. 
Vitamin E supplements are commonly used for anti-oxidant properties, although studies 
evaluating clinical benefit for various disorders show mixed results. Vitamin E is only 
synthesized by plants, and thus it is an essential nutrient for humans. The predominate 
bioavailable form of Vitamin E in humans is alpha-tocopherol. Approximately 90% of 
children and adults in the United States have inadequate intake of Vitamin E in their diet 
(11) and have insufficient plasma alpha-tocopherol concentrations (<30 μM) (12). The 
consequences of Vitamin E insufficiency are somewhat unclear, although true Vitamin E 
deficiency (plasma alpha-tocopherol concentration <12 μM) typically causes neurological 
deficits, although this only affects approximately 1% of the population (12; 13). 
Alpha-tocopherol crosses the blood-brain barrier, and Vitamin E supplementation improves 
neurological deficits in patients with Vitamin E deficiency (13). The range of Vitamin E 
supplementation doses is quite broad (from 6 IU daily recommended for infants to 1,500 IU 
daily considered as tolerated in adults, see Table 2), which is due to the very low side effect 
profile of Vitamin E and low risk of Vitamin E intoxication. The only recurrently-described 
side effect of Vitamin E is increased risk of bleeding in patients with risk factors (such as a 
bleeding disorder or receiving anticoagulation therapy). In general, Vitamin E doses up to 
400 IU daily are considered safe in adults and result in significant increases in plasma alpha-
tocopherol concentrations. Lower and higher doses of Vitamin E have been used in children, 
without reported adverse events, although these were relatively small studies. No studies 
have evaluated Vitamin E supplementation in patients with HI/HA. 
7.1.1 Dosing 
Subjects will be given Vitamin E supplements and instructed to take their dose once daily 
with a fat-containing meal.  Study dose will be below the upper acceptable limit for their age 
(Table 3), based on Institute of Medicine recommendations (Table 2). Capsules are 
available in the 200 IU dose, so can be used for subjects who will receive 600 IU daily. A 
liquid formulation (50 IU/mL) will be used for subjects who will receive lower doses, or any 
subjects who prefer liquid to capsules. 
7.1.2 Supplement Accountability 
Adequate records of study supplement receipt and disposition will be maintained by the 
study team and will be based upon the recommendations provided by the Investigational 
Drug Service. Investigational supplement orders, dispensing records, and disposition forms 
will be examined during the course of the study. The Vitamin E is to be provided by the 
Investigator or designee and may not be used for any purpose other than that described in 
this protocol. At study completion, all supplement supplies including partially used and 
empty containers must be returned to the Investigator so that they may be disposed of via 
manufacturer recommendations by the study team. Documentation of destruction will be 
captured via the Drug Accountability Log. 
 

 
  
 
 
 
19
8 
SAFETY MANAGEMENT 
8.1 
Clinical Adverse Events 
An adverse event is any untoward medical occurrence in a subject who has received an 
intervention (drug, biologic, or other intervention).  The occurrence does not necessarily 
have to have a causal relationship with the supplement.  An AE can therefore be any 
unfavorable or unintended sign (including an abnormal laboratory finding, for example), 
symptom, or disease temporally associated with an intervention, whether or not considered 
related to the supplement. 
The occurrence of an AE will be determined on the basis of observed or volunteered signs 
and symptoms and changes in the subject’s physical examination and laboratory test results. 
All AEs will be evaluated by the Investigator for seriousness, severity, relationship to study, 
and outcome. 
Collection of clinical information will begin after the subject’s written informed consent to 
participate in the study has been obtained.  AEs may be either spontaneously reported or 
elicited during questioning and examination of the subject. All identified AEs must be 
recorded and described in the subject’s source records. If known, the diagnosis of the 
underlying illness or disorder should be recorded, rather than its individual symptoms. 
8.2 
Adverse Event Reporting 
Unanticipated problems related to the research involving risks to subjects or others that 
occur during the course of this study (including SAEs) will be reported to the IRB in 
accordance with CHOP IRB SOP 408: Unanticipated Problems Involving Risks to Subjects. 
AEs that are not serious but that are notable and could involve risks to subjects will be 
summarized in narrative or other format and submitted to the IRB at the time of continuing 
review. 
8.3 
Definition of an Adverse Event 
An adverse event is any untoward medical occurrence in a subject who has received an 
intervention (drug, biologic, or other intervention).  The occurrence does not necessarily 
have to have a causal relationship with the treatment.  An AE can therefore be any 
unfavorable or unintended sign (including an abnormal laboratory finding, for example), 
symptom, or disease temporally associated with the use of a medicinal product, whether or 
not considered related to the medicinal product. 
All AEs (including serious AEs) will be noted in the study records and on the case report 
form with a full description including the nature, date and time of onset, determination of 
non-serious versus serious, intensity (mild, moderate, severe), duration, causality, and 
outcome of the event. 
8.4 
Definition of a Serious Adverse Event (SAE) 
An SAE is any adverse drug experience occurring at any dose that results in any of the 
following outcomes: 
• death, 

