Will Gene Therapy Cure HIV?

Published in: May-June 2011 issue.

THE CREATION of new drugs to treat HIV/AIDS has just about run its course. The next generation of therapies will involve modulating the body’s own immune system to better control the infection, and modifying its cells to make them more resistant to continued assault by the virus. The most advanced example of this line of research was recently presented at the Conference on Retroviruses and Opportunistic Infections, held in Boston. It is the world’s premier meeting on HIV science.

The study involved just six patients, but it demonstrated the proof of concept that it is possible to change the DNA of a person’s CD4+T cells so that they no longer express the CCR5 molecule that the virus uses to enter cells. The modified immune cells can be put back into a patient and they appear to thrive for at least three months and counting. Just how long they might last and how well they function has yet to be determined.

CCR5 Genetic Component

Genes affect our individual vulnerabilities to specific infections and responses to them. CCR5 is crucial because it is the preferred molecule that HIV uses as a coreceptor to enter cells. Some persons carry a naturally occurring variation in the DNA region that codes for CCR5. This is a twenty-base-pair genetic deletion known as the delta-32 mutation. Persons inheriting this mutation from both parents do not produce any CCR5 molecules on the surface of their cells, so the virus cannot get in. A mutant contribution from just one parent means fewer CCR5 molecules on the cell surface, greater resistance to HIV infection, and slower disease progression if the person does become infected with the virus.

The evolving consensus is that the mutation arose thousands of years ago near the Baltic Sea. Only an estimated one percent of Caucasians have inherited a double delta-32 mutation from their parents, and it is rarer still in other populations. CCR5 appears to be largely redundant in that most cell processes that use the receptor also can use another receptor. The initial thought was that persons with the delta-32 variant were perfectly healthy. However, that was tempered by later findings that they suffered more severe disease when infected with West Nile virus. There also is the suggestion from work in cell cultures that the receptor is important for the immune response to other tropical diseases. But the number of persons with the delta-32 variant living in tropical climates and exposed to those diseases is so small that it is impossible to tell. It may be that the delta-32 mutation is rare in the tropics because tropical diseases killed off carriers of that trait soon after it arose.

Pharmaceutical companies have used this information to create the entry inhibitor class of antiretroviral therapies that artificially block the function of the CCR5 receptor with small molecule drugs to prevent viral entry into cells. The best known of these is maraviroc (Selzentry®), which has been on the market since 2007. Now researchers are extending that line of thinking. Rather than administer a drug on a daily basis, they are seeking to modify host genetics, in this case the patient’s, to see if they might inhibit the CCR5 molecule on cells for a longer period of time, or perhaps even permanently.

The Berlin Patient

In 2007, German researcher Gero Hütter was treating a patient on therapy for HIV infection who had developed acute myeloid leukemia. Treatment for leukemia involves eradicating the existing immune system with radiation and chemotherapy, followed by a bone marrow transplant containing stem cells to build a new immune system. Hütter was intrigued by the possibility of using a bone marrow graft from a donor who carried the double CCR5 mutation. His patient embraced the experiment but initially requested anonymity and become known as “the Berlin Patient.” Last year he spoke publicly for the first time, revealing his identity as American Timothy Ray Brown.

The German registry of voluntary bone marrow donors is the largest in the world—2.5 million persons—and it sits amid the largest concentration of the delta-32 mutation. Yet among that huge dataset there was just one person who was both a good HLA (human leukocyte antigen) tissue match to the patient (necessary to prevent transplant rejection) and a carrier of the rare CCR5 double mutation.

The arduous treatment and transplantation had to be repeated when the leukemia reappeared. But eventually the transplanted stem cells took hold and repopulated a new immune system. Only then did Brown stop his anti-HIV drugs.

And nothing happened.

What typically occurs when even a person with undetectable levels of the virus stops an HIV drug regimen is that the virus quickly rebounds to about the level it was at prior to starting that therapy. Hütter drew a blood sample each month and anxiously awaited the lab results on his patient’s viral load. But as the months ticked by and each successive test came back negative for HIV RNA, he slowly came to believe that he had proven his hypothesis: Brown apparently had been cured of HIV.

Hütter published his findings in The New England Journal of Medicine in February 2009. Initial skepticism within the medical community slowly gave way to acceptance with publication of a follow-up paper in the journal Blood last December. The Berlin Patient was a fascinating proof of concept showing that perhaps, after all these years, a person really could be cured of HIV infection. Still, most scientists thought it was little more than a one-shot parlor trick that had little direct application for the real world.

