By Rex Graham
Pfizer (PFE) has the only FDA-approved drug in a class called CCR5 antagonists, and a “cure for HIV” will most likely focus on CCR5, top scientists agree.
If not Pfizer’s maraviroc, then a next-generation version of it could be part of the long-awaited cure for an infection that affects an estimated 33 million people worldwide.
This isn’t like fusion energy, where the breakthrough is always 30 years away. In this case, a conservative bet is more like 10 years. At most. Actually, a cure for HIV has been accomplished in one patient, which scientists agree amounts to a proof of concept of the CCR5 approach.
Mutations in human genes cause diseases from muscular dystrophy to cystic fibrosis, but a mutation in the human CCR5 gene has a huge health benefit: people with it are almost completely resistant to HIV infection. Since that discovery was made in 1996, large and small pharmaceutical companies and academic researchers have become focused on developing a treatment that mimics the effects of a CCR5 mutation.
But options other than CCR5 antagonists have been reported in a flurry of scientific publications and announcements in July, and many of them focus on CCR5.
The first solid evidence that the CCR5-focused approach would work came from a man dubbed the “Berlin patient.” Like 99 percent of the world’s population, Timothy Brown had a normal CCR5 gene. He was infected with HIV and was receiving highly active antiretroviral therapy (HAART), which is not a cure but kept Brown, former professional basketball star Magic Johnson and millions of other HIV-infected people healthy indefinitely. More than 35 anti-HIV drugs have been developed, including inhibitors of reverse transcriptase, protease, entry and fusion and integrase. Various combinations, tailored to individual patients have been highly effective.
“It was like a battlefield situation 25 years ago, but HAART therapy is allowing my HIV patients and me to grow old together, talking about things like blood pressure and smoking cessation,” said Douglas D. Richman, MD, director of UC San Diego’s Center for AIDS Research and a distinguished professor of Medicine and Pathology at the UCSD School of Medicine and VA San Diego Health System.
But experts like Richman know that HAART therapy is no cure, because HIV can develop resistance to the drugs in the cocktail and the drugs themselves can be toxic to patients.
Cure for HIV
In 2007, the Berlin patient was taking HAART therapy when he received X-ray ablation therapy that killed most of his immune cells, including those infected with HIV, and then received transplanted stem cells from a donor who had the CCR5 mutation. The risky procedure basically replaced most of the Timothy Brown’s white blood cells with uninfected, HIV-resistant white blood cells.
On the advice of his doctors, Brown stopped taking HARRT therapy.
“He has now been free of readily detect¬able virus in the absence of therapy for more than five years. In other words, he is cured,” Steven G. Deeks, a professor of medicine at the University of California, San Francisco, and Françoise Barré-Sinoussi, a professor of virology at the Pasteur Institute in Paris, France, wrote in a commentary in the July 19 issue of Nature. “His experience suggests that HIV infection might one day be curable.”
Deeks and Barré-Sinoussi agree that risky and expensive stem cell transplants are not the cure they’re expecting. The virus is able to integrate its DNA into the genomes of long-lived immune cells called memory CD4 T-cells, and there currently is no approved medicine to get rid of them. At any point, such as when HAART therapy is discontinued, HIV can re-emerge from its dormant or latent phase and roar back to cause acute symptoms.
“Determining how this ‘latency’ works, where infected cells reside and how latent infection could be reversed are the focal points of most ongoing cure research,” wrote Deeks and Barré-Sinoussi.
The federal National Institutes of Health, pharmaceutical companies and public and private research institutes have taken a three-pronged approach to defeating HIV: block viral attachment to the CCR5 co-receptor (and to the CXCR4 co-receptor, which is used by about 13 percent of HIV strains) on immune cells, prompt latent HIV genomes to emerge so the long-term reservoir can be eliminated, and a broad-spectrum HIV vaccine that keeps individuals from becoming infected in the first place.
HIV uses a glycoprotein on its exterior called gp120 to bind simultaneously to two receptors on the surface of human cells: the CD4 receptor and the normal, non-mutated CCR5 receptor.
Pfizer screened 500,000 candidates in its library of potential drugs and discovered that maraviroc is the first CCR5 antagonist that meets the approval criteria for therapeutic antiretroviral use, including the lack of unwanted side-effects. Maraviroc (Selzentry, made by Pfizer, NASDAQ: PFE) was approved by the FDA in 2007 to treat people –infected with the HIV strain that attaches to cells via CCR5.
Outwitting HIV resistance
HIV has a history of developing resistance to drugs, including those that bind to its gp120 surface protein; however, maraviroc is different. Instead of binding to the virus, maraviroc binds to HIV’s target: the CCR5 receptor.
However, once maraviroc was used in patients, the all-too-familiar chess match between virus and drug quickly appeared. Maraviroc-resistant HIV strains found in patients who had failed maraviroc therapy had cleverly substituted various amino acids in the gp120 glycoprotein in such a way that it continued to bind to the CCR5 receptor – with or without maraviroc.
So-called “escape viruses” arouse in tissue cultures in which maraviroc had been added, and those resistant strains had a similar pattern of genetic changes: all the resistant mutants had various amino acids substituted at seven positions in a variable loop of gp120. This same variable region, called V3, entertains amino acid substitutions in 15 or more other locations scattered throughout the V3 region.
Researchers are now painstakingly analyzing how HIV uses the infamous V3 loop as a template to continually spin out mutants that are resistant to CCR5 antagonists. For example, in addition to resistance to maraviroc, HIV strains have become resistant to vicriviroc (made by Schering-Plough, now Merck), Aplaviroc (GlaxoSmithKline), and other CCR5 antagonists.
