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HIV Hiding Inside Patient's Cells Can Now be Measured Accurately
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HIV Hiding Inside Patientís Cells Can Now be Measured Accurately

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Highlights:
  • Human Immunodeficiency Virus (HIV) can hide inside the cells of infected patients
  • HIV present inside these cells can now be accurately detected and measured
  • This would allow the precise assessment of antiviral therapies against HIV

Human Immunodeficiency Virus (HIV), which is capable of hiding itself within cells in the body can now be detected and quantified accurately, reveals a new study from the Howard Hughes Medical Institute, USA.

HIV can enter cells and integrate into the cell's genome and remain indefinitely in an inactive form. In this integrated dormant state, even antiretroviral drugs, which attack HIV, are not able to kill the virus.

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A novel genetic technique has been developed that can accurately measure the amount of the viral particles integrated within the cell's genome, which is critical for evaluating the efficacy of new antiretroviral therapies.

The study was led by Prof. Robert F. Siliciano, MD, PhD, who is an Investigator at the Howard Hughes Medical Institute and a Professor of Medicine at the Johns Hopkins University School of Medicine, Maryland, USA. He also holds joint appointments in the Departments of Molecular Biology and Genetics, Pharmacology and Molecular Sciences, and Biology, at the Johns Hopkins School of Arts and Sciences.
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The study has been published in Nature, the prestigious journal of Springer Nature Publishing, UK.

Challenges in Detecting and Quantifying HIV

During the replication cycle of HIV, it infects a population of immunological cells known as CD4+ cells, which circulate in the blood. Following infection, HIV continues to replicate within these CD4+ cells, resulting in the integration of the viral genes into the genome of these cells. Therefore, the viral genetic information remains stored in a latent state within the CD4+ cells, which make it extremely difficult to detect, much less quantify.

Trying to measure this 'latent reservoir' of HIV particles using conventional techniques has yielded erroneous results, sometimes overestimating up to 100-folds. This has necessitated the development of newer techniques that can not only detect, but also count the latent viral particles.

New Techniques for Detecting and Quantifying HIV

The development of the new technique occurred in three stages, over many years of experimentation. These are highlighted below:
  • Quantitative Viral Outgrowth Assay (QVOA): The basis of the new technique originated in 1995 when Siliciano and his team discovered the 'latent reservoir' of HIV by using a technique called Quantitative Viral Outgrowth Assay (QVOA), which was developed by them. However, a major drawback of this assay is that it is very difficult to grow the HIV-infected cells, coupled with the fact that it is very cumbersome and time-consuming, often taking several weeks to complete the assay.
  • Original PCR Technique: In order to circumvent the above problems, researchers prefer to use a simpler technique known as the Polymerase Chain Reaction (PCR), which is widely used in Molecular Biology labs. PCR detects the levels of viral DNA (deoxyribonucleic acid - the genetic code of an organism) in the CD4+ cells. But even this technique is not absolutely perfect for the accurate estimation of viral DNA, as 98 percent of the DNA is harmless. Only the remaining two percent viral DNA is harmful because it can reactivate to cause re-infection, and is thus relevant for monitoring the health status of the patient. Therefore, the original PCR technique also suffers from an overestimation of the viral particles present in CD4+ cells.
  • New PCR Technique: To overcome these drawbacks of the original PCR technique, Siliciano's team designed a new type of PCR that is capable of distinguishing between intact and defective segments in the viral DNA. This novel technique uses dual-colored fluorescent probes that are capable of binding to DNA regions that are highly prone to undergo mutations (changes), thereby causing genetic defects in the virus. A color read-out allows for quantification of the genetic defects in the viral DNA template. Therefore, this new technique gives an accurate measurement of the levels of harmful HIV particles present in the CD4+ cells. This can help monitor the patient's response to various antiviral therapies accurately without the risk of overestimation, which had plagued the previous techniques.

Concluding Remarks

The new technique is a landmark development in HIV diagnostics. Now, the efficacy of an antiretroviral drug or a combination of drugs can be assessed to see if they are able to destroy the HIV hiding within the cellular genome by measuring the activity of the virus.

"For decades, the field has been clamoring for an accurate measure for these hidden viral templates," Siliciano says. "Now, we have a good way to know if we are making a dent in their numbers." He adds: "We may still be a long way from a cure, but now at least we can measure our progress."

Reference :
  1. A Quantitative Approach for Measuring the Reservoir of Latent HIV-1 Proviruses - (http://dx.doi.org/10.1038/s41586-019-0898-8)


Source: Medindia

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