Gene therapy may be key to permanently putting HIV into dormant state

In a study of human immune cells infected with HIV, the virus that causes AIDS, scientists at Johns Hopkins Medicine say a molecule within HIV itself can be manipulated and amplified to force the virus into long-term dormancy, a state in which HIV does not replicate.

The Johns Hopkins team that conducted the new study had previously shown that the molecule of interest, an “antisense transcript,” or AST, is produced by HIV’s genetic material and is part of a molecular pathway that essentially puts the virus to sleep, a state known as viral latency.

The study’s leader, Fabio Romerio, Ph.D., associate professor of molecular and comparative pathobiology at the Johns Hopkins University School of Medicine, says the new findings add to a growing body of evidence that may help researchers develop a gene therapy that boosts AST production.

A report on the research is published in Science Advances.

An estimated 1.2 million people in the United States have HIV, for which there is no cure or vaccine, and 4,941 people in the U.S. die each year from AIDS, the disease caused by HIV, according to HIV.gov. Worldwide, there are 39.9 million people living with HIV, and 630,000 deaths from HIV-related illnesses each year, according to the World Health Organization.

Standard treatment for HIV involves taking daily antiretroviral therapy that stops the virus from making new copies of itself and from spreading. Antiviral medicines must be taken long term, and they carry short- and long-term side effects, whereas gene therapy would require as little as one dose.

Even after several years of antiretroviral therapy, the virus can remain in cells and tissues throughout the body, quickly spreading if the infected individual stops the therapy, Romerio says.

“Our aim is to find a way to provide a lasting, durable treatment for HIV,” says Rui Li, Ph.D., postdoctoral fellow in Romerio’s lab and first author of the paper.

To investigate the role of AST in viral dormancy, the scientists first turned to a human cell line of CD4⁺T cells, the immune cells HIV targets to insert its genome and make copies of itself. The scientists genetically engineered these T cells, infected with HIV, to boost production of AST by inserting a genetic element capable of generating many copies of AST.

The scientists then measured the rate of HIV transcription, the process the virus uses to create the genetic blueprint for copying itself. To measure transcription, the scientists tracked levels of GFP, a fluorescent protein that is used as a marker of HIV expression.

They found that levels of the GFP protein decreased to nearly undetectable levels once the cells continually produced AST, indicating that the virus in such T cells was dormant and unable to restart replicating.

Next, using a high-powered laser technique that measures the physical and chemical properties of cells as they pass through a fluid stream, the scientists set out to determine which parts of the AST molecule were most essential for binding to and recruiting proteins that promote HIV latency.

To do that, the researchers created several mutations of the molecule, which they inserted into the T cells, to determine the role that each component of the molecule plays in making the HIV virus dormant.

The scientists also studied AST in CD4⁺T immune cells collected with permission from 15 people with HIV. They did so by first poking small holes in the outer membranes of CD4⁺T cells. Then, they mixed the T cells with DNA that expressed the AST molecule, which was taken up by the T cells.

Using a method that can accurately measure whether HIV is asleep or awake, the scientists found that the virus remained dormant in all of the cells for a period of four days. After that, the AST-expressing DNA degraded within the T cells.

Romerio says the new findings could lead to gene therapies that permanently enhance production of AST in the T cells of people with HIV, placing the virus in a state of long-term sleep.

In a study of human immune cells infected with HIV, the virus that causes AIDS, scientists at Johns Hopkins Medicine say a molecule within HIV itself can be manipulated and amplified to force the virus into long-term dormancy, a state in which HIV does not replicate.

The Johns Hopkins team that conducted the new study had previously shown that the molecule of interest, an “antisense transcript,” or AST, is produced by HIV’s genetic material and is part of a molecular pathway that essentially puts the virus to sleep, a state known as viral latency.

The study’s leader, Fabio Romerio, Ph.D., associate professor of molecular and comparative pathobiology at the Johns Hopkins University School of Medicine, says the new findings add to a growing body of evidence that may help researchers develop a gene therapy that boosts AST production.

A report on the research is published in Science Advances.

An estimated 1.2 million people in the United States have HIV, for which there is no cure or vaccine, and 4,941 people in the U.S. die each year from AIDS, the disease caused by HIV, according to HIV.gov. Worldwide, there are 39.9 million people living with HIV, and 630,000 deaths from HIV-related illnesses each year, according to the World Health Organization.

Standard treatment for HIV involves taking daily antiretroviral therapy that stops the virus from making new copies of itself and from spreading. Antiviral medicines must be taken long term, and they carry short- and long-term side effects, whereas gene therapy would require as little as one dose.

Even after several years of antiretroviral therapy, the virus can remain in cells and tissues throughout the body, quickly spreading if the infected individual stops the therapy, Romerio says.

“Our aim is to find a way to provide a lasting, durable treatment for HIV,” says Rui Li, Ph.D., postdoctoral fellow in Romerio’s lab and first author of the paper.

To investigate the role of AST in viral dormancy, the scientists first turned to a human cell line of CD4⁺T cells, the immune cells HIV targets to insert its genome and make copies of itself. The scientists genetically engineered these T cells, infected with HIV, to boost production of AST by inserting a genetic element capable of generating many copies of AST.

The scientists then measured the rate of HIV transcription, the process the virus uses to create the genetic blueprint for copying itself. To measure transcription, the scientists tracked levels of GFP, a fluorescent protein that is used as a marker of HIV expression.

They found that levels of the GFP protein decreased to nearly undetectable levels once the cells continually produced AST, indicating that the virus in such T cells was dormant and unable to restart replicating.

Next, using a high-powered laser technique that measures the physical and chemical properties of cells as they pass through a fluid stream, the scientists set out to determine which parts of the AST molecule were most essential for binding to and recruiting proteins that promote HIV latency.

To do that, the researchers created several mutations of the molecule, which they inserted into the T cells, to determine the role that each component of the molecule plays in making the HIV virus dormant.

The scientists also studied AST in CD4⁺T immune cells collected with permission from 15 people with HIV. They did so by first poking small holes in the outer membranes of CD4⁺T cells. Then, they mixed the T cells with DNA that expressed the AST molecule, which was taken up by the T cells.

Using a method that can accurately measure whether HIV is asleep or awake, the scientists found that the virus remained dormant in all of the cells for a period of four days. After that, the AST-expressing DNA degraded within the T cells.

Romerio says the new findings could lead to gene therapies that permanently enhance production of AST in the T cells of people with HIV, placing the virus in a state of long-term sleep.