Summer 2010


Photographs by C&N Photography, Inc. GustoImages/Photo Researchers, Inc

For a woman with polycysticovary syndrome, life is full of unwelcome surprises. Starting at puberty, her body, surging with an unnatural burst of testosterone, will grow hair where it shouldn’t and produce acne and sweat. She may gain weight, often hurtling toward obesity despite her most fanatical efforts to shed pounds. She may become prone to diabetes and heart disease. But that’s not the worst of it. The cruelest blow is that all of this may happen without her knowing why. Though PCOS is the most common hormonal disorder among women of reproductive age, affecting as many as one in 10 women, it’s a tricky one for doctors to detect because its symptoms mimic many other ailments. Many women don’t discover they have PCOS until they try to get pregnant, their struggles to conceive only heightening their creeping doubt that something inside is wrong.

Short of a cure, what many women with PCOS hope for is a warning—a test that could alert future patients to the presence of the syndrome, giving them the head start they need to keep their symptoms in check. But no such test exists. PCOS involves multiple genes and an assortment of hormones that act on several different organs in the body. The best doctors can do now is diagnose PCOS by exclusion, ruling out other possible explanations in a process that can take months of testing.

But what if we knew what our bodies know? “Your body is very smart,” says Fariba Assadi-Porter PhD’94, an associate scientist in the CALS Department of Biochemistry. “It does really clever chemistry when it confronts disease. Before any physical signs show, your body is already adjusting its chemistry to defend itself.” Like sentinels prepared for combat, our body’s defenses react to conditions that we aren’t able to perceive. What we really need is news from the front—an alert that the enemies are massing at the gate.

Assadi-Porter is among a growing community of scientists who argue those alerts are all around us—in our blood, sweat, urine, tears and literally every breath we take. Those bodily fluids contain thousands of tiny molecules called metabolites, which are created when we digest foods, drugs or pollutants from the environment. By studying the profile of those metabolites, Assadi-Porter and other researchers hope to identify signals in the body’s internal chemistry that can help doctors diagnose hard-to-catch diseases like PCOS. Currently she is scouring blood, urine, sweat and breath samples from dozens of women with PCOS to look for metabolite profiles that are consistent with the syndrome. Once found, those telltale molecules could become the basis for a simple, noninvasive diagnostic test.

The project is a prime example of the promise of metabolomics, an exploding area of science that focuses on our chemical makeup at the most basic level. Smaller than cells, genes and proteins, metabolites are essentially the chopped-up products and by-products of our cells’ energy functions. Metabolic processes such as digestion create tiny fragments of foods and drugs, which float around as sugars or fatty acids inside us. Our bodies harbor at least 3,000 different types of metabolites, and their quantities are constantly changing, depending on factors such as diet, exercise and viral or bacterial infections.

Fariba Assadi-Porter prepares tubes of bodily fluids for analysis in a nuclear magnetic resonance machine (background), which reveals the identity and concentration of individual molecules in the samples.

Assadi-Porter says that shifting profile makes the metabolome—the term researchers use to describe the whole picture of our metabolites at any given moment—a compelling place to look for evidence of something new arising in our bodies. Her PCOS experiments—which won one of the first grants awarded by the university’s new research incubator, the Wisconsin Institutes of Discovery—are just the beginning. She predicts that within a decade a comprehensive screen of a patient’s metabolome will become a routine part of a trip to the doctor.

“This is very important for personalized medicine, to monitor peoples’ health status,” she says “With current technology we’re going to be able to do that. In the next ten years, we’re going to be there for sure.”

The idea behind metabolomics isn’t a new one. People have long understood that states of health and disease are somehow reflected in the concentrations of molecules inside our bodies. Physicians in ancient China used to set bowls of urine near colonies of ants to see if the insects swarmed. If they did, it meant the sample was full of sugar, confirming diabetes. Today doctors still look at sugar to diagnose the disease, measuring patients’ blood glucose levels. In the same way, they test cholesterol to monitor heart disease and urea and creatinine for kidney problems. Metabolomics is different mostly because of its scale: Instead of looking at the quantities of one or two isolated metabolites, it involves taking a broad view of scores or even hundreds of metabolites at once.

