Treating HIV infections remains one of the most formidable challenges in biomedicine, in part because cells harboring viral DNA in their chromosomes persist in the face of powerful drugs and immune responses. A research team has now, for the first time, isolated individual cells from these persistent viral reservoirs and characterized their gene activity, suggesting potential new treatment strategies.
“This is really exciting,” says Sharon Lewin, who heads the Peter Doherty Institute for Infection and Immunity and singled out the result as one of the most ground-breaking presented at the 24th International AIDS Conference which opened last week. “These single-cell advances are big.”
AIDS researchers have had many triumphs since the disease appeared 42 years ago, but only four people are considered cured and had cancers that required risky bone marrow transplants. The grafts reconstituted their immune systems with cells impervious to HIV infection.
Efforts to develop simpler and safer treatments for the other 38.4 million people living with the virus have been hampered by a fundamental obstacle: HIV persists in pockets of cells silently. After entering a human cell and integrating its DNA into the host’s chromosomes, HIV remains invisible to attack unless it starts producing new viruses. Antiretroviral therapy suppresses HIV replication, but sensitive tests show that even with the most effective treatments, small populations of white blood cells with the CD4 receptor harbor latent HIV DNA.
Researchers have used various compounds in what is called a shock-and-kill strategy, which awakens hidden viruses and either directly destroys host cells or allows the immune system to do the dirty work. This, in theory, should potentially reduce or even eliminate any remaining reservoirs. But people who stop antiretrovirals after taking these compounds usually have their HIV skyrocket to high levels in the blood within weeks.
At the AIDS conference, Eli Boritz, an immunologist at the National Institute of Allergy and Infectious Diseases (NIAID), described his team’s effort to better understand HIV’s hiding places by analyzing individual cells with latent viral DNA. Previous studies had isolated HIV inside individual cells in the reservoir, but scientists couldn’t assess the host cell’s gene activity because of a Catch-22: They could only identify whether a cell had been infected by prompting the virus to replicate. turn, likely altered cellular gene expression.
The new work avoided that dilemma by using a technique that isolates single, infected cells as tiny amounts of blood move through three microfluidic devices developed by physicist Adam Abate at the University of California, San Francisco and industrialist Iain Clark at UC Berkeley. In essence, the devices push blood through channels in microchips that trap individual cells in droplets, allowing them to be cut off so that other organs can read their genetic material.
“This is a technology that didn’t exist before” for HIV studies, says Mary Kearney, an HIV/AIDS researcher who focuses on the reservoirs. Lillian Cohn, who studies HIV reservoirs at the Fred Hutchinson Cancer Research Center, says developing this new technology required a “heroic effort” and predicts that many groups, including her own, will use it in the future.
Boritz and colleagues used the devices to compare the active genes in individual latently infected CD4 cells from three HIV-positive individuals with the CD4 cells of three uninfected individuals. When a gene is activated, its DNA is transcribed into a messenger RNA (mRNA) strand that is used to make a protein. In the CD4 cell comparison, the researchers analyzed the entire array of nearly 18,000 mRNAs—the transcriptome—and found two distinct patterns: The pool’s CD4 cells inhibited signaling pathways that normally lead to cell death and also activated genes that silenced the virus itself. .
“It’s remarkable that these cells are so distinct,” says Mathias Lichterfeld, an infectious disease clinician at Brigham and Women’s Hospital who studies HIV reservoirs in people who control their infections for decades without treatment.
Lewin says she is already looking for the genes Boritz’s team identified and wonders if a genome-editing method like CRISPR could destroy the reservoirs by, for example, crippling one of the CD4 genes that block its cell death pathway.
Lichterfeld says his lab has unpublished work that similarly suggests these infected reservoir cells have special properties that make them resistant to immune attack. “It’s really cool how we used completely different technological approaches but came to relatively similar conclusions,” he says.
Boritz, whose team spent 11 years on this project, says the results make “perfect sense for this nebulous phenomenon that we theorize about called virus latency.” He is particularly curious about what creates these patterns of gene expression. It could be that these CD4 cells are different types with special properties that allow them to survive infection longer than others. Or it may be that HIV infection transforms cells into long-lasting reservoirs. “It’s extremely important for us to do this,” says Boritz. “Maybe we could block this mechanism.”