Tissue Selective Estrogen Complex to Prevent Metabolic Dysfunction in Women
Phase: EARLY_PHASE1
Conditions: Menopause
Sponsor: Pennington Biomedical Research Center

1
March 18, 2020
PROTOCOL
Tissue selective estrogen complex to prevent metabolic 
dysfunction in women
(RISE)
Principal Investigator:
Eric Ravussin, Ph.D.
Medical Investigator:
Frank Greenway, M.D.
Version Number:
Eleven
Version Date:
March 18, 2020
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Table of Contents
BACKGROUND
3
STUDY DESIGN
4
STUDY OVERVIEW
4
HYPOTHESIS
4
POWER CALCULATION
4
TIMELINE
4
STUDY POPULATION
5
TARGET POPULATION
5
NUMBER OF PARTICIPANTS
5
INCLUSION CRITERIA
5
EXCLUSION CRITERIA
5
RECRUITMENT
5
STUDY DESCRIPTION
6
LOCATION
6
CONSENT PROCESS
6
SCREENING PROCESS
6
RANDOMIZATION
6
STUDY VISITS
6
STUDY PROCEDURES
7
DATA MANAGEMENT
8
DATA AND BIOSPECIMEN STORAGE
8
STATISTICAL ANALYSIS
9
CONFIDENTIALITY AND PRIVACY
9
BENEFITS, RISKS, AND SAFETY
9
POTENTIAL BENEFITS
9
RISKS
10
MONITORING SUBJECT SAFETY
12
COMPENSATION
12
WITHDRAWAL OF SUBJECTS
12
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BACKGROUND
With the dramatic increase in life expectancy, many women will spend a large part of their lives 
in a post-menopausal state. Apart from causing degeneration of the cardiovascular, skeletal and 
central nervous systems, estrogen deficiency also increases risk for metabolic syndrome, type 2 
diabetes (T2D) and non-alcoholic fatty liver diseases (NAFLD). Overall, the role of estrogen 
deficiency to the pathophysiology of chronic metabolic diseases in women is emerging as a 
novel therapeutic challenge that parallels the increased risk associated with conventional 
estrogen replacement protocols. It is against this backdrop that novel therapeutic strategies 
targeting estrogen receptor in non-reproductive tissues may offer therapeutic benefits that 
reduce metabolic dysfunction associated with both ovarian failure and estrogen deficiency.
Selective estrogen receptor modulators (SERMs) are compounds that exert tissue-selective 
estrogen receptor (ER) agonist or antagonist activity. For example, bazedoxifene (BZA) is a 
novel SERM that exhibits estrogen agonist activity in bone but estrogen antagonist activity in 
breast and uterus. Tissue-selective estrogen complexes (TSECs) are drugs in which a SERM 
and an estrogen are combined to produce mixed estrogen agonist and antagonist activity.7 The 
goal of a TSEC regimen containing BZA with conjugated equine estrogens (CE) is to provide 
the benefits of estrogen such as reducing hot flashes and vulvar–vaginal atrophy, preventing 
menopausal osteoporosis and promoting favourable effects on cardiovascular risk while 
simultaneously protecting the endometrium and breast from estrogen stimulation without the 
need for a progestin. The TSEC therapy combining CE and BZA was approved in September 
2013 by the Food and Drug Administration [Duavee™, Pfizer, Inc] for treatment of 
postmenopausal symptoms and prevention of osteoporosis.
Dr. Mauvais-Jarvis’ laboratory was a pioneer in demonstrating that estrogen protects pancreatic 
islet β-cell insulin secretion and synthesis, nutrient homeostasis, and survival via three estrogen 
receptors, ERα, ERβ and the G-protein coupled ER. They showed that ER actions were 
relevant to human islet function and survival, and had major clinical impacts in the diabetes field 
by enabling the design of a novel estrogenic peptide that targets β-cells without unfavorable 
gynecological effects in females. Most recent preliminary data, derived from a mouse model 
of post-menopausal metabolic syndrome, demonstrate that TSEC prevent estrogen 
deficiency-induced metabolic dysfunction – including obesity, T2D and NAFLD- as 
efficiently as CE alone and importantly, without causing endometrial hyperplasia. Because 
SERMs like Tamoxifen or Raloxifene have never shown such a beneficial metabolic effect in 
preclinical models and in humans, these data underscore the clinical significance of this line 
of investigation and they also provide a firm rationale for the next series of experiments that 
are proposed in this application. 
