Mysteries of immune system to be unraveled by new center

Norbert von der Groeben

Holden Maecker (left), Mark Davis and C. Garrison Fathman are some of the key players at the medical school's Human Immune Monitoring Center, which enables researchers to use a variety of tools to analyze blood samples in unprecedented detail.

Over the past few flu seasons, hundreds of people have trooped in to the medical center to get vaccinated, and then some. They’re getting blood draws both before and after their shot, because they’re participating in an ambitious clinical study. But this is bigger than a single study, however ambitious.

The study’s immediate goal is to tease out the ways in which the immune systems of people of different ages respond, or fail to respond, to specific influenza strains; that, in turn, could lead to scientists designing tailored vaccines that pack an additional punch for those whose immune systems need it.

But the flu-vaccine trial is part of a much more far-reaching effort to create the first-ever method capable of characterizing the human immune system under normal conditions — and thus knowing the multitude of minute changes that occur when we get sick, or successfully vaccinated, or old.

A key part of this endeavor is the Human Immune Monitoring Center, which allows researchers to analyze blood samples — such as those collected from the flu study participants — in exquisite detail. The center will play a crucial role in helping Stanford’s researchers understand the intricacies of the human immune system, said Mark Davis, PhD, director of the Institute for Immunology, Transplantation and Infection. To fund this larger endeavor, Davis has received close to $40 million from the National Institute of Allergy and Infectious Diseases as the project’s principal investigator.

“What we need is a scorecard: a routine, standardized, easily interpreted blood test you take before you get sick — analogous to the ones you get for cholesterol or glucose levels,” said Davis, the Bert and Marion Avery Family Professor in the Department of Microbiology and Immunology: “This would let you and your doctor know not only that right this minute what you’ve got is an allergy, not an infection, but also how well your immune system is functioning in general — and, if it’s malfunctioning, how, and with what consequences.”

From the standpoint of the practicing clinician, the immune system remains a black box, added C. Garrison Fathman, MD, the ITI institute associate director and professor of immunology and rheumatology. “If a patient were to ask me, ‘How’s my immune system doing today?’ I would have no idea how to answer that, and I’m an immunologist. None of us can answer that. Right now we’re still doing the same tests I did when I was a medical student in the late 1960s.”

In the last few decades a lot has been learned about the basic mechanisms of immune response — a super-smart system of sensors, cells and secretions that has evolved to guard us from invasion by pathogens or betrayal by our own tumor-prone tissues. We now know that cancer, autoimmune disease, infection, even chronic conditions such as heart disease once not recognized to have any immune connection — all involve a failure of some aspect of our immune defenses.

Still, the immune system remains a giant puzzle. Its complexity is overwhelming, comprising at least 15 different interacting cell types that spew dozens of different molecules into the blood in order to communicate with one another and to do battle. Within each of those cells sit tens of thousands of genes whose activity may be altered (or not) by age, exercise, infection, vaccination status, diet, stress, you name it.

Immunologists could be at the dawn of a new era, in which they determine what the differing levels of molecules and cells circulating in different people’s blood tell us about how the immune system works — and how to make it work better. The basis for these scientists’ hopes is the research enabled by the marriage between new or improved analytic instrumentation (much of it pioneered at Stanford) and the latest computing technology.

At the center of this effort sit a couple of clusters of world-class widgetry collectively called the Human Immune Monitoring Center at Stanford.

The HIMC operates according to a paradigm Davis only half-jokingly refers to as “ignorance-driven research.” The more formal name is systems biology, an information-technology-rich approach to dealing with complex systems of intensely interacting components.

“We can perturb the immune system all kinds of different ways, measure the levels of hundreds or thousands of different things in response to that, and figure out which ones go up or down with different states of health or non-health,” Davis said. “Anything that might affect the system — a vaccine, a disease, a drug — can tell you something.”

With systems biology, you don’t have to know what you’re looking for until you find it — some extremely high or low level of something (a cell count, a secreted immune protein, expression of a gene) that turns out to correlate with a disease or a vulnerability to it. You call that a “biomarker.” In a human blood sample, there’s an embarrassment of potential biomarkers to pick from. The HIMC is bringing new sophistication to the task.

In the past decade, scientists have steadily advanced a number of technologies capable of pinpointing biomarkers of immune status. These technologies can speedily record thousands of possible changes induced by a vaccine, a disease or aging. These changes might be in immune cells’ activity or numbers, in the amounts and types of molecules they secrete, or in which of their genes are idling or running in overdrive. Among the new techniques the HIMC employs are:

