Clinical trial NCT02457897
Conditions: Insulin Resistance Diabetes Mellitus Abnormal Glucose Metabolism
Keywords: Brown Adipose Tissue Liothyronine Insulin Receptors
Sponsor: National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Start date: 2015-04-17; Completion date: 2018-09-18
Enrollment: 7 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
Background:

\- Insulin receptor mutation causes high blood sugars and sometimes diabetes complications. Researchers want to see if thyroid hormone helps.

Objectives:

\- To see if thyroid hormone treatment changes how the body handles sugar in people with insulin receptor mutation and improves blood sugar in people with diabetes.

Eligibility:

\- People ages 12 65 with an insulin receptor mutation.

Design:

* Study part 1:19-day clinic stay. Participants will be monitored for 4 days. Then for 15 days they will take a thyroid hormone pill 3 times a day. Participants will have:
* Blood tests.
* Heart rate and skin temperature monitored.
* All their food provided.
* Two 5-hour sessions in a special room. They will wear special clothes and sometimes sit still.
* Two small tubes inserted in veins. One will deliver tiny amounts of sugar and fat with a non-radioactive tracer. Participants will also drink water with a tracer. The other tube will collect blood.
* A sweet drink. Participants may have finger stick blood sugar tests.
* Glucose-monitoring device inserted into body fat for two 24-hour periods.
* Adults may have samples of fat and muscle taken.
* Heart ultrasound.
* PET-CT scan in a machine. An intravenous catheter will be placed in an arm vein. A small amount of radioactive substance will be injected.
* DEXA scan of body fat and bone density.
* Participants with poorly controlled diabetes will then take thyroid hormone at home for 6 months. They will have blood drawn and sent to the study team monthly.
* After about 3 months, they will have an overnight visit. After 6 months, they will have a 4-day visit.

Detailed description:
Background

Patients with mutations of the insulin receptor have extreme insulin resistance. This frequently results in diabetes in childhood that is extremely difficult to manage with conventional diabetes therapies, including insulin at doses 10-50 fold higher than usual. Poorly controlled diabetes, in turn, leads to microvascular complications (e.g. blindness) and early death. Hyperthyroidism, whether endogenous (e.g. Graves' disease) or exogenous, increases energy expenditure, activates brown adipose tissue, and enhances skeletal muscle perfusion, leading to enhanced glucose disposal. In a single patient with mutation of the insulin receptor and poorly controlled diabetes despite maximal therapy, iatrogenic mild hyperthyroidism for treatment of thyroid cancer resulted in normalization of glycemic control, suggesting that thyroid hormone treatment could have therapeutic benefit in this rare disease.

Aim

The purpose of this study is to determine if treatment with thyroid hormone will increase glucose disposal in patients with mutations of the insulin receptor, and thereby improve glycemic control. The hypotheses to be tested are:

1. Thyroid hormone will increase whole-body glucose disposal in patients with insulin receptor mutations.
2. This increased glucose disposal will be mediated via increased glucose uptake in brow adipose tissue (BAT) and muscle.
3. Increases in glucose disposal will result in improved glycemic control.

Methods

This study is a non-randomized pre-post design, conducted in two sequential parts. Part 1 is a short-term (2 week) proof-of-principle study to test whether thyroid hormone will increase glucose disposal in patients with insulin receptor mutations (with or without diabetes), and the mechanisms by which increased glucose disposal occurs. Part 2 is a longer term (6 month) therapeutic study to test whether thyroid hormone will result in improved glycemic control in diabetic patients with insulin receptor mutations.

Arms: arm 1: None
Interventions: intervention 1: Liothyronine
Intervention descriptions: intervention 1: Oral supplement given every 8 hours

Protocol documents:
Prot_SAP_000.pdf: 
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Thyroid hormone to induce non-insulin mediated glucose disposal in 
patients with insulin receptor mutations 
 
Principal Investigator:  
 
*Rebecca J. Brown, M.D., M.H.Sc. 
  
Associate Investigators:  
 
Monica Skarulis, M.D. 
Corina Millo, M.D. 
*Phillip Gorden, M.D. 
*Elaine Cochran, M.S.N., C.R.N.P. 
Amber Courville, Ph.D., R.D. 
*Megan Startzell, R.N., M.P.H. 
Robert J. Brychta, PhD. 
Yevgeniya Kushchayeva, M.D. 
 
Non-NIH Collaborators: 
Morey W. Haymond, M.D. (Baylor College of Medicine) 
 
Shaji K. Chacko, Ph.D. (Baylor College of Medicine) 
 
*Indicates those authorized to obtained informed consent 
 
 
 
Protocol Overview 
Protocol Title:  
Thyroid hormone to induce non-insulin mediated glucose 
disposal in patients with insulin receptor mutations 
Abbreviated Title:  
T3 in insulin receptor mutations 
IRB:  
NIDDK/NIAMS  
Research Type:  
Phase II Clinical Trial 
Multi-site Collaboration:  
No 
Intramural Collaboration: 
No 
Ionizing Radiation Use: 
(X-rays, e.g., CT; radioisotope, e.g. PET; 
etc.)  
Research Indicated  
Investigational New Drug/Device:  
No 
Patient Self-Referral Allowed:  
Yes  
List Protocol On Web:  
Yes  
Is tissue being collected for research 
purposes:  
Yes 
 
Conflict of Interest  
The protocol involves no drugs/devices/products that may lead to payments and/or royalties to be paid 
to the investigators or the NIH. 
   
The investigators have no equity, consultative, or other financial relationship with a non-NIH source 
related to this protocol which might be considered a conflict of interest.  
 
Time Frame 
Start Date:  
12/1/2014 
End Date:  
11/30/2020 
 
  


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Table of Contents 
 
Precis – Abstract _______________________________________________ 3 
Study Objectives ________________________________________________ 4 
Introduction __________________________________________________ 4 
Subject Eligibility Assessment and Enrollment _____________________________ 5 
Study Design and Statistical Analysis __________________________________ 6 
Design ___________________________________________________________ 6 
Randomization______________________________________________________ 6 
Outcome Measures ___________________________________________________ 7 
Statistical Considerations __________________________________________ 8 
Sample Size Justification _______________________________________________ 8 
Study Implementation ___________________________________________ 10 
Diet ____________________________________________________________ 10 
Medications _______________________________________________________ 10 
Glucose Management ________________________________________________ 12 
Standardized Exercise ________________________________________________ 12 
Study Procedures ___________________________________________________ 13 
Follow-up ________________________________________________________ 18 
Human Subject Protection ________________________________________ 18 
Data and Safety Monitoring Plan _________________________________________ 18 
Rationale for Subject Selection __________________________________________ 19 
Risks/Benefits Analysis including Considerations of Alternatives to Participation _________ 20 
Financial Compensation ______________________________________________ 22 
Consent/Assent Procedures ____________________________________________ 23 
Research Use, Storage and Disposition of Human Subjects’ Samples and Data ___________ 23 
References _________________________________________________ 25 
 
  
 
 


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Precis – Abstract 
 
Background 
Patients with mutations of the insulin receptor have extreme insulin resistance.  This frequently results in 
diabetes in childhood that is extremely difficult to manage with conventional diabetes therapies, including 
insulin at doses 10-50 fold higher than usual.  Poorly controlled diabetes, in turn, leads to microvascular 
complications (e.g. blindness) and early death.  Hyperthyroidism, whether endogenous (e.g. Graves’ 
disease) or exogenous, increases energy expenditure, activates brown adipose tissue, and enhances 
skeletal muscle perfusion, leading to enhanced glucose disposal.  In a single patient with mutation of the 
insulin receptor and poorly controlled diabetes despite maximal therapy, iatrogenic mild hyperthyroidism 
for treatment of thyroid cancer resulted in normalization of glycemia control, suggesting that thyroid 
hormone treatment could have therapeutic benefit in this rare disease. 
 
Aim 
The purpose of this study is to determine if treatment with thyroid hormone will increase glucose disposal 
in patients with mutations of the insulin receptor, and thereby improve glycemia control.  The hypotheses 
to be tested are: 
• 
Thyroid hormone will increase whole-body glucose disposal in patients with insulin receptor 
mutations. 
• 
This increased glucose disposal will be mediated via increased glucose uptake in BAT and 
muscle. 
• 
Increases in glucose disposal will result in improved glycemia control. 
 
Methods 
This study is a non-randomized pre-post design, conducted in two sequential parts.  Part 1 is a short-term 
(2 week) proof-of-principle study to test whether thyroid hormone will increase glucose disposal in 
patients with insulin receptor mutations (with or without diabetes), and the mechanisms by which 
increased glucose disposal occurs.  Part 2 is a longer term (6 month) therapeutic study to test whether 
thyroid hormone will result in improved glycemia control in diabetic patients with insulin receptor 
mutations. 
 
 
  
Study Objectives 
 
Part 1 (2 week study):  The primary aim of this study is to determine if triiodothyronine (T3) treatment 
will increase glucose disposal in patients with mutations of the insulin receptor.  The null hypothesis is 
that T3 will have no effect on glucose disposal.  The alternative hypothesis is that T3 will increase 
glucose disposal. 
 
