Archived HIV-1 Drug Resistance Mutations: Role of Proviral HIV-1 DNA Genotype for the Management of Virological Responder People Living with HIV

Abstract

Despite its effectiveness in controlling plasma viremia, antiretroviral therapy (ART) cannot target proviral DNA, which remains an obstacle to HIV-1 eradication. When treatment is interrupted, the reservoirs can act as a source of viral rebound, highlighting the value of proviral DNA as an additional source of information on an individual's overall resistance burden. In cases where the viral load is too low for successful HIV-1 RNA genotyping, HIV-1 DNA can help identify resistance mutations in treated individuals. The absence of treatment history, the need to adjust ART despite undetectable viremia, or the presence of LLV further support the use of genotypic resistance tests (GRTs) on HIV-1 DNA. Conventionally, GRTs have been achieved through Sanger sequencing, but the advances in NGS are leading to an increase in its use, allowing the detection of minority variants present in less than 20% of the viral population. The clinical significance of these mutations remains under debate, with interpretations varying based on context. Additionally, proviral DNA is subject to APOBEC3-induced hypermutation, which can lead to defective, nonviable viral genomes, a factor that must be considered when performing GRTs on HIV-1 DNA.

Keywords: APOBEC; DNA GRT; HIV-1; NGS; Sanger sequencing; reservoir.

Figure 1

Main models proposed to explain the establishment of HIV-1 latently infected cells. Pre-activation latency: resting CD4+ T cells are refractory to HIV-1 infection, but under certain conditions such as the presence of IL-7, CCL-19, and CCL-20, the permissiveness of these cells to HIV-1 may increase, resulting in integrated proviruses. Post-activation latency: after activation, CD4+ T cells can become infected with HIV-1, but a subset of activated and infected CD4+ T cells can return to a resting state that allows integration and persistence of the viral genome. The presence of certain molecules can favor this transitional state. For instance, PD-1 dampens T-cell activation and the presence of myeloid dendritic cells (mDCs) can favor the transition to a post-activation latency state. mDCs: myeloid dendritic cells; PD-1: programmed cell death protein 1; TGF-β: transforming growth factor. Adapted from “The multifaceted nature of HIV latency” by Dufour et al. J ClinInvest. 2021; 131(11): e151380. doi: 10.1172/JCI151380 [4]. The above image was created with https://www.biorender.com/ (accessed on 28 July 2024).

Figure 2

Concordance/discordance between RNA and DNA genotypic resistance test in PLWH with at least one virological failure. In the early stages of virological failure, the detection of emerging drug resistance mutations (DRMs) is more sensitive in viral RNA than in proviral DNA. This is because the drug-resistant virus has not yet had time to be archived in the reservoir. However, if the drug-resistant virus has been circulating for a sufficient period, allowing resistant viral variants to be stored in the reservoir, there may be good concordance between RNA and DNA genotyping (panel A). When the ART regimen is changed, the resistant viral variants may only be found in the reservoir. Conversely, if the drug-resistant virus has not circulated long enough, it will not have been archived in the reservoir (panel B). In the image, PBMCs with drug-resistant provirus are represented with orange DNA, while wild-type PBMCs are represented with blue DNA. The above image was created with https://www.biorender.com/ (accessed on 3 August 2024).

Figure 3

Mechanism of action of APOBEC3 (A3) against HIV-1 infection and Vif countermeasure. In the absence of Vif, (Vif −) A3 proteins are incorporated into the new virions by binding HIV-1 genomic RNA and the nucleocapsid protein. RNase H-dependent degradation of the HIV-1 RNA template during the reverse transcription phase exposes newly synthesized single-stranded DNA (ssDNA). A3 proteins detach from the genomic RNA, bind the nascent ssDNA, and deaminate cytidine into uridine (C-to-U) on the new ssDNA inducing high rates of G-to-A mutation in the newly synthesized plus strand of viral DNA (panel A). In the presence of Vif (Vif +), the formation of an E3 ubiquitin ligase complex is promoted, which targets A3 proteins for proteasomal degradation (panel B). A3 is represented in yellow; Vif is represented in green. The above image was created with https://www.biorender.com/ (accessed on 12 August 2024).