Hybrid nanocarriers incorporating mechanistically distinct drugs for lymphatic CD4+ T cell activation and HIV-1 latency reversal.
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ABSTRACT:
SUBMITTER: Cao S
PROVIDER: S-EPMC6436934 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:HIV persists in latently infected CD4+ T cells during antiretroviral therapy (ART). Immune checkpoint molecules, including PD-1, are preferentially expressed at the surface of persistently infected cells. However, whether PD-1 plays a functional role in HIV latency and reservoir persistence remains unknown. Using CD4+ T cells from HIV-infected individuals, we show that the engagement of PD-1 inhibits viral production at the transcriptional level and abrogates T-cell receptor (TCR)-induced HIV reactivation in latently infected cells. Conversely, PD-1 blockade with the monoclonal antibody pembrolizumab enhances HIV production in combination with the latency reversing agent bryostatin without increasing T cell activation. Our results suggest that the administration of immune checkpoint blockers to HIV-infected individuals on ART may facilitate latency disruption.
| S-EPMC6379401 | biostudies-literature
Project description:To date, an affordable, effective treatment for an HIV-1 cure remains only a concept with most "latency reversal" agents (LRAs) lacking specificity for the latent HIV-1 reservoir and failing in early clinical trials. We assessed HIV-1 latency reversal using a multivalent HIV-1-derived virus-like particle (HLP) to treat samples from 32 people living with HIV-1 (PLWH) in Uganda, US and Canada who initiated combined antiretroviral therapy (cART) during chronic infection. Even after 5-20 years on stable cART, HLP could target CD4+ T cells harbouring latent HIV-1 reservoir resulting in 100-fold more HIV-1 release into culture supernatant than by common recall antigens, and 1000-fold more than by chemotherapeutic LRAs. HLP induced release of a divergent and replication-competent HIV-1 population from PLWH on cART. These findings suggest HLP provides a targeted approach to reactivate the majority of latent HIV-1 proviruses among individuals infected with HIV-1.
| S-EPMC10967673 | biostudies-literature
Project description:OBJECTIVE:To determine whether latency can be established and reversed in both proliferating and nonproliferating CD4+ T cells in the same model in vitro. METHODS:Activated CD4+ T cells were infected with either a nonreplication competent, luciferase reporter virus or wild-type full-length enhanced green fluorescent protein (EGFP) reporter virus and cultured for 12 days. The cells were then sorted by flow cytometry to obtain two distinct T-cell populations that did not express the T-cell activation markers, CD69, CD25 and human leukocyte antigen (HLA)-DR: CD69CD25HLA-DR small cells (nonblasts) that had not proliferated in vitro following mitogen stimulation and CD69CD25HLA-DR large cells (which we here call transitional blasts) that had proliferated. The cells were then reactivated with latency-reversing agents and either luciferase or EGFP quantified. RESULTS:Inducible luciferase expression, consistent with latent infection, was observed in nonblasts and transitional blasts following stimulation with either phorbol-myristate-acetate/phytohemagglutinin (3.8?±?1 and 2.9?±?0.5 fold above dimethyl sulfoxide, respectively) or romidepsin (2.1?±?0.6 and 1.8?±?0.2 fold above dimethyl sulfoxide, respectively). Constitutive expression of luciferase was higher in transitional blasts compared with nonblasts. Using wild-type full-length EGFP reporter virus, inducible virus was observed in nonblasts but not in transitional blasts. No significant difference was observed in the response to latency-reversing agents in either nonblasts or transitional blasts. CONCLUSION:HIV latency can be established in vitro in resting T cells that have not proliferated (nonblasts) and blasts that have proliferated (transitional blasts). This model could potentially be used to assess new strategies to eliminate latency.
| S-EPMC6319264 | biostudies-literature
Project description:Latency reversal agents (LRAs) have proven to induce HIV-1 transcription in vivo but are ineffective at decreasing the size of the latent reservoir in antiretroviral treated patients. The capacity of the LRAs to perturb the viral reservoir present in distinct subpopulations of cells is currently unknown. Here, using a new RNA FISH/flow ex vivo viral reactivation assay, we performed a comprehensive assessment of the viral reactivation capacity of different families of LRAs, and their combinations, in different CD4+ T cell subsets. We observed that a median of 16.28% of the whole HIV-reservoir induced HIV-1 transcripts after viral reactivation, but only 10.10% of these HIV-1 RNA+ cells produced the viral protein p24. Moreover, none of the LRAs were powerful enough to reactivate HIV-1 transcription in all CD4+ T cell subpopulations. For instance, the combination of Romidepsin and Ingenol was identified as the best combination of drugs at increasing the proportion of HIV-1 RNA+ cells, in most, but not all, CD4+ T cell subsets. Importantly, memory stem cells were identified as highly resistant to HIV-1 reactivation, and only the combination of Panobinostat and Bryostatin-1 significantly increased the number of cells transcribing HIV within this subset. Overall, our results validate the use of the RNA FISH/flow technique to assess the potency of LRAs among different CD4+ T cell subsets, manifest the intrinsic differences between cells that encompass the latent HIV reservoir, and highlight the difficulty to significantly impact the latent infection with the currently available drugs. Thus, our results have important implications for the rational design of therapies aimed at reversing HIV latency from diverse cellular reservoirs.