Part 2 (6 month study):  The primary aim of this study is to determine if triiodothyronine (T3) treatment 
will improve diabetes control, measured as hemoglobin A1c, in patients with mutations of the insulin 
receptor.  The null hypothesis is that T3 will have no effect on A1c.  The alternative hypothesis is that T3 
will decrease A1c. 
 
 
 


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Introduction 
 
Insulin receptor mutations 
Mutations of the insulin receptor result in extreme insulin resistance and dysglycemia in humans 1,2.  
These disorders have a spectrum of clinical severity linked to the degree of residual activity of the insulin 
receptor 3.  Three clinical syndromes have been described: infants with Donohue syndrome (also called 
leprechaunism) and children with Rabson-Mendenhall syndrome (RMS) have characteristic dysmorphic 
features, and frequently have fasting hypoglycemia coupled with postprandial hyperglycemia 4, while 
patients classified as Type A insulin resistance lack dysmorphisms 5.  The majority of patients who 
survive beyond two years of age (RMS and Type A insulin resistance) develop persistent hyperglycemia 
as endogenous insulin secretion declines5,6.  Hyperglycemia in patients with insulin receptor mutations is 
extremely difficult to treat 7, and patients are at risk for early morbidity and mortality from microvascular 
complications of diabetes 5. 
 
High doses of concentrated insulin have been used to treat patients with insulin receptor mutations in an 
effort to maximize any residual insulin receptor signaling.  Recombinant insulin-like growth factor-1 
(IGF-1) has also been tried in small numbers of patients, and appears to be most effective in those with 
the mildest form of the disease 8.  In our experience, IGF-1 has not been useful in the most severely 
affected patients, and if its action is through residual insulin receptor activity, it is more logical and safer 
to use high-dose insulin.  Other conventional glucose lowering therapies, such as metformin, may be 
modestly beneficial in patients with insulin receptor mutations, but additional glucose lowering treatments 
that do not require signaling through the insulin receptor are required for effective treatment of these 
conditions. 
 
Brown adipose tissue 
Brown adipose tissue (BAT) has recently gained attention as a potential target to increase energy 
expenditure in adult humans 9,10.  Whereas white adipose tissue (WAT) functions primarily as an energy 
storage organ, BAT burns energy for thermogenesis.  It does this through the action of uncoupling 
protein-1 (UCP-1), which allows leakage across the mitochondrial electrochemical gradient, causing the 
energy generated from substrate oxidation to be released as heat rather than used to form ATP.  Major 
activators of BAT include cold, beta-adrenergic receptor (primarily 3) stimulation, and thyroid hormone 
11.  Although the primary substrate utilized by BAT is free fatty acids, BAT also takes up glucose, and 
BAT activation has the potential to increase glucose utilization in diabetic individuals. 
 
 
Role of thyroid hormone in glucose uptake 
In 2010, Monica Skarulis and colleagues reported a remarkable clinical case12:  A 32 year old woman 
with homozygous mutation of the insulin receptor and poorly controlled diabetes despite up to 3000 units 
of insulin per day developed thyroid cancer.  As part of her cancer therapy, she was treated with 
levothyroxine (T4) at doses of 200-225 mcg per day to suppress her thyroid stimulating hormone.  Over 
the next 3 years, her hemoglobin A1c fell from 9.9 to 5.6%, despite discontinuing all diabetes medications 
and maintaining a stable body weight.  As expected based on her genetic defect, her insulin resistance did 
not change, with negligible insulin stimulated glucose disposal during a hyperinsulinemic euglycemic 
clamp (500 uU/m2/min insulin infusion) both prior to and after beginning T4 treatment.  PET scanning 
performed for cancer surveillance revealed activated BAT in the supraclavicular regions; this was no 
longer visible after 2 weeks of thyroid hormone withdrawal (in the hypothyroid state), and was once again 
visible 2 weeks following reinitiation of T4.  Biopsy of a supraclavicular focus of FDG uptake showed 
histologic and gene expression patterns consistent with BAT, and biopsy of subcutaneous adipose tissue 
(a classic WAT depot) also showed gene expression patterns consistent with BAT, suggestive of 


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“browning” of this WAT depot.  This case suggests that T4 treatment led to increased BAT activity and 
volume and possibly browning of WAT, which may have resulted in increased glucose uptake and the 
remarkable clinical improvement in glycemia observed in this patient.  This case study was limited as it 
did not incorporate measurements of glucose uptake by metabolically active fat and skeletal muscle.   
 
The role of thyroid hormone excess in glucose metabolism has been investigated in several studies in 
humans with normal insulin receptor function.  In general, patients with hyperthyroidism have increased 
blood glucose in the fasting and post-prandial states 13.  This worsening of glucose metabolism appears to 
be due to worsening of post-receptor insulin resistance, particularly at the level of the liver 14, resulting in 
increased hepatic glucose production 15.  In contrast, hyperthyroid patients (compared to euthyroid 
controls) have a 3-fold increase in glucose uptake in BAT, and 1.9-fold increase in glucose uptake in 
muscle; these normalize once patients return to a euthyroid state 16.   
 
Recently, Lahesmaa et. al., reported that BAT takes up 2.7 ± 2.3 umol/100g/min of glucose in the 
hyperthyroid state, and takes up 0.9 ± 0.1 umol/100g/min of glucose in the euthyroid state.  They also 
showed that in the hyperthyroid state, skeletal muscle takes up 1.9 ± 0.6 umol/100g/min of glucose, and 
in the euthyroid state, takes up 1.0 ± 0.3 umol/100g/min of glucose.  If we assume that an average human 
possesses ~50g of BAT, and ~30,000g of muscle, then BAT will take up an excess of 0.9 umol/minute of 
glucose in the hyperthyroid vs. the euthyroid state, and muscle will take up an excess of 270 umol/minute 
of glucose in the hyperthyroid vs. the euthyroid state.  Thus, muscle is anticipated to be the major organ 
taking up excess glucose in the presence of T3 treatment. 
 
Taken together, these data suggest that, in patients with fixed insulin resistance due to insulin receptor 
mutations, thyroid hormone might enhance glucose disposal without worsening insulin resistance, 
resulting in a net improvement in glucose homeostasis. 
 
 
Subject Eligibility Assessment and Enrollment 
 
2 week study: 
Inclusion Criteria 
• 
Mutation of the insulin receptor (either recessive or dominant negative).  If mutation status is 
not known prior to enrollment, subjects will undergo genotyping at enrollment.  In the 
unanticipated event that a patient does not have a mutation of the insulin receptor, he or she 
will not complete the study and his or her data will not be included in the analysis. 
• 
Age 12 to 65 years old 
 
Exclusion Criteria 
• 
Changes in doses of diabetes medications (including metformin, insulin, sulfonylureas, 
thiazolidinediones, leptin, GLP-1 agonists, DPP4 inhibitors, etc.) in the preceding 10 weeks. 
• 
Any medical condition or medication that will increase risk to the subject (e.g. ischemic or 
structural heart disease, congestive heart failure, uncontrolled hypertension, or arrhythmia) 
or that will interfere with interpretation of study data. 
• 
Disorders that would lead to erratic gastrointestinal absorption or loss of thyroid hormone 
from the gut (severe diarrhea, celiac disease, use of bile acid sequestrants, excessive 
consumption of soybean products). 
• 
Any form of endogenous hyperthyroidism or hypothyroidism at baseline. 
• 
Current or recent (past 8 weeks) use of thyroid hormone or anti-thyroid drugs. 


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• 
Extreme disorders of thyroid hormone binding to thyroid binding globulin (excess or 
deficiency) or protein loss (nephrotic range proteinuria) that would lead to difficulties 
achieving a consistent thyroid hormone level for study.  
• 
Known presence of a rare clinical disorder that leads to thyroid hormone insensitivity 
(known T3 receptor mutations, selenocysteine insertion sequence-binding protein 2 (SBP2) 
abnormalities, monocarboxylate transporter defects). 
• 
Current use of beta blockers 
• 
Pregnancy or breast feeding 
• 
Any EKG abnormality that could increase risk of T3 treatment (resting sinus tachycardia 
(age adjusted norms), atrial fibrillation, myocardial ischemia, left or right ventricular 
excitation block, left ventricular hypertrophy or extrasystoles) 
• 
Known allergy or hypersensitivity to any form of thyroid hormone 
• 
Known adrenal insufficiency 
• 
Dependence on oral anticoagulant medications (adults only) 
• 
Use of tricyclic anti-depressants, as transient cardiac arrhythmias have been observed with 
the concomitant use of thyroid hormone. 
• 
Use of cholestyramine. 
• 
History of clinically significant osteoporosis per investigator judgment (e.g. previous 
fragility fracture) 
 
6 month study: 
Patients must meet all inclusion and exclusion criteria for the short-term study, plus have poorly 
controlled diabetes, defined as a hemoglobin A1c ≥ 7%. 
 
