Levothyroxine Treatment and Cardiometabolic Outcomes in Adolescents With Down Syndrome
Phase: NA
Conditions: Down Syndrome, Subclinical Hypothyroidism
Sponsor: Children's Hospital of Philadelphia

 
 
Title: 
Levothyroxine treatment and cardiometabolic outcomes in 
adolescents with DS 
Short Title 
Aim 2 Levothyroxine and DS 
Drug Name(s): 
Levothyroxine 
 
 
Regulatory Sponsor: 
NIH 
eIRB Number 
12-009578 
Protocol Date: 
August 26, 2013 
Amendment 1 Date: 8/26/13 
Amendment 3 Date:11/14/2014 
Amendment 2 Date:9/22/14 
Amendment 4 Date: 9/23/2015 
Sponsor  
NIH 
 
Study Principal Investigator 
Andrea Kelly, MD, MSCE  
11 NW 30 Main Building, Children’s Hospital of Philadelphia, 34th St. & Civic Center Blvd. 
Philadelphia, PA, 19104 
Phone: 215-590-3420 
Email: kellya@email.chop.edu 
 
 

 
 
 
 
 
 
 
 
 
 
ii
TABLE OF CONTENTS 
Table of Contents ................................................................................................................... ii 
Abbreviations and Definitions of Terms............................................................................. iv 
Protocol Synopsis .................................................................................................................. vi 
Table 1: Schedule of Study Procedures: Both Groups ....................................................... x 
Figure 1: Study Diagram .................................................................................................... xiii 
1 
BACKGROUND INFORMATION AND RATIONALE ....................................................................... 1 
1.1 
INTRODUCTION ..................................................................................................................................... 1 
1.2 
NAME AND DESCRIPTION OF INTERVENTION ........................................................................................ 1 
1.3 
FINDINGS FROM NON-CLINICAL AND CLINICAL STUDIES ..................................................................... 1 
1.3.1 
Non-Clinical Studies:  N/A .............................................................................................................. 1 
1.3.2 
Clinical Studies ............................................................................................................................... 1 
1.4 
SELECTION OF DRUGS AND DOSAGES................................................................................................... 2 
1.5 
RELEVANT LITERATURE AND DATA ..................................................................................................... 3 
1.6 
COMPLIANCE STATEMENT .................................................................................................................... 4 
2 
STUDY OBJECTIVES .............................................................................................................................. 4 
2.1 
PRIMARY AIM ...................................................................................................................................... 4 
2.2 
SECONDARY AIM.................................................................................................................................. 4 
3 
INVESTIGATIONAL PLAN .................................................................................................................... 5 
3.1 
GENERAL SCHEMA OF STUDY DESIGN ................................................................................................. 5 
3.1.1 
Screening Phase .............................................................................................................................. 5 
3.1.2 
Phase 1 ............................................................................................ Error! Bookmark not defined. 
3.1.3 
Phase 2 ............................................................................................ Error! Bookmark not defined. 
3.1.4 
Follow-up Phase ............................................................................. Error! Bookmark not defined. 
3.2 
ALLOCATION TO TREATMENT GROUPS AND BLINDING ........................................................................ 7 
3.3 
STUDY DURATION, ENROLLMENT AND NUMBER OF SITES ................................................................... 8 
3.3.1 
Duration of Study Participation ...................................................................................................... 8 
3.3.2 
Total Number of Study Sites/Total Number of Subjects Projected .................................................. 8 
3.4 
STUDY POPULATION ............................................................................................................................. 8 
3.4.1 
Inclusion Criteria ............................................................................................................................ 8 
3.4.2 
Exclusion Criteria ........................................................................................................................... 8 
4 
STUDY PROCEDURES ............................................................................................................................ 9 
4.1 
SCREENING VISIT ......................................................................... ERROR! BOOKMARK NOT DEFINED. 
4.2 
SCREENING/BASELINE PHASE ...................................................... ERROR! BOOKMARK NOT DEFINED. 
4.3 
INTERIM PHASE ............................................................................ ERROR! BOOKMARK NOT DEFINED. 
4.3.1 
Visit 1 (3 Month) & Visit 2 (6 Month) ............................................. Error! Bookmark not defined. 
4.4 
RANDOMIZED PHASE .................................................................... ERROR! BOOKMARK NOT DEFINED. 
4.4.1 .............................................................................................................. Error! Bookmark not defined. 
4.4.2 
6 Month Visit and 12 Month Visit ................................................... Error! Bookmark not defined. 
4.5 
FOLLOW-UP PHASE ....................................................................... ERROR! BOOKMARK NOT DEFINED. 
4.5.1 
18 Month ......................................................................................... Error! Bookmark not defined. 
4.6 
TSH & T4 CHECK ........................................................................ ERROR! BOOKMARK NOT DEFINED. 
4.7 
CONCOMITANT MEDICATION ............................................................................................................. 11 
4.8 
SUBJECT COMPLETION/WITHDRAWAL ............................................................................................... 11 
5 
STUDY EVALUATIONS AND MEASUREMENTS ............................................................................ 12 
5.1 
SCREENING AND MONITORING EVALUATIONS AND MEASUREMENTS ................................................ 12 
5.1.1 
Laboratory Evaluations................................................................................................................. 14 

 
 
 
 
 
 
 
 
 
 
iii
5.2 
SAFETY EVALUATION ......................................................................................................................... 15 
6 
STATISTICAL CONSIDERATIONS .................................................................................................... 16 
6.1 
STATISTICAL METHODS...................................................................................................................... 16 
6.1.1 
Baseline Data ................................................................................................................................ 16 
6.1.2 
Efficacy Analysis ........................................................................................................................... 17 
6.1.3 
Safety Analysis .............................................................................................................................. 17 
6.2 
SAMPLE SIZE AND POWER .................................................................................................................. 17 
6.3 
INTERIM ANALYSIS ............................................................................................................................ 17 
7 
STUDY MEDICATION (INTERVENTION) ........................................................................................ 19 
7.1 
DESCRIPTION ...................................................................................................................................... 19 
7.1.1 
Packaging ...................................................................................................................................... 19 
7.1.2 
Labeling ........................................................................................................................................ 19 
7.1.3 
Dosing ........................................................................................................................................... 19 
7.1.4 
Treatment Compliance and Adherence ......................................................................................... 19 
7.1.5 
Drug Accountability ...................................................................................................................... 19 
8 
SAFETY MANAGEMENT ..................................................................................................................... 20 
8.1 
CLINICAL ADVERSE EVENTS .............................................................................................................. 20 
8.2 
ADVERSE EVENT REPORTING ............................................................................................................. 20 
8.3 
DEFINITION OF AN ADVERSE EVENT .................................................................................................. 20 
8.4 
DEFINITION OF A SERIOUS ADVERSE EVENT (SAE) ........................................................................... 20 
8.4.1 
Relationship of SAE to study drug or other intervention ............................................................... 21 
8.5 
IRB/IEC NOTIFICATION OF SAES AND OTHER UNANTICIPATED PROBLEMS ...................................... 21 
8.5.1 
Follow-up report ........................................................................................................................... 21 
8.6 
INVESTIGATOR REPORTING OF A SERIOUS ADVERSE EVENT TO SPONSOR .......................................... 21 
8.7 
MEDICAL EMERGENCIES .................................................................................................................... 22 
9 
STUDY ADMINISTRATION ................................................................................................................. 22 
9.1 
TREATMENT ASSIGNMENT METHODS................................................................................................. 22 
9.1.1 
Randomization............................................................................................................................... 22 
9.1.2 
Blinding ......................................................................................................................................... 22 
9.1.3 
Unblinding .................................................................................................................................... 22 
9.2 
DATA COLLECTION AND MANAGEMENT ............................................................................................ 22 
9.3 
CONFIDENTIALITY .............................................................................................................................. 23 
9.4 
REGULATORY AND ETHICAL CONSIDERATIONS.................................................................................. 24 
9.4.1 
Data and Safety Monitoring Plan ................................................................................................. 24 
9.4.2 
Risk Assessment ............................................................................................................................. 25 
9.4.3 
Potential Benefits of Trial Participation ....................................................................................... 28 
9.4.4 
Risk-Benefit Assessment ................................................................................................................ 28 
9.5 
RECRUITMENT STRATEGY .................................................................................................................. 28 
9.6 
INFORMED CONSENT/ASSENT AND HIPAA AUTHORIZATION ............................................................ 28 
9.6.1 
Consent for Down syndrome subjects 18 – 20 years of age .......................................................... 29 
9.6.2 
Assent Procedures ......................................................................................................................... 30 
9.7 
PAYMENT TO SUBJECTS/FAMILIES ..................................................................................................... 31 
10 
PUBLICATION ........................................................................................................................................ 32 
11 
REFERENCES ......................................................................................................................................... 32 
 
 
 

 
 
 
 
 
 
 
 
 
 
iv
 
Abbreviations and Definitions of Terms 
 
 
 
AAP 
 
American Academy of Pediatrics 
ABAS-II 
 
ANTITIPO 
ANTITG 
 
Adaptive behavior assessment system-Second edition 
parent form 
Thyroid peroxidase (microsomal) antibodies 
Thyroglobulin antibodies 
AE 
ASAQ  
 
Adverse events 
Adolescent Sedentary Activity Questionnaire 
BES 
 
Body esteem scale for children 
BG 
 
Blood glucose 
BP 
 
Blood pressure 
CEBQ 
CFQ 
 
Child eating behavior questionnaire 
Child feeding questionnaire 
CHOP 
 
The Children’s Hospital of Philadelphia 
CMR 
 
Cardiometabolic risk 
CNMC 
 
Children’s National Medical Center 
CPP 
 
Central pulse pressure 
CRP 
 
C-reactive protein 
CTRC 
 
Clinical and Translational Research Center 
DS 
 
Down syndrome 
DXA 
 
Dual energy x-ray absorptiometry 
EMR 
FRS 
 
Electronic medical record 
Stunkard figure rating scale 
HbA1c   
 
Hemoglobin A1c 
HDL-C 
 
High-density lipoprotein cholesterol 
HFI 
 
Home food inventory 
HOMA-IR 
 
Homeostasis model assessment – insulin resistance 
Hs-CRP 
 
High-sensitivity C-reactive protein 
IL-6 
 
Interleukin-6 
IWQOL-Kids 
 
Impact of weight on quality of life-Kids 
LDL-C 
 
Low-density lipoprotein cholesterol 
LV 
 
Left ventricular 
NMR 
 
Nuclear magnetic resonance 
OGTT 
 
Oral glucose tolerance test 

 
 
 
 
 
 
 
 
 
 
v
 
 
 
 
 
 
 
 
 
PedsQL 
 
Pediatric Quality of Life Inventory 
PeRC 
 
Pediatric Research Consortium 
PHI 
 
Protected Health Information 
PWA 
 
Pulse wave analysis 
PWV 
 
Pulse wave velocity 
QOL 
 
Quality of life 
RCT 
 
Randomized controlled trial 
SD 
 
Standard deviation 
T1DM 
 
Type 1 diabetes 
TC 
 
Total cholesterol 
TG 
TSH 
 
Triglycerides 
Thyroid stimulating hormone 
PAI-1 
 
Plasminogen Activator Inhibitor 1  
VLDL-C 
 
Very low-density lipoprotein cholesterol 
WC 
 
Waist circumference 
 
 
 
 
 
 

 
 
 
 
 
 
 
 
 
 
vi
PROTOCOL SYNOPSIS 
Study Title 
Levothyroxine treatment and cardiometabolic outcomes in 
adolescents with Down syndrome 
Funder 
National Institutes of Health (NIH) 
Clinical Phase 
N/A 
Study Rationale 
 The American Academy of Pediatrics (AAP) recommends annual 
screening for thyroid dysfunction in children and adolescents with 
Down Syndrome (DS)(1). Mild elevations in TSH are common in 
DS.  Recently, subclinical hypothyroidism has been associated with 
increased cardiovascular (CVD) risk in the general adult population, 
and debate rages as to whether subclinical hypothyroidism should 
be treated. Moreover, the potential benefits of treating subclinical 
hypothyroidism in DS are not known. This study will examine the 
effect of levothyroxine hormone treatment on body composition, 
cardiometabolic risk (CMR) factors, and quality of life (QOL) in 
children with DS and subclinical hypothyroidism. 
Study Objective(s) 
Primary  
 
To determine the effect of levothyroxine replacement on BMI-Z, 
fat mass, and CMR factors in pre-adolescents and adolescents 
with DS and subclinical hypothyroidism. 
 