| S-EPMC6715238 | biostudies-literature
Project description:HIV latency occurs predominantly in long-lived resting CD4+ T cells; however, latent infection also occurs in T cell subsets, including proliferating CD4+ T cells. We compared the establishment and maintenance of latent infection in nonproliferating and proliferating human CD4+ T cells cocultured with syngeneic myeloid dendritic cells (mDC). Resting CD4+ T cells were labeled with the proliferation dye eFluor 670 and cultured alone or with mDC, plasmacytoid dendritic cells, or monocytes in the presence of staphylococcal enterotoxin B (SEB). Cells were cultured for 24 h and infected with CCR5-tropic enhanced GFP (EGFP) reporter HIV. Five days postinfection, nonproductively infected EGFP- CD4+ T cells that were either nonproliferating (eFluor 670hi) or proliferating (eFluor 670lo) were sorted and cultured for an additional 7 d (day 12) with IL-7 and antiretrovirals. At day 5 postinfection, sorted, nonproductively infected T cells were stimulated with anti-CD3/CD28, and induced expression of EGFP was measured to determine the frequency of latent infection. Integrated HIV in these cells was confirmed using quantitative PCR. By these criteria, latent infection was detected at day 5 and 12 in proliferating T cells cocultured with mDC and monocytes but not plasmacytoid dendritic cells, where CD4+ T cells at day 12 were poor. At day 5 postinfection, nonproliferating T cells expressing SEB-specific TCR Vβ-17 were enriched in latent infection compared with non-SEB-specific TCR Vβ-8.1. Together, these data show that both nonproliferating and proliferating CD4+ T cells can harbor latent infection during SEB-stimulated T cell proliferation and that the establishment of HIV latency in nonproliferating T cells is linked to expression of specific TCR that respond to SEB.
| S-EPMC6103814 | biostudies-literature
Project description:During antiretroviral therapy (ART), human immunodeficiency virus type 1 (HIV-1) persists as a latent reservoir in CD4+ T cell subsets in central memory (TCM), transitional memory (TTM), and effector memory (TEM) CD4+ T cells. We have identified differences in mechanisms underlying latency and responses to latency-reversing agents (LRAs) in ex vivo CD4+ memory T cells from virally suppressed HIV-infected individuals and in an in vitro primary cell model of HIV-1 latency. Our ex vivo and in vitro results demonstrate the association of transcriptional pathways of T cell differentiation, acquisition of effector function, and cell cycle entry in response to LRAs. Analyses of memory cell subsets showed that effector memory pathways and cell surface markers of activation and proliferation in the TEM subset are predictive of higher frequencies of cells carrying an inducible reservoir. Transcriptional profiling also demonstrated that the epigenetic machinery (known to control latency and reactivation) in the TEM subset is associated with frequencies of cells with HIV-integrated DNA and inducible HIV multispliced RNA. TCM cells were triggered to differentiate into TEM cells when they were exposed to LRAs, and this increase of TEM subset frequencies upon LRA stimulation was positively associated with higher numbers of p24+ cells. Together, these data highlight differences in underlying biological latency control in different memory CD4+ T cell subsets which harbor latent HIV in vivo and support a role for differentiation into a TEM phenotype in facilitating latency reversal.IMPORTANCE By performing phenotypic analysis of latency reversal in CD4+ T cells from virally suppressed individuals, we identify the TEM subset as the largest contributor to the inducible HIV reservoir. Differential responses of memory CD4+ T cell subsets to latency-reversing agents (LRAs) demonstrate that HIV gene expression is associated with heightened expression of transcriptional pathways associated with differentiation, acquisition of effector function, and cell cycle entry. In vitro modeling of the latent HIV reservoir in memory CD4+ T cell subsets identify LRAs that reverse latency with ranges of efficiency and specificity. We found that therapeutic induction of latency reversal is associated with upregulation of identical sets of TEM-associated genes and cell surface markers shown to be associated with latency reversal in our ex vivo and in vitro models. Together, these data support the idea that the effector memory phenotype supports HIV latency reversal in CD4+ T cells.