 
Study Design and Statistical Analysis 
 
Design 
 
This is a prospective, non-randomized study.   
 
Randomization 
Although the optimal method to study short-term effects of thyroid hormone would be a randomized, 
double-blinded, placebo controlled, cross-over study, such a study design is not feasible in this 
population. This study design would require subjects to be hospitalized for more than 4 weeks (>2 weeks 
for each study period), and is unlikely to accrue sufficient subjects for successful completion in this 
extremely rare condition. 
 
Outcome Measures 
The primary aim of this study is to determine the effect of thyroid hormone on glucose disposal over 2 
weeks and A1c over 6 months in patients with mutations of the insulin receptor.  In addition to 
quantification of glucose disposal, we plan to study effects of thyroid hormone on energy expenditure, 
substrate utilization, sympathetic nervous system tone, and gene expression in adipose tissue and muscle. 
 
• 
Primary outcome:  
• 2 week intervention: Total body glucose disposal in the fasting state measured using isotopic 
tracers 
• 6 month intervention: Hemoglobin A1c 


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• 
Secondary outcomes: 
• 2 week intervention: 
• 
Glycemic outcomes: 
• 
Tissue-specific glucose disposal in the fasting state in BAT, WAT, and 
muscle quantified using positron emission tomography (PET) in adults only 
• 
Glucose area under the curve measured using 7 point daily plasma glucose 
measurement, continuous interstitial fluid glucose monitoring, and oral 
glucose tolerance testing 
• 
Integrated assessment of glycemia over 2 weeks using hemoglobin A1c, 
fructosamine, and glycated albumin  
• 
Quantification of fractional gluconeogenesis and glycogenolysis using 
isotopic tracers 
• 
Energy Intake/Expenditure: 
• 
Resting energy expenditure at thermoneutrality (measured at 25-27 °C using 
indirect calorimetry in the metabolic chamber)  
• 
Core and peripheral body temperature 
• 
Food intake measurement 
• 
Gene expression in WAT and muscle biopsies in adults only 
 
• 6 month intervention: 
• 
Glycemic outcomes: 
• 
Tissue-specific glucose disposal in the fasting state in BAT, WAT, and 
muscle quantified using positron emission tomography (PET) in adults only 
• 
Total body glucose disposal in the fasting state measured using isotopic 
tracers  
• 
Quantification of fractional gluconeogenesis and glycogenolysis using 
isotopic tracers 
• 
Glucose area under the curve measured using 7 point daily plasma glucose 
measurement, continuous interstitial fluid glucose monitoring, and oral 
glucose tolerance testing 
• 
Total daily dose of insulin 
• 
Energy Expenditure: 
• 
Resting energy expenditure at thermoneutrality (measured using indirect 
calorimetry in the metabolic chamber)  
• 
Core and peripheral body temperature 
• 
Food intake measurement 
 
 
Statistical Considerations 
Sample Size Justification 
 
2 week study 
The primary objective of the 2 week study is to assess the impact determine the effect of thyroid hormone 
on total body glucose disposal over 2 weeks in patients with mutations of the insulin receptor. Sample 
size is determined by the change in glucose disposal.  
 
The null hypothesis is that the glucose disposal before and after thyroid hormone treatment is same.  


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The alternative hypothesis is that the glucose disposal is changed after the treatment 
 
The sample size is calculated based on previous studies in literature. A similar study by Dimitriadis et al 
14 reported a difference in total body glucose uptake between the euthyroid and hyperthyroid conditions of 
0.39 mg·kg-1·min-1 with a standard deviation of <0.27.  With this mean change and SD, a sample size of 
only 6 subjects would give 80% power with a two-sided alpha of 0.05 for this outcome.  Based on the 
assumption that results for this study will fall in a similar ranges as those published in the literature, we 
calculate the sample size dependent on the glucose disposal change and its standard deviation, which are 
given in the following table. 
 
   
Glucose 
disposal 
change 
Power=80% 
Power=90% 
SD=0.1 
SD=0.2 
SD=0.3 
SD=0.4 
SD=0.5 
SD=0.1 
SD=0.2 
SD=0.3 
SD=0.4 
SD=0.5 
0.2 
5 
10 
20 
34 
52 
5 
13 
26 
44 
68 
0.3 
4 
6 
10 
16 
24 
4 
7 
13 
21 
32 
0.4 
3 
5 
7 
10 
15 
3 
5 
9 
13 
19 
0.5 
3 
4 
6 
8 
10 
3 
5 
7 
9 
13 
 
Here we note that the sample size determined is only for studying the hypothesis about the primary 
response, total body glucose disposal. An additional power analysis was performed for a key secondary 
endpoint: the change in muscle glucose uptake on PET.  Lahesmaa et al reported that the difference in 
muscle glucose uptake in the euthyroid versus hyperthyroid condition was 1.4 with a standard deviation 
of 0.6.  With this mean change and SD, a sample size of only 4 subjects would give 80% power with a 
two-sided alpha of 0.05 for this outcome.  Sample sizes for similar results are shown in the following 
table. For example, a sample size of 10 for this two treatment crossover study will have a probability of 
80% to detect a change of glucose disposal 1.0 with standard deviation of 1.0 at a two sided 5% 
significance level.  
 
Glucos
e 
disposa
l 
change 
Power=80% 
Power=90% 
SD=0.
4 
SD=0.
6 
SD=0.
8 
SD=1.
0 
SD=1.
2 
SD=0.
4 
SD=0.
6 
SD=0.
8 
SD=1.
0 
SD=1.
2 
0.5 
8 
14 
23 
34 
48 
9 
18 
29 
44 
63 
1.0 
4 
6 
8 
10 
14 
5 
7 
9 
13 
18 
1.5 
3 
4 
5 
6 
8 
4 
5 
6 
7 
9 
 
6 month study 
The primary objective of the 6 month study is to assess the impact determine the effect of thyroid 
hormone on A1c over 6 months in patients with mutations of the insulin receptor who have poorly 
controlled diabetes.  Recent data from our group in the same population (with a different intervention)17 
indicates the A1c change 6 month after treatment is about 2.1 with standard deviation of 1.4. If we expect 
similar A1c changes in the current study, then a sample size of 6 will lead to a power of 80% (at a two-
sided alpha of 0.05).  A sample size of 7 would allow us to detect an A1c change of 2.0 with a standard 
deviation of 1.5.  
 


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Based on all of the above, we will endeavor to recruit at least 10 patients with this extremely rare 
condition into the 2 week study, and have at least 7 progress into the 6 month study.  All patients who 
complete the 2 week study and who are eligible for the 6 month study will (if they consent) proceed 
immediately from the 2 week study into the 6 month study.  As not all subjects may complete the study, 
and the sample size calculations have some uncertainty, we will cap enrollment at 20 subjects.  Currently, 
we follow 17 patients with insulin receptor mutations, of whom 10 have poorly controlled diabetes.  In 
addition, we are in touch with other physicians throughout the world who follow similar patients who 
might be interested in participating in this study. 
 
Statistical Methods for Data Analysis 
 
The following analyses will be performed: 
 
• 
Descriptive statistics of response variables by visit. This includes calculation of means and 
standard deviations of the response variables at the pre and post treatment visits.  
• 
Paired comparison of measurements before and after the treatment for primary and secondary 
outcomes  
• 
Model analysis of selected response variables 
Primary Response Variable 
Primary response variable for the 2 week study is change in total body glucose disposal from baseline to 2 
weeks after the treatment. A paired t-test will be used to compare the glucose disposal before and after 
treatment. A p-value of 0.05 or less is considered significant.  
 
The primary response variable will also be analyzed using the Linear Mixed Model (LMM). This will 
allow for comparison of glucose disposal at different visit adjusting for additional covariates. The 
potential covariates include: 
 
• 
Probable covariates 
• 
Fasting blood glucose 
• 
Fasting insulin 
• 
T3 level 
• 
Lean mass 
• 
Possible covariates 
• 
Age  
• 
Sex 
• 
Mutation type (dominant versus recessive) 
 
The effect of covariates that could influence the primary response variable will be tested by including 
each of the covariates detailed above into a mixed model.  Each model tested will only include a single 
covariate due to small sample size. 
 
The primary response variable for the 6 month study is the change in A1c from baseline to 6 month after 
the treatment. Analyses of A1c measurements are similar to that for the primary response variable.  


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Secondary Response Variables  
Secondary outcomes are described above (Outcome Measures) and will be analyzed in a similar manner 
as the primary response variables. 
 
 
Study Implementation 
Subjects will be admitted to the Metabolic Research Unit in the NIH Clinical Center.  They will remain as 
inpatients throughout the 19 day duration of the short-term study.  Upon admission, they will undergo a 
history and physical examination, and baseline labs will be obtained.  Labs used to assess subject 
eligibility in all subjects include electrolytes, BUN, creatinine, CBC, and a pregnancy test for female 
patients of reproductive potential. Baseline serum 25-hydroxyvitamin D levels will be obtained in all 
subjects. In subjects with 25-hydroxyvitamin D levels below 20 ng/mL, vitamin D supplementation will 
be provided. An EKG and echocardiogram will be performed to assess eligibility.  Height, weight, blood 
pressure, resting pulse, temperature will be obtained. 
 