 
  
 
 
 
20
• a life-threatening event (at risk of death at the time of the event), 
• requires inpatient hospitalization or prolongation of existing hospitalization, 
• a persistent or significant disability/incapacity, or 
• a congenital anomaly/birth defect in the offspring of a subject. 
Important medical events that may not result in death, be life-threatening, or require 
hospitalization may be considered a serious adverse drug event when, based upon 
appropriate medical judgment, they may jeopardize the subject and may require medical or 
surgical intervention to prevent one of the outcomes listed in this definition. 
A distinction should be drawn between serious and severe AEs.  A severe AE is a major 
event of its type.  A severe AE does not necessarily need to be considered serious.  For 
example, nausea which persists for several hours may be considered severe nausea, but 
would not be an SAE.  On the other hand, a stroke that results in only a limited degree of 
disability may be considered a mild stroke, but would be an SAE. 
8.4.1 Relationship of SAE to study drug or other intervention 
The relationship of each SAE to the study intervention should be characterized using one of 
the following terms in accordance with CHOP IRB Guidelines: definitely, probably, 
possibly, unlikely or unrelated. 
8.5 
IRB/IEC Notification of SAEs and Other Unanticipated Problems 
The Investigator will promptly notify the IRB of all on-site unanticipated, serious Adverse 
Events that are related to the research activity. Other unanticipated problems related to the 
research involving risk to subjects or others will also be reported promptly. Written reports 
will be filed using the eIRB system and in accordance with the timeline below. External 
SAEs that are both unexpected and related to the study intervention will be reported 
promptly after the investigator receives the report. 
Type of Unanticipated 
Problem 
Initial Notification  
(Phone, Email, Fax) 
Written Report 
Internal (on-site) SAEs 
Death or Life 
Threatening 
24 hours 
Within 2 calendar days 
Internal (on-site) SAEs 
All other SAEs 
7 days 
Within 7 business days 
Unanticipated Problems 
Related to Research 
7 days 
Within 7 business days 
All other AEs 
N/A 
Brief Summary of 
important AEs may be 

 
  
 
 
 
21
reported at time of 
continuing review 
8.5.1 Follow-up report 
If an SAE has not resolved at the time of the initial report and new information arises that 
changes the investigator’s assessment of the event, a follow-up report including all relevant 
new or reassessed information (e.g., concomitant medication, medical history) should be 
submitted to the IRB. The investigator is responsible for ensuring that all SAE are followed 
until either resolved or stable. 
8.6 
Medical Emergencies 
Medical emergencies during the course of the study will be managed by physician-level 
members of the study team as per standard of clinical care. 
9 
STUDY ADMINISTRATION 
9.1 
Data Collection and Management 
1. Confidentiality and Security of Data. All information that is collected for this 
research protocol will be kept confidential. All subjects will be assigned a unique 
study identification number (study ID). This study ID will be used on all case report 
forms (CRFs). Data/CRFs will be stored in OnCore Clinical Trials Management 
System, a CHOP Research IS secured electronic data capture system. OnCore 
Clinical Trials Management System is a secure electronic data capture system with 
access controls and a data backup plan. The system is password protected. Only 
study team members will have access to subject data and case report forms stored in 
OnCore. Access to the system is monitored and logged for review if needed.  Exports 
from OnCore will be stored on a secure CHOP server. Data may also be stored on a 
secure CHOP server with password-protected files when possible. 
2. Anonymization, de-identification or destruction.  All data and records generated 
during this study will be kept confidential in accordance with institutional policies 
and HIPAA on subject privacy. De-identified data will be retained for 10 years 
from date of last treatment if subject is age 18 or over or an emancipated minor, or 
if the subject is a minor the data will be retained for 10 years from earlier of age 18 
or death. For subjects who consent to the use of their data for future research, PHI 
and the link between PHI and the study data will not be destroyed. A computer 
file, stored on one of CHOP’s secure servers, will be the site of storage. Each 
folder will be labeled with a note to date after which to destroy the data. “Destroy 
by….,” with the earliest dates at the front. 
3. The CHOP PI will monitor and review the study progress and the accuracy and 
security of the emerging data. 
9.2 
Confidentiality 
All data and records generated during this study will be kept confidential in accordance with 
Institutional policies and HIPAA on subject privacy.  The Investigators and other site 