Secondary 
 
 To determine the effect of levothyroxine replacement on QOL, 
lifestyle (physical activity, nutrition) and body image in pre-
adolescents and adolescents with DS and subclinical 
hypothyroidism. 
  
 
 
Test Article(s) 
 
Levothyroxine (starting dose 0.5 – 1 mcg/kg daily)  
 
Study Design 
 
Single blinded randomized controlled trial using switching 
replication design. 
Subject Population 
key criteria for 
Inclusion and 
Exclusion: 
Inclusion Criteria 
1. Males and females, ages 8 – 20 years 
2. Diagnosis of Down syndrome 
3. Subclinical hypothyroidism: TSH level between 5 – 10 
mIU/L, normal T4 

 
 
 
 
 
 
 
 
 
 
vii
 
4. Parental/guardian permission (informed consent) and if 
appropriate, child assent 
5. Females who are  11 years of age or who are menarchal 
must have a negative urine/serum pregnancy test 
6. Committed to adherence to levothyroxine treatment and 
study completion 
Exclusion Criteria 
1. Pregnancy 
2. Type 1/Type 2 diabetes 
3. Chronic medical conditions or medication use that can affect 
growth, nutrition, blood glucose, insulin secretion, or thyroid 
function (such as lithium or certain seizure medications) 
4. Current use of levothyroxine or anti-thyroid hormone 
5.   Cyanotic congenital heart disease, or pulmonary 
hypertension (as described by last echo report in subjects 
with CHD), or congenital heart disease considered medically 
unstable by the study cardiologists 
 
Number Of Subjects  
 
Total number of subjects overall and at CHOP: 29 enrolled for 20 
completers overall; 11 enrolled for 6 completers at CHOP 
Total number of study sites: 2: The Children’s Hospital of 
Philadelphia and Children’s National Medical Center 
Study Duration 
Participation will last 18  months 
All subjects will be observed for 6 months (observation period). 
Only subjects with persistently elevated TSH range (5-10 mlU/mL) 
at 6 months will be enrolled into 1 of 2 randomized groups.  
Group 1: 6 months of placebo followed by 6 months of 
levothyroxine. 
OR 
Group 2: 12 months of levothyroxine. 
 
Study Phases 
Screening 
Study Treatment 
Follow-Up   
 Screening & Recruitment: Subjects from Aim 1 (IRB# 9233) that 
qualify and consent to being contacted for future studies can be 
recruited into Aim 2.  
For subjects recruited from Aim 1, Aim 1 data will serve as baseline 
data and a screening visit is not necessary. 
Additional subjects will be recruited from CHOP’s Pediatric 
Research Consortium (PeRC), CNMC primary care clinics, DS 

 
 
 
 
 
 
 
 
 
 
viii
community events, other studies, local advertisements, local 
primary care practices, Trisomy 21 and Cardiology clinics, and 
Endocrinology clinic, where children with DS are routinely referred 
for thyroid function abnormalities.  
Subjects not recruited from Aim 1 will have an initial interview and 
a screening visit. 
We plan to recruit 29 subjects with the goal of 20 subjects who 
complete the 18 month study. 
Study Treatment: A single blinded randomized trial of 
levothyroxine treatment (starting dose 0.5 – 1 mcg/kg daily) using a 
switching replication design. Subjects will be randomized into 2 
groups of 10. 
(Group 1) Delayed treatment group: 6 months of placebo 
followed by 6 months of levothyroxine treatment 
(Group 2) Immediate treatment group: 12 months of 
levothyroxine treatment 
Follow-Up: Studies will be obtained at baseline, 6 months, 12 
months, and 18 months.  Interim thyroid studies will be checked at 3 
months, 6 weeks after randomization (month 7.5); month 13.5 and 6 
weeks after any dose adjustment.  
 
Efficacy Evaluations 
TSH 
T4 
Anthropometry (HT-Z, WT-Z, BMI-Z),  
body composition by DXA, 
PWV & PWA 
Blood lipid panel, non-HDL-C, lipoprotein subclass analysis, 
glucose, plasma insulin, HOMA-IR, hs-CRP, IL-6, PAI-1, 
adiponectin, leptin,  
QOL, Body Image, physical activity assessment.  
 
Safety Evaluations 
To monitor for safety, data will be made available to investigators 
every 3 months. The PI will not be blinded to the treatment arm, 
will be making dose adjustments, and for safety purposes, TSH and 
T4 data will be available to the PI as soon as it is available. 
Interview at 4 and 6 weeks after treatment initiation and at each visit 
to document intensity and frequency of any AE. 
Any AEs and SAEs will be reported to the CTRC and the IRB 
within the allotted time frame.   

 
 
 
 
 
 
 
 
 
 
ix
Statistical And 
Analytic Plan 
Standard mixed effects piecewise longitudinal data model (2) will be 
used to estimate in a single model both longitudinal (within subject) 
and cross-sectional (across subject) effects of the intervention. 
Cross-sectional comparisons will be possible for the second 6-
month period of observation when ½ of subjects will be switched to 
treatment while the other ½ will take a placebo.  
DATA AND SAFETY 
MONITORING PLAN 
The Biostatistics and Data Management Core (BDMC) at CHOP 
and the Investigative team will be responsible for data management. 
The Investigators will review TSH and T4 levels as they are 
available and other data every 3 months.  A data safety monitoring 
board (DSMB) will be established. The DSMB will consist of at 
least four members including a biostatistician, a developmental 
pediatrician from outside of The Children’s Hospital of 
Philadelphia, a pediatric endocrinologist, and a senior faculty 
member with experience in clinical research and in care of children 
and adolescents with complex chronic illness. The DSMB will 
choose its chair.  Specific rules and protocols for all reviews will be 
established in a formal protocol, and approved by the DSMB, prior 
to the first DSMB meeting.  The DSMB will work jointly with the 
trial statisticians and clinical investigators to establish specific 
criteria to accomplish its tasks. The Project PI will establish 
procedures for identifying classes of AEs, documenting them on an 
Adverse Experience Report (AER) form, and then reporting them 
regularly to the DSMB.    

 
x  
 
 
 
 
 
 
 
TABLE 1: SCHEDULE OF STUDY PROCEDURES: BOTH GROUPS 
Study Phase 
Observation  
Treatment 
Visit 
Study 
Visit #1  
(0 month) 
Blood Draw 
Visit A 
 (3 month) 
Study 
Visit #2  
(6 month) 
Randomiz
ation  
(no visit) 
Blood draw 
Visit B 
(7.5 month) 
Study 
Visit #3 
(12 month)  
Blood Draw 
Visit C 
(13.5 month)  
Study Visit 
#4  
(18 month) 
Study Days 
1 
 
1 
 
 
1 
 
1 
Informed 
Consent/Assent 
X (written 
consent 
for 
subjects 
not 
recruited 
from IRB 
#9233) 
X (verbal 
consent for 
subjects  
recruited 
from IRB 
#9233) 
X (written 
consent 
for 
subjects 
recruited 
from IRB 
#9233) 
 
 
 
 
 
Review 
Inclusion/Exclusion 
Criteria 
X 
 
 
 
 
 
 
 
Demographics/ Medical 
History 
X 
 
X 
 
 
X 
 
X 
Vital Signs: BP, HR, 
RR 
X 
 
X 
 
 
X 
 
X 
Height and Weight 
X 
 
X 
 
 
X 
 
X 
Pregnancy Test 
X 
 
X 
 
 
X 
 
X 
Prior/Concomitant 
Medications 
X 
 
X 
 
 
X 
 
X 
DXA Full Body  
X 
 
X 
 
 
X 
 
X 
Fasting blood glucose 
X 
 
X 
 
 
X 
 
X 
Fasting plasma insulin 
X 
 
X 
 
 
X 
 
X 
HOMA-IR 
X 
 
X 
 
 
X 
 
X 
PWV & PWA 
X 
 
 
 
 
 
 
 
Echocardiography 
X 
 
 
 
 
 
 
 

 
xi  
 
 
 
 
 
 
 
Study Phase 
Observation 
Treatment 
Visit 
Study 
Visit #1  
(0 month) 
Blood Draw 
Visit A 
 (3 month) 
Study 
Visit #2  
(6 month) 
Randomiz
ation  
(no visit) 
Blood draw 
Visit B 
(7.5 month) 
Study 
Visit #3 
(12 month)  
Blood Draw 
Visit C 
(13.5 month)  
Study Visit 
#4  
(18 month) 
Anthropometry 
X 
 
X 
 
 
X 
 
X 
QOL and Body Image 
questionnaires 
X 
 
X 
 
 
X 
 
X 
Pubertal Status 
X 
 
X 
 
 
X 
 
X 
Smoking status 
 
X 
 
 
 
 
 
 
 
Fasting lipid panel: TG, 
TC, HDL-C, LDL-C 
X 
 
X 
 
 
X 
 
X 
Non-HDL-C 
X 
 
X 
 
 
X 
 
X 
HbA1C 
X 
 
X 
 
 
X 
 
X 
PAI-1 
X 
 
X 
 
 
X 
 
X 
Hs-CRP 
X 
 
X 
 
 
X 
 
X 
IL-6 
X 
 
X 
 
 
X 
 
X 
Adiponectin 
X 
 
X 
 
 
X 
 
X 
Leptin 
 
X 
 
X 
 
 
X 
 
X 
Lipoprotein subclass 
analysis 
X 
 
X 
 
 
X 
 
X 
T4 
X 
X 
X 
 
X 
X 
X 
X 
TSH 
X 
X 
X 
 
X 
X 
X 
X 
ANTITPO 
 
X 
 
 
 
 
 
 
ANTITG 
 
X 
 
 
 
 
 
 
Physical activity 
armband (Worn for 7 
days after study visit) 
X 
 
X 
 
 
X 
 
X 
Randomization  
 
 
 
X  
 
 
 
 

 
xii  
 
 
 
 
 
 
 
Dispense study 
medication (placebo or 
levothyroxine) 
 
 
 
X  
 
 
 
 
Dispense levothyroxine 
– both groups 
 
 
 
 
 
X 
 
 
Dietary intake 
XXX 
 
XXX 
 
 
XXX 
 
XXX 
Drug Compliance 
 
 
 
 
X 
X 
X 
X 
Adverse Events  
 
 
 
 
X 
X 
X 
X 
 
 
 
 
 
 
 
 
 
 

 
page xiii 
Protocol No. XXXXX  
 
 
 
 
 
 
FIGURE 1: STUDY DIAGRAM 
 
 
 
 
 
 
                                                                     
 
 
                                                                      
 
 
 
 
 
 
 
0 month:  Study Visit #1 
3 month:  
Blood Draw Visit A 
6 month: Study Visit #2 
   
Delayed Treatment 
Group 
Immediate 
Treatment Group 
Dispense placebo  
(6-12 month) 
12 month: 
Study Visit #3  
Dispense levothyroxine (12-18 month) 
18 month: 
Study Visit #4 
Dispense 
levohyroxine  
(6-12 month) 
12 month: 
Study Visit #3 
Continue levothyroxine (12-18 month) 
18 month: 
Study Visit #4 
Subjects with persistently 
elevated TSH  
(5-10 mlU/mL) 
0-6 month: Observation Phase 
7.5 month:  
Blood Draw Visit B 
7.5 month: 
Blood Draw Visit B 
13.5 month: 
Blood Draw Visit B 
13.5 month: 
Blood Draw Visit B 
Randomization 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
1
1 
BACKGROUND INFORMATION AND RATIONALE 
 