We propose to assess the therapeutic potential of TSEC in preventing metabolic dysfunction in 
postmenopausal women and to explore the hypothesis that the metabolic effect of TSEC relies 
on the activation of the production and sensitivity of the hepatic hormone fibroblast growth factor 
21 (FGF21). These findings will have a direct and immediate scientific impact that fits with the 
LACaTS Center theme. The knowledge that will be generated from this work will fill key gaps in 
our understanding of the mechanism of estrogen action on metabolic diseases that will be used 
for therapeutic purposes. Thus, there is direct, clinically relevant translational impact to the 
proposed work. As part of this LACaTS grant, Dr. Mauvais-Jarvis will continue to investigate the 
potential molecular mechanisms of TSEC on the metabolic syndrome in mice only. 
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There are several innovative features to our proposed study. First, a novel feature of our 
approach is that translational studies of TSEC action in preventing metabolic dysfunction in 
postmenopausal women have never been performed. A second, innovation is to propose a 
novel paradigm in which the metabolic actions of TSEC are mediated via a stimulation of FGF21 
production and action.
STUDY DESIGN
Study Overview
The goal of this pilot clinical study is to perform a randomized crossover study to test the 
beneficial effect of 8 weeks of treatment with TSEC or BZA/CE/(a clinical image of Duavee™) 
vs. placebo on insulin sensitivity and energy metabolism in eight post-menopausal women. 
There will be a two-month washout period in-between the 2 treatments (TSEC vs. Placebo). The 
recruitment will be performed at the Pennington Biomedical Research Center with the help in 
recruiting from the Baton Rouge General Obstetrics and Gynecology Physicians (see below). 
The drugs will be provided by Pfizer, Inc. without charge.
Objective:  Assess the therapeutic potential of TSEC in preventing metabolic dysfunction in 
postmenopausal women.
Primary Endpoint:  Insulin sensitivity by a 2-step hyperinsulinemic euglycemic clamp
Secondary Endpoints:   a) Resting metabolic rate by a metabolic cart (Deltatrac) and;
      b) Body composition by DXA and MRS
Exploratory Endpoints:  Molecular work (at Tulane) in animal model only.
Hypothesis
TSEC treatment will improve both whole body insulin sensitivity (high insulin dose) and liver 
insulin sensitivity (suppression of HGP) in postmenopausal women.
Power Calculation
Although a formal power analysis was not feasible in this pilot study, in light of the high 
precision that is achieved using the clamp, we can expect similar improvements to be 
detectable with up to 10 or less completers for whole body and hepatic insulin sensitivity 
respectively in this randomized crossover trial. 
Timeline
After passing the screening process, the expected duration of an individual subject’s 
participation in the study is about 26 weeks.
We anticipate that it will take up to 6 months to complete enrollment for the study.  
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STUDY POPULATION
Target Population
Post-menopausal women (<5y post last period) aged 50-60 years old 
Number of Participants
We expect up to 10 post-menopausal women to complete this study.
Inclusion/Exclusion Criteria
Inclusion Criteria
Exclusion Criteria.
Post-menopausal women (<5y post last 
period)
Amenorrhea other causes (excess 
androgen)
Age between 50-60y
Diabetes mellitus
Symptomatic (hot flashes, vaginal 
dryness) or asymptomatic
Medications: (diabetes, antipsychotics, oral 
steroids, weight loss drugs†)
History of major depressive episodes 
requiring treatment
Tricyclic antidepressants (TCAs)
Current smokers, or having smoked within 
last 3 months
BMI 30-40 kg/m2
≤ 3 month washout of birth control pill, 
estrogen, and/or progestin
Hysterectomy (total and partial)
Normal mammogram past 12 months
Contraindications to estrogen treatment ††
Physician clearance (Ob/Gyn, Baton 
Rouge General Physician Primary Care 
Network, or PBRC)
Be willing to accept own blood during IV 
procedure
Unable or unwilling to do an MRS
Additional exclusion for optional BIA 
procedure only:

Medications: Diuretics, glitazones

Congestive heart failure

Chronic kidney disease

Pacemaker or other metal implants
† other chronic medications are acceptable as long as stable for 2 months, and may need MI 
approval before screening.