 
  
 
 
 
22
personnel will not use such data and records for any purpose other than conducting the 
study. Safeguards are described under Data Collection and Management. 
9.3 
Regulatory and Ethical Considerations 
9.3.1 Data and Safety Monitoring Plan 
There will be no added physical, psychological, economic, or societal risks due to this 
interventional study. The principal investigator will review the safety and progress of this 
study on a monthly basis.  Data reviewed will include laboratory results, OPTT results, 
tolerability questionnaire responses, adverse events, and data collected as per Section 5.4.4.  
The principal investigator or a physician-level study team member will be available during 
the OPTT to monitor and treat subjects in the event of hypoglycemia. The primary risk 
stems from breach of confidentiality. The Principal Investigator will provide oversight for 
risks associated with management of confidential data (see Data Collection and 
Management). 
9.3.2 Risk Assessment 
This study poses a minor increase above minimal risk with no prospect for direct benefit to 
the subject, but is likely to yield generalizable knowledge about the condition.  The study 
tests performed, including peripheral blood sampling and oral protein tolerance testing, are 
similar to those that HI/HA patients experience as part of their routine care.  The Visit 1 
OPTT can replace their annual OPTT. In the event that the Visit 1 OPTT cannot replace a 
subject’s annual OPTT, then two OPTTs will be completed for this research study. Vitamin 
E is a nutritional supplement available over-the-counter that may be taken in the dose ranges 
used in this study by the general population. The potential risks associated with the use of 
the Vitamin E supplementation include bruising and bleeding. To minimize these risks, the 
dose of Vitamin E will be tailored to the subject’s age, and baseline platelet counts will be 
measured. There is a potential risk of hypoglycemia during oral protein tolerance test. The 
frequent point-of-care glucose measurements, along with assessment of symptoms of 
hypoglycemia, during the OPTT will allow for rapid detection of hypoglycemia if it occurs. 
Potential hypoglycemia is readily reversible with oral carbohydrates or IV dextrose. 
There is potential risk of breach of confidentiality.  Measures will be taken to avoid breach 
of privacy and insure confidentiality (see Data Collection and Management). 
9.3.3 Potential Benefits of Study Participation 
There are no direct benefits to the study subjects. The knowledge gained from this study 
may lead to improved treatment of future patients with HI/HA syndrome. 
9.3.4 Risk-Benefit Assessment 
The potential benefits of advancement in medical knowledge outweigh the potential risk to 
subjects. Measures will be taken to minimize risks. 
9.4 Recruitment Strategy 
Subjects will be recruited prospectively through the Congenital Hyperinsulinism Center at 
CHOP. The Co-Investigator is the director of the center and the principal outpatient 
physician caring for these patients. The Primary Investigator, the Co-Investigator, or the 
study staff will approach the subjects in person, by telephone, mail, or email for enrollment. 

 
  
 
 