1.1 Introduction 
The AAP recommends yearly screening of thyroid studies in Down syndrome (DS)(1). 
Clinical experience suggests that TSH concentrations in the subclinical hypothyroid range 
(5-10 mIU/L) are not uncommon in DS, but the benefits and risks of treating subclinical 
hypothyroidism (SCH) in the DS population are not known. In adults, SCH has been 
associated with increased cardiometabolic risk and individuals with DS may be at increased 
cardiometabolic risk as well. Thus, this preliminary study will explore the effects of 
levothyroxine treatment of subclinical hypothyroidsim on cardiometabolic risk and 
quality of life in pre-adolescents and adolescents with DS.  This is part of the larger 
study, IRB #9233, investigating cardiometabolic risk in the DS population.  
1.2 Name and Description of Intervention 
 Levothyroxine: Group 1 (delayed treatment) and Group 2 (immediate treatment) will 
receive a starting dose of 0.5-1 mcg/kg/day. Group 1 will receive levothyroxine treatment at 
the 12th month and continue treatment until the 18th month. Group 1 will receive a total of 6 
months of levothyroxine treatment. Group 2 will receive levothyroxine treatment at the 6th 
month and continue treatment until the 18th month. Group 2 will receive a total of 12 months 
of levothyroxine treatment. 
Placebo: Group 1 (delayed treatment) will receive placebo for the first 6 months. 
Investigators will be unblinded to permit titration of levothyroxine doses. Dose adjustments 
will be made at the discretion of the PI. TSH will be repeated 6 weeks after any dose 
adjustment. 
1.3 Findings from Non-Clinical and Clinical Studies  
1.3.1 Non-Clinical Studies:  N/A 
1.3.2 Clinical Studies 
1.3.2.1 Human Pharmacokinetics: N/A 
 
1.3.2.2 Clinical Studies in Adults 
Subclinical hypothyroidism in adults is associated with increased risk of incident ischemic 
heart disease and related mortality, an effect that may be attenuated by levothyroxine 
treatment(3). The mechanisms linking subclinical hypothyroidism and CVD remain to be 
delineated but have been reviewed by Biondi and Cooper(4). Studies of CVD risk in patients 
with subclinical hypothyroidism have shown: 1. arterial stiffness identified by pulse wave 
analysis was increased in adults(5); 2. endothelial dysfunction was present (6, 7), which 
improved with levothyroxine treatment(8); 3. and elevated CRP was present in some(9-11) but 
not all(12) studies. Data on lipogenic profiles were also conflicting: LDL and total 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
2
cholesterol(10, 13, 14) were higher in some studies (an effect potentially mediated through 
insulin resistance(15)) but not in other studies(12, 14). In an RCT, levothyroxine treatment 
didn’t improve total cholesterol and LDL, but a sub-analysis suggested benefit for subjects 
with TSH>12 mIU/L or with elevated LDL(16). Debate remains whether adults with 
subclinical hypothyroidism should be treated(17). 
 
1.3.2.3 Clinical Studies in Children 
Data in children with subclinical hypothyroidism are limited. Despite the recommendations 
to screen for thyroid dysfunction, evidence to guide management of marginally elevated 
TSH (with normal T4) in children with DS is equally sparse. In non-DS children, TSH>4.65 
mIU/L was associated with lower HDL(18). One year of levothyroxine treatment in children 
with subclinical hypothyroidism and short stature improved growth velocity(19). Left 
ventricular function and LV mass (by echocardiography) was not different in 16 children 
with DS and subclinical hypothyroidism (TSH>6.5 mIU/L; mean TSH = 7.8 mIU/L) vs. 25 
children with DS and normal TSH. However, these findings may be limited by the small 
sample size. An intervention study of 7 subjects age 2-42 years with DS and 
“hypothyroidism” defined as low T4 and normal or elevated TSH (0.2-18.9 mIU/L) on 8 
weeks of levothyroxine treatment did not improve developmental or functional outcomes. 
Anthropometrics and CMR factors were not examined (20). In contrast, increased TSH in the 
absence of overt congenital hypothyroidism is common in neonates with DS(21) and 
prompted an RCT in 181 neonates with DS. TSH-directed levothyroxine treatment was 
associated with better growth, weight gain, and motor development after 24 months 
compared to placebo(22). These findings highlight that the “asymptomatic” component of 
subclinical hypothyroidism may have medically-relevant effects. 
 
1.4 Selection of Drugs and Dosages 
Levothyroxine (T4) is a synthetic form of thyroxine, an endogenous hormone secreted by 
the thyroid gland. T4 is converted to its active metabolite, L-triiodothyronine (T3). Thyroid 
hormones (T4 and T3) then bind to thyroid receptor proteins in the cell nucleus and exert 
metabolic effects through control of DNA transcription and protein synthesis. Thyroid 
hormones are involved in normal metabolism, growth, and development, promote 
gluconeogenesis, increase utilization and mobilization of glycogen stores, stimulate protein 
synthesis, and increase basal metabolic rate.  Onset of action for the oral form of 
levothyroxine is 3-5 days.  Oral tablets are available in a number of doses:  25 mcg, 50 mcg, 
75 mcg, 88 mcg, 100 mcg, 112 mcg, 125 mcg, 137 mcg, 150 mcg, 175 mcg, 200 mcg, 300 
mcg.  In children, pills are frequently cut in half to further refine doses:  37.5 mcg, 44 mcg, 
56 mcg, 62.5 mcg, 68.5 mcg.  Younger children tend to require higher doses for body weight 
of thyroxine replacement than older children and adults with hypothyroidism (TSH >10 
mIU/L). 
Typical doses for individuals with hypothyroidism, based on age, are: age 6-12 year: 4-5 
mcg/kg/day; age > 12 years: 2-3 mcg/kg/day; adults and in older children in whom growth is 
complete: 1.7 mcg/kg/day.  For subclinical hypothyroidism, a starting dose of 0.5-1 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
3
mcg/kg/day is typical.  For the purposes of this study, subjects will be initiated on a dose 
between 0.5-1 mcg/kg/day, depending on the available tablet formulations and the PI’s  
medical discretion.TFTs will be checked 6 weeks after treatment initiation and titrated to 
achieve TSH 0.5-3 mIU/L range. 
Generic lactase tablets will be used as placebo and will be dispensed by the CHOP 
Investigational Pharmacy. 
 
1.5 Relevant Literature and Data 
The AAP recommends yearly screening of thyroid studies in DS(1)and reports a 4-18% risk 
of hypothyroidism, with risk increasing with age. The TSH threshold for defining 
hypothyroidism varies among studies(23). Symptoms suggestive of hypothyroidism--
constipation, dry skin, weight gain, and decreased growth velocity—are not uncommon in 
children with DS(23). Studies in DS suggest TSH is often higher than the normal population 
and occurs in the presence of normal thyroxine concentration, a phenomenon referred to as 
subclinical hypothyroidism or isolated hyperthyrotopinemia. The clinical relevance of 
isolated increased TSH is unknown and there is little data to guide clinical management. 
Rates of medically treated thyroid disease (>95% with thyroid hormone replacement) in 
children with DS enrolled in Tennessee Medicaid increased by 73% from 1995 to 2003 
(22.37/1000 child years) subsequent to the 2001 AAP guideline publication. They estimated 
a 10.8% population based incidence of thyroid dysfunction in 1257 children with DS(24). The 
prevalence of overt hypothyroidism was not reported, and the question remains whether 
children with DS and subclinical hypothyroidism are being treated.  
Subclinical hypothyroidism in adults is associated with increased risk of incident ischemic 
heart disease and related mortality, an effect that may be attenuated by levothyroxine 
treatment(3). As described in section (1.3.3.1), CVD risk may be higher in adults with SCH(4-
15), and levothyroxine treatment may reduce CVD risk in adults with SCH(16). 
 
Subclinical Hypothyroidism in Children and DS  
Data in children with subclinical hypothyroidism are limited. Despite the recommendations 
to screen for thyroid dysfunction, evidence to guide management of elevated TSH in 
children with DS is equally sparse. In non-DS children, TSH>4.65 mIU/L was associated 
with lower HDL(18). One year of levothyroxine treatment in short children with subclinical 
hypothyroidism and short stature improved growth velocity(19). Left ventricular function and 
LV mass (by echocardiography) was not different in 16 children with DS and subclinical 
hypothyroidism (TSH>6.5 mIU/L; mean TSH = 7.8 mIU/L) vs. 25 children with DS and 
normal TSH. However, these findings may be limited by the small sample size. An 
intervention study of 7 subjects age 2-42 years with DS and hypothyroidism, defined as low 
T4 and normal or elevated TSH (0.2-18.9 mIU/L) on 8 weeks of levothyroxine treatment did 
not improve developmental or functional outcomes. Anthropometrics and CMR factors were 
not examined(20). In contrast, increased TSH in the absence of overt congenital 
hypothyroidism is common in neonates with DS(21) and prompted an RCT in 181 neonates 
with DS. TSH-directed levothyroxine treatment was associated with better growth, weight 
gain, and motor development after 24 months compared to placebo(22). These findings 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
4
highlight that the “asymptomatic” component of subclinical hypothyroidism may have 
medically-relevant effects. This aim will provide potentially clinically relevant 
preliminary evidence for the treatment of subclinical hypothyroidism in DS. 
1.6 Compliance Statement 
This study will be conducted in full accordance with all applicable Children’s Hospital of 
Philadelphia and Children’s National Medical Center Research Policies and Procedures and 
with all applicable Federal and state laws and regulations including 45 CFR 46, 21 CFR 
Parts 50, 54, 56, 312, 314 and 812 and the Good Clinical Practice: Consolidated Guideline 
approved by the International Conference on Harmonisation (ICH). 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 and Children’s National 
Medical Center IRB 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 
 
2.1 Primary Aim 
 To determine the effect of levothyroxine replacement on non-HDL-C levels in pre-
adolescents and adolescents with DS and subclinical hypothyroidism. Primary hypothesis: 
We hypothesize levothyroxine treatment of subclinical hypothyroidism will decrease non-
HDL-cholesterol in DS  
 