†† for unusual vaginal bleeding, blood clots, hepatic disease, bleeding disorder, past/present 
history of breast or uterine cancer, pregnant, breastfeeding 
Recruitment
Baton Rouge General Obstetrics and Gynecology Physicians will be the primary site for 
recruiting patients for the study. Gynecology and the Baton Rouge General Physician Primary 
Care Network will also be asked to provide additional recruits through a series of educational 
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presentations by Dr. Evelyn K. Hayes, MD, FACOG, NCMP. At these clinics, patients are seen 
for routine wellness exams in addition to problem visits. Women in the target population for 
recruitment frequently present with symptoms of estrogen deficiency. Over the past 12 months, 
the Baton Rouge General Obstetrics and Gynecology practice has seen over 800 women aged 
50-60 with a BMI between 30 and 40 and with menopause according to the definition adopted in 
this study. Further delineation of such group can identify those women who are naïve to 
estrogen and symptomatic with hot flashes and/or vaginal dryness. After an explanation of the 
study by Dr. Hayes or her staff, interested participants who meet initial screening criteria (date 
of final menstrual period, last mammogram, confirmation of medication use, age, BMI,) will be 
cleared by the primary-care Obstetrician-Gynecologist and a referral form will be sent by 
REDCap to PBRC. All this information will be transferred to a study coordinator at PBRC in 
accordance with HIPAA and eligible patients will be contacted to schedule a screening visit at 
PBRC. 
Additionally, PBRC will be a secondary site for recruitment. Participants will be recruited 
continuously by our recruitment core via media advertising, listserv, health promotion events, 
databases, and referral sources. Similarly, once potential participants are identified via PBRC 
recruitment methods and recorded in PBRC tracking databases, PBRC staff will schedule a 
screening visit at PBRC. We anticipate no problem in rapidly recruiting the 8 women needed to 
complete the study.
STUDY DESCRIPTION
Location
After referral of potential participants from Dr. Hayes (and her staff) or participant recruitment via 
PBRC, the screening visit will be conducted in the Outpatient Unit at PBRC.  
The Insulin sensitivity testing (clamp), energy expenditure testing, blood drawing will be 
performed in the Inpatient Unit at PBRC. 
Consent Process
The informed consent process will be conducted in the Outpatient Unit at PBRC during the  
screening visit and will be conducted primarily by the study coordinator, but also on occasion by 
the study PI or by a trained staff member of the Outpatient Clinic. Written informed consent will 
be obtained before any procedures are performed. Potential subjects will be given ample time to 
read the Informed Consent and to ask questions during the screening visit.
Screening Process
Prospective volunteers will be both referred from Dr. Evelyn Hayes or via PBRC recruitment 
efforts (see above) and invited to the Center for a complete screening visit conducted in the 
morning after an overnight fast. Height, metabolic weight, waist and hip circumference as well 
as blood pressure will be measured. This will be followed by a complete medical history 
including reproductive function and medications, a physical exam, and an ECG. If eligible, a 
blood sample (CBC, chemistry panel with electrolytes, total testosterone, estradiol and FSH) 
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and a urine sample (urine analysis for general health) will be collected. Potential participants will 
then be interviewed by trained staff that is overseen by a licensed Clinical Psychologist to 
identify potential barriers to adherence and study completion. We will work with potential 
participants to determine if the study is consistent with their lifestyle/time demands and if 
adhering to and completing the study is feasible. The final enrollment will then be determined on 
the basis of the laboratory results (blood and urine).
Randomization 
Participants will be randomized to the order in which they complete the two parts of the study 
(Active drug (TSEC) and placebo). 
Study Visits
The study visits are described in this section, while the study procedures that will be performed 
during these visits are detailed in the latter section “Study Procedures”.
