Abstract
The persistence of a reservoir of latently infected CD4 T cells remains one of the major obstacles to cure HIV. Numerous strategies are being explored to eliminate this reservoir. To translate these efforts into clinical trials, there is a strong need for validated biomarkers that can monitor the reservoir over time in vivo. A comprehensive study was designed to evaluate and compare potential HIV-1 reservoir biomarkers. A cohort of 25 patients, treated with suppressive antiretroviral therapy was sampled at three time points, with median of 2.5 years (IQR: 2.4–2.6) between time point 1 and 2; and median of 31 days (IQR: 28–36) between time point 2 and 3. Patients were median of 6 years (IQR: 3–12) on ART, and plasma viral load (<50 copies/ml) was suppressed for median of 4 years (IQR: 2–8). Total HIV-1 DNA, unspliced (us) and multiply spliced HIV-1 RNA, and 2LTR circles were quantified by digital PCR in peripheral blood, at 3 time points. At the second time point, a viral outgrowth assay (VOA) was performed, and integrated HIV-1 DNA and relative mRNA expression levels of HIV-1 restriction factors were quantified. No significant change was found for long- and short-term dynamics of all HIV-1 markers tested in peripheral blood. Integrated HIV-1 DNA was associated with total HIV-1 DNA (p<0.001, R² = 0.85), us HIV-1 RNA (p = 0.029, R² = 0.40), and VOA (p = 0.041, R2 = 0.44). Replication-competent virus was detected in 80% of patients by the VOA and it correlated with total HIV-1 DNA (p = 0.039, R² = 0.54). The mean quantification difference between Alu-PCR and VOA was 2.88 log10, and 2.23 log10 between total HIV-1 DNA and VOA. The levels of usHIV-1 RNA were inversely correlated with mRNA levels of several HIV-1 restriction factors (TRIM5α, SAMHD1, MX2, SLFN11, pSIP1). Our study reveals important correlations between the viral outgrowth and total and integrated HIV-1 DNA measures, suggesting that the total pool of HIV-1 DNA may predict the size of the replication-competent virus in ART suppressed patients.
Highlights
Current antiretroviral therapy (ART) successfully suppresses viral replication and reduces HIV-1 transmission, but fails to completely eliminate HIV-1 [1]
Our study reveals important correlations between the viral outgrowth and total and integrated HIV-1 DNA measures, suggesting that the total pool of HIV-1 DNA may predict the size of the replication-competent virus in ART suppressed patients
Total HIV-1 DNA estimates may be biased by unintegrated HIV-1 DNA, the correlation observed in our study indicates that total HIV-1 DNA could be a good surrogate maker for integrated HIV-1 DNA in patients on stable ART as reported earlier [18]
Summary
Current antiretroviral therapy (ART) successfully suppresses viral replication and reduces HIV-1 transmission, but fails to completely eliminate HIV-1 [1]. Different markers to monitor changes in the size and composition of the viral reservoir are being explored [5]. In this context, it is crucial to clearly define the characteristics of the reservoir that causes viral rebound upon treatment discontinuation. It is well known that only a small fraction of integrated HIV-1 DNA is composed of intact HIV-1 DNA that is capable to produce replication-competent virus. This small fraction of replication-competent virus is the sole contributor to viral persistence and sole source of viral rebound after treatment discontinuation. An effective biomarker needs to correlate with the changes in the fraction of the replication-competent reservoir and should not be biased by the levels of archival, dead-end integrated proviruses [5]
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