 
2.2 Secondary Aim 
1) To explore the effect of levothyroxine replacement on small LDL-P, fat mass, BMI Z 
score, and inflammation in pre-adolescents and adolescents with subclinical 
hypothyroidism and DS. Hypothesis: We hypothesize thyroxine treatment of 
subclinical hypothyroidism will decrease small LDL-P, decrease fat mass, decrease 
BMI Z score, and decrease inflammation 
2) To determine the effect of levothyroxine replacement on QOL, lifestyle (physical 
activity, nutrition) and body image in pre-adolescents and adolescents with DS and 
subclinical hypothyroidism.  Hypothesis: Treatment of subclinical hypothyroidism 
will be associated with improved QOL , physical activity, and body image in pre-
adolescents and adolescents with DS and subclinical hypothyroidism. 
 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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3 
INVESTIGATIONAL PLAN 
3.1 General Schema of Study Design 
This randomized, single-blinded, intervention study will take place at the CHOP outpatient 
CTRC and at Children’s National Medical Center. Subjects with Down syndrome and 
subclinical hypothyroidism (TSH: 5-10 mIU/L) ages 8-20 years will be recruited for a trial 
of levothyroxine treatment, using a switching replication design. All subjects will be 
observed for 6 months. TSH and T4 will be obtained at the 3rd and 6th month of the 
observation period. If a subject has a TSH which has reverted to normal at 3 months, they 
will continue on to the 6-month check.  Only subjects with persistently elevated TSH (5-10 
mIU/mL) at 6 months will be enrolled in the randomized phase. While there is no standard 
definition of “persistently elevated,” in this study, we consider TSH level that is 5-10 mIU/L 
at 0 and 6 months as “persistently elevated.” If TSH is < 5 mIU/L at 0 months and the 
subject has a history of TSH 5-10 mIU/L, a repeat TSH may be drawn, at the discretion of 
the principle investigator, within 3-6 months; if TSH is < 5 mIU/mL at 6 months, a repeat 
TSH may be drawn, at the discretion of the investigator, within 1-2 weeks. If the repeat TSH 
is < 5 mIU/L, the subject will not be eligible to continue. If TSH is > 10 mIU/L during the 
first 6 months, a repeat TSH will be drawn within 1-2 weeks. If the repeat TSH is > 10 
mIU/L, thyroid dysfunction will not be considered subclinical. The subject will be referred 
to an endocrinologist for definitive therapy. After 6 months of observation, subjects will be 
randomized to: Group 1 (6 months of placebo, followed by 6 months of levothyroxine) or 
Group 2 (12 months of levothyroxine). The initial levothyroxine dose will be 0.5 – 1 
mcg/kg/day, with a goal of TSH desired range of 0.5 – 3 mIU/L. Subjects/families will be 
blinded to the treatment arm. Investigators will be unblinded to permit titration of 
levothyroxine dose. TSH and T4 will be measured after 6 weeks (month 7.5) of 
levothyroxine/placebo commencement (as a measure of efficacy of levothyroxine dose), and 
then at month 12. At month 12, levothyroxine will be initiated in Group 1 (delayed 
treatment), allowing for 6 months of treatment. Group 2 (immediate treatment) will continue 
an additional 6 months of levothyroxine (12 months total). TSH will be checked at 6 weeks 
(month 13.5) and at month 18 in both groups. Dose adjustments will be made at the 
discretion of the PI. TSH will be repeated 6 weeks after any dose adjustments, which may be 
in addition to the time points above if necessary. This switching replication design will 
permit using each child to serve as his/her own control and provides a placebo control period 
for each treated child.  The goal is for 20 subjects to complete the 18 month study. 
3.1.1 Screening Phase 
Potential subjects will be identified using the protocol inclusion and exclusion criteria. With 
their permission, subjects may be screened and recruited from Aim 1: Cardio risk factors in 
DS youth (IRB #12-009233), which is recruiting male and female subjects with Down 
syndrome in the age range of 10-20 years of age. Additional children will be recruited from 
local primary care pediatric practices and from the Division of Endocrinology where 
children with DS are routinely referred for thyroid function abnormalities. A telephone 
interview and medical record review to determine eligibility will be conducted on subjects 
who are not recruited from #9233. 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
6
3.1.2 Observation Phase 
The first 6 months of the study are the Observation Phase of the study. Only subjects with 
persistently elevated TSH during the Observation Phase will proceed to the Treatment 
Phase. Subjects that have TSH >10 mIU/L during the 6 month Observational Phase will 
have a repeat TSH test within 1-2 weeks. If the repeat TSH is >10 mIU/mL, the subject not 
be considered to have subclinical hypothyroidism and will not qualify to continue the study. 
They will be referred to an endocrinologist for treatment. Subjects who have TSH <5 mIU/L 
at the 6 month visit may have a repeat TSH test within 1-2 weeks; this will be at the 
discretion of the study investigator. If TSH or repeat TSH (if applicable) is <5 mIU/L, the 
subject will be considered to have normal levels of TSH and will not qualify to continue the 
study. Subjects who have a history of subclinical hypothyroidism, but at the 0 month visit 
have a TSH value in the normal range, may have a repeat TSH test within 3-6 months; this 
will also be at the discretion of the study investigator.  
 
Study Visit #1 (0 Month):- For subjects recruited from #9233, #9233 data will serve as 
Study Visit #1 data. Study visit #1 will not be necessary for subjects recruited from #9233.  
Subjects who have not participated in Study #9233 will complete Study Visit #1. Subjects 
and their parents/guardians will come to the research facility following a 12 hour overnight 
fast. Parental/guardian/Legal Authorized Representative consent and child assent (if child is 
deemed capable of assent) will be obtained prior to any study procedures being performed. 
Females will receive a urine pregnancy test. Females found to be pregnant will be 
disqualified from the study and will receive appropriate counseling and referral. Study 
participants will have the following measures: fasting blood samples, TSH, T4, 
anthropometric measurements, whole body DXA scan, blood pressure, pubertal assessment, 
pulse wave velocity & pulse wave analysis, echocardiography and questionnaire 
assessments. After the study visit, subjects will be asked to wear a physical activity arm 
band at home for 7 days, and a 3 day dietary recall will be done by telephone. 
Blood Draw Visit A(3 Month):– During this observation time frame, subjects will have a 
blood draw to check TSH and T4 levels. This blood draw can be done at the research facility 
or at other hospital-affiliated sites. 
Study Visit #2 (6 Month): – Subjects and their parents/guardians will present to the 
research facility following an overnight fast. At this study visit, the subject will have the 
following measures: fasting blood samples, TSH, T4, anthropometric measurements, whole 
body DXA scan, blood pressure, pubertal assessment and questionnaire assessments. 
Females will receive a urine pregnancy test. Females found to be pregnant will be 
disqualified from the study and will receive appropriate counseling and referral.  After the 
study visit, subjects will be asked to wear a physical activity arm band at home for 7 days, 
and a 3 day dietary recall will be done by telephone.  
3.1.3 Treatment Phase 
Randomization and blinding -  

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
7
Only subjects with persistently elevated TSH (5 – 10 mIU/mL) will be randomized into the 
following 2 groups of 10: Group 1 (Delayed treatment) or Group 2 (Immediate treatment).A 
3 month supply of placebo will be dispensed to subjects that have been randomized into 
Group 1 and a 3-month supply of levothyroxine will be dispensed to subjects randomized 
into Group 2.  
Blood Draw Visit B  (7.5 Month): TSH and T4 will be measured 6 weeks after the Study 
Visit #2 (approximately 7.5 months from the start of the study). Subjects will obtain a blood 
draw. Dose adjustments will be made for Group 2 as needed.  Another 3-month supply of 
placebo or levothyroxine (dose adjusted as necessary) will be mailed to the home.  Subjects 
requiring dose adjustments will have TSH and T4 in another 6 weeks to confirm/assure TSH 
is in the desired range. 
Study Visit #3 (12 Month): – Subjects and their parents/guardians will present to the 
research facility following a 12-hour overnight fast. At this study visit, the subject will have 
the following measures: fasting blood samples, TSH, T4, anthropometric measurements, 
whole body DXA scan, blood pressure, pubertal assessment and questionnaire assessments. 
Females will receive a urine pregnancy test. Females found to be pregnant will be 
disqualified from the study and will receive appropriate counseling and referral.  After the 
study visit, subjects will be asked to wear a physical activity arm band at home for 7 days, 
and a 3 day dietary recall will be done by telephone. TSH and T4 will be reviewed and 
levothyroxine will now be dispensed to both Groups 1 and 2.  Group 2 will continue on 
current dose if TSH within range; otherwise dose adjustments will be made as necessary. 
Blood Draw Visit C (13.5 Month): TSH and T4 will be measured 6 weeks after the 12 
month visit (approximately 13.5 months from the start of the study). Subjects will have 
blood drawn.  A subsequent blood draw 6 weeks later will be required for any dose 
adjustments. 
Study Visit #4 (18 Month): – Subjects and their parents/guardians will present for a final 
follow up at research facility following a 12-hour overnight fast. At this study visit, the 
subject will have the following measures: fasting blood samples, TSH, T4, anthropometric 
measurements,whole body DXA scan, blood pressure, pubertal assessment and 
questionnaire assessments. Females will receive a urine pregnancy test. Females found to be 
pregnant will be disqualified from the study and will receive appropriate counseling and 
referral.  After the study visit, subjects will be asked to wear a physical activity arm band at 
home for 7 days, and a 3 day dietary recall will be done by telephone. 
 
3.2 
Allocation to Treatment Groups and Blinding 
Randomization and allocation concealment: Children will be randomized to two groups. 
Both will be observed for 6 months. One will be switched to the intervention at 6 months, 
and continued thereafter. The other will be switched to placebo at 6 months and to the 
intervention at 12 months. Randomization will be performed centrally by the project 
statisticians and implemented using opaque sealed envelopes to ensure allocation 
concealment. Balance between treatment arms will be maintained by use of randomly 
permutated blocks with unequal block sizes and with stratification by patient gender (two 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
8
strata, male and female and age (two strata, ages 8-14 and ages 14-20). After envelopes are 
opened, treatment assignment will be blinded to the subjects (and their parents). 
3.3 
Study Duration, Enrollment and Number of Sites 
3.3.1 Duration of Study Participation 
The total study duration per subject will be between 6 to 18 months.  
3.3.2 Total Number of Study Sites/Total Number of Subjects Projected 
The study will be conducted at 2 study sites, The Children’s Hospital of Philadelphia and 
Children’s National Medical Center.  Interim blood draws may take place at satellite 
Children’s Hospital of Philadelphia and Children’s National Medical Center sites, if it is 
more convenient for the family. 
It is expected that approximately 29 subjects will be enrolled to yield 20 study completers.   
3.4 Study Population 
3.4.1 Inclusion Criteria 
Inclusion Criteria 
1. Males and females, ages 8 – 20 years 
2. Diagnosis of Down syndrome 
3. Subclinical hypothyroidism: TSH level between 5 – 10 mIU/L, normal T4 
4. Parental/guardian permission (informed consent) and if appropriate, child assent 
5. Females who are  11 years of age or who are menarchal must have a negative 
urine/serum pregnancy test 
6. Committed to adherence to levothyroxine treatment and study completion 
3.4.2 Exclusion Criteria 
Exclusion Criteria 
1. Pregnancy 
2. Type 1/Type 2 diabetes 
3. Chronic medical conditions or medication use that can affect growth, nutrition, blood 
glucose, insulin secretion, or thyroid function (such as lithium or certain seizure 
medications) 
4. Current use of levothyroxine or anti-thyroid hormone 
5.   Cyanotic congenital heart disease, or pulmonary hypertension (as described by last 
echo report in subjects with CHD), or congenital heart disease considered medically 
unstable by the study cardiologists 
Subjects that do not meet all of the enrollment criteria may not be enrolled. Any violations 
of these criteria must be reported in accordance with IRB Policies and Procedures.  

