H1N1 flu-related research at Stanford

- By Bruce Goldman

Faculty members at the Stanford University School of Medicine -- a powerhouse of research in immunology, microbiology and infectious diseases -- are exploring a variety of paths to better understand and treat H1N1 influenza. Their expertise ranges from cutting-edge vaccine approaches, to current and projected public-health measures, to providing an overall historical context for the current influenza pandemic. Below are brief summaries of their work.

For more information about these researchers, please contact Bruce Goldman at (650) 725-2106 or goldmanb@stanford.edu.

1. Universal influenza-vaccine target? Structure-based tools for vaccine development

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Theodore Jardetzky

Researcher: Theodore Jardetzky, PhD, professor of structural biology; member, Bio-X and the Stanford Cancer Center.

A viral protein's structure and exposure to the immune system determines the immune response to it. Because the influenza virus is capable of mutating rapidly, changing the structure of its surface proteins in ways that force the immune system to start from scratch, new viral variants (such as H1N1) can continue to puzzle the immune system and delay its response, so that an infection becomes more severe.

A key influenza-virus surface protein, hemagglutinin, comes in 16 different subtypes. However, an important stretch of this protein seems to be highly conserved in evolution -- it is essentially unchanged across all known strains of influenza virus -- suggesting that altering the structure of this portion of the critical protein may interfere with its function. But in the natural course of infection, this portion of the protein is difficult for the immune system to "see."

Might this evolutionary conserved invariant site be rendered more visible to the immune system and enlisted as a universal vaccine target -- a "sitting duck" -- eliminating the need to adapt influenza vaccines from one season to the next? Jardetzky, an expert on the molecular mechanisms by which viruses infect host cells, and others are working to characterize the site. He is examining the detailed molecular structure of hemagglutinin, which the influenza virus uses to attach to and penetrate our cells. His work could help guide immunological approaches that home in on invariant and functionally important regions of this protein to improve vaccines.

2. Quick, safe, adaptable, large-scale vaccine production? Customized virus-like particles as a new vaccine platform

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James Swartz

Researcher: James Swartz, D.Sc., the Leland T. Edwards Professor in the School of Engineering and professor of chemical engineering and of bioengineering; member, Bio-X and Stanford Cancer Center.

A cell-free system for assembling complex proteins can be used to make exact replicas of viral surface proteins. Even as these proteins are being produced, they self-assemble to form generic virus-like particles (or VLPs) that lack the nucleic acids necessary for replication in our bodies, so they can't cause infection. Yet they can be decorated with additional proteins to stimulate strong immune responses against particular pathogens, such as the H1N1 virus. They can even be further modified (for example, loaded internally or coated externally with "cargoes" of immunostimulatory substances) to heighten that immune response.

VLPs have been used successfully in vaccines targeting two other pathogens: the human papilloma virus and the hepatitis B virus. Swartz, who spent 18 years at biotechnology giant Genentech before coming to Stanford, is working on a high-yield, versatile VLP version that, he believes, can be used to rapidly produce large amounts of an effective influenza vaccine -- a serious bottleneck in today's conventional production approaches.

It may be possible to stockpile the naked VLPs and the immune stimulators. Then, only those viral proteins that have mutated to form a new strain would need to be produced and mixed with the ready-made components in order to assemble a full-fledged VLP tailored to the latest influenza strain. Alternatively, it may prove enough simply to employ these cell-free systems to produce similarly souped-up viral-protein fragments -- such as the invariant protein portion Jardetzky is studying (see item #1). Such a vaccine could be generally protective against most influenzas. Perhaps someday we won't need a new flu shot every year.

3. More bang for the dose? A powerful substance boosts vaccines' power

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David Lewis

Researcher: David Lewis, MD, professor of pediatric immunology.

An adjuvant is a substance that increases the potency of a vaccine in which the adjuvant is included. To date, there is no influenza vaccine containing any adjuvant licensed for use in the United States. Moreover, the sole adjuvant approved for U.S. vaccine use -- aluminum hydroxide, also known as alum -- stimulates production of antibodies, but has a limited ability to boost the proliferation of key immune cells called cytotoxic T cells. While antibodies are effective in preventing viruses from infecting cells, only T cells can ferret out already-infected cells (such as those in the lung and upper respiratory tract) and destroy them, preventing further spread of the virus to other cells. Plus, T cells can recognize a broader assortment of viral features than antibodies do, conceivably improving a vaccine's ability to protect against more than one strain of influenza virus.

Evidence from mouse experiments suggests that a novel adjuvant, cationic liposome-DNA complexes or CLDC, may induce more-effective subtypes of antibodies than those stimulated by alum. Lewis is examining CLDC's ability to arouse durable T-cell responses to influenza. His preliminary results in mice suggest that this adjuvant may be prove to be substantially more potent than any other now approved for clinical use or in clinical trials. This could reduce required vaccine dosages and, in a mass-immunization scenario, help stretch limited vaccine supplies to reach more people. Another positive feature of CLDC is that, unlike currently licensed adjuvants, it is quickly metabolized and doesn't accumulate in the body.

4. When a pandemic strikes: Modeling public-health responses in urban environments

Researcher: Nayer Khazeni, MD, instructor in pulmonary and critical care medicine; associate, Center for Health Policy.

Which public-health responses would be most effective if the H1N1 influenza pandemic hits major metropolitan areas in force, as seems likely? The answer to this question takes on immense importance, considering that the current H1N1 influenza wave may well burgeon in the United States before sufficient vaccine supplies are available to saturate the population.