There is the practical matter of finding the right donor match. In this instance, the odds were like those of winning the lottery, one in 2.5 million, with little hope that it might get significantly easier with more patients. Then there are the medical and ethical concerns about destroying a patient’s immune system with radiation and chemotherapy if it is not absolutely medically necessary. While bone marrow transplantation is now in its fifth decade, the one-year survival odds are not much better than fifty-fifty. Neither physicians who undertake such work nor investigational review boards that approve clinical trials are eager to embrace those risks unless there is another justification, such as leukemia. Clearly other approaches were needed in order to build upon the glimmer of hope sparked by the Berlin Patient.

The Sangamo Study

Researchers at Sangamo BioSciences, a biotech company in Richmond, California, have developed a proprietary “zinc finger technology” that can target and snip or disrupt a specific region of DNA in cells. Left alone, some of the cell DNA might properly repair itself, while other DNA might do so improperly, creating a shorter, nonfunctioning segment in its place. Conversely, it is possible to snip the DNA and insert another small segment of genetic molecules that confer other functions to that gene. The company has applied this technology to plants, animals, and modified cell lines for the production of pharmaceutical-grade proteins, and they are pursuing its use to treat a handful of diseases in humans.

Sangamo chose to focus its HIV work on CD4+ T cells. The cells express the CD4 receptor on their surface and are a key component of the immune system. They also are the favorite target for HIV to infect and are the principle site of viral replication that sustains the infection. Working with these T cells rather than with stem cells present in bone marrow also avoided the issue of needing to destroy the existing immune system. And by using the patient’s own cells and manipulating them outside the body, it avoided issues of making a tissue match and the potential for rejection that is common when the donation comes from another person.

The phase 1 safety study conducted by Quest Clinical Research for Sangamo enrolled six male patients aged 48 to 55 who had been infected with HIV for at least twenty years and were on therapy suppressing the virus below the level of detection. Their CD4+ T cells were showing the long-term effects of that struggle: there were between 200 and 500 copies, less than half of what is considered normal.

The patients were hooked up to a machine that filtered their blood for CD4+T cells and returned all other components of blood to their body in a continuous loop, in a process known as apheresis. The gathered T cells were sent to a central facility where they were processed with the zinc finger technology to induce a break in the DNA at the point of the delta-32 mutation. Cells that lost the CCR5 function were separated out and stimulated to grow and divide into large numbers. Finally, thirty million of these modified CD4+ cells were infused back into the bloodstream of the original patient.

The results were quickly apparent. Study leader Jay Lalezari says there was “a significant engraftment and expansion of the cells, a three-fold increase over what would have been predicted.” The total CD4+ T cell count increased by at least 100 in five of the six patients. The changes persisted, with 67 percent of CD4+ cells in the blood still showing the modification three months after treatment. The researchers took numerous other measures during the study and all seemed to move in a positive direction. Lalezari says it is “probably as good as we could have hoped for in this population.”

One Patient’s Experience

Matt Sharp is one of the six patients in the trial. When Lalezari told the San Francisco resident and longtime AIDS treatment activist about the study he was putting together, Sharp initially was hesitant to join. “I’ve done some pretty wild things, but I didn’t think that was for me,” he remarked. But he slowly came around, entering the study last summer and receiving the modified CD4+ cells back into his body in early September.

The first blood work came back a few weeks later showing “an immediate doubling of T-cells to about 550. … I hadn’t seen my numbers in this range in twenty years.” He says his CD4+ blood work has fluctuated between 450 and 600 over the last six months, while his HIV viral load remains undetectable.

The crucial event for the Berlin Patient was when he interrupted his haart (highly active antiretroviral therapy) regimen and the virus did not return. Some patients in the Sangamo study anticipate doing the same thing as a further test of the experimental therapy. They want to see if the virus has been cleared from their body, or, more reasonably, to what extent their new CD4+ cells alone might be able to control the infection. But Sharp is not yet eager to join them. He is concerned about “the fragile situation” of his immune system, which has been ravaged by the virus. His nadir T cell count was once so low that he had to take prophylactic drugs to avoid opportunistic infections.

Other patients in the study never lost so many T cells and probably have a greater reserve in their immune function, a greater margin for error, and the ability to recover if a planned interruption does not follow the beneficial trajectory of the Berlin Patient. Sharp wants to see that data first. If others do well on their treatment interruption, then he might reconsider. He also wants to see data on the functionality of the zinc finger-modified CD4+ cells. They seemed to work fine in the lab, but experience has taught us that lab experiments don’t always translate well into the human body. There is not yet any data from humans as to how well those cells perform their immune tasks. Nor is there a sense as to how long the modified T cells might survive in the body, or whether subsequent rounds of infusions of additional modified T cells are possible or advisable.