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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4 
STUDY PROCEDURES 
4.1  Screening Phase 
4.1.1 Telephone screening and medical record review 
A telephone screening and medical record review will be done to assess eligibility for the 
study. 
4.2 Observation Phase 
4.2.1 Study Visit #1 (0 Month), Blood Draw Visit A (3 Month) & Study Visit #2 (6 
Month) 
The purpose of the 3 Observation Phase visits is to observe TSH and T4 levels over a period 
of 6 months. All subjects will get a blood draw for TSH and T4 labs 3 months after the 
Study Visit #1. The TSH and T4 at 0 and 6 months will be done as part of Study Visit #1 
and #2.At the PI’s discretion, additional blood draws following the 0 and 6 month visits may 
take place to assess TSH levels. 
The following visits and procedures occur during the Observation Phase: 
Study Visit #1 (0 month) = 1 visit lasting approximately 4.5 hours 
 Informed Consent: 30 minutes/as long as needed for parent/guardian/participant to 
fully comprehend the study involvement 
 Fasting blood draw: 15 minutes 
 Pubertal Status Exam: 5 minutes 
 Vital Signs: 10 minutes 
 DXA scan: 10 minutes 
 Anthropometric measurements: 15 minutes 
 PWV and PWA: 30 minutes 
 Echocardiography: 20 minutes 
 Family history: 10 minutes 
 Questionnaire assessments (PedsQL, IWQOL, BES, FRS, ASAQ, CEBQ, CFQ, 
ABAS-II, HFI): 1.5 hours, or as long as needed by subject and parent/guardian. HFI 
will be mailed to family and completed prior to visit. 
 Physical activity armband (worn at home for 7 days) 
 3 day dietary recalls over the phone from home = Approximately 30 minutes for 
each diet recall. 
Blood Draw Visit A (3 month) = Approximately 15 minutes for blood draw 
 TSH &T4 blood draw: 15 minutes 
 
Study Visit #2 (6 month) = 1 visit lasting approximately 4 hours 
 Fasting blood draw: 15 minutes 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
10
 Pubertal Status Exam: 5 minutes 
 Vital Signs: 10 minutes 
 DXA scan: 10 minutes 
 Anthropometric measurements: 15 minutes 
 Family history: 10 minutes 
 Questionnaire assessments (PedsQL, IWQOL, BES, FRS, ASAQ, CEBQ, CFQ, 
ABAS-II, HFI: 1.5 hours, or as long as needed by subject and parent/guardian. HFI 
will be mailed to family and completed prior to visit. 
 Physical activity armband (worn at home for 7 days) 
 3 day dietary recalls over the phone from home = Approximately 30 minutes for 
each diet recall. 
4.3 Treatment  Phase 
4.3.1 Randomization 
After Study Visit #2 (6 months), only subjects with persistently elevated TSH (5-10 
mlU/mL) will be randomized into two groups of 10. Randomization will be performed 
centrally by the project statisticians and implemented using opaque sealed envelopes to 
ensure allocation concealment. Balance between treatment arms will be maintained by use 
of randomly permutated blocks with unequal block sizes and with stratification by patient 
gender (two strata, male and female) and age (two strata, ages 8-14 and ages 14-20). After 
envelopes are opened, treatment assignment will be blinded to the subjects and their 
parents/guardians. 
The following visits and procedures will occur during the Treatment Phase: 
Blood Draw Visit B (7.5 month): Approximately 15 minutes for blood draw 
TSH and T4 blood draw will be done 6 weeks after the 6 month visit (at 7.5 months) as a 
measure of efficacy of levothyroxine dose. 
Study Visit #3 (12 month) = 1 visit lasting approximately 4 hours 
 Fasting blood draw: 15 minutes 
 Pubertal Status Exam: 5 minutes 
 Vital Signs: 10 minutes 
 DXA scan: 10 minutes 
 Anthropometric measurements: 15 minutes 
 Family history: 10 minutes 
 Questionnaire assessments (PedsQL, IWQOL, BES, FRS, ASAQ, CEBQ, CFQ, 
ABAS-II, HFI): 1.5 hours, or as long as needed by subject and parent/guardian. HFI 
will be mailed to family and completed prior to visit. 
 Physical activity armband (worn at home for 7 days) 
 3 day dietary recalls over the phone from home = Approximately 30 minutes for 
each diet recall. 
Blood Draw Visit C (13.5 month):  Approximately 15 minutes for blood draw 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
11
TSH and T4 blood draw will be done 6 weeks after the 12 month visit (at 13.5 months) as a 
measure of efficacy of levothyroxine dose. 
Study Visit #4 (18 month) = 1 visit lasting approximately 4 hours 
 Fasting blood draw: 15 minutes 
 Pubertal Status Exam: 5 minutes 
 Vital Signs: 10 minutes 
 DXA scan: 10 minutes 
 Anthropometric measurements: 15 minutes 
 Family history: 10 minutes 
 Questionnaire assessments ((PedsQL, IWQOL, BES, FRS, ASAQ, CEBQ, CFQ, 
ABAS-II, HFI): 1.5 hours, or as long as needed by subject and parent/guardian. HFI 
will be mailed to family and completed prior to visit. 
 Physical activity armband (worn at home for 7 days) 
 3 day dietary recalls over the phone from home = Approximately 30 minutes for 
each diet recall. 
4.4 Blood Draw Visits B & C 
Randomized subjects will be asked to have a blood draw to check TSH and T4 levels 6 
weeks after the 6 month and 12 month visits. TSH will be repeated 6 weeks after any dose 
adjustment. The purpose to for this check is to adjust the levothyroxine dose if the PI deems 
necessary. 
4.5 Concomitant Medication 
After the participant has completed the screening and interim visits, the participant will be 
randomized to take an oral placebo or levothyroxine treatment. The initial levothyroxine 
dose will be 0.5-1 mcg/kg/day. Participants are required to bring in their full or partially full 
bottles of placebo/levothyroxine to each study visit (Study Visits #3 and 4).  Subjects can 
continue other medications; these will be recorded at each study visit. 
4.6 Subject Completion/Withdrawal 
Subjects may withdraw from the study 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 treatment, visit schedules, or AEs.  The Investigator or the Sponsor (if 
applicable) 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 CRF. 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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5 
STUDY EVALUATIONS AND MEASUREMENTS 
 
5.1 Screening and Monitoring Evaluations and Measurements 
 Fasting blood draw: This will be completed by nursing staff. A total of 
approximately 12.5 mL (approximately 2½ teaspoons) of blood will be obtained. 
 
 Pubertal Status Exam: Tanner staging of puberty will be performed by a pediatric 
endocrinologist team member. This exam will take place in a private exam room. 
 
 Vital Signs: Blood pressure, heart rate, and respiratory rate will be taken by the 
nursing staff. The blood pressure will be taken 3 times and averaged. Vital signs will 
be collected after the blood draw. 
 
 Full Body DXA Scan: The DXA scan will be performed by trained . The scan 
measures BMC, BMD, BMD-Z, fat mass, lean body mass, total mass, fat-free mass 
and percent fat.   
 
 Anthropometry: Weight will be measured by digital electronic scale (Scaletronix) 
and stature on a stadiometer (Holtain). Body proportions and fat distribution 
measurements (sitting height, arm and waist circumference, and skin fold thickness 
at triceps, biceps, subscapular, and suprailiac sites) will be measured. 
 
 Pulse Wave Velocity (PWV) and Pulse Wave Analysis (PWA): Aortic PWV and 
PWA will be measured by a trained technician. PWV and PWA will only be done at 
Study Visit #1. 
 
 Echocardiography: LV mass will be measured by echocardiography by a trained 
technician. Echocardiography will only be done at Study Visit #1. 
 
 Family History: The study coordinator/appropriate member of the investigative team 
will administer the CRF to collect relevant patient medical history, family history of 
CVD/dyslipidemia/T2DM.  
 
 Questionnaire Assessments: The following questionnaires: (PedsQL, IWQOL, BES, 
FRS, ASAQ, CEBQ, CFQ, ABAS-II, body satisfaction scales) will be administered 
by the study coordinator/appropriate member of the investigative team. Each 
questionnaire will be verbally communicated to the parent/participant, and the team 
member will show the questions as they are read, as appropriate. The HFI and 
physical activity/armband diary will be self-administered at home. 
 
 PedsQL - The Pediatric Quality of Life Inventory uses a modular 
approach to measuring health related QOL in healthy children, 
adolescents, and those with acute and chronic health conditions. Scales 
include: 1) physical functioning, 2) emotional functioning, 3) social 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
13
functioning, and 4) school functioning, which yield summary scores of a 
total scale score, a psychosocial health summary score, and the physical 
health summary score. 
 
 IWQOL-Kids - The Impact of Weight on Quality of Life – Kids 
(IWQOL-Kids) © is a validated self-report measure of weight-related 
quality of life for youth ages 11-19. This condition-specific QOL 
assessment is a 27 item questionnaire that yields a total score and 4 
domain scores, including physical comfort, body esteem, social life, and 
family relations. It has been used in children, adolescents, and in special 
populations. 
 
 IWQOL-Kids – Parent Form - The Impact of Weight on Quality of Life 
Kids – Parent Form (IWQOL-Parents) is a measure of the parent 
perception of weight-related quality of life. This condition-specific QOL 
assessment is a 27 item questionnaire that yields a total score and 4 
domain scores, including physical comfort, body esteem, social life, and 
family relations. 
 
 BES – the Body Esteem Scale for Children is a 20-item, yes/no 
questionnaire designed to assess children’s attitudes and feelings about 
their body and appearance.  
 
 FRS  – The Stunkard Figure Rating Scale depicts 9 male and 9 female 
figures, ranging in size from very thin to very overweight. It is used to 
assess perceptions of current and ideal body size. 
  
 ASAQ -   The Adolescent Sedentary Activity Questionnaire is a reliable 
assessment of a comprehensive range of sedentary behaviors that occur in 
school-aged young people. In this measure, the parent will estimate the 
amount of time spent engaging in various sedentary activities during each 
day of a typical school week and weekend. 
 
 CEBQ – The Child Eating Behavior Questionnaire is a reliable and valid, 
35 item questionnaire measuring appetite and eating style (satiety 
responsiveness, food enjoyment, food responsiveness, slowness in eating, 
food fussiness, desire to drink, emotional over-eating and emotional 
under-eating). 
 
 CFQ – The Child Feeding Questionnaire measures parental feeding 
practices, assessing parental beliefs, attitudes, and practices regarding 
child feeding. It is designed for parents of children in the age range of 2-
11 years of age. 
 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
14
 
 ABAS-II – The Adaptive Behavior Assessment System-Second Edition is 
widely used to evaluate individuals with intellectual and developmental 
disabilities measuring daily living skills (what people actually do, or can 
do, without the assistance of others). It assesses adaptive behavior in 
individuals 5-21 years of age. 
 
 Body Satisfaction Scales – The body satisfaction scales measure the level 
of satisfaction with body size, shape and weight. 
 
 HFI – The Home Food Inventory is a valid assessment of the home food 
environment. Families will be mailed the HFI prior to the study 
appointment and will be asked to bring the completed survey to the study 
appointment. 
 
 Physical activity/armband diary – Participants will be asked to keep a 
record of the times that they did not wear the physical activity armband 
and of their activities on the days that they wore the armband. The diary 
packet will be sent home with each participant, along with a postage-paid 
envelope to return the diary.  
 
 Sense Wear® Armbands (Body Media, Inc): Participants will pick up the Sense 
Wear armbands at the visit. The subject will be asked to wear the armband 
accelerometer for 7 consecutive days, 24 hours per day. The exception of wearing 
the armband is when the subject is bathing/showering or if participating in water 
activities, such as swimming. Information collected will include the amount and 
intensity of physical/sedentary activities and sleep.  
 
 3 day dietary recalls: After the study visit, a nutritionist will contact the participant 
and guardian by phone to collect information as to what the participant ate and drank 
within the last 24 hours. This will be done three times (2 weekdays and 1 weekday). 
5.1.1 Laboratory Evaluations 
A fasting blood sample will be performed for the laboratory evaluations described in section 
5.1.1.1. 
 