• Tetramer profiling: This technique, pioneered by Davis, deftly detects attractions between members of a class of immune cells and the foreign or altered biochemical entities they target. This technique, for instance, can measure the presence, or changes in the number, of specialized immune cells targeting such entities as viral or bacterial components — changes that could signal a state of immune readiness, or sluggishness.
• Mass-spectrometer flow: This was developed in large part in the lab of Garry Nolan, PhD, professor of microbiology and immunology. It involves an instrument — one of five in the world — that busts a single cell’s protein molecules into tiny pieces and, effectively, flings them at a wall; different metal tags attached to as many as 30 or more proteins enable researchers to track levels of these proteins in individual blood cells. Knowing in such sensitive detail how individual cells’ protein-contents are altered by challenges with drugs or disease may allow the detection of important events in our bodies (such as the onset of illness) well before they become obvious.
• Luminex panel: This method uses beads carrying fluorescent barcodes to quickly determine, for almost 100 different blood samples at a crack, which and how much of 51 different important immune-signaling molecules called cytokines — far more varieties than were known to exist a couple of decades ago — reside in each sample. Viral infection, bacterial infection, cancer, immune deficiency and traumatic injury, to name a few “perturbations” life visits on us, all result in different “cytokine signatures” — characteristic alterations in absolute and relative amounts of these molecules in our blood. These signatures are allowing scientists to create a map that could contribute to the effort to do a speedy check of a person’s immune health.

“What they’re doing at Stanford is truly unique,” said University of Washington immunology professor Jerry Nepom, MD, PhD, who is past president of the Federation of Clinical Immunology Societies, a 40,000-member organization of clinical immunologists that Fathman founded almost a decade ago. “They’re bridging the gap between these new technologies and a major unmet need: We don’t have any standard clinical validated test to monitor your immune system and tell us how it’s doing. They’re developing the tools that will get us there.” (Nepom is one of many outside researchers providing the HIMC with samples.)

The HIMC started life in 2005 as a bootstrap operation. “We were parceling out samples from my clinic to five different laboratories to get them analyzed,” Fathman recalled. “It was getting crazy, and I thought, ‘Why don’t we create a one-stop shop — put all the bells and whistles of immune profiling into a single unit? That way, a clinical investigator who doesn’t have a lab, and who may not even know what all these tests are, can simply ask to have an immune profile run on his or her patients.’” Fathman persuaded a philanthropic group, the Hedco Foundation, to give him $1.5 million in start-up funds, to buy equipment. But there was no place to put it.

At that moment, Phillip Pizzo, MD, dean of the School of Medicine, was launching the ITI institute with Davis at the helm. Davis was already convinced of the field’s need to adopt a systems-biology approach. He requested space for an industrial-strength facility, and Pizzo provided it. The HIMC opened for business at the beginning of 2007.

About 60 different projects large and small are under way right now at the center, which employs 12 people. “We’re probably processing at least 10,000 samples per year,” said HIMC director Holden Maecker, PhD. The search is on for immune biomarkers of aging, Alzheimer’s, autoimmune disease, cancer, chronic pain, rejection in organ transplantation and viral infection — both acute (influenza) and chronic (HIV).

A survey Maecker took recently showed that researchers and clinicians from more than a dozen medical center departments and divisions have teamed up with the HIMC to study everything from anesthesia’s impact on wound healing to opioids’ effects on immune function to potential immune biomarkers of eating disorders and depression.

Davis, for instance, is conducting the flu-vaccine trial in close collaboration with Cornelia Dekker, MD, professor of pediatrics and medical director of the Stanford-Lucile Packard Children’s Hospital Vaccine Program. This season, 240 people made three visits apiece for the study. Each of the resulting 720 blood draws was distributed into about 10 vials. Those samples are being analyzed from every angle through the HIMC’s lens in a bid to determine why this vaccine prevents many people from contracting influenza while failing to do so for others. In particular, why does it have about 85 percent efficacy for those under 65 years, but less than 50 percent for those older?

Fathman also noted that 90 percent of the HIMC’s work is done with frozen blood samples, which makes possible longitudinal studies. “Over time, as people whose blood was collected years ago start to manifest symptoms of autoimmune disease or cancer or an inability to mount an immune response to a germ or a vaccine, you can pick through the data to see whether any telltale biomarkers, way back when, were quietly predicting all along that this would happen,” he said.

Fathman is collaborating with William Robinson, MD, an associate professor of immunology and rheumatology, who has access to stored blood samples from an Armed Forces repository and is trying to create tests to allow early prediction of autoimmune disease. “As we start to mine those data, we’re starting to recognize that there are interesting changes in patients with various diseases that, early on, distinguish one disease from another,” Fathman said. “We know, 10 years after they were taken, who in that group developed rheumatoid arthritis or lupus. So we can start looking at those stored serum samples to see if there were any telltale biomarkers in their blood that would have allowed someone to spot the disease closer to its inception, long before obvious signs and symptoms emerged.”

If you’re collecting about 3,000 data points per blood sample, from hundreds of patients per year, you’re going to pile up a staggering amount of raw data. So, taking the systems-biology tack, you hand the entire database to the computer guys and let them sift through it, asking questions like: What’s different between samples from, say, older versus younger people or people whose flu shots worked versus those who got sick anyway? Which differences appear to be medically important? Which are the most reproducible? Which could be used in a diagnostic test? Which ones point to mechanisms that might actually be responsible for immune dysfunction?

Maecker, the HIMC director, regularly meets with investigators to help them plan studies, determine their needs (such as what samples to take, how to bank those samples and which assays to apply to them), and interpret their data.

“For them, it’s like going to the best restaurant,” Maecker said. “Everything’s on the menu.”