How close are we to an HIV cure? What have we learned from the precious few "functional cure" cases out there, and what other interventions are being developed?
To answer these questions, Joseph Eron, M.D., director of the University of North Carolina Center for AIDS Research, provided an overview of HIV cure research at IDWeek 2012. In his presentation, he reviewed the barriers to a cure, while highlighting the current and potential strategies being researched.
Three Patients "Functionally" Cured
Eron started by highlighting the known cases of patients who appear to have been "functionally" cured of their HIV infection. Timothy Brown, whose case has been widely reported, received two stem-cell transplants for leukemia in 2006. His CD4+ cells were replaced by the donor's CD4+ cells, which lacked the CCR5 receptor that HIV primarily attaches to, effectively making him immune to most forms of HIV. He has been off therapy for over five years without the virus rebounding.
Similarly, two other patients show no traces of HIV in their blood after receiving stem-cell transplants. Unlike Timothy Brown, these two patients received donor cells that did not lack the CCR5 receptor. However, because they were on antiretroviral therapy during the transplant period, the donor cells were not infected with HIV. Although the two patients are still on treatment, at 1,300 days post-transplant, no virus can be detected, even with a single-copy assay.
Eron noted that stem-cell transplants are too toxic, too dangerous and too expensive for most individuals living with HIV. However, he suggested that for HIV-infected individuals who need a stem-cell transplant, curing HIV should also be a goal. Regardless, he said, any intervention should be time limited and tolerable, with only moderate risk and a measurable level of success -- it doesn't have to be 100%, Eron said, but we have to start somewhere.
Cure Barriers: Why HIV Persists Despite Treatment
The first and biggest hurdle to achieving a cure for HIV is an infected person's pool of latently infected cells, specifically resting CD4+ cells, Eron pointed out. These cells would be part of hidden HIV "reservoirs" that continue to evade current antiretrovirals. There are different reservoir sites in the body, including the blood, gut, central nervous system and kidneys. They consist of resting CD4+ cells, but could include other cell types. Targeting and eradicating the HIV reservoir remains the number one challenge, Eron said.
Eron also briefly mentioned the role that may be played by long-lived cells, such as macrophages and microglia. "Turns out if you get chemotherapy and radiation, your microglia in your brain actually turn over. That's perhaps one reason why people who get transplants don't relapse in the brain with HIV," he stated. While residual HIV replication may exist in the body, Eron hypothesized that it probably exists in a low level and wouldn't be a major barrier to a cure.
The last barriers Eron listed were HIV-specific immunity loss and generalized immune dysfunction among patients who have been virally suppressed for a long time.
HIV's Evolution in the Body
After patients start treatment, HIV doesn't seem to evolve or diversify for at least six years among those who stay on treatment, Eron said, referencing work done by Mary Kearney, Ph.D., and John Coffin, Ph.D. However, Eron raised a finding he found particularly disturbing: HIV "clone cells" that were showing up five or six years after treatment initiation. "There are things called predominant plasma clones. This is a clone of the virus that's almost identical. There are some cells that are producing identical virus over long periods of time," Eron explained.
"That either means there's one cell producing a giant amount of virus that lives for a long time, or what I think is going on: There's probably a cell that's divided -- maybe it was a stem cell and then it proliferated -- and it continues to produce virus. I think those may be another hurdle we may run into," he continued.
Persistent HIV Production and CD4+ Cell Infection Despite Antiretroviral Therapy
Despite technically being virally suppressed (typically defined as a viral load below 50 copies/mL), most HIV-infected patients are found to still have small but measurable amounts of virus in their blood when using more sensitive assays. About 70% of suppressed patients showed detectable virus using a single-copy assay, Eron said, referencing a study by Frank Maldarelli, M.D., and others.
While there are debates about where this extremely low-level viremia is coming from, Eron believes that it's almost certainly coming from the latent HIV reservoir, with HIV replicating from resting CD4+ cells.
The reservoir turns out to be incredibly stable, Eron said: Even if a patient had only one million resting CD4+ cells, it would take about 73 years of perfect treatment to eradicate the HIV within it, based on calculations by Robert Siliciano, M.D., and others.
Potential Cure Strategies
Targeting the Reservoir
Awakening the latently infected cells and getting them to express virus was the first step Eron listed that would bring us to a cure. "We want to use anti-latency serum therapy, something that you can do to get the virus out of latency, and then have the patient remain on therapy so that new infections are blocked. These cells [would] become productively infected and then die," he said.
"But there are challenges. If you produce these infected cells, something's got to clear them. You've got to clear the virions. You've got to block new infection of these cells. You've got to figure out how to do this," he added.
There are many potential ways to disrupt latently infected cells, Eron pointed out, including stimulating the chromatin, P-TEFb, or Tat and NF-kB proteins. Chromatin is the mass of DNA and proteins that condense to form chromosomes, located within a cell's nucleus. P-TEFb, Tat and NF-kB all play essential roles in HIV transcription.
Out of all of these ideas, Eron thinks the one that's gone the farthest is unwinding the chromatin. "HIV lives within chromatin, and the chromatin tips the balance away from activation and expression," he explained. "There are arguments about how much HIV is bound up in chromatin. If it's deacetylated, it's closed. If it's acetylated, it's open and could be transcribed. One thing you could use is a histone deacetylase [HDAC] inhibitor."