5.1.1.1 Table: Clinical Laboratory Tests 
Tests 
 
Lipid panel (TG, TC, 
HDL-C, LDL-C) 
 
Measured at CHOP Clinical Lab 
Non-HDL cholesterol 
Calculated as Total cholesterol – HDL-C from the lipid 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
15
panel 
Lipoprotein subclass 
analysis 
By LipoScience, Inc. (Raleigh, NC) using NMR 
spectroscopy 
Blood glucose 
Measured in duplicate at study site on NOVA Biomedical 
STAT strip meter 
Plasma immunoreactive 
insulin 
Measured in duplicate by ELISA in the CHOP CTRC 
Biochemistry Core Lab 
Insulin resistance 
Fasting insulin and glucose used to calculate Homeostasis 
Model Assessment-Insulin Resistance Index (HOMA-IR): 
[fasting insulin (ulU/mL) x fasting BG(mmol/L)]/22.5  
Thyroglobulin antibody 
Thyroid peroxidase 
antibody 
Measured at CHOP Clinical Lab 
Measured at CHOP Clinical Lab 
High sensitivity C-reactive 
protein (hs-CRP) 
Measured in the Penn CTRC TCL 
Interleukin 6 (IL-6)  
Measured at CHOP CTRC Biochemistry Core Lab using  
R&D kits 
PAI 1 
Measured at CHOP CTRC Biochemistry Core Lab using 
R&D kits 
HbA1c 
Measured at CHOP Clinical Lab 
 
 
 
 
Adiponectin & Leptin 
Both measured in duplicate at CHOP CTRC Biochemistry 
Core Lab using Alpco kits 
TSH & T4 
Measured at CHOP Clinical Lab 
5.1.1.2 Pregnancy Testing 
A urine pregnancy test will be performed for female subjects who are ≥ 11 years old or who 
are menarchal during the following visits: screening/baseline, 6 month, 12 month, and 18 
month. A positive pregnancy result will be disclosed to the participant only. Pregnancy 
results will be disclosed to the parent/guardian if the subject gives the investigators 
permission. Subjects found to be pregnant during the visit will not be able to enroll in this 
study. The investigators will counsel the subject and guide them to seek the appropriate care. 
5.2 Safety Evaluation 
Subject safety will be monitored by adverse event reporting, vital signs, physical 
examinations, and TSH levels. This protocol involves a very small radiation exposure, 
which is unlikely to cause any untoward effects to the participant. However, because the 
protocol may be unsafe to an unborn child, pregnancy is an exclusion criterion of the study. 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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6 
STATISTICAL CONSIDERATIONS 
 
6.1 Statistical Methods 
6.1.1 Baseline Data  
Data management will proceed as with Aim 1, with particular attention to obtaining all 
information on each subject at each encounter. Subject retention in our previous studies of 
children with DS has been excellent.  
Estimates will be based on a standard mixed effects piecewise longitudinal data model(2), 
implemented in the Stata v 12 program “xtmixed”, to estimate in a single model both 
longitudinal (within subject) and cross-sectional (across subject) effects of the intervention. 
Cross-sectional comparisons will be possible for the second 6-month period of observation 
when ½ of subjects will be switched to treatment while the other ½ will take a placebo. A 
piecewise model allows for a change in trajectory of an endpoint upon start of the 
intervention and for a possible shift in pattern of endpoint values soon after intervention, 
thus allowing for contrasts of both levels and trajectories. After descriptive statistics, mixed 
effects linear regression, with random intercepts for subject-specific baseline z-score, and 
random slopes (for inter-subject variation in trajectories over time), will model and compare 
outcomes over time. Inclusion of quadratic terms for time, and implementation of splines, 
will avoid unrealistic assumptions of linear changes in outcomes(25, 26). Baseline covariates 
and auxiliary variables that predict dropout will be included to adjust for any residual 
imbalance and to meet the model assumption of dropout at random. Multiple imputations 
will be used for any missing covariates(27).  
 
Primary comparisons will assume complete treatment adherence. Using the methods of 
Nagelkerke(28) and Small(29), we will estimate the effect of the intervention among those 
subjects who adhered to treatment. Robust confirmatory analysis will use generalized 
estimating equations weighted to adjust for dropout(30).  
 
Key contrasts will compare the level of each endpoint before and after treatment by 
comparing differences of model-based expected values at the end of treatment and expected 
values at baseline; expected values arise from implementation of mixed effects models. This 
approach permits adjustment for relevant changes in patient characteristics over time (e.g. 
changes in growth and pubertal status) and for any baseline factors predicting dropout. 
Alternative contrasts will compare slopes of endpoints over time prior to the intervention 
against the slopes after treatment initiation. Pre-intervention measures will improve 
statistical power.  
 
Sensitivity analyses will examine the potential impact of informative dropout(31), although 
this source of bias will be minimized through subject retention and follow-up after 
dropout(32). Supplementing models will be complete descriptive reports, including graphical 
displays of cholesterol and BMI-Z trajectories for each subject.  
 
Potential Problems/Feasibility: Dosing of levothyroxine treatment tends to be weight-
based and is not an exact science. The starting daily dose of 0.5-1 mcg/kg/day is less than 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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full replacement doses since TSH will be in the 5-10 mUI/mL range and we wish to avoid 
“over-supplementing” and suppressing TSH. TSH will be rechecked 6 weeks after 
levothyroxine initiation to confirm TSH is in the desired range. After 6 weeks some subjects 
may not have “optimal” TSH levels and dose adjustments will be required. We expect that 
by 3 months following levothyroxine initiation, all subjects will have TSH in the desired 
range, allowing an additional 3 months for the effect of intervention to be realized; in fact, 
the immediate treatment group will be followed for 12 months, over which time effects upon 
body composition and CMR factors should be apparent. This study might find that the 
relationship between subclinical hypothyroidism and CMR factors is pure association, an 
important finding as the medical community looks for evidence upon which to base 
management of “mild” TSH elevations in DS. In fact, this finding could avert unnecessary 
treatment of subclinical hypothyroidism in adolescents with DS.  
 
6.1.2 Efficacy Analysis 
Key contrasts will compare the level of each endpoint before and after treatment by 
comparing the differences of model-based expected values at the end of treatment and 
expected values at baseline. These expected values will arise out of the implementation of 
mixed effects models that will adjust for any relevant changes in patient characteristics over 
time (such as changes in growth and pubertal status) as well as for any baseline factors that 
predict dropout. Alternative contrasts will compare the trajectories (slopes) of endpoints 
over time prior to the intervention against the slopes after the start of treatment.    
6.1.3 Safety Analysis 
To monitor for safety, data will be made available to investigators every 3 months. Since the 
PI will not be blinded to treatment arm and will be making dose adjustments, TSH data will 
be available to the PI as soon as it is available.  
6.2 Sample Size and Power 
Target sample size is 20, but 29 subjects will be enrolled to allow for dropout. Assuming 
that the dependent variables are continuous, the power calculations using nonparametric 
tests that would be suitable for both continuous and ordered data are listed below: 
Correlation over 
time 
Effect size of 
interaction (SD) 
Power  
0.5 
0.7 
0.84 
0.6 
0.6 
0.71 
0.8 
0.4 
0.82 
  
6.3 Interim Analysis 
Interim efficacy or safety analyses will be done to monitor TSH levels in all subjects. 
Subjects with persistently elevated TSH (5-10 mlU/mL) will be randomized for the 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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treatment phase. If TSH is >10, a repeat test will be done within 1-2 weeks. If TSH remains 
>10, the subject will not be randomized. Subjects that have TSH <5 may have a repeat TSH 
test within 1-2 weeks; this will be at the discretion of the study investigator. If TSH or repeat 
TSH (if applicable) is <5, the subject will be considered to have normal levels of TSH and 
will not qualify to continue the study. 
 
 

 
 
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7 
STUDY MEDICATION (INTERVENTION) 
7.1 Description 
Levothyroxine sodium tablets, USP contain synthetic crystalline L-3,3',5,5'-
tetraiodothyronine sodium salt [levothyroxine (T4) sodium]. Synthetic T4 is identical to that 
produced in the human thyroid gland.   50 mcg levothyroxine tablets will be prescribed for 
anticipated doses ranging from 25 – 100 mcg/day (0.5-1 mcg/kg/day).  
The placebo is generic lactase manufactured by Major Pharmaceuticals. Each caplet 
contains 3,000 FCC units of lactase enzyme (active ingredient). Inactive ingredients are: 
crospovidone, dicalcium phosphate, magnesium stearate, mannitol, microcrystalline 
cellulose, polyalditol, silicon dioxide, sodium citrate. 
7.1.1 Packaging 
A 3 month supply of tablets will be packaged in a child safe prescription container. 
7.1.2 Labeling 
Both levothyroxine and the generic lactase tablets are available commercially and will be 
labeled appropriately by the manufacturer. The investigational pharmacy will transfer a 3-
month supply of tablets into a child-resistant vial and label with a patient-specific label that 
complies with all the federal, state and CFR requirements. 
7.1.3 Dosing 
Levothyroxine daily dose of 0.5 – 1 mcg/kg/day by mouth once per day. 
7.1.4 Treatment Compliance and Adherence 
Pill counts, monthly calendars for charting, and completion of a Supplement Adherence 
Questionnaire, a semi-structured interview, will be used to assess treatment compliance. 
Study staff will contact the families weekly to review adherence. The study psychologist 
will address with families issues that may be interfering with adherence.  
 
7.1.5 Drug Accountability 
Adequate records of study drug receipt and disposition will be maintained by the CHOP 
Investigational Pharmacy.  The purpose of these records is to ensure regulatory authorities 
and the Sponsor that the investigational new drug will not be distributed to any person who 
is not a study subject under the terms and conditions set forth in this protocol.  The study 
medication is to be prescribed by the Investigator or designee and may not be used for any 
purpose other than that described in this protocol. At study completion, all drug supplies 
including partially used and empty containers must be returned to the Study Coordinator. 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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8 
SAFETY MANAGEMENT 
 
8.1 Clinical Adverse Events 
Clinical adverse events (AEs) will be monitored throughout the study. Following 
randomization, adverse events will be monitored using: (a) Home diary of adverse events 
(AE), rated by intensity (mild, moderate, severe). (b) Interview at each study visit to 
document intensity and frequency of AE. 
8.2 Adverse Event Reporting 
The Investigator is responsible for recording and reporting unanticipated problems related to 
research that occur during and after study treatment. All on-site SAEs will be reported to the 
IRB in accordance with IRB policies. AEs that are not serious 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, 
 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. 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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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 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 Investigator Reporting of a Serious Adverse Event to Sponsor 
Reporting must be consistent with regulatory, sponsor or GCRC requirements (if applicable) 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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8.7 Medical Emergencies  
Medical emergencies that might develop during the course of the study would be referred to 
the patient’s local emergency room. 
9 
STUDY ADMINISTRATION 
9.1 Treatment Assignment Methods 
9.1.1 Randomization 
Only subjects with persistently elevated TSH (>5mlU/mL) at 6 months will be recruited for 
the randomization phase. After 6 months of observation, subjects will be randomized to one 
of two treatment arms. 
1) 6 months of placebo followed by 6 months of levothyroxine 
2) 12 months of  levothyroxine  
9.1.2 Blinding 
Subjects/families will be blinded to treatment arm. Investigators will not be blinded to 
permit titration of levothyroxine dose.   
9.1.3 Unblinding 
At the conclusion of the study, subjects/families will be notified of the treatment arm they 
received.  
9.2 Data Collection and Management 
The CHOP CTRC Informatics Core will create case report forms (CRF) and a REDCap 
database for data capture. REDCap is an NIH-supported web-based data management 
software designed by Vanderbilt University investigators. Using REDCap checking tools, 
the BDMC will confirm completeness and valid values and will inform the investigative 
team of any errors or omissions for prompt resolution. Data entered in REDCap will be 
exported for use with Stata and SAS statistical packages as needed for immediate access to 
the research team.   
The PIs are responsible for the accuracy and completeness of data collection and 
management. The PIs may designate qualified individual(s) to collect data and manage data. 
Only investigators and research staff that have completed appropriate IRB training and 
approval and are listed on the IRB approved protocol are eligible to collect and work on 
information from the study. Future studies that may use patients or data collected from this 
study must have separate approved IRB protocols and consent forms, if applicable. 
Recruitment data will be recorded onto the screening questionnaire after the verbal consent 
is recorded. Original data will be recorded directly onto CRFs by the study coordinator or a 
study investigator. Copies of laboratory, physical exam, anthropometric, DXA, PWV, and 
PWA results will be received through inter-office mailing, picked up directly, and 
sometimes through email. This information also will be recorded onto CRFs while the 
originals may be kept at the testing site (CHOP Outpatient CTRC) or with the study 
investigative staff. CRFs will be kept in a locked filing cabinet in a locked room at all times. 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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All information will be transferred to a REDCap  password-protected database, located on a 
secure sever supported by the CHOP Research Institute. The password to log onto the 
database will be unique to each member of the study team. CNMC study staff will be 
assigned to a REDcap user group that will restrict their access so they may edit solely the 
information of subjects enrolled at CNMC. The CNMC Principle Investigator will have 
access to view/edit data of all subjects. Note that only limited PHI (date of visit, date of 
birth) is included in the REDcap database. CHOP will have read-write access for data 
collected at both sites. CHOP will require access to edit records for both sites because 
several evaluations will be analyzed at CHOP, and reports including data from both CHOP 
and CNMC sites will be forwarded to the CHOP team, who will then import or enter the 
data into REDCap. Written informed consent granting permission for other site(s) to access 
PHI will be obtained from all subjects. The CRF and REDcap database will be designed by 
the CTRC Informatics Services. 
All data and records generated during this study will be kept confidential in accordance with 
institutional policies and on HIPPA subject privacy. The investigators/study team 
members/site personnel will not use such data and records for any purpose other than for 
conducting the study. 
As a way to minimize the chance of PHI (protected health information) from being 
disclosed, a unique identification code will be used for each participant. The key to this code 
will be kept in a locked file in the PI/Study Coordinator’s office.  
If any publications result from this research, the participant will not be identified by 
name/PHI. 
The information collected as part of this study will be kept for 6 years or until the 
completion of the study (whichever is longer). At that time, the information collected will be 
destroyed or all identifiable information will be removed. All keys will be destroyed at this 
time, as well. 
 
