The search for a vaccine to combat the human immunodeficiency virus (HIV), a pathogen “notorious for its rate of mutation and variability,” in the words of Larry Corey, MD, principal investigator for the HIV Vaccine Trials Network (HVTN), was never going to be simple or straightforward.
In a recent confirmation of this challenge, the HVTN, an international collaboration of scientists and educators that conducts clinical trials around the world, ran squarely into failure last year when its multisite, US-based study, HVTN 505, was forced to stop after data showed that the vaccine neither prevented infection nor reduced viral load in those infected with HIV.
“The vaccine failures we’ve experienced have given us humility and served as a reality test,” observes Anthony Fauci, MD, the longtime director of the National Institute of Allergy and Infectious Diseases (NIAID), which sponsors the HVTN. “The development of the vaccine does not follow the usual paradigm, in which we’re taught to look at infection and response and then mimic what has been induced by natural infection. Not only does the body not produce an adequate response to HIV, there are documented cases of superinfection. This means that someone who is already infected gets infected with HIV again, indicating that the original infection clearly did not induce an immune response that could protect the person from getting a repeat infection.”And yet acquired immunodeficiency syndrome (AIDS) researchers and policy experts are far from disheartened.
Fauci says the disappointments represented by trial failures such as HVTN 505 coincide with important scientific and technical breakthroughs on several fronts that are now leading research in promising directions. He describes a 3-pronged approach to vaccine development that includes an all-out research effort into what he calls “the most satisfying solution,” a vaccine based on broadly neutralizing antibodies with potency against the widest possible array of HIV-1 strains; further research into a nonneutralizing antibody that prevented infection in about a third of enrollees in a recent clinical trial; and a “third path” that induces powerful CD8-positive T-cells that can destroy infected cells and clear the virus.
The most effective HIV vaccine could potentially include a combination of these approaches, he and other researchers say. Over the past 3 years, newly gleaned insights into broadly neutralizing antibodies, including breakthrough discoveries enabling researchers to identify, isolate, and characterize the evolutionand behavior of both the antibodies and their antigens, has changed the landscape of HIV research. “A B-cell-lineage design approach that selects out B cells that can recognize epitopes or antigens on the HIV envelope and stimulate the B cells to make broadly neutralizing antibodies (bNAbs) would be the most widely effective tool,” Fauci says. “This is difficult, but we’re working on a good, solid scientific basis.”
One of the principal challenges is that while some people infected with HIV produce these antibodies naturally, they have proved largely ineffective in fighting the virus. “Twenty percent of people infected with HIV can make bNAbs, but only after at least 2 years of stimulation by the virus. At this point, the cells are very mutated. They have developed affinity maturation, and although they are broadly neutralizing, they often don’t help much, as the person has lived through at least 2 years of infection and the virus is firmly established in the body,” Fauci notes. Another long-standing frustration of B-cell research has been the difficulty in reaching the regions on the viral envelope that would be susceptible to an orchestrated attack.
“There are only a few vulnerable areas on the HIV envelope protein—these are conserved regions where antibodies can bind and inactivate the virus. Current HIV vaccines are unable to induce antibodies to these vulnerable regions of the viral envelope,” says John Mascola, MD, director of the Dale and Betty Bumpers Vaccine Research Center (VRC) at NIAID, adding that the epitopes “are particularly difficult to reach because the viral envelope is covered by glycans (sugar molecules).”
Indeed, proving bNAbs existed at all was a critical first step. “We started by studying the serum taken from a large group of people in the early stages of HIV infection. This allowed us to identify donors with potent serum-neutralizing antibody titers against most strains of HIV,” he recounts. “We screened the sera against a large panel of viruses and found that about 20% of people make broadly neutralizingantibodies and we focused on the best of these donors. By sorting HIV-specific single B cells, we were able to isolate HIV monoclonal antibodies with broad and potent neutralizing activity.”
Since then, research by Barton Haynes, MD, director of the Duke Human Vaccine Institute at Duke University School of Medicine, Mascola, and others has mapped the evolutionary paths of both antibodies and different virus strains.1 “We performed B-cell genetic analyses early after an infection and then tracked how antibodies evolve over the course of 2 years. It takes months for the immune system to generate broadly active antibodies against the virus, because it takes that amount of time for antibodies to hone in on a conserved area of the virus,” Mascola explains.