Diet 
After baseline phenotyping, subjects will be placed on a metabolic diet with a fixed food 
quotient, consisting of 40% carbohydrates, 20% protein, and 40% fat.  Subjects will be asked to 
consume all food at meals, and snacks (of similar macronutrient composition to meals) will be 
provided ad libitum.  All food will be provided by the metabolic research kitchen.  Uneaten food 
will be weighed, and daily food intake (total kcal and macronutrient content) will be calculated. 
 
Medications 
 
Pre-study Medications 
With the exception of insulin (discussed below) subjects will continue their pre-admission medications 
throughout the study.  This includes oral hypoglycemic agents, and other medications either related or 
unrelated to mutation of the insulin receptor or its complications. 
 
Liothyronine (T3) 
T3 is the active form of thyroid hormone. Our rationale for choosing liothyronine (T3) over levothyroxine 
(T4) for this study is for two reasons.  First, due to the short duration of the initial proof of principle study 
(two weeks), we would prefer to use a formulation of thyroid hormone with a short half-life that can be 
rapidly titrated.  The half-life of liothyronine is a few hours, versus levothyroxine which is one week, thus 
allowing us to rapidly titrate the dose.  Secondly, we prefer to use a formulation of thyroid hormone that 
will maximally stimulate BAT without the need for conversion from T4 to T3 by type 2 deiodinase.  By 
using liothyronine as opposed to levothyroxine, we are able to study the direct effect of T3 on BAT, and 
are essentially eliminating an additional step.   
 
Replacement doses of T3 to achieve euthyroidism in hypothyroid adults are approximately 0.57 
mcg/kg/day 18.  Stable plasma T3 levels can be achieved with q8 hour T3 dosing 19. In this study, the goal 
will be to achieve peak plasma T3 levels between 125% and 150% of the upper limit of normal (200 
ng/dL in the NIH Clinical Center laboratory).  The estimated T3 dose to achieve this level of plasma T3 is 
2 times the replacement dose, or 1.14 mcg/kg/day.  In a 50 kg individual, this equals 57 mcg.  T3 is 
available in dosing increments of 5 mcg; thus doses will be rounded (lower dose will be used if calculated 
dose is 0 to <2.5 mcg above a dose divisible by 5; higher dose will be used if dose is 2.5 to <5 mcg above 
a dose divisible by 5). By comparison, T3 at doses of 150 mcg per day (~2.1 mcg/kg/day, or 


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approximately 4 times the replacement dose) was given to healthy, euthyroid adults for 2 weeks, raising 
plasma T3 levels from 125 to 507 ng/dL 14.  This treatment was well tolerated, with an increase in heart 
rate from 65 to 78 beats per minute, and weight loss of 2 kg, and the development of “mild to moderate 
symptoms of enhanced appetite, nervousness, headache, or palpitations”.  In a similar study, T3 at doses 
of 75 to 100 mcg per day (1.1 to 1.4 mcg/kg/day) was given to healthy, euthyroid adults for 2 weeks, 
raising plasma T3 levels from 110 to 350 (range, 190-520) ng/dL, with no reported adverse events. 
 
The dose titration scheme for T3 is as follows: 
• 
Day 1: 1x replacement (0.57 mcg/kg/day) 
• 
Day 2: 1.5x replacement (0.855 mcg/kg/day) 
• 
Days 3-14: 2x replacement (1.14 mcg/kg/day) 
 
Plasma T3 peak (2-4 hours, with a goal of 3 hours, after morning dose) and trough (just before morning 
dose) levels will be measured on a daily basis for the first week, and every other day thereafter.  If the T3 
peak or trough exceeds targets below, the peak and trough will be repeated the following day (or the same 
day at the investigators’ discretion), and doses will be reduced if trough or peak levels exceed targets on 
both measurements. 
• 
Target trough T3 = 150-250 ng/dL (25% above or below the upper limit of normal of 200 
ng/dL) 
• 
Target peak T3 = 250-300 ng/dL 
 
In addition, if subjects meet one or more of the following clinical and biochemical criteria the dose will 
be reduced to avoid toxicity: 
• 
Tachycardia, defined as heart rate consistently over 110 over a 24 hour period 
• 
Symptomatic palpitations 
• 
Anxiety or nervousness that are not tolerable to the patient 
• 
Development of systolic hypertension (defined as >130 mmHg AND a ≥ 10 mmHg increase 
over baseline) for a 24 hour period) 
• 
Chest pain 
• 
Congestive heart failure 
• 
Diarrhea that is distressing to patient 
• 
Weight loss > 4 kg or >10% of body weight (whichever is smaller) 
• 
Peak T3 > 350 ng/dL 
 
After the conclusion of the 2 week T3 treatment, subjects with poorly controlled diabetes will continue 
long-term (6 month) management with T3, at the maximum dose tolerated during the inpatient study (up 
to 1.14 mcg/kg/day).  Because key safety monitoring and dose titration occur during the 2 week study, 
subjects must participate in the 2 week study in order to proceed to the 6 month study.  During the next 6 
months, doses will be reduced if the following toxicity criteria are met: 
• 
Symptoms of thyroid hormone excess that are intolerable to the subject are reported by the 
patient, or elicited on monthly telephone screening, including: 
• 
Symptomatic palpitations (sensation of racing heart) 
• 
Changes in sleep, mood or behavior, school/work performance 
• 
Tremulousness, anxiety, sweating, diarrhea, or heat intolerance. 
• 
Signs of thyroid hormone excess are detected at the 3 month NIH follow-up visit, including: 
• 
Tachycardia, defined as heart rate consistently over 110 over a 24 hour period 
• 
Development of systolic hypertension (defined as >130 mmHg AND a ≥ 10 mmHg 
increase over baseline) for a 24 hour period) 


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• 
Weight loss > 4 kg or >10% of body weight (whichever is smaller), AND body mass 
index less than 25 kg/m2 (adults) or less than the 50th centile (children).  This will 
permit healthy or desirable weight loss in overweight subjects. 
• 
Trough T3 or free T4 more than 50% above the upper limit of normal on monthly safety 
laboratory evaluation 
 
Special consideration will be provided for females taking oral contraceptive pills, as levels of thyroid 
hormone may appear falsely elevated though the patients remains clinically euthyroid.  These patients 
may require further increases in doses of T3 to achieve desired peak levels. 
 
Glucose Management 
Subjects who take insulin or insulin secretagogues may be at a small risk of hypoglycemia after initiation 
of thyroid hormone due to improvements in glucose disposal.  Bedside blood glucose monitoring will be 
performed prior to meals and at bedtime (or more frequently if clinically indicated) in diabetic subjects.  
Insulin doses will be reduced on an as-needed basis to minimize hypoglycemia.  Decision making 
regarding insulin dose reduction must be individualized for each patient based on clinical expertise, as 
patients with insulin receptor mutations may have fasting or postprandial hypoglycemia even in the 
absence of exogenous insulin treatment. 
 
Standardized Exercise 
In order to maintain physical fitness during the hospitalization, sedentary subjects will engage in 30 
minutes of walking on a treadmill each day (during both Periods 1 and 2) at a mild to moderate self-
selected pace.  In subjects who habitually exercise, the duration and intensity of exercise will be increased 
to be consistent with their typical exercise routine.  This activity will be supervised by medical staff. 
 
 
 
 
 
 
 
 
Study Procedures 
 
Short-term study: 
 
Off T3 
On T3 
 Study Day 
0  1 
2 
3 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 11 12 13 14 15 
EKG & 
Echocardiogram 
X  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
OGTT 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Glucose & lipid 
turnover 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Metabolic 
Chamber  
 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
DEXA (body 
composition) 
 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
DEXA (bone 
 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


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density) 
PET-CT (age ≥ 
18y) 
 
 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
Adipose biopsy 
(age ≥ 18y) 
 
 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Muscle biopsy 
(age ≥ 18y) 
 
 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Continuous 
glucose 
monitoring 
 
 
X X  
 
 
 
 
 
 
 
 
 
 
 
X 
X 
 
Temperature 
monitoring 
 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
7 point plasma 
glucose 
 
 
X X  
 
 
 
 
 
 
 
 
 
 
 
X 
X 
 
Bedside glucose 
monitoring 
X X X X X X X X X X X X X X 
X 
X 
X 
X 
X 
Fasting research 
blood tests  
X  
 
X X  
 
 
 
 
X X  
 
 
 
 
X 
X 
Thyroid function 
X  
 
 
X X X X X X X X  
X 
 
X 
 
X 
X 
Food intake 
measurement 
X X X X X X X X X X X X X X 
X 
X 
X 
X 
X 
BP & heart rate  
X X X X X X X X X X X X X X 
X 
X 
X 
X 
X 
Body weight 
X X X X X X X X X X X X X X 
X 
X 
X 
X 
X 
Pregnancy test 
X  
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
  
 
 
 
 