This study involves is a randomized crossover study with two 8-week treatment period 
separated by an 8-week washout period. Each participant will participate in this randomized 
scheme with two evaluation periods (see Figure 1)
a) End of 8 weeks of treatment with PLACEBO or TSEC
b) End of 8 weeks of treatment with TSEC or PLACEBO
Figure 1.  TSEC Experimental Protocol
Screening
8 week
washout
1 2
1 2
1 2
1 2
8 week 
treatment
TSEC
TSEC
Placebo
Placebo
8 week 
treatment
Randomization
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After screening, enrollment and 
randomization (Week 0) , participants will 
come for a brief (<1 hour) clinic visit after 1, 
3 and 5 weeks (± 3 days) into the first 
treatment phase (TSEC or Placebo) to get 
their pills, record weight, waist and hip, and 
vital signs and collect adverse events. 
During the eight week (Day 53 & 54 ± 2 
days), participants will be admitted to our 
inpatient unit at ~4pm for measures of total 
body fat (DXA) and ectopic fat (intra-hepatic 
and intra-myocellular fat) by 1H Magnetic 
Resonance Spectroscopy (1H –MRS) as 
well as visceral fat (VAT by MRI). These 
measures will be followed by a standard 
dinner (see Study procedures below) at ~7pm. The following morning at ~4.45am, an IV infusion 
of D2-glucose will be started to measure splanchnic glucose output (SGO) over 3 hours 
followed by a 150 min low dose insulin clamp followed by a 2-hour high dose insulin clamp 
(Figure 2). Also, at this time metabolic weight, blood pressure, and pulse will be collected. At 
around 6am, during the baseline of the clamp procedure, an adipose and skeletal muscle biopsy 
will be collected. Additionally, an optional Bioelectrical Impedance Analysis procedure will be 
offered and, if consent is given, will be completed following voiding and metabolic weight. 
Participants will be discharged ~2 hours after the end of the clamp procedure.
After an 8 week washout period, the exact same 8-week treatment and clinic visit protocol will 
be repeated, this time with the opposite treatment (Placebo or TSEC).
Study Procedures
Body Composition and Ectopic Fat
DXA scans will be performed using a GE Lunar iDXA whole-body scanner. The scan takes 
about 10 minutes and the radiation dose is less than 1 mrem, equal to about 12-h of 
background radiation. The scans are analyzed with the QDR for Windows V11.1 software. We 
will also use the DXA data to estimate muscle mass. 
Visceral (VAT) and subcutaneous (SAT) adipose tissue (MRI):  Determination of body fat, 
visceral fat (intraperitoneal and retroperitoneal fat), and subcutaneous fat will be obtained using 
a 3T GE Signa magnet with measurements from a series of 8 T1-weighted MRI slices 
positioned axially across the abdomen. Measuring VAT and SAT from only 8 non-contiguous 
slices has been shown in our hands to be as accurate as measuring it from a much larger 
number of contiguous slices  
Intrahepatic and intramyocellular lipid by 1H –MRS: The magnetic resonance scans will be 
performed at the Imaging Facility of the Center (part of the NORC, NIDDK funded Center Grant; 
PI Ravussin). The subject will be asked to lie supine on the patient table of the 3.0 T magnet for 
approximately 20 min.  A 1H body coil will be used to measure intrahepatic fat stores. A single 
PRESS box (30 x 30 x 30 mm) will be collected in an area of the liver that is free from heavy 
vascularization. Similar measures will be acquired from 3 voxels in the soleus muscle. Data are 
analyzed using the jMRUi software package.
2-Step Hyperinsulinemic (20 & 120 mU.m-2.min-1) Clamp at 90 mg/dl Glucose with RMR
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The glucose clamp will be performed at the end of the two 8-week treatment periods as shown 
in Figure 2. At approximately 7pm the night before, participants will eat a standard meal (30% of 
energy requirements calculated on the basis of their (Harris-Benedict predicted RMR x 1.4; 50% 
CHO, 30% fat, and 20% protein). Between 4am and 7am, a catheter will be inserted into a hand 
vein heated at 56oC to obtain arterialized blood samples and into an antecubital vein of the 
contralateral arm for infusion of labeled glucose. Approximately 30 minutes after inserting the IV 
lines, a skeletal muscle biopsy and an abdominal adipose tissue biopsy will be collected. After 
baseline blood samples are obtained, [6,6-2H2]glucose (22 µmol/kg prime and 0.22 µmol.kg-
1.min-1 constant infusion basal) will be started between 4am and 7am. Three hours later, the 2-
step euglycemic (20 and 120 mU m-2 min-1) hyperinsulinemic clamp is initiated and continued for 
5 hours. Euglycemia (90 mg/dl) is achieved by a variable infusion of 20% dextrose enriched to 
approximately 2.0% with [6,6-2H2]-glucose to minimize changes in glucose isotopic enrichment. 