One HDAC inhibitor that's currently being studied for latency disruption is vorinostat, or suberoylanilide hydroxamic acid (SAHA), a drug approved in the U.S. for the treatment of cutaneous CD4+ cell lymphoma.
"It clearly inhibits a bunch of HDACs that are in the body. And it definitely works in vitro: You can show expression of HIV from latently infected cells across multiple labs," Eron said. This was first shown in a proof-of-concept study by David Margolis, M.D., Eron himself and others.
"In eight out of eight patients, HIV was expressed from these resting CD4+ cells. [Vorinostat] was given in vivo, cells taken in vivo. These cells were expressing [HIV] RNA. So it was successful disruption of latency in people," Eron noted.
However, he added, "We didn't show that you could actually reduce the reservoir. We didn't show that these cells died. We did show that there were no adverse events, which was good, but these were only single doses." Still, Eron said, this study marked the first successful in vivo demonstration of an HDAC inhibitor disrupting latency.
To see whether the size of the reservoir can be reduced, further studies using HDAC inhibitors in multiple doses are ongoing or being developed.
Are HDAC Inhibitors the Answer?
While HDAC inhibitors show promise, Eron remained hesitant. "Will HDAC inhibitors be enough? Are we done? I bet not. Not all studies show induction of viral expression by HDAC inhibitors ex vivo. There are some studies that show over time you get less induction ex vivo," he cautioned.
Instead, Eron suggested, "It's possible the combination of anti-latency compounds with different mechanisms might be more effective. Where have we heard that before? Combinations with different mechanisms." Undoubtedly, Eron was referring to the evolution of HIV antiretroviral therapy, which started off with single drugs and mediocre results. It wasn't until the advent of combination antiretroviral therapy, which combined drugs with different mechanisms of action, that patients began to see more potent treatment and widespread success.
"What if latently infected cells produce virus, but then they don't die?" Eron asked aloud. He then once again highlighted work by Siliciano's team: "They actually showed that when you stimulated resting cells with vorinostat, ex vivo, they actually didn't die," Eron noted. "They produce virus, but then they didn't die. Prior to treatment, they're dosed with vorinostat. Then they look at infectious units per million, and it hasn't changed. So virus was expressed, but perhaps those latently infected cells just became quiescent again."
Kick and Kill
While getting latently infected cells to express virus is a key first step, researchers also need a way to actually eradicate those cells once latency has been disrupted. Hence, what is now being called the "kick and kill" approach.
"You could give an HIV-specific vaccine to wake up the immune system and then give the kick," Eron hypothesized. "Perhaps you could manipulate CD8 cells ex vivo, then give them back and 'kick' to see if they get killed. Pablo Tebas and others are working on CD4+ cell receptor enhancement to try to improve CD8 cells. Maybe you could infuse a broadly neutralizing antibody, or antibody prime for antibody-dependent cell-mediated cytotoxicity [an immune response in which an infected target cell that is coated with antibody is destroyed by an immune cell] to kill the cells."
Eron also noted that the immune system, over time, becomes exhausted trying to control HIV. In the case of CD4+ and CD8 cells, when PD-1 (a key inhibitory receptor) attaches to the PD-L1 receptor of antigen-presenting cells, the CD4+ and CD8 cells become less effective at killing invading microbes. Therefore, according to Eron, "The idea is that you block this [PD-1/PD-L1 pathway], you reinvigorate the T cell and perhaps you could clear cancer cells -- which has been shown -- and HIV-infected cells expressing virus, which of course hasn't been shown."
Additional Cure Strategies
Eron pointed out other cure strategies currently being worked on, including genetic modification of CD4+ cells to limit or stop HIV replication. In particular, using a zinc finger nuclease to artificially disrupt the CCR5 receptor on CD4+ cells in an effort to make them resistant to HIV infection. "That's been done by a company called Sangamo and multiple investigators. The issue is they've never tried to do this in stem cells. They've only done it in peripheral CD4+ cells. I think to effect a cure, you have to do it in stem cells," Eron said.
He also noted two other strategies. "Perhaps there's specific activation of HIV-infected cells using targeted nanoparticles," Eron offered; he said researchers were working on that idea. "There may be some cytokines like interleukin-7 (IL-7) and interleukin-16 (IL-16) that could be given with [antiretroviral therapy] intensification to effect a cure," he added. "There are other ways to go forward, but we've a long way to go."
Treatment During Acute Infection
While not technically a cure, patients who start treatment immediately after becoming infected show a better response to HIV. Notably, a group of patients in France known as the Visconti cohort showed an ability to control HIV without treatment, much like elite controllers, Eron said. This was after they started treatment within 10 weeks of infection and stayed on treatment for an average of three years.
These patients present an interesting alternate road to a potential cure, Eron said. "Perhaps patients treated during acute infection might be the opportune patients to at least be the first ones to be cured. The reasons I say that is most individuals are infected with a single variant, and if you put them on therapy soon enough, that single variant does not get a chance to diversify."
Eron continued: "CTL [cytotoxic T cell] responses early are robust and could be preserved, as might be CD4+ cell responses. Persistent activations may be less. The size of the reservoir is smaller. We have seen that. So it's possible that these people treated during acute infection and suppressed for a long period might be ideal candidates for a cure soon."