In a May 2013 paper published in Science, Mascola, working with Peter Kwong and NIH colleagues, detailed an important advance in identifying serum-neutralizing antibodies’ responses that could potentially guide future vaccine development.2 The tool they developed, known as “neutralization fingerprinting,” allows researchers to identify which types of bNAbs exist in an individual blood sample and to measure which virus strains these particular antibodies block and with what intensity.
“The next step in developing a vaccine is designing an immunogen that mimics the right viral structure. From everything we know about vaccinations and viruses, if we are able to effectively stimulate the right type of antibody, it is likely that the vaccine could work to block HIV infection,” Mascola notes. Technical advances are playing a key role in what is increasingly interdisciplinary vaccine research. Working together, a group of biochemists, immunologists, and computer scientists at Duke University, for example, were able to determine the structure of a part of the HIV envelope protein, the gp41 membrane proximal external region (MPER), which may prove to be an important target for a broadly neutralizing antibody. Further, they were able to capture the conformational changes in the viral protein as it prepares to insert its genetic material into the host cell, and pinpoint what they believe is an optimal point in that process for a vaccine to bind.
“There has been an evolution in thinking about how gp41 operates, making it important to identify the key intermediate steps in the path gp41 takes when the virus merges with the cell’s membrane in order to develop an effective antibody,” notes Leonard Spicer, PhD, a professor of biochemistry and radiology at Duke and senior author of a study the group published in the Proceedings of the National Academy of Sciences. “As it turns out, 2 of the neutralizing MPER antibodies do not bind well with the resting phase of the virus, but rather at a site where the activity is, and so the intermediate we constructed had to be a dynamic one. The proteins that act on the virus must have a dynamic nature and make conformational changes. The idea is to catch them in the process and to bind at a point in the virus’s action.”3
In order to better observe the virus’s structural changes and its interaction with the antibody, Patrick Reardon, PhD, until recently a student in Spicer’s lab and the first author of the study, engineered a protein that incorporated the HIV MPER and associated directly with the membrane surface. “The MPER is an important target for vaccine development because it is a conserved region that binds several broadly neutralizing antibodies,” says Reardon, now a Wiley postdoctoral fellow at the Environmental Molecular Sciences Laboratory at the Department of Energy’s Pacific Northwest National Laboratory. “The question was whether the neutralizing antibodies would recognize our construct, which was designed to mimic the dynamic intermediate stage that we thought was occurring during viral fusion.” To their satisfaction it did, the study’s authors reported.
“A vaccine must mimic the virus in several ways. It must be a structure recognized by B cells and have properties that induce antibody development customized to the native behavior of the virus,” Spicer said. “In the case of the gp41 envelope protein, we know dynamics are involved—that’s the nature of the viral fusion machine.”
Spicer, a physical biochemist, previously worked with Haynes to develop another domain of the envelope protein, the V3 loop in gp120, with structural and dynamic characteristics that look like those on the viral protein. While bNAbs research is generating excitement in the scientific community, HIV policy experts say it is also important to conduct follow-up studies on the DNA prime/protein boost vaccine model tested in a study known as RV144 study, a collaboration between the Thai Ministry of Health and the US Military HIV Research Program. The vaccine used in that study produced some degree of protection—an overall efficacy rate of 31.2%—without eliciting broadly neutralizing antibodies, although its mechanisms are still unclear, researchers say.
“One of the main missions of the HVTN is to initiate what others have called the RV144 follow-up studies. The HVTN is part of a public/private partnership that aims to improve the performance of such a regimen,” says Corey, who is also the head of the University of Washington’s Virology Division and the Fred Hutchinson Cancer Research Center’s Program in Infectious Diseases. “HIV vaccine efficacy trials that are being conducted in Sub-Saharan Africa with vaccines designed specifically for these trials are in the late stages of planning. The HVTN is also testing other regimens in recognition of the fact that there may be other product combinations that will perform equally well through different pathways.”