 
23
An estimated 40 subjects over the course of 1 year will be eligible for inclusion based on 
clinical experience from recent years. 
Informed consent will be obtained prior to completion of eligibility screening. 
9.5 Informed Consent/Assent and HIPAA Authorization 
The principal investigator or a physician-level investigator on the study team will obtain 
informed consent from the subject’s family in person at the subject’s bedside at CHOP or a 
private conference room at CHOP, or by telephone. If the consent discussion takes place via 
phone, then the physician investigators will obtain a signed consent form from the subject 
and their parent/guardian at the first in-person visit, prior to the start of any study 
procedures. Two parent/guardian signatures will be required for the enrollment of a minor in 
this research study. The study will be explained in full detail. The subject’s family will have 
the opportunity to contact the staff at CHOP at any time to clarify any questions that they 
have. Subjects’ family will be informed that their participation in the study is voluntary and 
will not affect the medical care the subject receives, that there will be no direct benefit of 
participation, and that the primary risk is breach of confidentiality. The informed consent 
form will be kept in the investigator’s locked office cabinet. A combined consent-
authorization document will be used. 
9.6 Payment to Subjects/Families 
9.6.1 Payments to the family for time and inconvenience, parking (i.e. 
compensation) 
This study requires that subjects and their family must take time out of their schedules in 
order to participate.  Therefore, time and inconvenience compensation of $50 per outpatient 
CHPS visit, and $25 for completing the whole study (maximum $125/subject) will be 
provided. For subjects <18 years of age, the subjects will receive one-third and the 
parents/legal guardians will receive two-thirds of the compensation. Subjects >18 years of 
age will receive the total compensation amount. Subjects/parents or legal guardians will 
receive this compensation only once (even if they must return to CHOP for further testing) 
at the completion of the study.  If for some reason, the subject cannot complete the study at 
the time of study visit, they will still receive the time and inconvenience compensation.  This 
compensation will be in the form of a Participant Reimbursement Card. 
9.6.2 Payment to the family for transportation and lodging. 
Subjects will be reimbursed for transportation and lodging up to a maximum of $1,500.00 
for the entire study.  Receipts for travel and lodging will be required for reimbursement.  
This reimbursement will be in the form of a check and will be paid at the end of their study 
participation.  If a subject is under the age of 18, the compensation will be paid to the parent 
or legal guardian of the subject. 
 
PUBLICATION 
The investigators intend to present the results at national conferences and to publish the 
results in peer-reviewed medical journals. 

 
  
 
 
 
24
 
 
 

 
  
 
 
 
25
10 REFERENCES 
1. Palladino AA, Stanley CA: The hyperinsulinism/hyperammonemia syndrome. Rev 
Endocr Metab Disord 2010;11:171-178 
2. Snider KE, Becker S, Boyajian L, Shyng SL, MacMullen C, Hughes N, Ganapathy K, 
Bhatti T, Stanley CA, Ganguly A: Genotype and phenotype correlations in 417 children with 
congenital hyperinsulinism. The Journal of clinical endocrinology and metabolism 
2013;98:E355-363 
3. Hsu BYL, Kelly A, Thornton PS, Greenberg CR, Dilling LA, Stanley CA: Protein-
sensitive and fasting hypoglycemia in children with the hyperinsulinism/hyperammonemia 
syndrome. The Journal of Pediatrics 2001;138:383-389 
4. Stanley CA, Baker L: Hyperinsulinism in infancy: diagnosis by demonstration of 
abnormal response to fasting hypoglycemia. Pediatrics 1976;57:702-711 
5. Yildizdas D, Erdem S, Kucukosmanoglu O, Yilmaz M, Yuksel B: Pulmonary 
hypertension, heart failure and neutropenia due to diazoxide therapy. Advances in therapy 
2008;25:515-519 
6. Hu S, Xu Z, Yan J, Liu M, Sun B, Li W, Sang Y: The treatment effect of diazoxide on 44 
patients with congenital hyperinsulinism. Journal of pediatric endocrinology & metabolism : 
JPEM 2012;25:1119-1122 
7. Li C, Allen A, Kwagh J, Doliba NM, Qin W, Najafi H, Collins HW, Matschinsky FM, 
Stanley CA, Smith TJ: Green Tea Polyphenols Modulate Insulin Secretion by Inhibiting 
Glutamate Dehydrogenase. Journal of Biological Chemistry 2006;281:10214-10221 
8. Li C, Matter A, Kelly A, Petty TJ, Najafi H, MacMullen C, Daikhin Y, Nissim I, Lazarow 
A, Kwagh J, Collins HW, Hsu BYL, Nissim I, Yudkoff M, Matschinsky FM, Stanley CA: 
Effects of a GTP-insensitive Mutation of Glutamate Dehydrogenase on Insulin Secretion in 
Transgenic Mice. Journal of Biological Chemistry 2006;281:15064-15072 
9. Li M, Smith CJ, Walker MT, Smith TJ: Novel inhibitors complexed with glutamate 
dehydrogenase: Allosteric regulation by control of protein dynamics. The Journal of 
biological chemistry 2009;284:22988-23000 
10. Institute of Medicine: Dietary Reference Intakes for Vitamin C, Vitamin E, Selenium, 
and Carotenoids. Washington, DC, The National Academies Press, 2000 
11. McGuire S: Scientific Report of the 2015 Dietary Guidelines Advisory Committee. 
Washington, DC: US Departments of Agriculture and Health and Human Services, 2015. 
Advances in Nutrition: An International Review Journal 2016;7:202-204 
12. McBurney MI, Yu EA, Ciappio ED, Bird JK, Eggersdorfer M, Mehta S: Suboptimal 
Serum α-Tocopherol Concentrations Observed among Younger Adults and Those 
Depending Exclusively upon Food Sources, NHANES 2003-2006(1-3). PloS one 
2015;10:e0135510 
13. Ulatowski L, Manor D: Vitamin E Trafficking in Neurologic Health and Disease. 
Annual Review of Nutrition 2013;33:87-103 
14. Pfeiffer CM, Sternberg MR, Schleicher RL, Haynes BMH, Rybak ME, Pirkle JL: CDC’s 
Second National Report on Biochemical Indicators of Diet and Nutrition in the US 
Population is a valuable tool for researchers and policy makers(). The Journal of nutrition 
2013;143:938S-947S 
15. Ferslew KE, Acuff RV, Daigneault EA, Woolley TW, Stanton PE: Pharmacokinetics 
and Bioavailability of the RRR and All Racemic Stereoisomers of Alpha-Tocopherol in 
Humans After Single Oral Administration. J Clin Pharmacol 1993;33:84-88 