| S-EPMC6880164 | biostudies-literature
Project description:Herpes simplex virus type 2 (HSV-2) coinfection is associated with increased HIV-1 viral loads and expanded tissue reservoirs, but the mechanisms are not well defined. HSV-2 recurrences result in an influx of activated CD4+ T cells to sites of viral replication and an increase in activated CD4+ T cells in peripheral blood. We hypothesized that HSV-2 induces changes in these cells that facilitate HIV-1 reactivation and replication and tested this hypothesis in human CD4+ T cells and 2D10 cells, a model of HIV-1 latency. HSV-2 promoted latency reversal in HSV-2-infected and bystander 2D10 cells. Bulk and single-cell RNA-Seq studies of activated primary human CD4+ T cells identified decreased expression of HIV-1 restriction factors and increased expression of transcripts including MALAT1 that could drive HIV replication in both the HSV-2-infected and bystander cells. Transfection of 2D10 cells with VP16, an HSV-2 protein that regulates transcription, significantly upregulated MALAT1 expression, decreased trimethylation of lysine 27 on histone H3 protein, and triggered HIV latency reversal. Knockout of MALAT1 from 2D10 cells abrogated the response to VP16 and reduced the response to HSV-2 infection. These results demonstrate that HSV-2 contributes to HIV-1 reactivation through diverse mechanisms, including upregulation of MALAT1 to release epigenetic silencing.
| S-EPMC10232005 | biostudies-literature
Project description:Cells that are latently infected with HIV-1 preclude an HIV-1 cure, as antiretroviral therapy does not target this latent population. HIV-1 is highly genetically diverse, with over 10 subtypes and numerous recombinant forms circulating worldwide. In spite of this vast diversity, much of our understanding of latency and latency reversal is largely based on subtype B viruses. As such, most of the development of cure strategies targeting HIV-1 are solely based on subtype B. It is currently assumed that subtype does not influence the establishment or reactivation of latent viruses. However, this has not been conclusively proven one way or the other. A better understanding of the factors that influence HIV-1 latency in all viral subtypes will help develop therapeutic strategies that can be applied worldwide. Here, we review the latest literature on subtype-specific factors that affect viral replication, pathogenesis, and, most importantly, latency and its reversal.
| S-EPMC6950696 | biostudies-literature
Project description:The human immunodeficiency virus (HIV-1) causes a progressive depletion of CD4+ T cells, hampering immune function. Current experimental strategies to fight the virus focus on the reactivation of latent HIV-1 in the viral reservoir to make the virus detectable by the immune system, by searching for latency reversal agents (LRAs). We hypothesize that if common molecular pathways elicited by the presence of LRAs are known, perhaps new, more efficient, "shock-and-kill" strategies can be found. Thus, the objective of the present study is to re-evaluate RNA-Seq assays to find differentially expressed genes (DEGs) during latency reversal via transcriptome analysis. We selected six studies (45 samples altogether: 16 negative controls and 29 LRA-treated CD4+ T cells) and 11 LRA strategies through a systematic search in Gene Expression Omnibus (GEO) and PubMed databases. The raw reads were trimmed, counted, and normalized. Next, we detected consistent DEGs in these independent experiments. AZD5582, romidepsin, and suberanilohydroxamic acid (SAHA) were the LRAs that modulated most genes. We detected 948 DEGs shared by those three LRAs. Gene ontology analysis and cross-referencing with other sources of the literature showed enrichment of cell activation, differentiation and signaling, especially mitogen-activated protein kinase (MAPK) and Rho-GTPases pathways.
| S-EPMC7601709 | biostudies-literature
Project description:Therapeutic strategies that target the latent HIV-1 reservoir in resting CD4+ T cells of infected individuals are always administered in the presence of combination antiretroviral therapy. Using a primary cell of HIV-1 latency, we evaluated whether different antiviral drug classes affected latency reversal (as assessed by extracellular virus production) by anti-CD3/CD28 monoclonal antibodies or bryostatin 1. We found that the nonnucleoside reverse transcriptase inhibitors efavirenz and rilpivirine significantly decreased HIV-1 production, by ≥1 log.
| S-EPMC5328526 | biostudies-literature