November 3, 2012
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.
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.
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."
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."
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