The plasma insulin concentrations achieved with these insulin infusion rates provide optimal 
ranges for evaluating insulin’s effect on hepatic glucose production (HGP) (20 mU m-2 min-1) and 
glucose uptake by skeletal muscle (120 mU m-2 min-1). The infusion of [2H2]-glucose will be 
decreased by 50% (0.11 µmol.kg-1.min-1) of basal during stage 1 and completely during stage 2 
(full suppression of HGP). Three blood samples will be taken during the last 30 min of each step 
and at each step including baseline, a blood sample will be drawn to be archived (20 ml at 
baseline and 10 ml at steps 1 and 2).  
Calculations.  At low dose insulin infusion, plasma insulin levels are expected to increase to ~50 
U/ml, and HGP is expected to be ~50% suppressed in these participants. During this steady 
state (Rd), peripheral glucose uptake equals the glucose infusion rate (GINF) plus the 
measured residual HGP (Rd = St-state GINF + residual HGP).  At 120 mU m-2 min-1 of insulin, 
HGP will probably be totally suppressed, and Rd = Steady State GINF. At each step, insulin 
sensitivity (SI) will be calculated using the formula: SI = Rd / (St-state insulin level - basal insulin 
level).
Resting metabolic rate (RMR) will be measured 3 times during the clamp procedure. At each 
step, O2 consumption and CO2 production will be measured by indirect calorimetry for 40 min 
(last 30 min will be used for calculations) using a metabolic cart (DeltaTrac, SensorMedics, 
Yorba Linda, CA). Patients will void before the test, and urine will be collected to determine 
urinary nitrogen and substrate oxidation.
Fat and muscle biopsy
Fat biopsy will be collected during the baseline of the clamp for cell sizing and 
numbering using osmium fixation, and to extract DNA and RNA for future work. The biopsy will 
be obtained as follows: After cleansing the skin on the right side of the abdomen with povidone-
iodine solution, and placing a sterile drape, topical anesthesia will be administered with a 
mixture containing 50%/50% lidocaine 2% and bupivacaine 0.5%. A 0.75-cm incision will be 
made in the skin and a Bergstrom needle inserted to collect, under aspiration, approximately 
500-750 mg of adipose tissue. The sample will be washed in sterile PBS and 75% of it snap 
frozen in liquid nitrogen, the remaining placed in osmium tetra oxide. Upon completion of the 
biopsy, the incision will be closed with a sterile bandage, and antibiotic ointment / sterile 
dressing applied.  
Muscle biopsy.  Similarly, after the adipose tissue biopsy, a Vastus Lateralis muscle 
biopsy will be performed using the technique of Bergstrom in order to measure the impact of 
treatment on insulin signaling pathway gene expression, measure in vitro the impact on glucose 
uptake, and to look at skeletal muscle morphology after TSEC vs. placebo treatment. After 
cleansing the skin with povidone-iodine solution, the skin, adipose tissue and skeletal muscle 
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fascia of the right leg will be anesthetized using a 50%/50% mixture of bupivicaine 0.5% and 
lidocaine 2%. The skin will be incised (0.75cm) with a #11 scalpel. The fascia fibers are 
separated with the blunt edge of the scalpel and the Bergstrom needle inserted into the vastus 
lateralis. After suction is applied, approximately 100 mg of tissue is cut and removed.  Several 
passes will be used to obtain at least 250mg of muscle.  Pressure is applied and the skin is 
closed with sterile tape. After cleaning the sample, the muscle will be snap frozen in liquid 
nitrogen and stored for subsequent analysis.  