 
  
 
 
 
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16. Tousoulis D, Antoniades C, Tentolouris C, Tsioufis C, Toutouza M, Toutouzas P, 
Stefanadis C: Effects of combined administration of vitamins C and E on reactive hyperemia 
and inflammatory process in chronic smokers. Atherosclerosis 2003;170:261-267 
17. Miller ER, Iii, Pastor-Barriuso R, Dalal D, Riemersma RA, Appel LJ, Guallar E: Meta-
analysis: High-dosage vitamin e supplementation may increase all-cause mortality. Annals 
of Internal Medicine 2005;142:37-46 
18. Sesso HD, Buring JE, Christen WG, Kurth T, Belanger C, MacFadyen J, Bubes V, 
Manson JE, Glynn RJ, Gaziano JM: Vitamins E and C in the Prevention of Cardiovascular 
Disease in Men: The Physicians’ Health Study II Randomized Trial. JAMA : the journal of 
the American Medical Association 2008;300:2123-2133 
19. Bakaltcheva I, Gyimah D, Reid T: Effects of α-tocopherol on platelets and the 
coagulation system. Platelets 2001;12:389-394 
20. Pastori D, Carnevale R, Cangemi R, Saliola M, Nocella C, Bartimoccia S, Vicario T, 
Farcomeni A, Violi F, Pignatelli P: Vitamin E Serum Levels and Bleeding Risk in Patients 
Receiving Oral Anticoagulant Therapy: a Retrospective Cohort Study. Journal of the 
American Heart Association: Cardiovascular and Cerebrovascular Disease 2013;2:e000364 
21. Dereska NH, McLemore EC, Tessier DJ, Bash DS, Brophy CM: Short-term, Moderate 
Dosage Vitamin E Supplementation May Have No Effect on Platelet Aggregation, 
Coagulation Profile, and Bleeding Time in Healthy Individuals. Journal of Surgical 
Research 2006;132:121-129 
22. Woestenenk JW, Broos N, Stellato RK, Arets HG, van der Ent CK, Houwen RH: 
Vitamin E intake, alpha-tocopherol levels and pulmonary function in children and 
adolescents with cystic fibrosis. Br J Nutr 2015;113:1096-1101 
23. Ghaffari J, Farid Hossiani R, Khalilian A, Nahanmoghadam N, Salehifar E, Rafatpanah 
H: Vitamin e supplementation, lung functions and clinical manifestations in children with 
moderate asthma: a randomized double blind placebo- controlled trial. Iran J Allergy 
Asthma Immunol 2014;13:98-103 
24. Mustafa Nachvak S, Reza Neyestani T, Ali Mahboob S, Sabour S, Ali Keshawarz S, 
Speakman JR: alpha-Tocopherol supplementation reduces biomarkers of oxidative stress in 
children with Down syndrome: a randomized controlled trial. Eur J Clin Nutr 2014;68:1119-
1123 
25. Yousefichaijan P, Kahbazi M, Rasti S, Rafeie M, Sharafkhah M: Vitamin E as adjuvant 
treatment for urinary tract infection in girls with acute pyelonephritis. Iran J Kidney Dis 
2015;9:97-104 
26. Cazeau RM, Huang H, Bauer JA, Hoffman RP: Effect of Vitamins C and E on 
Endothelial Function in Type 1 Diabetes Mellitus. Journal of diabetes research 
2016;2016:3271293 
27. Melis D, Minopoli G, Balivo F, Marcolongo P, Parini R, Paci S, Dionisi-Vici C, Della 
Casa R, Benedetti A, Andria G, Parenti G: Vitamin E Improves Clinical Outcome of 
Patients Affected by Glycogen Storage Disease Type Ib. JIMD Rep 2016;25:39-45 
28. Treberg JR, Clow KA, Greene KA, Brosnan ME, Brosnan JT: Systemic activation of 
glutamate dehydrogenase increases renal ammoniagenesis: implications for the 
hyperinsulinism/hyperammonemia syndrome. American journal of physiology 
Endocrinology and metabolism 2010;298:E1219-1225 
 
