Using magnets to find new drugs: Inside UAB's high-field nuclear magnetic resonance facility

Most high-end lab equipment is inaccessible to the public eye, but one of UAB's most powerful drug-discovery tools is clearly visible from the Campus Green. The Central Alabama High Field Nuclear Magnetic Resonance Facility occupies a gleaming ground-floor space in the Chemistry Building. Its massive magnets give researchers invaluable insight into disease-causing proteins — and the data they need to find new ways to stop them.

The cover story of the latest issue of UAB Magazine features the Alabama Drug Discovery Alliance, a partnership between UAB and Southern Research Institute that aims to accelerate high-potential discoveries from the lab to patient-ready treatments. One key tool in that process is the Central Alabama High Field Nuclear Magnetic Resonance Facility, which opened in 2013. The Mix takes a closer look in this new feature.

Spin This Way

Each of the facility's NMR machines specializes in a different type of job, but the basic functioning is the same, explains NMR director N. Rama Krishna, Ph.D., UAB professor in the Department of Biochemistry and Molecular Genetics. The machines generate strong magnetic fields that polarize the tiny magnets in the nuclei of hydrogen atoms. “Then, using radiofrequency pulses, you can count all of the individual hydrogen atoms in a sample, which tells you what amino acids are present and how they are arranged in space,” Krishna says. And that’s precisely the information you need to create a detailed picture of a protein’s structure.

Mapping a protein's structure is crucial to understanding its function — and to finding ways to alter that function to treat disease. For instance, locating suitable "binding pockets" on a protein linked to brain cancer tells medicinal chemists how to design a drug to block (or enhance) that protein. "That's why NMR is one of the most versatile tools for drug-discovery research," Krishna says.

The bigger your magnet, the better images you can get. The centerpiece of the NMR facility is an 850 MHz Bruker BioSpin model, one of the largest in the South, which allows scientists to analyze structural data on even the largest proteins.

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Building a Better Drug

The 850 MHz machine can also accelerate the drug-discovery process "by allowing researchers to rapidly test new compounds they've developed in the lab," Krishna adds. Using a technique called saturation transfer difference NMR (STD-NMR), Krishna and his team can register the minute changes in signals from hydrogen atoms that occur when a compound binds to a protein. It would be nearly impossible to capture this interaction directly, he points out, because "it may last only a few microseconds." With STD-NMR, researchers can screen a number of potential drugs at once, then focus on the ones that show signs of binding to the target protein.

UAB's Rama Krishna and scientists from Southern Research
Institute have collaborated in developing a novel high-field
NMR-based protocol for determining the binding of
allosteric ligands to target proteins. They used the kinesin-5
protein Eg5 (a cancer target) and its inhibitor monastrol
 as an example (see above) for this protocol.

Using other techniques, researchers can analyze the disease-causing interaction between two proteins, and then find the right location to dock an inhibitor that would prevent the proteins from coming together. Or they could do the opposite, in an approach dubbed “fragment-based discovery” — using NMR data to identify two compounds that bind close together on a protein and “cross link” them to significantly improve their binding.

Krishna uses these techniques in his own National Cancer Institute-funded research to find new treatments for pancreatic cancer. Other UAB investigators are using the NMR facility to further their drug-discovery efforts in Parkinson's disease, brain tumors, breast cancer, heart disease, HIV and more. And as word of these capabilities has spread, researchers at institutions across the South have begun sending in samples to the NMR facility for evaluation.

Early Warning Signs

NMR is useful for many applications beyond drug discovery, Krishna adds. The facility's 600 MHz machine specializes in a hot area of medicine known as metabolomics, which studies the way the body processes everything from food to medicines.

"If you are taking a drug that is toxic to the liver, the body will generate some small molecules — known as metabolites — associated with liver damage,” Krishna explains. "We can detect these molecules in the NMR spectra of biofluids such as urine and blood plasma and say, 'Aha, after this patient started taking the drug, we can see an increase in these signals, so something is going wrong." That can warn researchers of side effects from new drug treatments "long before there is any major problem," Krishna says.

"The range of applications in this facility is amazing," adds Krishna. "It is a unique platform for everything from basic science to translational research.”

Grad student receives national award for new insight on alcohol and liver damage

Uduak Udoh

By connecting the dots between chronic drinking, molecular clocks, and energy storage patterns, UAB doctoral student Uduak Udoh has identified a potential new approach to target alcoholic liver disease. The work has also earned her a top honor from the Research Society on Alcoholism (RSA) and the National Institute on Alcohol Abuse and Alcoholism (NIAAA), and kudos from former NIAAA director Enoch Gordis, M.D.

Udoh, a fifth-year doctoral student in Pathobiology and Molecular Medicine, received the Enoch Gordis Research Recognition Award during the RSA's annual scientific meeting this summer. At the meeting, she presented results from her dissertation project, "Hepatic Glycogen Metabolism Is Impaired by Alcohol Consumption: Possible Role of the Liver Molecular Clock."