 
Long-term study: 
 Study Month 
1 
2 
3 
4 
5 
6 
Thyroid function (home labs) 
X X  
X X  
Pregnancy test (females) 
X X X X X X 
Telephone contact (safety) 
X X  
X X  
NIH visit 
 
 
X  
 
X 
Pill count 
 
 
X  
 
X 
Fasting research blood tests  
 
 
X  
 
X 
Thyroid function 
 
 
X  
 
X 
Vital signs (BP, heart rate, temperature, body weight)  
 
X  
 
X 
Metabolic Chamber  
 
 
 
 
 
X 
DEXA (body composition) 
 
 
 
 
 
X 
DEXA (bone density) 
 
 
 
 
 
X 
OGTT 
 
 
 
 
 
X 
PET-CT (age ≥ 18 years) 
 
 
 
 
 
X 
Glucose & lipid turnover 
 
 
 
 
 
X 


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Bedside glucose monitoring 
 
 
 
 
 
X 
Food intake measurement 
 
 
 
 
 
X 
Echocardiogram 
 
 
 
 
 
X 
 
During the 6 month follow up period, safety labs will be obtained on a monthly basis, either locally near 
the subject’s home or at NIH.  Safety labs will consist of trough T3 level, free T4, TSH, and pregnancy 
test (females).  Monthly telephone assessments will be conducted during the months that no NIH visit is 
scheduled. This will include review of compliance, as well as screening for symptoms of 
hyperthyroidism, including changes in sleep, mood or behavior, school/work performance, tremulousness, 
anxiety, sweating, diarrhea, heat intolerance, and patient self-measurement of pulse rate and weight.  
 
For those subjects where it is not feasible (due to distance from the NIH or inconvenience) for them to 
return to the NIH for a 3 month follow-up visit, the research team will communicate with the subject’s 
primary doctor (or endocrinologist) and see if they are willing to evaluate them.  The local physicians 
would not be engaged in human subject research because they would be providing medical services that 
would typically be performed as part of routine clinical monitoring.  The physician would be requested to 
obtain the following labs: thyroid function tests, fasting glucose, insulin, c-peptide, lipid panel, free fatty 
acids and high-sensitivity C-reactive protein.  Research samples for storage would not be obtained as it 
would be difficult to ensure consistency in processing.   Additionally the clinic visit would include vital 
signs, weight and a conversation about their medication compliance.  All results would need to be faxed 
to the research team at the NIH.   
 
 
Metabolic Testing 
 
• 
DEXA scan for total body composition:  Measurements of fat-free mass will be obtained and 
used to normalize energy expenditure measurements. A DEXA scan (iDXA, GE Healthcare, 
Madison WI) will be performed to determine total and regional body fat and lean soft tissue 
masses, bone mineral content and density. DEXA produces photons at two different energy 
levels, 40 and 70 KeV. The photons pass through tissues and attenuate at rates related to 
elemental composition. Bone mineral, with highly attenuating calcium and phosphorous, is 
readily distinguished from soft tissues. The different elemental profiles of fat and bone-mineral 
free lean components allows for the analysis of soft tissue fat content, so that bone mineral, fat, 
and bone mineral fat-free lean components may be resolved.  
 
• 
Skin temperature: Wireless probes (iButtons, Maxim Inc., Sunnyvale,CA) will record skin 
temperature with data reported each minute.  The positions of the skin sensors at five sites 
(deltoid, hand, pectoralis major, anterior thigh, and shin) will be according to the ISO standards21.   
 
• 
Metabolic chamber: Study volunteers will enter the metabolic chamber at 08:00 AM after a 
minimum 8-hour fast and voiding. The temperature of the chamber will be set at between 25-27 C 
.  Study volunteers will wear standard clothing (long-sleeved hospital scrub shirt and pants, cotton 
t-shirt, subject’s own underwear; CLO value 0.55) and will be instructed to be minimally active 
while staying awake in the metabolic chamber at two predefined time periods (10:30-11:00AM 
and 11:30AM-12:00PM). To help with the compliance and occupying the time in the chamber, 
each subject will be allowed to watch television (sports, movies, series, etc), play non-active 
video games, read, or perform deskwork. Subjects will exit the chamber at 1 pm, after a 5 hour 
stay. Details of the chamber have been published previously22,23.  
 


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The metabolic chamber is a specially constructed room to assess energy expenditure.  Designed 
as a walk-in “pull” calorimeter, it is an open circuit unit that draws conditioned room air into the 
chamber at the same flow rate as it is extracted into the gas analysis system.24 The room is 
equipped with toilet and sink with privacy screen, treadmill, bed, desk, and computer with access 
to television and other forms of entertainment.  Physical activity level is measured continuously 
through a wall mounted monitoring device (microwave sensor).  Calibration tests over the past 
three years at temperature settings from 15.3 to 32.1C have demonstrated an accuracy of 
98.4±2.5% for EE as compared to a weighed propane standard oxidized by combustion.   
 
The relative humidity of each chamber is controlled between 30-50% for all temperatures. The 
overhead circulation fan unit mixes the air of the chamber uniformly. At the bed and the recliner 
where the subjects will be spending the 5 hour resting periods, the wind speed is less than 0.4 m/s 
(0.8 mile/hour), making the windchill and humidex (heat index) factors less than 0.2C. We will 
measure chamber wall temperature using iButtons,25 and calculate the exact “operational 
temperature” as defined previously26. 
 
• 
PET-CT:  
In adult subjects, dynamic PET image data of the upper torso will be acquired by a Siemens mCT 
PET/CT scanner (Siemens Medical Solutions, Hoffman Estates, IL).  A 5 mCi dose of [18F]FDG 
will be injected IV at approximately 7:30 am after a 12 hour fast.  Patients will be instructed to 
keep movement to a minimum until after the scan is completed.  The temperature in the PET 
scanner room is maintained at normal room temperature (~20-22 C), and will be monitored and 
recorded.  First, an attenuation CT scan of the upper torso will be performed, followed by 
dynamic PET image of the same area (usually requiring 2 levels, each of 22 cm) initiated at the 
time of F18-FDG IV injection and acquired as described below, up to about 65 min post tracer 
administration. 
 
The field of view (usually 2 levels, lower-mid chest and upper chest-neck) will include:  
• 
Brown adipose tissue (cervical/supraclavicular) 
• 
White adipose tissue (subcutaneous in the chest wall) 
• 
Muscle (periscapular region) 
• 
Heart/left ventricle (for calculation of arterial input functions, described below) 
 
Data acquisition will be performed alternating between the 2 levels of the torso described above, 
with a total of ten 30-second frames, ten 3-minute frames, and six 5-minute frames, for a total of 
65 minutes from the time of FDG administration.  To facilitate parametric modeling of the BAT, 
WAT, and muscle tracer kinetics, arterial input functions will be derived from left ventricular 
(LV) region of interest (ROIs) drawn on the dynamic PET images. The LV ROIs will be defined 
from a sum of the early 30-second PET frames, with the aid of the accompanying CT scan, being 
careful to minimize the quantitative bias caused by partial volume effects.  Tracer time-activity 
curves for the regions of interest will be then obtained for the entire duration of the PET study. 
These will be used to calculate the fractional rate of FDG uptake (Ki). 
 
Regions of interest for quantification of tissue glucose uptake will be defined as follows: 
• 
BAT will be defined visually in on-treatment scan, when BAT is anticipated to be active 
in all subjects due to stimulation by T3.  The BAT ROI will then be mapped to the pre-
treatment scan, so that glucose uptake is quantified in the same region for both studies.  
• 
WAT will be defined based on CT imaging.  In lean subjects, it may not be possible to 
define acceptable ROIs for WAT, as the small size of these depots may result in 


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unacceptable volume averaging effects.  In subjects with adequate WAT, the same ROI 
will be evaluated in the pre- and post-treatment scans. 
• 
Skeletal muscle ROIs will be visually defined based on CT imaging.  The same ROI will 
be evaluated in the pre- and post-treatment scans. 
 
 
The rate of glucose uptake will be calculated by multiplying Ki by the plasma glucose 
concentration of the study subject, and dividing this by the lumped constant (LC) of adipose 
tissue of 1.14 27 (for BAT and WAT), or skeletal muscle of 1.2 28. 
 
• 
Glucose and Lipid Turnover: Stable isotope tracers will be used to measure glucose and lipid 
turnover.  Endogenous glucose production will be measured using steady-state infusion of [6,6-
2H2]glucose tracer.  The fractional rate of gluconeogenesis will be measured using steady-state 
oral dosing of deuterated water (2H2O) 29. The rate of lipolysis will be measured using steady-
state infusion of deuterium-labeled 2H5-glycerol, and the fatty acid turnover rate will be measured 
using steady-state infusion of [U-13C16] palmitate.  Subjects will fast after 8 pm.  A total of 3 
grams per kg lean body mass of 2H2O will be given in four divided doses every two hours at 9 
pm, 11 pm, 1 am, and 3 am in order to enrich the subject’s body water pool to approximately 
0.5% 2H2O.  Beginning at ~5 am,  a primed [6,6-2H2]glucose infusion will be given for 3 hours, 
after which blood samples to measure isotope enrichment will be measured over a period of 30 
minutes at steady state.  Two hours after the [6,6-2H2]glucose infusion begins, a primed 2H5-
glycerol infusion, and unprimed [U-13C16] palmitate infusion will be given for one hour, and 
isotope enrichment will be measured over a period of 30 minutes at steady state.  Subjects will 
empty their bladders immediately prior to the 30 minute steady state period, and urine will be 
collected at the conclusion of the steady state, for calculation of urinary glucose excretion during 
the steady-state period. 
 