9.3 Confidentiality 
All data and records generated during this study will be kept confidential in accordance with 
Institutional policies and HIPAA on subject privacy and the Investigator and other site 
personnel will not use such data and records for any purpose other than conducting the 
study.  
No identifiable data will be used for future study without first obtaining IRB approval. The 
investigator will obtain a data use agreement between the provider (the PI) of the data and 
any recipient researchers before sharing a limited dataset (PHI limited to dates and zip 
codes).  

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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9.4 Regulatory and Ethical Considerations 
9.4.1 Data and Safety Monitoring Plan 
Study progress and safety will be reviewed quarterly (and more frequently if needed) by the 
PI. Review of the rate of subject accrual, adherence to inclusion/exclusion criteria will occur 
quarterly to assure that participants meet the eligibility criteria. There will be ongoing 
collection of data on adverse events and compliance to the treatment protocol throughout the 
study by research staff, which will be reviewed monthly by the study team and PI. If there is 
an incidental finding deemed by the PI to be clinically relevant, the primary care provider 
will be notified. The parent/guardian will be notified and asked to follow up with the 
subject’s primary care provider if during the visit a subject is noted to have blood pressure 
that is ≥ the 95th percentile for age, gender, and height. We will notify the 
parent/guardian/primary care provider with clinically relevant abnormal labs. Specifically, 
these include: lipid levels/insulin levels/blood glucose levels /TSH and T4.  Note that some 
labs, such as lipids and insulin, are run in batches and reporting of results may be delayed. 
Data Monitoring Board (DSMB)  -- Interim monitoring and analysis 
A data safety monitoring board (DSMB) will be established: (1) to protect study patients, (2) 
to safeguard their interests, (3) to monitor the overall conduct of the trial, (4) to help to 
protect the integrity of the trial, and (5) to supervise the conduct of any interim analyses.   
Specific rules and protocols for all reviews will be established in a formal protocol, and 
approved by the DSMB, prior to the first DSMB meeting.  The DSMB will work jointly 
with the trial statisticians and clinical investigators to establish specific criteria to 
accomplish its tasks. The DSMB will consist of at least four members including a 
biostatistician, a developmental pediatrician from outside Children’s Hospital of 
Philadelphia, a pediatric endocrinologist, and a senior faculty member with experience in 
clinical research and in care of children and adolescents with complex chronic illness. The 
DSMB will choose its chair.   
The Project PI will establish procedures for identifying classes of AEs, documenting them  
on an Adverse Experience Report (AER) form, and then reporting them regularly to the 
DSMB(33) . 
The DSMB will meet annually to review efficacy issues and adverse events, and will have 
additional conference calls as needed.  Annual reports will be prepared for them by the 
CHOP Data Management Core. The DSMB will independently evaluate whether adverse 
events constitute grounds to discontinue the study. 
The DSMB will report to the trial principal investigators within one month of the start of the 
DSMB meeting, or earlier if patient safety becomes an issue.   Early stopping determinations 
will be limited to considerations of safety and adverse events; owing to the small sample 
size, there will be no formal early stopping rule for superiority or futility.    
 
DSMB reporting; Independence of DSMB and study trial leadership 
All data and the interim analyses will be conducted by the DSMB and the project 
statisticians, independently from the trial leadership and staff.    The DSMB report will 
consist of the following elements: 
 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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DSMB monitoring plan and summary of report 
Major protocol changes 
Study accrual by month 
Any violations of eligibility  
Demographics 
Adherence to medication 
Dropout and loss to follow up 
Analysis of endpoints for safety  
Report of any adverse events  
Formal recommendation on early stopping 
Recommendations for notification to IRB 
 
It will be the responsibility of the trial leadership to notify the IRB or IRBs involved of any 
issues that are relative to patient safety or to early stopping of the study.  
9.4.2 Risk Assessment 
The risks of participating in this study are relatively small and the investigators believe the 
overall study is of minimal risk to the participants. 
Expected risks to the subject are as follows: 
 Blood draw may result in temporary discomfort from the needle stick, bruising, 
fainting, weakness, and rarely an infection at the site 
 Anthropometric measurements and pubertal status exam pose minimal risks 
 Exposure to radiation during DXA scan 
 Sharing of private health information (PHI) including dietary intake, demographic 
information, health history and medical information 
 Pulse wave velocity and pulse wave analysis pose minimal risks 
 12 hour overnight fasting may cause hunger pangs, upset stomach, headache, or 
light-headedness 
 Blood pressure measurement by auscultation may cause temporary 
numbness/tingling in the arm 
 SenseWear® Armbands worn for 7 consecutive days may cause skin irritability  
 Sensitive psychosocial issues may arise during questionnaire assessment 
A small amount of blood (about 3 teaspoons) will be drawn at major assessment points. 
Adolescents may experience some degree of discomfort, bruising, or lightheadedness 
with fasting blood draws. There is also a risk of infection when blood is drawn. These 
occurrences are rare, generally mild, and respond to conservative treatment. 
Participation in the study involves risks associated with the small amount of radiation 
exposure associated with the DXA whole body scan for body composition assessment; 
the total radiation EDE from DXA scans is less than 3 μSv (or 1 mrem). This total 
amount of radiation is less than the exposure to daily background radiation at sea level 
(3,000 μSv per year) and is therefore considered minimal risk. Females of childbearing 
potential undergo a urine pregnancy test prior to the DXA scans. Pregnant females do 
not have DXA scans performed in order to protect the unborn fetus. 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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The anthropometric assessment involves measurements of height, weight, 
circumferences and skinfold thickness. There is a very minor possibility of bruising 
from the skinfold thickness measurements. The exam will be performed by trained 
anthropometrists experienced in obtaining measurements in children at all levels of 
cognitive ability. The measurements are obtained in a private room. The parent is 
permitted to stay with the child if it increases their comfort with the exam. 
The puberty assessment will be performed by a pediatric endocrinologist in a private 
setting. The procedure will be explained to the child in advance, and the parent is 
permitted to be present if preferred by the child. The exam is performed by highly 
experienced personnel who are familiar with minimizing distress associated with the 
exam. 
SenseWear® Armbands (Body Media, Inc.). There is little to no risk associated with 
measuring physical activity, sedentary behavior, and sleep with armband accelerometers. 
These accelerometers fit comfortably on the participant’s arm and can easily be removed 
should he/she become uncomfortable. Some patients may experience skin irritation, 
particularly when sweating, with the devices in which case they can adjust them or in 
rare instances, remove them if necessary. To prevent irritation, parents will be instructed 
to clean the armband with mild soap and water when the child removes it to bathe or 
shower. If irritation does occur, parents will be instructed to use Aquaphor or Vaseline 
where the fabric of the band is to treat and prevent further irritation. If a child is allergic 
to metal they will not be required to complete this assessment. 
A licensed clinical psychologist with expertise in working with children and 
adolescents, will meet with families concerning sensitive psychosocial issues that may 
arise during assessments or intervention. Any emotional upset will be handled with 
appropriate support to the participants and their caregivers. Further, if a behavioral 
health concern arises during the study needing further evaluation and treatment, the 
clinical psychologist will assist with a referral. 
If a subject shows any signs of clinical instability or definitively decides to discontinue 
participation, the study visit will end. 
Protection against risks associated with thyroid hormone treatment 
 
Children and adolescents with subclinical hypothyroidism are commonly treated with 
thyroid hormone replacement, even in the absence of data supporting treatment.  
Similarly, subclinical hypothyroidism is frequently treated in the setting of DS 
despite a dearth of evidence to support or negate intervention.  The primary risk of 
thyroid hormone treatment in this 6 month observation followed by 6-12 month 
intervention study is excessive replacement leading to relative hyperthyroidism with 
TSH suppression; short-term symptoms include restlessness, irritability, tachycardia, 
palpitations, poor sleep, and weight loss. To minimize these risks: 
 
 The family will be instructed to observe and report these symptoms/signs to the 
study team 
 TSH and T4 will be checked if signs/symptoms develop 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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 TSH and T4 will be checked 6 weeks after levothyroxine initiation, the time 
required to reach steady state;  the dose will be titrated to maintain TSH in the normal 
range 
 If the initial TSH is within range, TFTs will be checked in another 14 weeks, sooner 
if symptoms arise 
 If a TSH is elevated or suppressed, the dose will be adjusted and TFTs repeated in 6 
weeks to reassess thyroid status 
 
The dose of levothyroxine replacement (0.5-1 mcg/kg/dy) is not expected to cause TSH 
suppression and symptoms of hyperthyroidism.  A dose of 1 mcg/kg/day is used in the 
treatment of subclinical hypothyroidism in adults and is much lower than typical full 
replacement doses of 4-5 mcg/kg/day for age 6-12y; 2-3 mcg/kg/day for age>12 y but 
growth and puberty incomplete; and 1.7 mcg/kg/day for children in whom growth and 
puberty are complete.  In fact, chronic suppressive doses of levothyroxine are routinely 
used following thyroidectomy and radioactive iodine ablation for thyroid cancer in adults 
and children.  For children, this treatment means TSH is maintained in the low range (TSH 
< 0.1 mIU/L).(34); the major concern is the impact of years of suppressive therapy upon 
bone health. In this study, TSH will be measured and dose adjustments made for low TSH; 
thus, any subclinical hyperthyroidism induced by levothyroxine therapy will be brief.  
Morbidity from acute levothyroxine intoxication requires massive doses of levothyroxine 
(> 5-10 mg) and is not anticipated.  The study team will remind families that, as with all 
medications, administration should be supervised by a parent and medications maintained 
out of reach of children. 
The study team will review potential side effects and toxicities with subjects by telephone 4 
weeks after treatment initiation.  If symptoms are present, TSH and T4 will be checked and 
dose adjustments made if necessary.  If symptoms are absent, TSH and T4 will be checked 
as scheduled and symptoms reviewed at 6 weeks.  All symptoms will be recorded.  The 
Investigative team will have immediate access to TSH and T4 when these data are available 
from the lab. Thyroid function data will also be reviewed monthly by the study team, to 
assess study dosing practice. 
For subjects enrolled in this study, any subject who develops TSH>10 mIU/L during the 
course of the study will have a repeat TSH test done within 1-2 weeks. If the repeat TSH 
remains >10 mIU/mL, the subject will be withdrawn from the study and referred to 
endocrinology for management of hypothyroidism.  If a suppressed TSH (TSH <0.1 
mIU/L) is identified in a child on placebo, the TSH will be repeated within 2-3 weeks.  If 
the TSH remains suppressed the subject will be referred to endocrinology for management 
of possible hyperthyroidism.  If a suppressed TSH (TSH <0.1 mIU/L) is identified in a 
child on levothyroxine, the levothyroxine dose will be decreased and the lab repeated 
within 4-6 weeks.  If the TSH remains suppressed or child has symptoms consistent with 
hyperthyroidism, the TSH will be discontinued and the TSH repeated in 6 weeks.  If the 
TSH remains suppressed the subject will be referred to endocrinology for management of 
possible hyperthyroidism, 
At the completion of the study, serum TSH and T4 will be measured, thyroid hormone 
replacement will be discontinued, and thyroid hormone results from the study reported to 
the PMD.  The subject will be directed to follow-up with the primary care provider or with 
pediatric endocrinology to determine if continued thyroid hormone replacement is 
appropriate. 
 