Liver cells, like almost all human cells, have a built-in circadian clock—a set of genes that control metabolism and other biological processes in a daily cycle. Udoh's research shows that chronic alcohol consumption disrupts the liver's normal pattern of creating glycogen, a storage form of glucose.

"Glycogen in the liver is an important fuel reserve that the body uses in between fasting and eating," explains Udoh, who is a member of the lab of Shannon Bailey, Ph.D., in the Division of Molecular and Cellular Pathology. Previous research has shown that the liver clock controls glycogen synthesis in a regular rhythm throughout the day. Emerging studies show that alcohol can disrupt the liver clock's timing, just as it disturbs the main circadian clock in the brain to disturb sleep and other behaviors.

Udoh's work connects these two observations. In mouse models, "we normally see a nice diurnal rhythm to glycogen content in the liver, which makes sense because metabolic needs vary throughout the day," Udoh says. But chronic alcohol consumption brings a significant change in that pattern, and a corresponding decrease in glycogen levels, Udoh found. She also demonstrated that alcohol disrupts signaling genes and proteins regulated by the liver clock that control glycogen metabolism.

Without sufficient glycogen, the liver may lack the energy to repair alcohol-generated damage, contributing to alcoholic liver disease. Ultimately, Udoh's research "highlights the molecular clock as a novel therapeutic target for alcoholic liver disease," says Bailey. (Learn more about Bailey’s own research into chronic alcohol consumption and the liver clock in this Mix podcast.)

Udoh's work was one of only six selected for presentation at the Research Society on Alcoholism meeting from hundreds of applications. Judges selected her for the Gordis award, which recognizes outstanding research among graduate students and postdoctoral fellows, based on her oral presentation and research poster session. One of the highlights of the event was discussing her work with Dr. Gordis himself, Udoh says. "It's a great honor."

How do mom's microbes affect pregnancy outcomes? UAB research aims to find out

As a baby slides out of the birth canal, on the way to its first breath, its body becomes coated in its mother’s microbes. This first interaction with outside organisms could be key to shaping the development of the baby’s immune system.

Our microbes, collectively called the microbiome, most often live in harmony with our bodies. They support the immune system, help to digest food and keep the metabolism on track, and fight off disease-causing bacteria. But researchers suspect that mom’s microbiome could play a role in when her children are born, and what happens to them as they grow.

“Most people know about the microbes that colonize the gut,” says Rodney Edwards, M.D., an associate professor in UAB’s Department of Obstetrics and Gynecology. “But there are bugs in and on us in many other sites—our skin, our mouths, our noses, our genitalia.”

During pregnancy, it turns out, the new needs and demands of a woman’s body change the numbers and types of these microbes. Alterations in how the body divvies up nutrients, stores fat, and produces hormones shift the properties of the microbes’ environments. But exactly how the microbiome changes over this nine-month period varies between pregnancies. And these variations, researchers are discovering, could impact not only the well being of a pregnant women herself, but the likelihood of pregnancy complications and the long-term health of a baby.

The Mix Quiz: Are You Smarter Than a Medical Resident?

Just like you, doctors love their smartphones. And they like playing games. But a new game pioneered at the UAB School of Medicine has lots more ROI than Farmville.

It's called Kaizen, a word borrowed from the Japanese auto industry that means something like "continuous improvement." This Web-based quiz game challenges medical residents at the School of Medicine's campuses in Birmingham and Huntsville with two questions every day. They're brief scenarios meant to highlight key practice skills and new evidence-based findings from a range of specialties.

Learn all about Kaizen in this feature from UAB Magazine. And test your medical knowledge with our five-question quiz.

Cable guys: Inside UAB's high-tech, custom-built approach to eye science

This machine, designed and built by UAB vision researcher Crawford Downs, is producing ultra-clear images of a key structure implicated in glaucoma, the world's second leading cause of blindness. See the machine in action in a video below.

The miracle of sight relies on a masterpiece of wiring. More than a million individual nerve cells scattered around the eye convert visual information into electricity. Then these individual cells are bundled together at the back of the eye into the optic nerve, which carries the signal to the brain.

Problems with this central cable are at the root of glaucoma, the world's second leading cause of blindness, after cataracts. The underlying causes of this optic nerve deterioration are still poorly understood. But a pioneering group of researchers and clinicians at UAB are exploring a new paradigm that could revolutionize our understanding of glaucoma and other eye conditions, including myopia and keratoconus.

The approach, known as ocular biomechanics, applies engineering principles to the eye. By creating detailed models of key eye structures, then stress-testing them in computer simulations, the scientists aim to identify the features of individual eyes that lead to glaucoma.

The work is led by J. Crawford Downs, Ph.D., vice chair of basic science research in the UAB Department of Ophthalmology and director of the new UAB Ocular Biomechanics and Biotransport Program, and Christopher Girkin, M.D., chair of the UAB Department of Ophthalmology. It has attracted the attention of the National Eye Institute, which awarded Downs and Girkin a $1.125-million grant in early 2013.

