Dormant HIV Copies Are Mostly Defective

A recent study published in Nature Medicine showed that more than 90% of latent proviruses are damaged and no longer function, indicating the need for new ways to count only non-damaged proviruses.

Since the introduction of antiretroviral therapies (ART), the treatment has been an effective way to treat HIV. However, the virus is only able to be completely cleared because it’s able to successfully hide inside certain immune system cells, and become active when ART is stopped.

Efforts have been made to knock out the latent virus reservoirs, but have been unsuccessful. While new HIV experimental drugs are being tested for efficacy in eliminating the provirus, researchers typically take snapshots of the provirus reservoir levels, before and after treatment.

The most common technique is called polymerase chain reaction (PCR), which allows researchers to quantify the presence of certain HIV genes, but unfortunately it counts defective proviruses too.

“To cure HIV, you want to get rid of the proviruses without defects,” said senior study author Robert Siliciano, MD, PhD. “But our work shows that the standard assays used to do that are measuring forms of the virus that are not really relevant to these cure strategies.”

Previously, researchers had not taken issue with the PCR technique.

“Most of us in the field assumed that the initial pool of latent proviruses, immediately after infection, contained entirely functional copies of the virus, so we also assumed that PCR — at least for patients treated with ART early in the course of infection – would mostly be counting proviruses that had the ability to cause disease again,” said first study author Katherine Bruner.

A different method called a quantitative viral outgrowth assay (QVOA) is a laborious technique considered to be the best way to measure the size of the reservoir. To determine whether or not the 2 techniques were accurately counting the proviruses, researchers sequenced the full genomes of a large number of viruses from 19 men and women of various races.

Participants were between the ages of 20- and 76-years-old, and had started ART either less than 100 days after infection or more than 180 days after. All participants had been on ART for a minimum of 8 months.

The results of the study showed that between 90% and 98% of proviruses from any individual patient were defective, and unable to replicate in order to become an active virus. The results remained, regardless pf whether patients started ART within weeks or years of contracting the disease.

“Even in patients treated really early, if you’re using a PCR-based approach to measure the reservoir, you’re really just detecting a lot of defective proviruses,” Bruner said.

Researchers compared the full sequencing data with the analyses of the same patients’ proviruses using QVOA or PCR. The results showed that among patients who started ART more than 6 months after being infected, the PCR technique overestimated the actual size of the latent reservoir by a median of 188-fold in patients, and 13-fold in patients who began treatment earlier.

The QVOA method was found to underestimate the reservoir by a median of 27-fold in patients who started treatment late and 25-fold in patients who began treatment early.

“(The full sequencing method) is very helpful from a research standpoint but time-consuming, expensive, and impractical for wide clinical use,” Bruner said. However, the 2 inaccurate standbys are all that’s available to researchers for now, she noted.

Study authors hope their findings will provide further understanding of what the PCR and QVOA results actually mean.

“With current counting methods, it looks like you can bracket the true size range,” Siliciano said. “But we’d like to figure out better ways to do the measurements.”