This expansion has been made possible by major advances in the field’s two workhorse technologies: nuclear magnetic resonance, or NMR, and mass spectrometry. Both techniques can reveal information about the mass and structure of individual molecules, as well as the composition of complex molecular mixtures. Over the past couple decades, these machines have become significantly more powerful, capable of detecting more metabolites in a sample, while requiring smaller sample volumes.

But the human metabolome has remained a relative scientific frontier. Unlike in genetics, where efforts such as the Human Genome Project led to vast libraries of freely accessible data as early as 2003, scientists have had few resources to make sense of metabolites. The equipment necessary to measure and analyze them is large and expensive, and the resulting data streams can overwhelm even the best-equipped lab. Only in the past five years have scientists begun to piece together a roadmap, assembling databases of known metabolites to aid researchers in making sense of their data.

One of first researchers to join in that quest was John Markley, a biochemistry professor and director of the Nuclear Magnetic Resonance Facility at Madison (NMRFAM). Housed in the basement of the Biochemistry Addition, NMRFAM looks like a set from a James Bond movie, a vast, hangar-like room lined with gleaming domed machines. That equipment offers researchers the power and sensitivity to break a sample of blood or urine down into a roster of metabolites. Recognizing this unique capacity, Markley applied in 2004 for funding from a special National Institutes of Health Roadmap initiative called Metabolomics Technology Development to begin building tools to advance the field.

“We proposed that one of the major roadblocks in the field was the lack of a database containing data about pure, bona fide metabolites, as well as a lack of methods to rapidly collect and analyze data,” says Markley. “So that’s what we’ve been doing ever since.”

NMRFAM has now run more than 700 pure metabolites through its machines, compiling the data in a free, online database. Scientists are beginning to use the data—and NMRFAM’s technology—for a range of applications that extend well beyond human health. The aim of one of the facility’s projects is to compile a database of all the molecular constituents found in the plant cell wall, to aid researchers trying to unlock new forms of renewable energy from plants.

“Our major emphasis has been to get the technology in hand and get our database set up,” says Markley. “What excites me now is being able to apply the technology that we’ve developed to studies that are well-defined, and where we can use this approach to get solid information.”

Assadi-Porter’s PCOS project is the second such study to emerge from Markley’s lab. She first explored the power of metabolomics to monitor the progression of sepsis, a type of bacterial infection that sparks a dangerous, whole-body inflammatory response. She chose sepsis because the current testing technology is woefully inadequate. “By the time a doctor determines a person has sepsis,” she explains, “they are on the knife’s edge.”

Using mass spectrometry, researcher Amy Harms is searching for a chemical signature in body tissue that could alert doctors to the onset of colon cancer.

With animal sciences professor Mark Cook and zoologist Warren Porter, Assadi-Porter began analyzing metabolites in the breath associated with sepsis. In experiments with mice, the team was able to detect sepsis two hours after the onset of infection, hours earlier than previously possible. They later found the same results in rats and chickens. The team patented its “breathalyzer” technology and then founded a medical devices company to develop it into a viable product for at-risk patients.

Assadi-Porter’s sepsis project highlights one of the main advantages of metabolomics: its acute sensitivity to what’s happening inside the body at a given moment. And that plasticity has some scientists saying that metabolomics could turn out to be the missing link in delivering on the promise of personalized medicine.

The idea at the root of personalized medicine is that every body functions a little differently, and what works for one may not at all work for another. Many people believed that the sequencing of the human genome would unlock this great vault of individuality, yielding a master guide that would tell us how to diagnose conditions and prescribe therapies that optimally fit each person’s unique genetic makeup. But while genes reveal a surfeit of information about inherited conditions—such as a patient’s predisposition to breast or colon cancer—most of our day-to-day maladies are not hard-wired into our genetic code. To get a complete understanding of the processes that govern our bodies, we need to look not just at our genome, but at the other –omes: the transcriptome, which describes all of the protein-encoding RNA molecules in our cells; the proteome, our complete set of proteins; and the metabolome.

