Origin of Rebound Plasma HIV Includes Cells with Identical Proviruses That Are Transcriptionally Active before Stopping of Antiretroviral Therapy

Abstract

Understanding the origin of HIV variants during viral rebound may provide insight into the composition of the HIV reservoir and has implications for the design of curative interventions. HIV single-genome sequences were obtained from 10 AIDS Clinical Trials Group participants who underwent analytic antiretroviral therapy (ART) interruption (ATI). Rebounding variants were compared with those in pre-ART plasma in all 10 participants and with on-ART peripheral blood mononuclear cell (PBMC)-associated DNA and RNA (CA-RNA) in 7/10 participants. The highest viral diversities were found in the DNA and CA-RNA populations. In 3 of 7 participants, we detected multiple, identical DNA and CA-RNA sequences during suppression on ART that exactly matched plasma HIV sequences. Hypermutated DNA and CA-RNA were detected in four participants, contributing to diversities in these compartments that were higher than in the pre-ART and post-ATI plasma. Shifts in the viral rebound populations could be detected in some participants over the 2-to 3-month observation period. These findings suggest that a source of initial rebound viremia could be populations of infected cells that clonally expanded prior to and/or during ART, some of which were already expressing HIV RNA before treatment was interrupted. These clonally expanding populations of HIVinfected cells may represent an important target for strategies aimed at achieving reservoir reduction and sustained virologic remission. IMPORTANCEAntiretroviral therapy alone cannot eradicate the HIV reservoir, and viral rebound is generally rapid after treatment interruption. It has been suggested that clonal expansion of HIV-infected cells is an important mechanism of HIV reservoir persistence, but the contribution of these clonally proliferating cells to the rebounding virus is unknown. We report a study of AIDS Clinical Trials Group participants who underwent treatment interruption and compared rebounding plasma virus with that found within cells prior to treatment interruption. We found several incidences in which plasma HIV variants exactly matched that of multiple proviral DNA copies from infected blood cells sampled before treatment interruption. In addition, we found that these cells were not dormant but were generating unspliced RNA transcripts before treatment was interrupted. Identification of the HIV reservoir and determining its mechanisms for persistence may aid in the development of strategies toward a cure for HIV. (This study was presented in part at the Conference on Retroviruses and Opportunistic Infections, Seattle, WA, February 23 to 26 2015.)

Defective HIV-1 proviruses produce novel protein-coding RNA species in HIV-infected patients on combination antiretroviral therapy

“…However, there is minimal evidence for transcription of defective proviruses (14,27). Here, we were able to identify the presence of clonally expanded defective provirus populations and to demonstrate ongoing transcription of these expanded "defective" proviral clones in two different patients (Pts 5 and 7) with pVL < 40 copies per milliliter (clones ii and vi in Fig.…”

No evidence of HIV replication in children on antiretroviral therapy

“…To look for evidence of ongoing viral replication during ART, HIV populations in samples taken at baseline were compared with the populations present after long-term ART. Three measures of evolution were assessed for significant change over time: (a) HIV-1 genetic diversity, measured as the average pairwise distance (APD), from early infection to long-term infection; (b) genetic divergence from the founder virus(es) using a test for panmixia (3,12,26,27); and (c) root-to-tip distances in maximum-likelihood (ML) trees (3,12,27). ML trees were rooted in 2 ways: first, on an outgroup (HIV subtype C consensus; www.hiv.lanl.gov), and second, on the majority sequence of the baseline sample (Table 1).…”

“…More recently, expanded clones harboring replication-competent proviruses were identified by sequence matches between viruses recovered in multiple different viral outgrowth cultures, revealing that most of the HIV reservoir is maintained by cellular proliferation (4,5,41). Furthermore, proviruses with identical sequences, and thus likely to be in cell clones, were reported to be a source for rebound viremia after stopping ART (12). Collectively, these studies, along with the data shown here, demonstrate that proliferating, infected cells are a common mechanism of reservoir persistence that needs to be targeted to cure HIV-1 infection.…”

“…net). Shifts in population structure were calculated using a subdivision test for panmixia, with a P value of less than 10 -3 as the significance cutoff (as described by the original report and other publications) to account for the high number of comparisons between sequences and nucleotide sites (3,12,26). The test was derived from a geographic population structure test proposed by Hudson et al (26).…”

“…Prior studies addressing the question of ongoing replication have analyzed the evolution of virion RNA or proviral DNA sequences in chronically infected adults as a function of the duration of ART (3,10,(12)(13)(14)(15). Such studies have ascribed the success of ART to its ability to effectively prevent the infection of new cells by reducing the in vivo reproductive rate to less than 1 and effectively halting viral evolution and preventing the associated development of drug resistance (3,16,17).…”