 
 

 
  
 
 
 
27
11 APPENDIX 1 
Medications Known to Adversely Interact With Vitamin E 
Orlistat 
Risk Category D 
Tipranavir 
Risk Category D 
*Note: As this list is subject to change over time, the Lexicomp database will be referenced 
for the most up-to-date drug interaction information during inclusion/exclusion screening. 
Medications known to adversely interact with Vitamin E are those listed as Risk Category D 
(“consider therapy modification”) or Risk Category X (“avoid combination”) in the 
Lexicomp database. 
 
 

 
  
 
 
 
28
12 APPENDIX 2 
Baseline Symptom Questionnaire 
Event 
Severity 
Number of events in the 
past 14 days 
Severity Definitions: 
None – event did not occur 
Mild – minimal symptoms, no treatment needed 
Moderate – symptoms requiring treatment at home or as an outpatient 
Severe – symptoms requiring hospitalization or emergency room visit or 
life-threatening or potentially life-threatening symptoms 
Seizure 
 None 
 Mild 
 Moderate 
 Severe 
 
Headache 
 None 
 Mild 
 Moderate 
 Severe 
 
Vision change/blurred 
vision 
 None 
 Mild 
 Moderate 
 Severe 
 
Weakness 
 None 
 Mild 
 Moderate 
 Severe 
 
Fatigue 
 None 
 Mild 
 Moderate 
 Severe 
 

 
  
 
 
 
29
Nausea 
 None 
 Mild 
 Moderate 
 Severe 
 
Vomiting 
 None 
 Mild 
 Moderate 
 Severe 
 
Diarrhea 
 None 
 Mild 
 Moderate 
 Severe 
 
Stomach pain 
 None 
 Mild 
 Moderate 
 Severe 
 
Constipation 
 None 
 Mild 
 Moderate 
 Severe 
 
Bruising 
 None 
 Mild 
 Moderate 
 Severe 
 
Bleeding 
 None 
 Mild 
 Moderate 
 Severe 
 
Rash 
 None 
 

 
  
 
 
 
30
 Mild 
 Moderate 
 Severe 
Itching 
 None 
 Mild 
 Moderate 
 Severe 
 
 
LIST ANY OTHER SYMPTOMS YOU EXPERIENCED SINCE STARTING VITAMIN 
E 
IN THE BOXES BELOW 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 

 
  
 
 
 
31
 
 

 
  
 
 
 
32
13 APPENDIX 3 
Tolerability Questionnaire 
Event 
Severity 
Number of events since 
starting Vitamin E 
Severity Definitions: 
None – event did not occur 
Mild – minimal symptoms, no treatment needed 
Moderate – symptoms requiring treatment at home or as an outpatient 
Severe – symptoms requiring hospitalization or emergency room visit or 
life-threatening or potentially life-threatening symptoms 
Seizure 
 None 
 Mild 
 Moderate 
 Severe 
 
Headache 
 None 
 Mild 
 Moderate 
 Severe 
 
Vision change/blurred 
vision 
 None 
 Mild 
 Moderate 
 Severe 
 
Weakness 
 None 
 Mild 
 Moderate 
 Severe 
 
Fatigue 
 None 
 Mild 
 Moderate 
 Severe 
 

 
  
 
 