• 
Oral glucose tolerance test:  After a 12-hour fast preceding the test, subjects will be given 75 
gram (1.75gm/kg in patients less than 40kg) oral glucose solution. Venous glucose, plasma 
insulin and C-peptide will be obtained from blood samples drawn at -10, 0, 30, 60, 90, 120 and 
180 minutes during the oral glucose tolerance test.  The area under the glucose, insulin and C-
peptide will be evaluated.  
 
• 
Continuous glucose monitoring: A continuous glucose monitoring device will be inserted 
subcutaneously to measure interstitial fluid glucose every 5 minutes.  Area under the curve for 
glucose will be calculated using the trapezoidal method. 
 
• 
7-point plasma glucose: Plasma glucose will be measured prior to each meal, 2 hours after each 
meal, and at bedtime.  Area under the curve for glucose will be calculated using the trapezoidal 
method. 
 
• 
Fasting research blood tests:  Fasting glucose, insulin, C-peptide, lipid panel, free fatty acids, and 
high-sensitivity C-reactive protein will be measured every other day. Additional blood samples 
will be stored for future analysis. 
 
• 
Muscle biopsies: Muscle biopsies will be performed twice (once in the off-T3 period, and once in 
the on-T3 period) in adult subjects to study expression of molecules important in glucose 
homeostasis. This will be performed in adult subjects only, and subjects will have the ability to 
opt out of participating in the biopsies.  If participants choose not to participate in the biopsies, 


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they can still continue with participation in the other elements of the protocol. Muscle biopsies 
(100-250 mg) will be obtained under local anesthesia from the vastus lateralis using a Bergstrom 
needle. Connective tissue will be removed. A representative portion of the tissues will be 
formalin fixed, paraffin embedded, sectioned, and stained with hematoxylin and eosin using 
standard histological methods. The remaining tissue will be separated into aliquots of 
approximately 50 mg for RNA isolation and protein extraction, flash frozen in liquid nitrogen and 
stored at -80ºC.  Analyses in muscle will include measurement of gene expression (mRNA) for 
glucose transporters, irisin, UCP-3, and mitochondrial glucose-3-phophate dehydrogenase.  In 
addition, RNAseq analyses will be performed as a hypothesis-generating experiment to look for 
changes in gene expression profile in the off versus on-T3 periods.  Subjects with low platelet 
counts (<100 K/uL) and/or abnormal coagulation profiles (INR>2.0), or subjects on anti-
coagulants, will be excluded from the muscle biopsies. 
 
• 
Adipose tissue biopsies: Subcutaneous adipose tissue and muscle biopsies will be performed 
twice (once in the off-T3 period, and once in the on-T3 period) in adult subjects to study 
expression of molecules important in glucose homeostasis. This will be performed in adult 
subjects only, and subjects will have the ability to opt out of participating in the biopsies. If 
participants choose not to participate in the biopsies, they can still continue with participation in 
the other elements of the protocol.  Approximately one gram of adipose tissue will be removed 
from the abdominal or gluteal region by aspiration with a 16-gauge needle under local anesthesia 
(2% xylocaine). Slides will be examined visually to assess cell populations/tissue composition. 
The remaining tissue will be separated into aliquots of approximately 500 mg for RNA isolation 
and protein extraction, flash frozen in liquid nitrogen and stored at -80ºC for future studies.  
Analyses in fat will include measurement of gene expression (mRNA) for markers of 
beige/brown adipose tissue, including UCP-1, PRDM16, DIO2, etc. Subjects with low platelet 
counts (<100 K/uL) and/or abnormal coagulation profiles (INR>2.0), or subjects on anti-
coagulants, will be excluded from the adipose tissue biopsies. 
 
Thyroid Evaluation 
• 
Thyroid function will be assessed as serum levels of total and free thyroxine, triiodothyroinine 
(T3), and thyroid stimulating hormone (TSH).   
• 
Thyroid biomarkers: SHBG, osteocalcin, lipid NMR 
 
Safety Testing 
• 
EKG will be performed at baseline to rule out any rhythm abnormalities that might increase risk 
associated with T3 treatment. 
• 
Echocardiogram will be performed at baseline to evaluate structural abnormalities that would 
increase risk associated with T3 induced heart rate increase.  Examples of structural abnormalities 
that would exclude patients include valvular heart disease, left ventricular hypertrophy, and 
hypokinesis suggestive of ischemia. Repeat echocardiogram will be performed after 6 months to 
evaluate any changes that may have occurred with T3 treatment. 
• 
Blood pressure and heart rate will be measured 3 times daily during the 2 week study and during 
each inpatient visit (at 3 and 6 months) during the 6 month study.  Blood pressure (right arm) and 
heart rate will be measured using an automated device after subjects have been seated in quiet 
room for 10 minutes.  Measurements will be repeated at 2 minute intervals for a total of 3 
measurements, and the mean of the last 2 measurements will be recorded (excluding the first 
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• 
Body weight will be measured during inpatient visits in the morning before breakfast and after 
emptying the bladder.  Three body weight measurements will be obtained, with the subject 
stepping off the scale between measurements.  The mean of the 3 measurements will be recorded. 
• 
Urine or serum pregnancy testing will be obtained on female subjects with reproductive potential 
after admission, and will be repeated prior to radiation exposure as needed and on a monthly basis 
during the 6 month follow up.  Effective contraception (including abstinence, barrier methods, 
hormonal contraception, or IUD, will be required for female subjects with reproductive potential.) 
• 
DEXA for bone density:  Because hyperthyroidism is associated with decreased bone mineral 
density, DEXA for bone density at the AP spine, hip, and 1/3 radius will be performed at baseline 
and after 6 months of T3 in patients with poorly controlled diabetes participating in the long-term 
component of the study.  Bone density will not be assessed after 2 weeks of T3 in patients 
participating in only the 2-week portion of the study, as it is extremely unlikely that changes will 
be seen with this short duration of hyperthyroidism. 
 
Follow-up 
Post-Study Treatment 
After completion of this study, subjects may continue to receive care through NIH via protocol 76-DK-
0006 (Studies of Molecular Genetics of Insulin Secretion, Insulin Action and Diabetes Mellitus) or 
protocol 03-DK-0257 (Effect of Leptin Therapy in the Treatment of Severe Insulin Resistance). 
 
Post-Study Obligations 
There are no anticipated post-study obligations. 
 
Human Subject Protection 
 
Data and Safety Monitoring Plan 
The collection, monitoring and analysis of adverse events will be the responsibility of the Principal 
Investigator and the investigative team.  Overall accrual and adverse event information will be reported to 
the NIDDK/NIAMS IRB annually.  Study procedures will be subject to audits and/or monitoring visits to 
ensure compliance with the protocol and applicable regulatory requirements consistent with the NIDDK 
quality assurance program plan.  Audit and/or monitoring visit results will be reported to the Principal 
Investigator for further reporting as appropriate.   Study documents and pertinent hospital or clinical 
records will be reviewed to verify that the conduct of the study is consistent with the protocol plan.  The 
collection, monitoring and analysis of adverse events will be the responsibility of the Principal 
Investigator and the investigative team. 
 
Adverse Events, Protocol Deviations, and Unanticipated Problems 
Adverse events, protocol deviations, unanticipated problems (UP), serious adverse events, sponsor and 
serious, are defined as described in NIH HRPP SOP 16 (“Reporting Requirements for Unanticipated 
Problems, Adverse Events and Protocol Deviations.”). All adverse events occurring during the study, 
including those observed by or reported to the research team, will be recorded and submitted to the 
NIDDK/NIAMS IRB at each annual review. Serious unanticipated problems and serious protocol 
deviations will be reported to the IRB and CD as soon as possible but not more than 7 days after the PI 
first learns of the event. Not serious unanticipated problems will be reported to the IRB and CD as soon as 
possible but not more than 14 days after the PI first learns of the event. Non-serious protocol deviations 
will be reported to the IRB (as soon as possible but not more than 14 days after the PI first learns of the 
event) only if they represent a departure from NIH policies for the conduct of human subjects research, 
adversely affect the health care of the subject(s) or compromise the interpretation or integrity of the 


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research.  Non-serious protocol deviations that result from normal subject scheduling variations or 
technical issues associated with sampling that does not impact the health of the subject or the 
interpretation of the study data will not be reported. 
 
Deaths will be reported to the Clinical Director and IRB within 7 days after the PI first learns of the 
event.  All deaths that have occurred among study participants since the previous review will be 
summarized at the time of continuing review. 
 