A team of researchers led by Rogier W. Sanders, a Cornell microbiology professor, wrote in the June 2012 issue of Virology that CCR5 antagonists “may favor the expansion of pre-existing minor variants” in the virus’s gp120 protein, each with a degree of natural resistance. For example, 350 changes in amino acids in the protein’s V3 region have been identified in various HIV strains. Some resistant strains actually require a CCR5 antagonist to efficiently infect immune cells.
“Knowing that the various drugs might lead to different mechanisms of resistance should assist the design and development of the next generation CCR5 antagonists and anticipate the viral escape pathways,” Sanders’ team wrote.
In the Sanders team’s paper in Virology, the first comprehensive study of HIV resistance to CCR5 antagonists, the team noted that some amino acid positions in the V3 region are “highly conserved.” This knowledge could be leveraged in the improved design of the next generation CCR5 antagonists, which have factored in, or “anticipated” all mutational tricks at the virus’s disposal.
Several CCR5 antagonist candidate drugs have entered clinical development, only to be dropped due to poor results related to toxicity or other factors. The short list of survivors Include the following:
UCSD’s Richman said in a telephone interview that while progress on CCR5 antagonists and other drugs is promising, “we also don’t want to raise false hopes” about how soon a cure could be available.
Latency is another riddle in HIV’s bag of tricks that may have to be solved to effect a cure.
HIV can wait out almost any therapy or immune response because its genome becomes stably integrated in resting memory CD4 T cells. HIV-infected patients must remain on HAART therapy because the latent viral genomes can become activated and set off new viral replication. Deeks and others argue that any cure would involve a “shock and kill” approach that includes the ability to coax HIV out of its latency stage and eliminate all the virus particles subsequently produced.
A team of researchers led by David M. Margolis, a professor of medicine at the University of North Carolina at Chapel Hill, reported in the July 26 issue of Nature that an anti-cancer drug developed by Merck called vorinostat activates genes of HIV that had been dormant and inactive.
Deeks argued in another commentary in Nature that the stimulation of HIV replication by vorinostat, a type of compound called a histone deacetylase inhibitor, is as important as the findings in 1986 that AZT reduced HIV replication in people. The vorinostat study, said Deeks, “provides a rationale for an entirely new approach to the management of HIV infection.”
Now, the search is on at Merck and other companies and university laboratories for histone deacetylase inhibitors that are better than vorinostat in terms of specificity for HIV latency, potency, and safety profiles.
What’s hot: Zinc fingers
The Berlin patient was cured with hematopoietic stem cells with a deletion of 32 base pairs in the CCR5 gene in both pairs of chromosomes. The shortened CCR5 protein is still present on the surface of immune cells, but the viral gp120 protein can no longer bind to it effectively.
Scientists at The Scripps Research Institute reported in the July 1 issue of Nature Methods a new technique capable of precise gene-cutting. The researchers, led by Carlos F. Barbas III, a professor of molecular biology and chemistry, used zinc finger nuclease (ZFN) proteins to cut the human CCR5 gene. They added the ZFN proteins directly to human T cells in a culture dish, and within hours, a significant fraction of the cells showed sharp reductions in CCR5 gene activity.
“This work removes a major bottleneck in the efficient use of zinc finger proteins as a gene therapy tool in humans,” Michael K. Reddy said in a news release. He oversees transcription mechanism grants at the National Institutes of Health’s (NIH) National Institute of General Medical Sciences.
CIRM approach: RNA interference
The California Institute for Regenerative Medicine (CIRM) in San Francisco is focusing on CCR5 and recently handed out more than $40 million in grants to HIV researchers at UCLA, the City of Hope National Medical Center in Duarte, Calif., and UC Davis.
“It seems that the Berlin patient was cured via the CCR5 pathway, and our phase I and II studies will be really interesting because they are focused on targeting the CCR5 gene,” said Alan Trounson, PhD, the president of CIRM who has a family member infected with HIV.
One of the CIRM grants is helping UCLA researcher Irvin Chen design a RNA interference therapy that would require only a single treatment of stem cells. RNA interference can be used to selectively suppress any specific gene in a cell. In theory, the approach would prevent the CCR5 protein from appearing on the surface of white blood cells.
If Chen is successful, HIV in the blood, including virus particles emerging over many years from long-lived memory CD4 T cells, would die out because they won’t encounter any CCR5 receptor proteins.
Final exit: HIV vaccines
A new $77 million grant to the Scripps Research Institute from the U.S. National Institutes of Health (NIH) announced July 11 is aimed at developing a vaccine to prevent HIV infections from occurring in the first place.
Dennis Burton, PhD, a Scripps Research professor and leader of the new center, says that HIV and other highly variable viruses, including influenza and hepatitis C, have “highly conserved exposed sites” that vaccines could be designed to exploit.
“HIV is highly evolved to fight off antibodies,” Burton said in a telephone interview. “The classic approaches that worked for polio and small pox won’t work because of this extreme variability.”
Burton said a few HIV-infected people were found to make antibody molecules that target conserved exposed sites on the virus; these antibodies act broadly against many strains of HIV. “These antibodies provide a lot of clues for vaccine candidates that could be designed,” he said. “It’s part of a rational approach for vaccine development.”
Burton’s lab at Scripps is not currently collaborating with a biotech or pharmaceutical company, but Scott Forrest, vice president of business development at Scripps Research, said the institute “will attempt to commercialize promising technology that arises.”
Forrest said that companies focused on HIV research are investigating drugs rather than vaccines. “I think it’s more philanthropy than corporate investment that will push a vaccine forward to help those in the developing world, where the cost of drug cocktails is still an issue.”