http://www.medicalnewstoday.com/articles/279945.php

HIV 'eliminated' from cultured human cells for the first time

Rearchers from Temple University School of Medicine in Philadelphia, PA, claim to have found a way to eliminate HIV permanently from human cells. The team describes its findings in the Proceedings of the National Academy of Sciences (PNAS).

The researchers constructed HIV-deleting molecular tools from "a DNA-snipping enzyme" and a strand of gRNA used to "hunt down" the genome of the virus.

According to the Centers for Disease Control and Prevention (CDC), there are currently more than 1 million people in the US who have HIV, with 50,000 Americans contracting the virus each year.

Highly active antiretroviral therapy (HAART) has been the main treatment for controlling HIV-1 - the most easily transmitted form of the virus - for the last 15 years. As we reported yesterday, antiretroviral therapies have had a significant worldwide impact in extending the lifespan of people infected with HIV.

However, even though HAART controls the replication of HIV-1, the presence of the virus in the infected patient may still contribute to health problems, such as weakening of the heart muscle, bone disease, kidney disease and neurocognitive disorders.

"The low-level replication of HIV-1 makes patients more likely to suffer from diseases usually associated with aging," says Kamel Khalili, PhD, professor and chair of the Department of Neuroscience at Temple. "These problems are often exacerbated by the toxic drugs that must be taken to control the virus."

Crafting molecular tools to 'delete' a virus

The Temple team instead investigated methods to "delete" the HIV-1 DNA from cells. With this in mind, they constructed molecular tools from the combination of a nuclease - which they describe as "a DNA-snipping enzyme" - and a strand of gRNA used to "hunt down" the genome of the virus.

They based this two-part "HIV editor" on a bacterial defense mechanism that evolved to protect against infection.

The team engineered a strand of gRNA consisting of 20 nucleotides (the building blocks of DNA and RNA) as a targeting system for the editor. This gRNA strand looks for the "long terminal repeats" (LTRs) of the HIV-1 genome - identical sequences of DNA that repeat many times at either end of a viral genome.

When the gRNA locates an end of LTR, the nuclease - called Cas9 - makes an incision. With both ends of the 9,709-nucleotide HIV-1 genome snipped, the host cell's gene-repair mechanisms take over and "solder" the loose ends of the genome back together. The result is a virus-free cell.

To prevent the engineered gRNA from binding with any part of the patient's genome, the researchers ensured that the nucleotide sequences they selected do not appear in the code of human DNA.

Testing the mechanism in cultures of human cells, the team found the tool was effective in eliminating HIV-1 from several varieties of cell known to harbor the virus. These include microglia, macrophages and T cell lymphocytes - the main cell types targeted by HIV-1.

More research is needed before the concept can go into the clinic

"This is one important step on the path toward a permanent cure for AIDS," says Dr. Khalili. "It's an exciting discovery, but it's not yet ready to go into the clinic. It's a proof of concept that we're moving in the right direction."

The next step is for the team to devise a method of delivering their HIV editor to every single infected cell within a host. In addition, the team has to consider that the editor may need to be individualized for the unique viral sequences of each patient, as HIV-1 is prone to mutation.

"We are working on a number of strategies so we can take the construct into preclinical studies," Dr. Khalili says. "We want to eradicate every single copy of HIV-1 from the patient. That will cure AIDS. I think this technology is the way we can do it."

Dr. Khalili and his colleagues also believe that their technique could be adapted to remove a variety of viruses from patients' cells.