Khazeni and her colleagues have done extensive modeling of both the impact of a pandemic on urban populations and the probable outcomes associated with different public-health measures, such as distributing masks, closing schools and stockpiling antivirals. Khazeni's recently published meta-analysis of studies comparing two front-line antivirals for influenza, oseltamivir (Tamiflu) and zanamivir (Relenza), shows a rough equivalence in their efficacy and also points out some limitations of studies done so far. Her team is currently modeling strategies to see how to make the best use of the H1N1 vaccine as it becomes available in the fall.

5. Breathing hard: An inexpensive, disposable ventilator for one-off use in a pandemic

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Matthew Callaghan

Researcher: Matthew Callaghan, MD, surgical innovation fellow, Stanford Biodesign Program.

A critical piece of equipment, should H1N1 become prevalent, is the ventilator, which helps patients in severe respiratory distress to breathe. Government studies have estimated a national shortage of almost 750,000 ventilators in the event of a serious influenza pandemic. These devices typically cost between $30,000 and $60,000 apiece; even stripped-down portable models found in ambulances run between $4,000 and $6,000.

A team led by Callaghan is developing a bell-and-whistle-free ventilator prototype -- now entering animal testing -- that can be assembled from off-the-shelf components and could sell for as little as $300 to $600 per unit. That would allow hospitals to stockpile hundreds of them and to use them in a disposable manner: one patient, one ventilator. Callaghan estimates the shelf life of such a ventilator to be about three years.

6. What mice won't tell you about your immune response: Beyond animal models to large-scale monitoring of human immune biomarkers

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Mark Davis

Researcher: Mark Davis, PhD, the Burt and Marion Avery Family Professor of Immunology and professor of microbiology and immunology; director, Stanford Institute for Immunology, Transplantation and Infection.

The inbred laboratory mouse's relative immunological simplicity and lack of genetic diversity makes it easy to study, but far from a perfect model system for humans, whose complexity and genetic diversity work to confound easy extrapolations from results of mouse studies. "The mouse is an excellent animal model -- for a mouse," says Davis, who has spearheaded efforts to collect, on a large scale, blood samples obtained from clinical trials, hospitalized patients or anyone else from whom blood is drawn. These samples are systematically analyzed in Stanford's bioinformatics-rich Human Immune Monitoring Core for differences between normal and diseased subjects, between people with one disease versus those with another, between similarly afflicted patients of different ages, and among those with differing versions of particular genes.

Davis is a principal investigator in a study now under way at Stanford to learn more about why older people's immune responses to influenza vaccines is generally weaker than younger people's. Many hundreds of biomarkers isolated from trial subjects' blood are quantified and compared for differences that correlate with the relative strengths of people's immune responses to seasonal influenza, or to the vaccine for it. (Samples from H1N1 influenza vaccine recipients and from those who contract this new strain will be analyzed in a similar fashion.) One goal: to be able to predict individuals' vulnerability to specific infectious organisms, the better to either provide more-tailored instruction for avoiding disease or to more effectively combat disease when it occurs. Another goal: to guide the development of more-effective vaccines.

7. On the front lines: Speeding vaccines through clinical trials

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Cornelia Dekker

Cornelia Dekker, MD, professor of pediatric infectious diseases; medical director, Stanford-LPCH Vaccine Program.

Dekker is responsible for initiating and carrying out clinical trials of vaccines at Stanford. An ongoing trial is examining age-related differences in response to the seasonal influenza vaccine. Groups of adult (ages 18-64), "young-old" (65-80) and "old-old" (80+) subjects are being vaccinated for seasonal influenza, and numerous biomarkers from blood samples will be correlated with measures of the cellular and antibody immune response. This is also the first year of studies to examine the effects of genetics on the immune response, by immunizing young and old twins with licensed influenza vaccines and investigating the same responses.

Before joining the Stanford faculty, Dekker spent during several years at Chiron Corp. (before it was bought by Novartis), where she was involved in the development of a new vaccine adjuvant, MF-59, now part of a seasonal vaccine approved in Europe. She is a recognized vaccine expert, currently serving on the National Vaccine Advisory Committee. Dekker is also the principal investigator of a National Institutes of Health-sponsored clinical trial at Stanford, concerning the safety and effect on immune response of a vaccine targeting the novel H1N1 strain. Stanford's trial will be unique in that it will test a version of the vaccine that contains an adjuvant.

8. Influenza 101: A seasoned researcher offers the long view

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Harry Greenberg

Researcher: Harry Greenberg, MD, the Joseph D. Grant Professor in the School of Medicine; senior associate dean for research and training.

The H1N1 influenza strain is but the latest in a series of new variations served up by the virus' tremendous potential to mutate. "If diseases were rock stars," says Greenberg, "influenza would be Elvis. In the pantheon of pathogens, influenza is the King." Greenberg was instrumental in the development of the FDA-licensed, nasally delivered, seasonal influenza vaccine containing live, attenuated virus. He knows first-hand all the ins and outs of bringing an influenza vaccine from the concept stage to regulatory approval.

Greenberg is a world-acknowledged authority on vaccine research and development, and is the past president of the American Society of Virology. He can offer a seasoned perspective on the shifts that the influenza virus has undergone and is likely to undergo, as it evolves in response to the pressures placed on it by our attempts to defang or defeat it.

About Stanford Medicine

Stanford Medicine is an integrated academic health system comprising the Stanford School of Medicine and adult and pediatric health care delivery systems. Together, they harness the full potential of biomedicine through collaborative research, education and clinical care for patients. For more information, please visit med.stanford.edu.

2023 ISSUE 1

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