Bioelectrical Impedance Device (BIA) (optional procedure)
Participants should remain resting in the supine position for 10 minutes prior to testing, after 
voiding, in the fasting state. Once a participant’s Subject ID, date of birth, age, sex, height, and 
weight are entered into the device, the procedure can be conducted. The BIA device 
automatically tests and analyzes with imbedded software. BIA will be performed according to 
the recommendations (ESPEN Guidelines, Kyle et al. Clin Nutr 2004). Two disposable 
BioTekna electrodes will be applied to the dorsal surface of the right hand: the first (injector 
electrode) on the distal end of the third metacarpal bone between the 2nd and 3rd finger 
(metacarpophalangeal joint), and the second (sensor electrode) between the distal prominences 
of the radius and ulna of the wrist (radio-ulnar joint). Another couple of electrodes will be placed 
on the dorsal surface of the right foot: the first electrode on the distal end of the third metatarsal 
bone between the 2nd and 3rd toe (metatarsophalangeal joint), and the second electrode) 
between the medial and lateral malleolus of the ankle (tibiotarsal joint).
The distance between the electrodes is 5 cm. Two pairs of cables will be connected to the right 
hand (it is irrelevant which of the two pairs of cables). Similarly, the other pair of cables will be 
connected to the right foot. The red alligator clip shall be connected to the injector electrode 
(metatarsophalangeal joint and metacarpophalangeal joint), while the black clip to the sensor 
electrode (tibiotarsal joint and radio-ulnar joint). Note: the two pairs of cables should not touch or 
cross each other, or touch the floor. Failure to observe these precautions may distort the test 
results.
DATA MANAGEMENT
Data and Biospecimen Storage
Electronic data will be stored securely on computers at PBRC. Paper forms will be kept securely 
in locked storage at PBRC. Blood samples will be stored in the Clinical Chemistry Core at 
PBRC. Specimens will be labeled with a unique de-identification code if transported outside of 
Pennington Biomedical.  
Statistical Analysis
Using SAS Version 9.4 and/or other statistical software, the data will be analyzed using mixed 
model or other appropriate statistical approaches and using post-hoc comparison adjustments for 
multi-testing. Descriptive analysis will also be performed. Statistical significance is set at α=0.05.
Confidentiality and Privacy
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Records that identify study participants will be kept confidential as required by law, and every 
effort will be made to maintain the confidentiality of participants’ study records. Except when 
required by law, study participants will not be identified by name, social security number, 
address, telephone number, or any other direct personal identifier in records disclosed outside 
of Pennington Biomedical Research Center.  
All biological specimens will be stored and analyzed at Pennington by the Clinical Chemistry 
Core, and will be labeled with a unique code number (“A number”). Some samples may be 
analyzed by outside labs or outside personnel. All samples analyzed externally from Pennington 
Biomedical will be sent with no identifying information and in compliance with Pennington 
Biomedical policies and procedures.  
To protect subjects’ privacy interests and to make subjects feel at ease, all participants will be 
assured of their anonymity and confidentially. In addition, the amount of personal information 
collected by the study will be minimized and limited to the study’s endpoints; all data will be 
used only for research purposes.  
Access to participants’ data and biological specimens will be limited only to the study’s 
investigators, including investigators external to Pennington Biomedical, to clinical support staff 
for this study, and to the overseers of clinical facilities at Pennington. This will be ensured 
through stringent security measures (assured by the Research Computing Core), including the 
use of subject ID numbers, storage of medical records in locked areas, and electronic security 
systems required by HIPAA.
BENEFITS, RISKS, AND SAFETY
Potential Benefits
Given the short duration of the study, we do not promise any benefits to participants from 
participating in this study. Possible benefits to participants include learning about their current 
health status.  
Risks
This study involves the following risks to subjects:

Risks associated with TSEC (Bazedoxifene/Conjugated Estrogens (BZA/CE): The most 
common side effects reported in at least 1 out of 100 postmenopausal women receiving 
BZA/CE include abdominal (stomach) pain, vaginal discharge, vaginal yeast infection, and 
leg cramps. Other adverse events reported in less than 1 out of 100 subjects were gall 
bladder disease and increase in liver function tests. More serious but rare side effects 
associated with the use of BZA/CE included venous thromboembolic events (blood clot in a 
vein of the legs or lungs), reported in less than 1 out of 100 subjects. If you experience 
sudden pain in legs or difficulty breathing, notify the Medical Investigator immediately.