The HVTN currently has 16 vaccine trials open, including those in various follow-up stages, while none are phase 3. But Corey says a phase 3 trial is planned, and will take place depending on the outcome of a phase 1 trial slated to open in the first quarter of 2015. This planned phase 3 trial in southern Africa will likely use a regimen similar to the RV144 trial, adapted to the endemic HIV strains in the area. “RV144 is the only trial so far that has shown any vaccine efficacy, so we are excited about this particular trial,” he notes. The development of a vaccine that induces a swift, powerful, and sustained T-cell response is yet a third promising avenue of research.
Louis Picker, MD, associate director of the Oregon Health & Science University’s Vaccine and Gene Therapy Institute, recently reported notable results from his animal trial of a T-cell—based vaccine based on a modified version of another virus, cytomegalovirus (CMV), and engineered to express the SIV protein that causes AIDS in monkeys. The vaccine completely cleared the virus from half of the monkeys in the trial, and is now proceeding to human trials.4
“Previous T-cell targeted vaccine strategies have been designed to generate a response composed of resting memory T-cells that upon pathogen infection proliferate and mobilize an effective army of T-cells to attack the invading virus. If the pathogen is defeated, the response would return to a resting state. Some pathogens, however, and HIV in particular, can within the time frame from detection to mobilization massively replicate and mutate, and thus escape the immune system’s response when it comes.”
“So 15 years ago, I chose CMV as a persistent vector that elicits and maintains a high-frequency mobilization of antiviral T-cell response all of the time, and therefore, its army of antiviral effectors is present and available at the very outset of infection and can control a pathogen like HIV before it is able to effectively escape,” he explains, adding that CMV is a large DNA virus that has evolved with humans and so most of the world has an immune response to it.
“CMV promotes immune response at the right level, with a high initial response, but it keeps itself in check after and sticks around at low levels,” he notes. In the study, whose results were published last year in the journal Nature, the T-cells eliminated the SIV virus over time. “Early on after the infection, we could still find the virus, but after a year and a half, we looked carefully and it was gone,” he says. “The T-cells work by early interception and control, which prevents the virus from spreading around. There is a reservoir level, but that is eventually eliminated. Because the vector maintains the troops, they slowly pick off the virus.” Going forward, he says, “We need to try to get the second half of the group, maybe with a higher level of the vector,” while adding that a “beefed-up version” of RV144 able to reach 40% of patients could possibly be combined with his T-cell vaccine to reach an efficacy rate of 80%, at least to create a “potentially effective vaccine.”
“It is also possible for antibodies to work together with killer T-cells; the antibodies can limit the amount of virus infection so that the infected cells can be killed by T-cells,” Mascola notes. With funding from NIH and the Bill and Melinda Gates Foundation, Picker is now planning a prototype vector phase 1 human trial, designed for safety, and another phase 1 trial to follow, in sub-Saharan Africa. Amid its successes and failures, the search for a vaccine has been aided by coordination around the globe of research, resources, and testing protocols, as well as the speedy dissemination of information from trials. “What we are able to do—the questions we ask and can now answer—has evolved in depth and complexity over the years. We have developed a highly sensitive and standardized array of laboratory assays, as well as innovative computational biology and statistical models. As a result, we are able to tease apart finer details of immune responses and safety data for the products we are testing,” Corey says. “Our advantage as a network is that, with so many studies under our belt, we can now look across multiple study results and draw conclusions about vaccine methods and schedules that are worth pursuing versus those that are not so promising.”
Researchers caution, however, that despite breakthroughs, a deployable vaccine is likely still years away. “It has been an extraordinary couple of years, but from the standpoint of an actual vaccine candidate we’re so far away we still don’t know what it is going to be,” Fauci says, adding that while researchers now know the conformation of the epitopes that broadly neutralizing antibodies bind to, developing a robust immunogen to stimulate them will be a challenge. “Because the broadly neutralizing antibodies we have identified are hyper-mutated, we may need to coax the immune system along, possibly stimulating a succession of antibodies that recognize epitopes and neutralize them in a series of vaccinations. The challenge is how to induce the correct B-cells and stimulate them to produce antibodies with a high degree of affinity maturation, because in the case of HIV, we must do better than what the natural infection is able to do.”
The solution, he adds, “may likely be a combination. We can’t ignore the efficacy of the RV144 vaccine trial, as modest as the efficacy was (31%); there is a role for non-neutralizing antibodies. And we also don’t yet know what role T-cells will play. They may be better in a therapeutic vaccine, however, rather than a preventive one.”