Mark Feinberg
is associate director of the Emory/Atlanta Center for AIDS Research. He has been a national leader in HIV research since the earliest days of the epidemic, serving as medical officer for the Office of AIDS Research at the NIH.

by Holly Korschun

Just two decades ago, most vaccines were still being developed through empirical methods - shot-in-the-dark experiments to see what might work, rather than well-theorized strategies based on knowledge of the complexities of the immune system. Scientists tried this and that to evoke an immune response - dead viruses, live but weakened viruses, and animal viruses that were harmless to humans.

While these approaches worked beautifully for many diseases - yellow fever, polio, rubella - no one has ever really fully understood why. Scientists now have come to realize that this hit-or-miss approach is unlikely to succeed against infections that hit back, such as HIV.

"Clearly, the days of empirical vaccine development are over," says Emory AIDS researcher Mark Feinberg. "Now we need to understand much more about how the immune system works to improve on immune responses."

An immune-focused approach is just the one being followed at the Emory Vaccine Center, located in a new, three-story building attached to laboratories at the Yerkes Regional Primate Research Center. Here, a comprehensive vaccine program is taking shape that includes basic research, vaccine design, preclinical testing, and clinical trials in humans. While investigators eventually will tackle persistent diseases like tuberculosis, malaria, influenza, measles, and flu, at present, their sights are set uniformly on finding a vaccine for HIV.

Preserving Immune Memory

Rafi Ahmed
is director of the Emory Vaccine Center, and the Georgia Research Alliance Eminent Scholar in Vaccine Research. He is recognized internationally for his expertise on viral pathogenesis and immune memory resulting from viral infection or vaccination.

Understanding the intricacies of long-term immune memory is key to this effort. Rafi Ahmed, director of the vaccine center, is considered an international expert on this topic.

There are actually two types of viral immunity, Ahmed explains - humoral and cellular, each with its own capacity for long-term memory and each with a role to play in vaccine development. In humoral immunity, B cells serve as the first line of defense by producing antibodies to prevent viral infection. By contrast, in cellular immunity, T cells activated by viral antigens kill cells already infected.

In mouse studies, Ahmed recently found that B cells live for much longer than previously believed, perhaps even for the entire life of an organism. Until now, scientists had thought B cells were short-lived and that other factors, such as re-exposure or chronic infection, were necessary to maintain a long-term antibody response. If B cells in humans also persist for long periods, this may explain why humoral immunity can be long lasting.

Cellular immunity can be long lasting too. Ahmed and colleagues have discovered that CD8 T cells respond at their peak for only a few weeks. After this peak, about 5% to 10% of the T cells become memory cells capable of mounting a stronger and more rapid immune response if reintroduced to the original virus. The number of T cells involved in the initial response seems to dictate the number of memory cells that remain. Most scientists now believe HIV should be attacked with strong antiviral drugs as soon as possible after infection to give T cells their best shot at producing a good long-term response.

Improving the immune response to HIV through effective drugs is part and parcel of the search for an effective vaccine. "At this point, I don't think anyone is optimistic about eradicating the AIDS virus right away through combination drug therapy or producing sterilizing immunity through a vaccine," says viral immunologist Jeff Safrit. "But studies are pretty clear that if you can keep the viral load down below a certain level for a long time, HIV becomes a chronic, manageable disease. We are accomplishing this with antiviral drugs, but these are not affordable in most parts of the world. We have to come up with a good vaccine that can limit the infection early on, then let the immune response take care of it."

Working with mathematical biologist Rustom Antia, Ahmed has demonstrated that the total pool of memory T cells may be dictated by the biological principle of homeostasis. If the number of immune cells is in fact constant and homeostatic, then long-term cellular immunity may be regulated by competition for space by memory cells. In other words, as an individual is exposed to new pathogens, some memory cells may need to get out of the way to make room for new ones. Since the total number of memory cells is quite large, normally the immune system is capable of maintaining immunity to many pathogens at once. The impact of new pathogens on homeostasis, however, could explain the eventual loss of immune memory to certain viruses. And understanding the mechanisms behind the loss of immune memory is a first step toward figuring out how to prevent it.

Measure for Measure

John Altman
came to Emory from Stanford and has become a household name among immunologists as co-developer of a new assay for detecting T cell response to viruses and vaccines. Altman's tetramer assay is said to have "revolutionized the field of immunology."

Just as important as understanding immune memory is the ability to measure the immune system's response to viruses. Until recently, this process was murky and laborious, with scientists relying on a time-consuming method called limiting dilution analysis (LDA) and waiting weeks before they had results. That test had led investigators to believe that only a small fraction of activated CD8 T cells responded to specific viral antigens and that most of the response was nonspecific.

Enter John Altman and his tetramer staining assay, a revolutionary new test that has demonstrated a specific T cell response in mice 100 times greater than that shown by the LDA, prompting Nobel laureate Peter Doherty to dub it "the new gold standard" for measuring specific immune responses to viruses.

In 1996, Rafi Ahmed was at Stanford University to deliver a lecture in the immunology department where Altman was a postdoctoral fellow. Ahmed had heard about Altman's new assay for identifying antigen-specific responses of CD8 T cells and asked him about a research collaboration.

"I told him what I was really interested in was a job," recalls Altman, who would join the Emory team not long thereafter. "What Rafi was building with the vaccine center was very attractive to me. I knew the technology I had developed would be helpful, and it was exciting to come into an environment where we can do excellent basic science and apply what we learn to real-world problems.

"The beauty of the tetramer technology," Altman says, "is that it can be applied to T cell responses to any pathogen, from influenza to HIV to malaria. Tetramers give you both a more accurate and a much more rapid picture of the immune response."