Rationale for Subject Selection 
Subjects eligible for this study include adults and children who are 12 years or older with mutations of the 
insulin receptor.  Children less than 12 years of age will be excluded because they are unlikely to possess 
the emotional maturity needed to undergo the intense testing regimen, and because young children with 
insulin receptor mutations are less likely to have overt diabetes or complications of diabetes. Adolescents 
age 12-17 will be included because overt diabetes (and its complications) commonly develops during 
puberty in this patient population, and they are likely to be able to comply with study procedures.  
Because each subject will be compared to him/herself in this within-subjects design, the effects of inter-
subject heterogeneity will be less important. 
 
For adult participants, the short-term component of this study is of greater than minimal risk, and has no 
prospect of direct benefit. For adult participants, the long-term component of this study is of greater than 
minimal risk, and with prospect of direct benefit to subjects.  For minor participants, the short-term 
component of this study involves a minor increase over minimal risk and has no prospect of direct benefit, 
but is likely to yield generalizable knowledge about the subjects’ disorder or condition. For minor 
participants, the long-term component of this study is of greater than minimal risk, and with prospect of 
direct benefit to subjects. 
 
Withdrawal Criteria 
• 
Withdrawal of consent.  A subject wishes to withdraw from the study as stated in the informed 
consent (all subjects reserve the right to withdraw from the study without prejudice). 
• 
Adverse event.  A subject experiences an adverse event that in the investigator’s opinion 
necessitates withdrawal from the study.  For example, the investigators would withdraw a subject 
if he/she developed intolerable symptoms of hyperthyroidism despite T3 dose reduction. 
• 
Investigator decision.  An investigator feels it is in the subject's best interest to terminate 
participation. The detailed reasoning behind this decision will be documented. 
• 
Protocol violation.  Includes subject noncompliance, pregnancy, study entry criterion violation, 
or start of an unacceptable concomitant medication. 
 
Risks/Benefits Analysis including Considerations of Alternatives to Participation 
 
Benefits 
There are no direct medical benefits to either minor or adult subjects from participating in the initial 2 
week T3 treatment.  For subjects participating in the 2 week study, study procedures represent a minor 
increment over minimal risk, and are likely to yield generalizable knowledge about the subject’s 
condition.  Subjects with diabetes who continue T3 for 6 months have the potential to experience 
clinically meaningful improvements in diabetes control, which could reduce the risk of vascular 
complications of diabetes. 
 
Risks/Discomforts 


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• 
General:  Some patients may find the time needed to complete the research studies an 
inconvenience in their routine lives. 
 
• 
T3:  
• 
Hyperthyroid symptoms: Subjects treated with supraphysiologic doses of T3 are likely to 
experience some symptoms of hyperthyroidism.  These include palpitations, sleep 
disturbance, weight loss, anxiety, mood change, tremor, sweating, heat intolerance, 
menstrual irregularities, and diarrhea. In most cases, these symptoms are expected to be 
mild.  If these symptoms are not tolerable to subjects, or exceed defined toxicity criteria, 
the dose of T3 will be reduced.  Likewise, subjects may experience increases in systolic 
blood pressure or heart rate.  If these exceed toxicity criteria, the dose of T3 will be 
reduced.  Because the T3 dose will be titrated to avoid toxicity during the 2 week study, 
the risk of meeting toxicity criteria (especially for blood pressure and heart rate) during 
the 6 month study is minimized.  Thus, subjects will not be required to conduct home 
blood pressure or heart rate monitoring during the 6 month study. 
• 
Decreased bone mineral density:  Hyperthyroidism increases bone turnover, which over 
the long term can result in decreased bone mineral density.  This effect is not different in 
children versus adults.  This is unlikely to result in clinically meaningful increases in 
fracture risk over a 6 month period.  If the 6 month treatment with T3 results in clinically 
meaningful improvements in diabetes control, the benefits of T3 would likely outweigh 
the risks related to decreased bone density.  Bone density changes of the 6 month 
treatment period will be monitored using DEXA.  If dietary intake is inadequate or 
Vitamin D levels are low, subjects will be prescribed appropriate supplementation. 
Subjects participating in the short-term part of the study will undergo one DEXA for 
bone mineral density, and those participating in the 6 month study will have a total of two 
DEXA scans for bone mineral density. 
• 
Cardiovascular disease:  Long-term hyperthyroidism may place patients at increased risk 
of cardiovascular events.  If the 6 month treatment with T3 results in clinically 
meaningful improvements in diabetes control, the benefits of T3 would likely outweigh 
the risks related to cardiovascular disease. Our rationale for this is that patients with 
insulin receptor mutations likely have a decreased risk of macrovascular complications of 
diabetes, because the insulin receptor mutations protects them from de-novo lipogenesis 
in the liver, resulting in a favorable lipid phenotype (high HDL and low triglycerides), in 
opposition to what is seen in typical type 2 diabetes mellitus and the metabolic syndrome.  
We do not have good data on the actual incidence of macrovascular disease in this 
population, as most of these patients unfortunately don’t live long enough to develop 
macrovascular complications.  Thus, due to the favorable lipid phenotype and the severity 
of this life-limiting disease, we believe that the benefits of T3 treatment outweigh the 
risks.  Patients with baseline EKG or echocardiographic abnormalities that might place 
them at risk for acute cardiovascular events will be excluded.  Changes in cardiac 
function will be monitored via echocardiography after 6 months. 
 
• 
Blood Sampling:  Peripheral blood draws (venipuncture) performed during this study for research 
will not exceed 10.5 mL/kg, or 550 mL (whichever is smaller) per 8-week period for adults.   For 
pediatric patients, blood draws will not exceed 5 mL/kg in a single day, or 9.5 mL/kg or 550 mL 
(whichever is smaller) per 8 week period.  Patients may experience some discomfort at the site of 
the needle entry, and there is a risk of bruising at the site.  There is a remote risk of fainting or 
local infection.   
 


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• 
Studies involving radiation: Adult patients will undergo up to 318F-FDG PET-CT scans (2 in the 
short-term study and 1 in the long-term study) , each using 5 mCi of 18 fluorodeoxyglucose and 
an attenuation CT scan of the upper torso.  Adult and pediatric subjects will undergo up to 5 
DEXA scans (3 in the short-term study and 2 in the long-term study), each having an effective 
radiation dose of 0.00003 rem. Children under age 18 will only undergo DEXA scans and will not 
undergo the 18F-FDG PET-CT scans. The total radiation exposure for different groups of subjects 
is as follows: 
• 
Adults, short-term and long-term study: 1.83 rem (3 PET scans + 5 DEXA scans) 
• 
Adults, short-term study only: 1.22 rem (2 PET scans + 3 DEXA scans) 
• 
Minors, short-term and long-term study: 0.00015 rem (5 DEXA scans) 
• 
Minors, short-term study only: 0.00009 rem (3 DEXA scans) 
These are below the guideline of 5 rem per year allowed for adult research subjects or 0.5 rem per 
year allowed for pediatric research subjects by the NIH Radiation Safety Committee.  The 
average person in the United States receives a radiation exposure of 0.3 rem per year from natural 
sources, such as the sun, outer space, and the earth's air and soil.  Aside from the radiation 
exposure, the risks/inconveniences associated with the PET scan are related to claustrophobia and 
intravenous injection. The discomfort of lying in a supine position for an extended period of time 
may exist. 
 
• 
Glucose and Lipid Turnover: The stable isotopes 13C is not associated with any toxicity at the 
doses used in these studies 30.  Deuterium at the doses given may cause temporary dizziness.  This 
risk will be minimized by dividing the 2H2O dose into four aliquots, and by giving the doses at 
night while subjects lie in bed.  If subjects need to get out of bed (e.g. to use the bathroom) the 
patient will first sit upright in bed for a few minutes prior to standing, and a nurse will be 
available to supervise and assist.  Palmitate must be complexed to albumin prior to delivery, and 
thus carries the risks associated with use of human blood products.  To minimize exposure to 
human albumin, uniformly labeled 13C16 palmitate will be used, permitting use of the minimum 
possible dose of this tracer. 
 
• 
Metabolic Chamber: Besides inconveniences that can reasonably be expected as a result of 
spending an extensive time (up to 5 hours) in the live-in room calorimeter, the serious risk to 
subjects' health is minimal. 
 
• 
Fat biopsy: This will be performed only in adult subjects.  The major risks/discomforts include 
pain, bruising, hematoma, infection, scarring, and localized lipodystrophy.  Patients on anti-
platelet agents, anti-coagulants or aspirin for cardioprotection will be excluded from both the 
skeletal muscle and fat biopsy. Patients will be asked to refrain from taking aspirin or NSAIDS 
(for analgesia) for 10 days before the biopsy. The procedure will be performed under sterile 
technique to minimize the chances of infection. Local anesthetic will be used to minimize pain.  
Ice will be applied to the site immediately after the procedure to limit bruising, swelling and 
tenderness. After subcutaneous tissue biopsy, patients will be monitored by nurses. Post-biopsy 
the incision site will be cleaned and closed with adhesive wound closures and covered with gauze 
and translucent dressing tape. Study participants will be instructed to report to the clinical staff 
any changes at the biopsy site including bleeding, secretion, erythema, pain, and signs and 
symptoms of infection. Study participants will be instructed to self-monitor the incision site after 
discharge from the Clinical Center. 
 