As a treatment activist, Sharp remembers how an earlier attempt at increasing the number of CD4+ cells turned out. Researchers had discovered that administering the cytokine IL-2, an immune system signaling molecule, could boost the number of CD4+ cells. They hoped that using IL-2 to modulate the immune response might become an important part of HIV therapy. The esprit study set out to prove that hypothesis. It enrolled over 4,000 patients in 25 countries, randomizing them to receive placebo or alternating cycles of the cytokine in addition to a haart regimen. But when the study was unblinded in January 2009, researchers were shocked to learn that even though IL-2 increased the number of T cells, there was no clinical benefit from those increased numbers. Whether they received IL-2 or not, both groups of patients had similar rates of disease progression and death.

Donald Abrams, the lead investigator for the study, said at the time that it was likely that “the CD4+ cells expanded by IL-2 are not functionally equivalent to the CD4+ cells arising as a result of viral-load suppression” with antiretroviral therapy. Or it could be that IL-2 also generates harmful effects through other pathways, which cancel out the benefits. The study also noted toxicities associated with receiving IL-2.

So Sharp’s caution is warranted. It was echoed by Scott Hammer, an AIDS researcher at Columbia University and a vice chair of the retroviral conference where the Sangamo study was presented. He reminds us: “This is early work that takes molecular biology into the clinic. … We shouldln’t be raising the flag to say we’ve solved the problem yet.” Lalezari agrees, emphasizing that this proof-of-concept study demonstrates that it is technically feasible to do the work, and it is safe. The procedure’s real test will come when patients stop their therapies. Will the virus remain suppressed or will they have to go back on a drug regimen?

Sangamo is planning a number of additional small studies in HIV patients with less advanced disease, including some who have not yet started antiretroviral drugs, and with different doses of the modified CD4+ cells. It also anticipates that the process might have to be repeated periodically as the modified cells wear out.

Other Potential Therapeutic Targets

Many researchers believe that a true long-term solution, and possibly a cure, will come only through modifying stem cells to be resistant to HIV, as was done for the Berlin Patient with the bone marrow transplant. Stem cells hold the potential to generate the complete lineage of immune cells, not just CD4+ cells. We understand some of the contributions that other types of immune cells make in controlling HIV infection, but that knowledge is probably incomplete. It is also likely that some rare subsets of specialized immune cells are wiped out during the first few weeks or months of HIV infection. The only way to restore those components of immune function is to regenerate them de novo from stem cells.

CD4+ cells circulate freely in the blood, and in large numbers, so it is easy to identify and separate them out for manipulation in the lab. Stem cells are orders of magnitude fewer in number and tend to be rooted in tissue sites rather than circulating in the blood. Assuming that the problems of isolating stem cells, modifying them to be resistant to HIV, and expanding their number sufficiently can be overcome, there still remains the issue of the established immune system. Will radiation and chemotherapy be required to kill it off? Some researchers believe that lower doses of those toxins to “condition” or partially knock down existing immune function might be sufficient to allow the new modified stem cells to flourish. They are working out the details.

Another step that may be required for a “cure,” at least for some patients, is developing a parallel genetic “fix” for CXCR4, another coreceptor that HIV sometimes uses to enter cells. At the Boston meeting, researchers from UCLA presented early work demonstrating that it is possible to modify these receptors in mice. But the CXCR4 molecule probably is a riskier target to manipulate. Unlike the CCR5 delta-32 mutation there are no naturally occurring experiments demonstrating that the human body can function well without the CXCR4 receptor molecule. We already know that CXCR4 has more uses and purposes than CCR5, and some functions probably remain unknown.

HIV “elite controllers” offer clues to other potential sources of immune control that might be therapeutically manipulated to better rein in infection. Those persons have become infected with the virus, but their disease progresses very slowly and sometimes apparently not at all. Some have been infected with HIV for two decades or more, have never taken an antiretroviral drug, and still have a low viral load or in some instances a viral load that can only be detected using an ultra-sensitive research assay. Detailed analysis of their DNA has teased out some factors contributing to this immune control, such as presence of the HLA B*5701 genetic variant that codes for part of their immune system. It is present in about ninety percent of elite controllers. HIV can reproduce in the presence of this genetic factor, but in order to do so the virus must change in other ways that reduce its fitness.

This is only one of what may be dozens of genetic factors that, alone or in combination, influence how rapidly or slowly HIV disease progresses. Each could be a potential target for a therapeutic intervention ranging from immune modulation to genetic modification. This is the complicated future of HIV therapy.


Bob Roehr is a biomedical writer based in Washington, DC. Parts of this article were adapted from earlier work in Scientific American online, BMJ (British Medical Journal), Medscape, and The Bay Area Reporter.



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