With funding from the NIH, Altman has established a national tetramer core facility at Emory that prepares custom tetramers for researchers around the country. He and Jeff Safrit also have a grant to develop another, more specific assay to measure function of the T cell response along with quantity. Their hope is eventually to design a panel of tetramers able to follow the breadth of an individual's immune response.

From Mice to Monkeys to Men

Harriet Robinson
is the first scientist to demonstrate that purified DNA could be used as a safe and effective vaccine. She came to Emory in 1997 from the University of Massachusetts Medical Center to become chief of microbiology and immunology at Yerkes.

Developing an AIDS vaccine has proved particularly difficult not only because the virus is able to elude the immune antibody response but also because HIV destroys the helper T cells (CD4 cells) needed for a killer T cell response. Also, HIV can establish a latent reservoir of proviral DNA that re-emerges to cause infection after the initial infection has been contained. Most scientists now think an AIDS vaccine will be completely effective only if it involves both the humoral antibody response and the cellular T cell response to deal with this spectrum of viral trickery.

As immunologists work to transform this knowledge into development of an HIV vaccine, they also must make the important step of moving from mice to humans, says Feinberg. "Although Rafi and John's work in mice has transformed the way people think about immune responses, unfortunately, there is very little information about how the human immune system responds to antigens," he says.

To get this information, researchers often turn to macaque monkeys as the interim step between mice and humans.

Until recently, only vaccines using live, attenuated viruses with the potential risk of causing disease have been successful in protecting macaques against HIV. However, now scientists are testing several vaccines that are safer because they use only pieces of the virus to stimulate an immune response. Vaccines that use purified proteins from the viral envelope have generated short-lived antibody protection against one specific protein. Some vaccines using live vectors from animal viruses, such as a recombinant avian or cow pox virus, have raised both antibody and cellular immune responses to HIV. These live vectors are desirable because they contain powerful viruses that elicit a strong immune response but are unable to replicate in humans.

Much of the national cutting-edge vaccine research is now using tiny pieces of DNA in various combinations with viral proteins or viral vectors to induce a synergistic immune response. DNA vaccines with recombined pieces of the virus expressing specific viral proteins also have elicited both antibody and cellular responses, and they have protected against a highly attenuated strain of HIV-1 in chimpanzees.

Yerkes microbiologist Harriet Robinson - a DNA vaccine pioneer - was lead author of last year's comprehensive American Society of Microbiology guide to DNA vaccines. In a recent successful clinical trial in macaques, she first primed the immune system with DNA coding for HIV or SIV (simian immunodeficiency virus) proteins, then followed with a boost from HIV viral proteins administered through an avian pox viral vector.

When the macaques were challenged with both a nonpathogenic combination of SIV and HIV (SHIV) and a highly virulent strain of the same virus, they demonstrated immune protection. Although the macaques' T cell immune response was able to control the virus, blood taken from some of the protected animals was still able to transmit SHIV to other animals, showing that the T cell response had not prevented the establishment of long-term latent infection.

"Although these cell-mediated responses have the potential for stemming the spread of AIDS, they will not prevent the establishment of low levels of long-term proviral DNA," says Robinson.

Robinson's next trial will use a DNA vaccine primer in macaques followed by a booster with modified vaccinia ankara - a derivative of the vaccinia virus used in the eradication of smallpox. Three other groups nationally have demonstrated success with a similar vaccine. For example, John Altman, working on one such trial with David Watkins at the Wisconsin Primate Center, used the tetramer assay to demonstrate that 12% to 18% of all CD8 T cells in the animals' blood were specific for SIV.

Emory investigators also plan to vaccinate human volunteers with proven vaccines like yellow fever, then study their immune responses. "If we could get an effective vaccine like yellow fever or chicken pox to express HIV antigens, we could have a safe vaccine that would raise immunity against the backbone vector as well as HIV," says Jeff Safrit.

Countdown to a Trial

The timetable for progressing from animal to small-scale human studies at Emory hinges partly on obtaining regulatory approval and on developing vaccine manufacturing capability. Vaccine testing must include a pathway of logical steps, including measures of the magnitude and breadth of the immune response, tests first in mice, then in monkeys, and finally, in humans, after which researchers will need to return to the lab to fine-tune their effort.

Vaccine production is a relatively new concept for academic researchers. Traditionally, vaccines have come from large companies. But most large companies do not view HIV vaccines as particularly lucrative investments. They also would be unlikely to produce the small quantities of vaccine required for intensive, small-scale testing. Emory investigators and administrators have considered developing a small production facility, perhaps at the Biotechnology Development Center at the new Emory West property near campus. Within the next six months, Emory scientists plan to begin human trials of HIV vaccines developed elsewhere. And within two years, they plan to be testing their own vaccines.

As in building any program, recruiting has been the key to the Emory Vaccine Center's accomplishments. "Our vaccine work has progressed faster than we expected in many ways because of our recruiting successes," says Ahmed. "In a very short time, we have a great team."

The interest in vaccines is infectious, too. At the inaugural meeting of the Vaccine Dinner Club last January, 172 people gathered on a rainy night to hear about vaccine research in the area. Sponsored by the Emory/Atlanta Center for AIDS Research, the CDC, and the University of Georgia, the club continues to grow, as colleagues from near and far come to hear speakers such as Ahmed discuss the cutting edge of vaccine development.

Feinberg is not surprised. "I know of no other city in the world where there are more people interested in vaccine issues than Atlanta," he says.

Holly Korschun is science managing editor for the Woodruff Health Sciences Center Communications Office.


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