Zooming In on Glaucoma
Downs' efforts are focused on the lamina cribrosa, which acts as a mechanical seal at the optic nerve head where the optic nerve passes out of the back of the eye on its way to the brain. "The optic nerves go through pores in that structure," Downs explains. "It's also the place where the nerves get damaged in glaucoma. We want to understand the mechanics of the lamina cribrosa and what it looks like in three dimensions. That’s a key to understanding glaucoma biomechanics."

The problem is that this tiny structure—"it's about the size of a pencil lead," Downs explains—doesn't respond well to conventional microscopic imaging techniques. "Every time you put a section of the tissue on a slide for imaging, it's always warped or folded or stretched, so you can’t stack up successive images into a 3D structure" Downs says. So he built his own machine to do the job.

(See video below.)





This “fluorescent three-dimensional histologic reconstruction device” slices away tissue 1.5 micrometers at a time (about 1/100th the diameter of a human hair), snapping high-resolution pictures of the remaining tissues as it goes. With a volumetric resolution about 5 million times better than the best MRI, "I can see cell bodies with this technique," Downs says.

He reveals an engineer's pride in the clever details of his creation. For instance, the device automatically e-mails him a picture every 100 frames and texts him if it runs into problems. That way he can monitor the process remotely and allow the machine to run 24 hours a day. "There are only two in the world—the one here at UAB for eyes and one we built for colleagues at Imperial College London to study osteoarthritis in mouse knees," Downs says.

An individual image from Downs' machine

Image to Insights
Downs's first 3D rendering of the lamina, built from around 1,500 individual images, is just the beginning. Because everyone's lamina cribrosa is different, he is building a library of digitized models of laminas from the eyes of patients with and without glaucoma, as well as laminas from patients of different ages and ethnicities. "We can put the models in a computer, apply pressure to them, and simulate what happens mechanically," Downs says.

3D rendering of the lamina cribrosa

Downs was one of the first biomedical engineers to take up the study of the eye; now he is making UAB the hub of the rapidly growing field of ocular biomechanics. He has already recruited a team of fellow bioengineers to tackle complex problems in glaucoma and other eye diseases. Raphael Grytz, Ph.D., is studying the growth and remodeling of the sclera and lamina cribrosa; Massimo Fazio, Ph.D., is developing new, ultra-precise tools to measure scleral deformations with pressure and track deformations in images; and Vincent Libertiaux, Ph.D., is simulating how the optic nerve head reacts to different intraocular pressures. "We're one of the biggest groups in the world," Downs says.

Left to right: Massimo Fazio, Crawford Downs, Vincent Libertiaux, and Raphael Grytz

Defining Disparities
The gulf between surgeons and basic scientists isn't so wide in a specialty such as ophthalmology, where clinicians are used to doing their diagnosis at the tissue level. "I like to say that ophthalmologists are in vivo histopathologists," Girkin says.

Girkin's research is focused on health disparities in glaucoma, particularly in identifying why African Americans are at increased risk. Research by Downs and Girkin has helped uncover "some fundamental structural differences between the eyes of individuals with sub-Saharan African ancestry and those of individuals with European ancestry that may account for this elevated risk of glaucoma," Girkin says. "If we can define these differences we can target not just African Americans but anyone who is going to get glaucoma."

This basic research complements UAB's glaucoma service, which is among the nation’s busiest, Girkin notes. In addition to evaluating new treatment options, including laser therapy and minimally invasive surgeries, “our clinical research is looking at detection methods to allow us to find glaucoma earlier than ever, along with discovering novel pathways to treat this blinding disease," Girkin says.
In a pilot program led by Girkin, UAB's Department of Ophthalmology has installed sophisticated imaging devices in the offices of two central Alabama independent optometrists who are located adjacent to Walmart Vision Centers, with a centralized image-reading center housed at UAB.

(Learn more about the program in the video below.)




Toward Early Detection
The optical coherence tomography machines provide high-resolution images of the back of the eye. An optometrist can detect the earlier stages of glaucoma in those images, even before symptoms appear. Images of a patient’s eyes are electronically transmitted from the imaging machines at the optometrist’s office to the UAB center for confirmation of the diagnosis. UAB’s trained glaucoma specialists can then confer with the optometrist on complex cases to determine an appropriate treatment regimen. Patients who undergo the glaucoma testing also receive a dilated comprehensive eye exam and educational materials about glaucoma.

“This is an excellent example of the value of translating technology that has been evaluated and fine-tuned in the research setting and employing it in the field for the betterment of patients,” Girkin says. “This provides better access to care and better delivery of care within these hard-to-reach populations.”

The ultimate goal is to develop a noninvasive, image-based test "that a clinician can do in five minutes," Downs says. A human trial is at least a decade away, he predicts, but success would bring dramatic benefits: "You could cut the costs of treating glaucoma in half," saving billions of dollars per year.


Learn More
UAB Department of Ophthalmology

Ocular biomechanics at UAB

Research areas, UAB Department of Ophthalmology