 
 
Protocol No. XXXXX 
 
 
 
 
 
 
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9.4.3 Potential Benefits of Trial Participation 
There may be no direct benefit and benefits will be to future patients, science, or society. 
The significant direct potential benefit from participation in this trial would be that subjects 
found to have subclinical hypothyroid disease (elevated TSH >5-10 mlU/mL) will be treated 
with levothyroxine. If a subject’s TSH is > 10 mlU/L during the observation period, a repeat 
TSH test will be done within 1-2 weeks. If the repeat TSH is >10 mIU/mL, the thyroid 
dysfunction will not be considered subclinical and the subject will be referred to an 
endocrinologist for definitive therapy.  Participants may indirectly benefit from 
identification of abnormalities such as diabetes, dyslipidemia, and hypothyroidism from the 
fasting blood draw. If clinically relevant abnormalities are found, the family will be notified. 
With the consent from the family, clinically relevant tests results will be shared with the 
subject’s primary care physician. Only studies performed by CLIA certified labs will be 
disclosed. Participants found to have an impaired fasting glucose will be referred 
appropriately for further management and treatment. 
9.4.4 Risk-Benefit Assessment 
The benefits to participation in this trial outweigh the potential risks. Levothyroxine is used 
in treating subclinical hypothyroidism with few side effects.  
9.5 Recruitment Strategy 
Subjects from the corresponding study, IRB #9233 who consent to screened for this study 
(IRB #9578) may be recruited if they meet eligibility criteria based on the information 
collected at the #9233 study visit. Subjects may also be recruited from CHOP’s Pediatric 
Research Consortium (PeRC) and Recruitment Enhancement Core, the CHOP Clinical 
Research Finder, CNMC Primary Care sites, Endocrinology and Trisomy 21 Clinics, 
Cardiology Clinics, DS community events, other studies, and through local advertisements. 
The NIH DS-Connect recruitment will e-mail registry participants the IRB-approved 
recruitment flyer.  
9.6 Informed Consent/Assent and HIPAA Authorization 
For subjects recruited from IRB #9233: 
The first #9578 visit for these subjects recruited from IRB #9233 will be Blood Draw Visit 
A. Prior to Blood Draw Visit A, verbal consent from the parent/guardian/legal authorized 
representative will be obtained and documented by the study coordinator/study investigator.  
A description of the procedures involved in Blood Draw Visit A, as well as the 
risks/benefits, will be provided verbally as part of this process. Additionally, it will be 
stressed that any questions are appropriate and that all aspects of the study are voluntary. 
Assent may not be obtained for screening purposes if the child subject is not available at the 
time of the phone call or is not cognitively able. 
If the subject is eligible to continue to Study Visit #2, written informed consent for the entire 
study will be obtained prior to conducting Study Visit #2 procedures. The written informed 
consent process will be the same as described below for subjects who are not recruited from 
IRB #9233.  

 
 
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For subjects not recruited from IRB #9233: 
Subjects not recruited from IRB #9233 will have an initial telephone screening interview; 
those who are eligible based on the study inclusion/exclusion criteria will be invited to 
participate. 
Prior to conducting the screening interview, verbal consent from the parent/guardian/legal 
authorized representative will be obtained and documented by the study coordinator/study 
investigator. A description of the procedures involved in the study, as well as the 
risks/benefits will be provided verbally as part of this process. Additionally, it will be 
stressed that any questions are appropriate and that all aspects of the study are voluntary. 
Assent may not be obtained for screening purposes if the child subject is not available at the 
time of the phone call or is not cognitively able. 
Prior to conducting the study visit procedures, written informed consent will be obtained 
from the parents/guardians of children and adolescents with DS. Assent will be obtained 
from children who, by parent report, are at a first grade or 6 year level of development or 
higher. . Study participants will be shown a picture book that depicts the study visit and at-
home procedures.  If the study participant is able to communicate that they understand the 
procedures, then written assent will be obtained if the participant is able to write, and verbal 
assent will be obtained if they are not able to write. Assent will not be obtained from 
subjects who do not have sufficient capacity.  
The consent/assent process will take place in a private consent room. It will be stressed 
again that participation in the study is voluntary and that any questions can and should be 
raised. Consent and assent will be documented by the parent/guardian/participant’s 
signatures on the approved consent documents. A description of the procedures involved in 
the study, as well as the risks and benefits, will be provided verbally as part of the consent 
process.  
Consent/assent documents will be maintained in the participant’s study file and documented. 
The parent/guardian will receive a copy of the signed document(s).   
9.6.1 Consent for Down syndrome subjects 18 – 20 years of age 
For the screening and telephone verbal consent processes, consent will not be obtained for 
Down syndrome subjects 18-20 years of age.  
Consent will be obtained from Down syndrome subjects aged 18-20, when possible, during 
the in-person written informed consent process that will occur at either Study Visit #1 or Study 
Visit #2.  Ability to provide consent will be ascertained by the study team. Initially, the team 
will assess the participants’ ability by engaging them in conversation to obtain a brief, general 
sense of their ability to comprehend and communicate. If ability to provide consent seems 
likely, the team will then explain the following to confirm that the study participant 
understands all of the essential elements of the consent form: 
a. The purpose of the research study. We will explain that the study is trying to 
understand if thyroid pill treatment will help improve the health of children 

 
 
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and adolescents who have DS and SCH. We will ask them to explain in their 
own words what the study is about. 
b. Study procedures will be explained with the aid of a picture book that depicts 
the study visit and at-home procedures. We will then ask them to explain in 
their own words what each procedure will be like. 
c. The staff will clearly explain that the study participant does not have to 
participate in the study, that they can change their mind about being in the 
study at any time, and that no one will be disappointed, upset or consider it a 
failure if they decide they do not want to continue with the study. Subjects 
will be asked to explain in their own words that they understand that they do 
not have to participate in the study and that there are no consequences if they 
choose not to.  
d. We will explain that their information will be kept private to the best of our 
ability. We will explain the risks of each procedure. We will also explain that 
they may learn more about their health from the study, and that their health 
may improve from taking the thyroid hormone pills, but that it is also 
possible that they may not benefit from the study. Participants will be asked 
to explain in their own words what they understand about the risks and 
benefits of the study. 
If it is deemed by the study team at any of the above points that the subject is unable to 
consent for themselves, the subject’s legally authorized representative/health care 
representative will provide the written informed consent on the subject’s behalf. 
Assent Procedures 
Assent will be obtained and documented on the consent form for children capable of 
assenting.  DS subjects age 18-20 years of age that are not capable of consenting for 
themselves will have assent documented in the consent form. If the capability of some of the 
participants is limited in comprehending the study and that they cannot reasonably be 
consulted, assent will not be obtained in these cases; the investigators will document it on 
the consent form. Investigators will obtain assent whenever possible. 
9.6.2 Waiver of Documentation of Consent 
Waiver of documentation of consent is being sought for screening procedures, as the 
parent/guardian/legal authorized representative may be contacted by telephone and obtaining 
a signature will not be feasible. As described in (9.6), verbal consent will be obtained and 
documented by the study coordinator/study investigator prior to conducting the screening 
interview.  
 
9.6.3 Waiver of Assent 
Waiver of assent is being sought for screening procedures, as subjects may not be present 
when parents/guardians/legally authorized representatives are contacted 

 
 
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9.7 Payment to Subjects/Families 
All study procedures and expenses will not be billed to the participant. The total 
compensation can be up to $815 for the entire study. 
Study Visit #1 (0 month): The total reimbursement for Study Visit #1 is $150.  The 
parent/guardian will receive a total of $100 in a pre-paid bankcard to offset the burdens of 
transportation/parking/time off work/babysitting fees/meals. The bankcard will be loaded 
with $50 immediately after the study visit and with an additional $50 once all post-visit 
procedures are complete.  The participant will receive a $50 gift card for their efforts toward 
the study. 
Blood Draw Visit A (3 month): For Blood Draw Visit A, the subject will receive $25 in a 
pre-paid bankcard for their time and efforts.   
Study Visit #2 (6 month): The total reimbursement for Study Visit #2 is $105. The 
parent/guardian will receive a total of $75 in a pre-paid bankcard to offset the burdens of 
transportation/parking/time off work/babysitting fees/meals. The bankcard will be loaded 
with $50 immediately after the study visit and with an additional $25 once all post-visit 
procedures are complete.   The participant will receive a $25 gift card plus a small gift worth 
$5.00 for their efforts toward the study. 
Blood Draw Visit B (7.5 month): For Blood Draw Visit B, the subject will receive $25 in a 
pre-paid bankcard for their time and efforts.   
Study Visit #3 (12 month): The total reimbursement for Study Visit #3 is $105. The 
parent/guardian will receive a total of $75 in a pre-paid bankcard to offset the burdens of 
transportation/parking/time off work/babysitting fees/meals. The bankcard will be loaded 
with $50 immediately after the study visit and with an additional $25 once all post-visit 
procedures are complete.  The participant will receive a $25 gift card plus a small gift worth 
$5.00 for their efforts toward the study. 
Blood Draw Visit C (13.5 month): For Blood Draw Visit C, the subject will receive $25 in a 
pre-paid bankcard for their time and efforts.   
Study Visit #4 (18 month): The total reimbursement for Study Visit #4 is $105. The 
parent/guardian will receive a total of $75 in a pre-paid bankcard to offset the burdens of 
transportation/parking/time off work/babysitting fees/meals. The bankcard will be loaded 
with $50 immediately after the study visit and with an additional $25 once all post-visit 
procedures are complete.  The participant will receive a $25 gift card plus a small gift worth 
$5.00 for their efforts toward the study. 
Additional  TSH & T4 Checks: In the case of dose adjustments or TSH >10, subjects will 
receive $25 in a pre-paid bankcard for each time they have their blood drawn to check TSH 
and/or T4 levels. Up to 3 TSH and T4 checks may be necessary in the event of dose 
adjustment(s) or TSH > 10. Subjects will receive up to $75.00 for these TSH & T4 checks. 

 
 
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In rare circumstances, participants may not be able to complete all study procedures on the 
date of their study visit (for example, if a machine is broken). If this occurs, we would like 
to compensate the parent/guardian with an additional $50 in a bankcard. This additional 
compensation will be an option for the 4 study visits.  
 
10 PUBLICATION 
The results of this study may be submitted for consideration for presentations at national 
meetings and/or publication in academic journals. At no time will any PHI from this study 
be disclosed for any presentation(s) or journal article(s). 
 
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