Single-cell studies offer new insight into how HIV infections persist — and may be cured | Science

Treating HIV infections remains one of the greatest challenges in biomedicine, in part because the cells that hold the 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 single cells from these persistent viral reservoirs and characterized their gene activity, suggesting potential new treatment strategies.

“It’s really exciting,” said Sharon Lewin, who heads the Peter Doherty Institute for Infection and Immunity and highlighted the result as one of the most groundbreaking presented at the 24th International AIDS Conference, which began 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 have had cancers that required risky bone marrow transplants. The transplants restore their immune system with cells impervious to HIV infection.

Efforts to develop simpler, safer drugs for the remaining 38.4 million people living with the virus have been dogged by a major obstacle: HIV persists in pockets of cells, going silent. Once it enters a human cell and integrates its DNA into the host’s chromosomes, HIV remains invisible to attack unless it starts producing new viruses. Antiretroviral treatment suppresses HIV replication, but sensitive tests show that even with the most effective treatments, small populations of white blood cells studded with the CD4 receptor contain latent HIV DNA.

The researchers used a variety of compounds in a so-called shock-and-kill strategy that wakes up hidden viruses and either destroys host cells directly or allows the immune system to do the dirty work. This, in theory, should greatly reduce or even eliminate all remaining tanks. But people who stop antiretroviral drugs after routinely receiving these compounds have HIV spike to high blood levels within weeks.

At the AIDS conference, Eli Boritz, an immunologist at the National Institute of Allergy and Infectious Diseases (NIAID), described his team’s efforts to better understand the hiding places of HIV by analyzing single cells with latent viral DNA. Previous studies had isolated HIV inside single reservoir cells, 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. which in turn likely altered cellular gene expression.

The new work avoids this dilemma by using a technique that isolates single, infected cells while passing small amounts of blood through three microfluidic devices developed by UC San Francisco physicist Adam Abate and UC Berkeley bioengineer Ian Clark. Essentially, the devices force blood through channels in microchips that trap individual cells in droplets, allowing them to be dissected so that other tools can read their genetic material.

“It’s a technology that didn’t exist before” for HIV research, 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 requires “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 people with the CD4 cells of three uninfected people. When a gene is turned on, its DNA is transcribed into a strand of messenger RNA (mRNA) that is used to make a protein. In their comparison of CD4 cells, the researchers analyzed the entire set of nearly 18,000 mRNAs—the transcriptome—and found two distinct patterns: the reservoir CD4 cells inhibited signaling pathways that normally lead to cell death, and they also activated genes that silenced the virus itself.

“It’s remarkable that these cells are so different,” said Matthias Lichterfeld, an infectious disease clinician at Brigham and Women’s Hospital who studies HIV reservoirs in people who have controlled their infections for decades without treatment.

Lewin says he is already looking at the genes Boritz’s team identified and wonders whether a genome-editing method like CRISPR could destroy reservoirs, such as crippling one of CD4 genes that block its cell death pathway.

Lichterfeld says his lab has unpublished work that similarly suggests that these infected reservoir cells have special properties that make them resistant to immune attack. “It’s actually very nice how we used completely different technological approaches but came to relatively similar conclusions,” he says.

Boritz, whose group spent 11 years on this project, says the results make “perfect sense for this nebulous phenomenon we theorize about called viral latency.” He is particularly curious about what creates these patterns of gene expression. It is possible that these CD4 cells are different types with special properties that allow them to survive infection longer than others. Or it could be that HIV infection transforms cells into long-lasting bunkers. “It’s extremely important for us to understand this,” Boritz says. “Perhaps we could interfere with this mechanism.”

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