Precautions
You may be allergic to conjugated estrogens or bazedoxifene. If you experience allergy 
symptoms such as rash, hives, or itching, notify your study doctor immediately.
     All estrogens and SERMS should be stopped prior to and during prolonged
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immobilization (such as bed rest, surgery or long-distance travel). It is recommended that 
study drug be discontinued four to six weeks prior to prolonged immobilization.
     You may not use hormone products during the study; including any oral, transdermal, 
vaginal or other products containing estrogens, progestins, androgens, testosterone, DHEA 
or SERMs.

Blood Draws:  There is the possibility of discomfort, pain, and bruising at the arm vein or 
site where the needle is inserted. There is also a small risk of bleeding and a very small risk 
of infection at the site of the blood draw. Aseptic (sterile) technique and trained personnel 
minimize these risks.
 

DXA: The DXA scan involves extremely low levels radiation exposure. This technique has 
been demonstrated to be safe and approved for children. There are no known risks of the 
body scan.

MRI & MRS: There are no known serious risks associated with MR studies provided that 
prescreening is performed to exclude individuals with internal ferromagnetic foreign bodies. 
MR prescreening will be performed according to PBRC policy and procedure and as part of 
the inclusion exclusion criteria. PBRC policy includes informed consent prescreening, a pre-
screen by the study coordinator, a screening questionnaire reviewed by the MR technologist 
with the participant, and a hand wand search for ferromagnetic bodies where necessary by 
procedure. Claustrophobia is a potential non-serious adverse event. The proposed RF 
energy deposition is within limits set forth by the FDA [calculated on the MR instrument as 
SAR].

Two-Step Clamp Procedure: The use of sterile procedures and good technique will 
minimize the risks. The intended site of catheterization will be cleaned thoroughly with an 
antiseptic and, after insertion, the area will be dressed so that it is not in contact with the 
outer environment. There has been no incidence of catheter infection in any of the hundreds 
of experiments we have already performed using this technique. A risk during the 
measurement of insulin sensitivity is hypoglycemia (low blood sugar) which will be 
minimized by: 1) constant supervision by a qualified physician at all times during the 
procedure; 2) constant monitoring of blood glucose during the procedure (every 5 min) and 
after (every 15 min for 2 hr); and 3) immediate availability of corrective measures (IV 50% 
dextrose). There is no risk to the measurement of RMR by indirect calorimetry during the 
different steps of the clamp.

Fat biopsy: There is a small risk of infection, bleeding, and scarring of the skin at the site of 
the biopsy.  Our inpatient medical team has performed more than 500 subcutaneous 
adipose biopsies.

Muscle biopsy: Muscle biopsy risks include: pain from the injecting local anesthesia and 
cramping during the biopsy, bleeding, scarring of the skin at the site of the biopsy, 
cutaneous anesthesia from cutting a subcutaneous sensory nerve (1 in 70), the latter is 
almost always temporary but occasionally can become permanent.  Our inpatient team has 
performed more than 1,000 muscle biopsies with no serious complications.
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
Bioelectrical Impedance Analysis (BIA): There is no risk associated with the BIA 
measurement provided that prescreening is performed to exclude individuals with medical 
implants such as pacemakers or metal joint replacements. Measurements will not be 
performed on any subject who is pregnant, and all females should inform the technologist if 
there is any possibility that they are pregnant.
In addition to the risks listed above, participants may experience a previously unknown risk or 
side effect.
Monitoring Subject Safety
Participants will be asked about any adverse events following each stay in the respiratory 
chamber. Any instances adverse events and safety concerns will be documented and reported 
to the PBRC IRB per standard procedures.  
Once every 4 months, the PI will review the progress and safety of the study with the MI, and 
write a summary of the results of those discussions as applicable.
Compensation
Participants will be compensated $1000 if they complete the entire study. $350 of the 
compensation will be paid at the end of their first phase treatment and the remaining $650 at the 
end of the study. There is no compensation available for research-related injury.
Withdrawal of Subjects
There is no risk associated with withdrawal from TSEC. Participants may be withdrawn from the 
study if either the PI or MI feels that their continued participation would jeopardize either the 
subject’s health or the results of the study.
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