 
33
Nausea 
 None 
 Mild 
 Moderate 
 Severe 
 
Vomiting 
 None 
 Mild 
 Moderate 
 Severe 
 
Diarrhea 
 None 
 Mild 
 Moderate 
 Severe 
 
Stomach pain 
 None 
 Mild 
 Moderate 
 Severe 
 
Constipation 
 None 
 Mild 
 Moderate 
 Severe 
 
Bruising 
 None 
 Mild 
 Moderate 
 Severe 
 
Bleeding 
 None 
 Mild 
 Moderate 
 Severe 
 
Rash 
 None 
 

 
  
 
 
 
34
 Mild 
 Moderate 
 Severe 
Itching 
 None 
 Mild 
 Moderate 
 Severe 
 
 
LIST ANY OTHER SYMPTOMS YOU EXPERIENCED SINCE STARTING VITAMIN 
E 
IN THE BOXES BELOW 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 

 
  
 
 
 
35
 
 
 
 
 
 
 
 
 
14 APPENDIX 4 
Study Staff Telephone Script 
Hello [name of contact], I am [name of caller], calling from CHOP regarding 
you/your child’s enrollment in the Vitamin E Supplementation in Hyperinsulinism/ 
Hyperammonemia Syndrome study.  I am calling to check in and see how you are doing.  
The call will probably take around 15 minutes. 
Have you/your child had any new medical problems since you last spoke with our 
team? 
Have you/has your child been seen in the Emergency Department, Urgent Care 
Center, or by another health provider since you last spoke with our team? 
If yes to either of the above, what has happened? 
Have you/your child taken any new medications or supplements or stopped taking 
any previous medications or supplements since you last spoke with our team? 
Sometimes it is hard to remember to take a new a new supplement.  Since you were 
given vitamin E at the visit, how has it been going? 
At what time of the day do you take the vitamin E? 
How much do you take each time? 
Do you take the vitamin E with meals or with any other medications or supplements? 
Have you missed any doses of vitamin E? 
If yes, how many? 
Can you tell me about any new feeling or symptoms you/your child has experienced 
since beginning taking the vitamin E (or since our last call)?  Are there any new feelings or 
symptoms you are/your child is experiencing that you think may be side effects from the 
vitamin E supplementation? 
Have you/has your child had any of the following (if yes, was the severity mild, 
moderate, or severe, and, what is the number of events that have occurred since starting 
Vitamin E): 
 

 
  
 
 
 
36
 
 
 
 
 
 
 
 
 
Event 
Severity 
Number of events since 
starting Vitamin E 
Severity Definitions: 
None – event did not occur 
Mild – minimal symptoms, no treatment needed 
Moderate – symptoms requiring treatment at home or as an outpatient 
Severe – symptoms requiring hospitalization or emergency room visit or 
life-threatening or potentially life-threatening symptoms 
Seizure 
 None 
 Mild 
 Moderate 
 Severe 
 
Headache 
 None 
 Mild 
 Moderate 
 Severe 
 
Vision change/blurred 
vision 
 None 
 Mild 
 Moderate 
 Severe 
 
Weakness 
 None 
 Mild 
 Moderate 
 

 
  
 
 
 
37
 Severe 
Fatigue 
 None 
 Mild 
 Moderate 
 Severe 
 
Nausea 
 None 
 Mild 
 Moderate 
 Severe 
 
Vomiting 
 None 
 Mild 
 Moderate 
 Severe 
 
Diarrhea 
 None 
 Mild 
 Moderate 
 Severe 
 
Stomach pain 
 None 
 Mild 
 Moderate 
 Severe 
 
Constipation 
 None 
 Mild 
 Moderate 
 Severe 
 
Bruising 
 None 
 Mild 
 Moderate 
 Severe 
 

 
  
 
 
 
38
Bleeding 
 None 
 Mild 
 Moderate 
 Severe 
 
Rash 
 None 
 Mild 
 Moderate 
 Severe 
 
Itching 
 None 
 Mild 
 Moderate 
 Severe 
 
 
HAVE THERE BEEN ANY OTHER SYMPTOMS YOU EXPERIENCED SINCE 
STARTING VITAMIN E? 
(If yes, these should be recorded by the person administering the survey in the boxes 
below) 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 
 
 
 
 None 
 Mild 
 Moderate 
 Severe 
 

 
  
 
 
 
39
 
 
 
 None 
 Mild 
 Moderate 
 Severe