“The exciting part is not the metabolome, not the transcriptome and not the proteome,” says Mike Sussman, a biochemist and director of UW-Madison Biotechnology Center. “It’s the integration of them all.”

Figuring out how these four systems work together is one of the most pressing problems in systems biology, and billions of dollars are being invested to learn how their interconnection affects our health. One notable example comes from Sussman’s own lab, which is studying a special breed of rat to try to find the biochemical signals associated with colon cancer. Developed by UW-Madison oncologist William Dove, the rats have a genetic mutation that causes them to develop a rat form of the cancer. Scientists already know this mutation has a human analog, and patients who are missing the gene are more likely to develop colon cancer. But they can’t predict when in someone’s life that might happen. Sussman’s project is part of a larger effort to follow the chain from gene to RNA to protein to metabolite, which scientists hope will lead them to key signals that can sound the alarm when the cancer starts growing.

“Using metabolomics, we are trying to find a small molecule whose concentration precedes and predicts colon cancer,” he says. And success in this case would have a result that everyone over the age of 50 could appreciate. “People won’t have to get colonoscopies,” he says.

Unfortunately, the story for PCOS isn’t so simple. Despite exhaustive searches, scientists have yet to find a cause for the syndrome, which seems to arise from multiple layers of dysfunction. Current thinking is that the path to PCOS starts in the womb, when a fetus is exposed to a blast of testosterone—or possibly some other chemical signal—that permanently reprograms the genes her body will start expressing at puberty.
But what causes this blast in the first place? Because PCOS cases tend to cluster in families, signs point to some kind of heritable genetic factor, possibly a large group of problem genes that add up to initiate the syndrome. But very little is known about how this might work.
“PCOS behaves like it’s caused by a dominant gene that doesn’t always express itself, and that’s just baffled people for a long time,” says endocrinologist Dave Abbott, a professor of obstetrics and gynecology at the UW-Madison who has spent 18 years trying to understand the in utero conditions that trigger PCOS.

Peaks on graphic display of data tell scientists which molecules are present in a sample, offering a clue to what diseases look like on a molecular level.

When Abbott heard about Assadi-Porter’s PCOS project, he jumped at the chance to join the team, eager to approach the disease from a new angle. “The metabolome approach allows us to go from just having a diagnostic test to being a mechanistic cause investigation,” he says. “It may allow us to figure out what’s causing the metabolic [part of the] syndrome and lead to new therapeutic approaches that haven’t been applied because the knowledge isn’t there.”

But the clearest and most devastating calls for answers have come from patients themselves. From the moment the project was announced, Assadi-Porter says she has received numerous emails from women with PCOS, asking how they could participate in the search for a diagnostic test. And while the test wouldn’t help these women directly, they were eager to participate in something that could help future generations catch the disease early enough to intervene and keep the syndrome’s symptoms under control.

“These women were in the later stages and had so many symptoms,” says Assadi-Porter. “They were sending me their blood chemistry and asking, ‘Can we help you in any way? It’s so terrible to have this disease.’ They didn’t want their daughters to have to go through what they went through, should they have it.”

Each volunteer spent 12 hours inside a metabolic chamber at the UW Hospital, where they ate a prescribed dinner, ran on a treadmill and slept, all while machines recorded their breathing. Along the way, they gave samples of blood, urine and saliva that were later packaged and sent to Assadi-Porter at the NMRFAM for analysis.

From the vast pool of metabolites in these samples, Assadi-Porter has found a handful that rise to the surface as indicators of PCOS. If all goes well, this suite of metabolites will enable the creation of the medical community’s first-ever diagnostic test for the syndrome, which Assadi-Porter plans to undertake next.

The test would not merely save time, although that’s an important outcome for women who endure the guesswork currently involved in diagnosing PCOS. An immediate answer would eliminate mis-diagnosis, a common problem, and get women on therapies faster, before symptoms become severe. But mostly, an immediate answer would be just that: an answer. A way for a woman to know what her body is up against, deal with it and move on with her life.

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