Development of Novel Therapies for NIDDM

The overarching goal of this program project grant is the development of technologies that
lead to new methods for studying, detecting, and treating type 2 diabetes, and their
integration with hypothesis-driven diabetes research projects. The research will be led by an
Administrative Core team (Core C) under the direction of Dr. Newgard at the Sarah W. Stedman
Nutrition and Metabolism Center at Duke to ensure the integration of all analyses in the
Projects and Cores. Research within the PPG will be supported by two established scientific
core laboratories including the mass spectrometry (MS)-based metabolic profiling core (Core
B) of Duke Stedman Center Metabolomics Core Laboratory directed by James Bain, Ph.D. and the
Metabolomic flux/Imaging Core (Core A) at the Advanced Imaging Research Center (AIRC) of the
University of Texas Southwestern Medical Center under the direction of Dean Sherry, Ph.D.

Projects 1-3 of the current program continue the animal model work of the previous research
and Project 4, to be conducted at UTSWMC, adds a human studies component as well as
additional animal studies. The goal of Project 1 (led by Dr. Newgard/Duke) is to collaborate
with the other projects and cores to fully understand the metabolic and molecular changes
that lead to perturbed branched-chain amino acid (BCAA) homeostasis and loss of insulin
sensitivity in animal models, thereby leading to better understanding of possible
cause/effect relationships between BCAA and metabolic disease. The goal of Project 2 (led by
Dr. Muoio/Duke), also working with the other components of the program, is to test the
hypothesis that excessive mitochondrial catabolism of lipid and BCAA plays a central role in
triggering mitochondrial stress, insulin resistance and eventual metabolic failure in
skeletal muscle during the pathological progression of diet-induced obesity. Project 3 (led
by Dr. Burgess/UTSWMC) uses the tools in Cores A and B to help define the temporal sequence
of changes in mitochondrial metabolism in liver during development of hepatic insulin
resistance, and also dissects the contribution of key signaling events (insulin receptor
engagement, mTOR activation) and nutrients (BCAA, lipids) in this process. Project 4 (led by
Dr. Malloy at UTSWMC) will develop and apply novel high-field 7T NMR spectroscopy/metabolic
flux analysis technologies to help test three core hypotheses about the development of
insulin resistance emanating from Projects 1-3 (animal studies) in human subjects.

For project 4, the current request is to establish a protocol to vary the site of 13C
labeling in several physiological molecules: glucose, lactate, acetate, pyruvate, and
octanoate. All of these molecules can undergo oxidation in the citric acid cycle and all can
be safely administered to human subjects. The intent is to study a particular 13C labeling
pattern and molecule in 70 subjects to determine if downstream products (such as 13C
bicarbonate or 13C glutamate) due to oxidation in the mitochondria can be detected in
skeletal muscle or in blood by NMR analysis.

Additional work will include using 31P imaging to probe mitochondrial function by measuring
phosphorus-containing metabolites, as well as pH and metabolic flux activities
non-invasively.

Aim 3 of this project to commence in September of 2014 is a cross-sectional study in
overweight humans with measurements of branched-chain amino acids (BCAA) to test
mitochondrial function in skeletal muscle using 7T 31P imaging and an insulin clamp (infusion
of a glucose tracer). Subjects for the Aim 3 phase of the study will be 25 to 60 years of
age.

These new high-field MR technologies will be integrated with those largely in hand for
understanding mitochondrial function in liver and skeletal muscle in a project that
translates the biological/mechanistic findings of all these grant projects to human studies.

This protocol is primarily for technology development. Most of the subjects will undergo a 7T
MR exam as optimal information is expected from the stronger field strength. The 3T MR
scanner will be used once a protocol has been established for comparison.

Inclusion Criteria:

- Good general health (normal vital signs; no acute signs or symptoms of illness)

- Able to give informed consent

- Able to tolerate study procedures including insertion of an intravenous catheter and
approximately 90 minutes in the MRI scanner

- Aim 3 phase subjects will have BMI of 28 to 35 and a sedentary lifestyle; may be
pre-diabetic or have undiagnosed type 2 diabetes

Exclusion Criteria:

- Known diagnosed disease, including known heart rhythm disturbance, cancer or risk of
seizure disorder or other disorder that could create risk to the subject

- Pregnancy

- Subjects with implanted metal that would compromise safety in MR use

- Subjects not fluent in English will be excluded because the immediate ability to
respond to instructions in the scanner is required