• 
Muscle biopsy: This will be performed only in adult subjects. The major risks/discomforts 
include pain, bruising, hematoma, bleeding, infection, and scarring. The risk of excess bleeding is 


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increased if the subject takes aspirin, non-steroidal anti-inflammatory drugs (NSAIDS), anti-
platelet agents or anti-coagulants.  Patients on anti-platelet agents, anti-coagulants or aspirin for 
cardioprotection will be excluded from both the skeletal muscle and fat biopsy. Patients will be 
asked to refrain from taking aspirin or NSAIDS (for analgesia) for 10 days before the biopsy. The 
procedure will be performed under sterile technique to minimize the chances of infection. Local 
anesthetic will be used to minimize pain.  Ice will be applied to the site immediately after the 
procedure to limit bruising, swelling and tenderness. After muscle biopsy, patients will be 
monitored by nurses. The incision site will be cleaned and closed with adhesive wound closures 
and covered with gauze and translucent dressing tape. Direct compression will be applied for 10 
minutes. Study participants will remain on bed rest with bathroom privileges for 2-4 hours post-
procedure. Study participants will be instructed to report to the clinical staff any changes at the 
biopsy site including bleeding, secretion, erythema, pain, and signs and symptoms of infection. 
Study participants will be instructed to self-monitor the incision site after discharge from the 
Clinical Center.   
 
Alternatives to participation 
Participation in clinical trials is completely voluntary.  Refusal to participate will not affect a subject's 
ability to participate in other studies at NIH or elsewhere. 
 
Financial Compensation 
All subjects will receive financial compensation for their time per Clinical Center guidelines for on-site 
visits, and additional compensation will be provided for specific procedures based on inconvenience 
units, as follows: 
 
 
 
 
Short-term (2 
week) study 
3 month visit 
6 month visit 
Procedure 
Incon-
venience 
Units 
Compensation 
Count 
Total 
Count Total Count Total 
Inpatient stay 
n/a 
40 
18 
720 
2 
80 
4 
160 
Metabolic 
chamber stay 
2 
20 
2 
40 
 
 
1 
20 
PET/CT 
10 
100 
2 
200 
 
 
1 
100 
DEXA 
1 
10 
2 
20 
 
 
1 
10 
Tracer studies 
2 
20 
2 
40 
 
 
1 
20 
Adipose biopsy 
5 
50 
2 
100 
 
 
 
 
Body Sensor 
Measurements 
1 
10 
2 
20 
 
 
 
 
Heart Rate 
Monitoring 
1 
10 
2 
20 
 
 
 
 
Muscle biopsy 
5 
50 
2 
100 
 
 
 
 
OGTT 
4 
40 
2 
80 
 
 
1 
40 
Maximum Total (adults) 
Maximum Total (children) 
1340 
  860 
 
80 
80 
 
350 
250 
 
 
Consent/Assent Procedures 


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Written consent/assent will be obtained from each subject after detailed explanations of the planned 
procedures by the principal or an associated investigator. Consent for participation of minors will be 
obtained from two parents/guardians of minor subjects.  If both parents/guardians of minor subject cannot 
travel to the NIH Clinical Center, consent will be obtained via phone from the parent/guardian who is 
unable to travel.  Written assent will be obtained from all minor subjects after detailed explanations of the 
planned procedures by the principal or an associated investigator.  The consent/assent process will take 
place prior to any study procedures.  Subjects have the right to withdraw participation from this protocol 
at any time. We do not anticipate enrolling non-English speaking subjects, however, we will not exclude 
them either.  Should someone present who is non-English speaking and qualifies for enrollment in the 
study, we plan to request IRB approval for the use of the short form consent process (per NIH SOP 12) to 
obtain study consent for this patient population. 
  
 
Research Use, Storage and Disposition of Human Subjects’ Samples and Data 
For future reference and potential use, we will store all samples (blood or fluids) in our locked freezers 
for an unlimited period of time. Samples will be labeled with coded identifiers linked to patient identity 
only via a secured database. Research records and data with personal identifiers will be stored in our 
locked offices, the medical record department, and the electronic study database. This material will 
additionally be protected by medical record and computer access procedures.  Access to records and data 
associated with personal information will be restricted to the Principal Investigator, Co-Investigators, 
study support staff, and database support staff. 
 
Stored samples and/or data may be sent to outside collaborating laboratories, or shared with other NIH 
collaborating investigators, to study questions related to lipodystrophy or its complications (including, for 
example: glucose metabolism, diabetes, obesity, weight, appetite, steatohepatitis, and lipid metabolism). 
Samples may be sent to outside commercial laboratories for analysis.  Samples and data sent to outside 
laboratories and collaborators for analysis and/or testing will contain only coded numbers, without 
personal identifiers. Tech Transfer agreements will be completed before the exchange of samples and/or 
data with outside collaborators. 
 
Subjects may request that unused samples be removed from our freezers and returned to the subject, or be 
destroyed.  If no such request is made, we will keep samples until they are completely used or no longer 
of scientific value, at which time they will be destroyed. We do not plan to destroy personal medical 
information or stored data. The Principal Investigator will report loss or destruction of data or samples to 
the IRB.  
 
Material Transfer Agreement/Collaboration(s): 
Collaboration has been established with Dr. Morey W. Haymond, a board-certified pediatric 
endocrinologist and Professor of Pediatrics and Medicine at Baylor College of Medicine.  Our direct site 
contact will be Shaji Chacko, Ph.D.   The purpose of the collaboration is to assist in the conduct of the 
clinical studies determining the rates of glucose production and gluconeogenesis in subjects with 
mutations of the insulin receptor. This will include the analysis of coded human subject samples and 
interpretation of the results. 
 
 


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Appendix A:  Investigator Qualifications and Roles 
 
Rebecca J. Brown, M.D., M.H.Sc. is a board-certified pediatric endocrinologist, and an Assistant 
Clinical Investigator of the Diabetes Endocrinology, and Obesity Branch. Dr. Brown has six years of 
experience conducting clinical studies in children adults with diabetes, and has a Master’s degree in 
Clinical Research. She is the Principal Investigator of this study, and is responsible for the study design, 
implementation, and interpretation. 
 
Elaine K. Cochran, M.S.N., CRNP, is a nurse practitioner in the Diabetes, Endocrinology and Obesity 
Branch, and is lead associate investigator of the ongoing study of leptin treatment in lipodystrophy 
patients.  She is an expert in the administration of leptin to patients with lipodystrophy.  Her role in this 
project will be to assist in clinical management and education of patients. 
 
Amber Courville, PhD, RD, CSSD, is a registered dietitian working for the NIH Clinical Center.  Dr. 
Courville’s role in this project will be to assess subjects’ energy needs and coordinate the provision of a 
controlled metabolic research diet to the subjects during the study. 
 
Phillip Gorden, M.D. is a board-certified endocrinologist, Senior Investigator in the Diabetes, 
Endocrinology and Obesity Branch, and Director Emeritus of the NIDDK.  He has over four decades of 
experience conducting clinical studies in patients with rare disorders of extreme insulin resistance, and is 
Principal Investigator of the ongoing study of leptin treatment in lipodystrophy patients.  His role in this 
project will be to work with Dr. Brown on study strategy, design, and interpretation. 
 
Megan Startzell, RN, MPH is a research nurse in the Diabetes, Endocrinology and Obesity Branch.  Ms. 
Mattingly’s role in this project will be to assist Dr. Brown with the coordination of patient visits, patient 
education and communication. 
 
Corina Millo, M.D. is a board-certified radiologist specializing in nuclear medicine and Positron 
Emission Tomography (PET).  Her role in this project will be to work with Dr. Brown in the planning, 
implementation, and interpretation of PET scans for quantification of tissue glucose uptake. 
 
Monica Skarulis, M.D. is a board-certified endocrinologist, and Special Volunteer with the Diabetes, 
Endocrinology and Obesity Branch.  She has extensive expertise in the study and management of thyroid 
disorders.  Her role in this project is to work with Dr. Brown on study design and interpretation, and 
management of T3 treatment. 
 
Robert Brychta, Ph.D. holds a doctorate in biomedical engineering and is a staff scientist for the 
Diabetes, Endocrinology, and Obesity Branch. He has extensive expertise in the study and measurement 
of human metabolism. His role in the project will be to assist Dr. Brown in the measurement, analysis, 
and interpretation of energy expenditure using the metabolic chamber, body composition, and body 
temperature. 
 
Yevgeniya Kushchayeva, M.D. is a clinical fellow in Endocrinology with the Diabetes, Endocrinology 
and Obesity Branch.  Her role in this project is to work with Dr. Brown on the study implementation and 
interpretation. 
 


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