The Phillip T. and Susan M. Ragon Institute, a unique collaboration of engineers, biologists, and doctors, was recently established at Massachusetts General Hospital (MGH) in Boston with US$100 million in funding from technology magnate Phillip Ragon to explore how the immune system combats disease, with an initial focus on developing an AIDS vaccine. The gift is unprecedented for MGH and the newly established Ragon Institute will be headed by Bruce Walker, an immunologist and director of the Partners AIDS Research Center, which is now part of the Ragon Institute. 
Ragon, who has a degree in physics from MIT, became drawn to the field of AIDS vaccines after meeting Walker and hearing about his research. Two years ago Walker suggested that Ragon visit AIDS clinics in South Africa and this affected him deeply. “I began to talk with Bruce about what I could do to help,” says Ragon.

“What this money means is that we can launch new collaborations in new areas with people with new perspectives, and do that immediately,” says Walker. The funding will be used to attract researchers from MGH as well as Harvard University and the Massachusetts Institute of Technology (MIT). “What we are going to be able to do is track a lot of talented people and give them license with flexible funding—the license to be innovative and creative and to take some bold chances.”

The Ragon Institute is also partnering with IAVI to conduct preclinical and clinical evaluation of AIDS vaccine concepts developed at the Institute. —By Regina McEnery

The Indian Council of Medical Research and IAVI have launched a Phase I trial to test the safety and immune responses elicited by two AIDS vaccine candidates administered sequentially in a prime-boost regimen. The trial known as P001 will enroll 32 volunteers at clinical trial centers in Pune and Chennai to evaluate different doses and vaccination regimens of the vaccine candidates. One candidate, TBC-M4, utilizes a modified vaccinia Ankara virus vector to deliver non-infectious HIV fragments in the hope of inducing an immune response against HIV. The candidate was developed in collaboration with the National Institute of Cholera and Enteric Diseases in India and was tested previously in a Phase I trial conducted in Chennai. In this trial, administration of TBC-M4 will be preceded by a prime vaccination with ADVAX, a DNA-based vaccine candidate, which was developed at the Aaron Diamond AIDS Research Center in New York City in collaboration with Rockefeller University and IAVI. Neither of the candidates being tested in this trial can cause HIV infection.

IAVI is also planning to begin enrolling volunteers in a Phase I trial of its adenovirus serotype 35 (Ad35)-based vaccine candidate. The trial will enroll 42 volunteers at the University of Rochester Medical Center who will be randomly selected to receive either two intramuscular injections of the Ad35-based vaccine candidate or placebo at three different doses. Clinicians will first administer the lowest dose and will review the safety data before proceeding to the next higher dose.

Ad35 is a serotype or strain of the common cold virus that researchers are using as a vaccine vector in this candidate to shuttle non-harmful fragments of clade A HIV, which is the predominant strain circulating in East Africa. The prevalence of naturally circulating Ad35 is much lower worldwide than the prevalence of adenovirus serotype 5, which was the virus used as a vector in Merck's AIDS vaccine candidate that was tested in the STEP trial. By using Ad35, it may be possible to circumvent issues involving pre-existing immunity to the viral vector (see VAX February 2005 Primer on Understanding Pre-existing Immunity). —By Regina McEnery

What are the limitations of current methods used to analyze immune responses to AIDS vaccine candidates and what new strategies are being explored?

Researchers do not measure the efficacy of a vaccine candidate—its actual ability to protect against HIV infection or control disease progression in individuals who become HIV infected despite vaccination—until the candidate is tested in large trials that involve thousands of volunteers who are potentially at risk of acquiring HIV. Instead, during the early stages of clinical evaluation, researchers primarily evaluate the safety of the candidate as well as its ability to trigger an immune response against HIV. The ability of a candidate vaccine to induce immune responses is referred to as its immunogenicity, and evaluating immunogenicity is one way that researchers can determine which candidates are worth pursuing in larger trials.

Researchers utilize different tests known as assays to determine the immunogenicity of AIDS vaccine candidates and different types of assays are used to measure different types of immune responses. Antibodies—Y-shaped proteins that latch on to the virus and stop it from infecting human cells—are most commonly measured using an ELISA or enzyme-linked immunosorbent assay (for more on how ELISA works, see VAX August 2007 Primer on Understanding Immunogenicity).

But many of the vaccine candidates that are currently undergoing clinical testing induce primarily cellular immune responses—both CD4+ and CD8+ T cells—against HIV, and not antibodies. Researchers measure and categorize the cellular immune responses induced by a vaccine candidate in many different ways.

Cytokine secretion

To study HIV-specific CD4+ and CD8+ T-cell responses, researchers isolate these cells from blood samples taken from volunteers in AIDS vaccine trials who received the candidate vaccine. They then expose these cells to the HIV fragment, or antigen, that was included in the vaccine candidate. This stimulates some of the immune cells and causes them to secrete certain proteins, known as cytokines, which can then be measured.

There are many different cytokines that play an important role in the immune response against a virus or bacteria. Some have direct antiviral activity, while others work more indirectly by activating other types of immune cells.

The ELISPOT assay is used to detect secretion of a single cytokine by both CD4+ and CD8+ T cells that are induced by a vaccine candidate. It is most commonly used to measure the release of a specific cytokine called interferon-gamma (IFN-γ; see VAX August 2007 Primer on Understanding Immunogenicity).

Measuring multiple cytokines

Another assay that can measure the ability of CD4+ and CD8+ T cells to secrete a broad range of cytokines is known as multi-parameter flow cytometry. As its name suggests, multi-parameter flow cytometry has a distinct advantage over ELISPOT assays in that it can measure the secretion of multiple cytokines simultaneously. This helps researchers more thoroughly define the cellular immune responses induced by a vaccine candidate.

In flow cytometry, cells or parts of cells are tagged with fluorescent probes that then flow through a beam of light, usually from a laser. Cells with different characteristics scatter the light in different ways, allowing them to be analyzed and sorted based on their ability to secrete different cytokines.

Limitations

While ELISPOT and flow cytometry assays provide useful data, they are not perfect tools. There are some indications, based on the results of clinical trials, that the ability of a vaccine candidate to induce cells that secrete cytokines is not necessarily an accurate predictor of whether an AIDS vaccine candidate will be effective. For instance, in the recently conducted STEP trial that tested Merck’s adenovirus serotype 5-based vaccine candidate, the ELISPOT assay analysis showed the candidate induced high levels of T cells secreting the cytokine IFN-γ, but the vaccine was still found not to be effective in preventing or controlling HIV infection.

Functional assays

Another assay now being assessed in clinical trials measures the specific function of immune cells induced in response to an AIDS vaccine candidate, rather than cytokine secretion, which is just a signal of immune activation. One of these so-called functional assays is known as the viral inhibition assay. It measures whether CD8+ “killer” T cells taken from blood samples of volunteers who received an AIDS vaccine candidate in a clinical trial are actually capable of doing their job and killing HIV-infected cells. Researchers isolate CD8+ T cells from blood of a vaccinated trial volunteer and combine them with HIV-infected cells in the lab to see if they are able to inhibit the virus. This approach is just now starting to be utilized in clinical trials of AIDS vaccine candidates.

Since researchers do not know precisely what immune responses against HIV will help control the virus or prevent infection altogether, it is important to study several different assays to infer as much as possible about the immune responses induced by vaccine candidates.