Combinatorial latency reactivation for HIV-1 subtypes and variants.
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ABSTRACT:
SUBMITTER: Burnett JC
PROVIDER: S-EPMC2876650 | biostudies-literature | 2010 Jun
REPOSITORIES: biostudies-literature
Publications
Combinatorial latency reactivation for HIV-1 subtypes and variants.
Burnett John C JC Lim Kwang-Il KI Calafi Arash A Rossi John J JJ Schaffer David V DV Arkin Adam P AP
Journal of virology 20100331 12
The eradication of HIV-1 will likely require novel clinical approaches to purge the reservoir of latently infected cells from a patient. We hypothesize that this therapy should target a wide range of latent integration sites, act effectively against viral variants that have acquired mutations in their promoter regions, and function across multiple HIV-1 subtypes. By using primary CD4(+) and Jurkat cell-based in vitro HIV-1 latency models, we observe that single-agent latency reactivation therapy ...[more]
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Project description:Latency is the main obstacle towards an HIV cure, with cure strategies aiming to either elicit or prevent viral reactivation. While these strategies have shown promise, they have only succeeded in modulating latency in a fraction of the latent HIV reservoir, suggesting that the mechanisms controlling HIV latency are not completely understood, and that comprehensive latency modulation will require targeting of multiple latency maintenance pathways. We show here that the transcriptional co-activator and the central mediator of canonical Wnt signaling, β-catenin, inhibits HIV transcription in CD4+ T cells via TCF-4 LTR binding sites. Further, we show that inhibiting the β-catenin pathway reactivates HIV in a primary TCM cell model of HIV latency, primary cells from cART-controlled HIV donors, and in CD4+ latent cell lines. β-catenin inhibition or activation also enhanced or inhibited the activity of several classes of HIV latency reversing agents, respectively, in these models, with significant synergy of β-catenin and each LRA class tested. In sum, we identify β-catenin as a novel regulator of HIV latency in vitro and ex vivo, adding new therapeutic targets that may be combined for comprehensive HIV latency modulation in HIV cure efforts.
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Project description:DNA methylation of retroviral promoters and enhancers localized in the provirus 5' long terminal repeat (LTR) is considered to be a mechanism of transcriptional suppression that allows retroviruses to evade host immune responses and antiretroviral drugs. However, the role of DNA methylation in the control of HIV-1 latency has never been unambiguously demonstrated, in contrast to the apparent importance of transcriptional interference and chromatin structure, and has never been studied in HIV-1-infected patients. Here, we show in an in vitro model of reactivable latency and in a latent reservoir of HIV-1-infected patients that CpG methylation of the HIV-1 5' LTR is an additional epigenetic restriction mechanism, which controls resistance of latent HIV-1 to reactivation signals and thus determines the stability of the HIV-1 latency. CpG methylation acts as a late event during establishment of HIV-1 latency and is not required for the initial provirus silencing. Indeed, the latent reservoir of some aviremic patients contained high proportions of the non-methylated 5' LTR. The latency controlled solely by transcriptional interference and by chromatin-dependent mechanisms in the absence of significant promoter DNA methylation tends to be leaky and easily reactivable. In the latent reservoir of HIV-1-infected individuals without detectable plasma viremia, we found HIV-1 promoters and enhancers to be hypermethylated and resistant to reactivation, as opposed to the hypomethylated 5' LTR in viremic patients. However, even dense methylation of the HIV-1 5'LTR did not confer complete resistance to reactivation of latent HIV-1 with some histone deacetylase inhibitors, protein kinase C agonists, TNF-alpha, and their combinations with 5-aza-2deoxycytidine: the densely methylated HIV-1 promoter was most efficiently reactivated in virtual absence of T cell activation by suberoylanilide hydroxamic acid. Tight but incomplete control of HIV-1 latency by CpG methylation might have important implications for strategies aimed at eradicating HIV-1 infection.
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2010-06-06 | E-GEOD-1441 | biostudies-arrayexpress
Project description:Latency allows HIV-1 to persist in long-lived cellular reservoirs, preventing virus eradication. We have previously shown that the heat shock protein 90 (Hsp90) is required for HIV-1 gene expression and mediates greater HIV-1 replication in conditions of hyperthermia. Here we report that specific inhibitors of Hsp90 such as 17-(N-allylamino)-17-demethoxygeldanamycin and AUY922 prevent HIV-1 reactivation in CD4+ T cells. A single modification at position 19 in the Hsp90 inhibitors abolished this activity, supporting the specificity of the target. We tested the impact of Hsp90 on known pathways involved in HIV-1 reactivation from latency; they include protein kinase Cs(PKCs), mitogen activated protein kinase/extracellular signal regulated kinase/positive transcriptional elongation factor-b and NF-?B. We found that Hsp90 was required downstream of PKCs and was not required for mitogen activated protein kinase activation. Inhibition of Hsp90 reduced degradation of IkB? and blocked nuclear translocation of transcription factor p65/p50, suppressing the NF-?B pathway. Coimmunoprecipitation experiments showed that Hsp90 interacts with inhibitor of nuclear factor kappa-B kinase (IKK) together with cochaperone Cdc37, which is critical for the activity of several kinases. Targeting of Hsp90 by AUY922 dissociated Cdc37 from the complex. Therefore, Hsp90 controls HIV-1 reactivation from latency by keeping the IKK complex functional and thus connects T-cell activation with HIV-1 replication. AUY922 is in phase II clinical trial and, in combination with a PKC-? inhibitor in phase II clinical trial, almost completely suppressed HIV-1 reactivation at 15 nM with no cytotoxicity. Selective targeting of the Hsp90/Cdc37 interaction may provide a powerful approach to suppress HIV-1 reactivation from latency.
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2004-08-16 | GSE1441 | GEO
Project description:UnlabelledAntiretroviral therapy (ART) inhibits HIV-1 replication, but the virus persists in latently infected resting memory CD4(+) T cells susceptible to viral reactivation. The virus-encoded early gene product Tat activates transcription of the viral genome and promotes exponential viral production. Here we show that the Tat inhibitor didehydro-cortistatin A (dCA), unlike other antiretrovirals, reduces residual levels of viral transcription in several models of HIV latency, breaks the Tat-mediated transcriptional feedback loop, and establishes a nearly permanent state of latency, which greatly diminishes the capacity for virus reactivation. Importantly, treatment with dCA induces inactivation of viral transcription even after its removal, suggesting that the HIV promoter is epigenetically repressed. Critically, dCA inhibits viral reactivation upon CD3/CD28 or prostratin stimulation of latently infected CD4(+) T cells from HIV-infected subjects receiving suppressive ART. Our results suggest that inclusion of a Tat inhibitor in current ART regimens may contribute to a functional HIV-1 cure by reducing low-level viremia and preventing viral reactivation from latent reservoirs.ImportanceAntiretroviral therapy (ART) reduces HIV-1 replication to very low levels, but the virus persists in latently infected memory CD4(+) T cells, representing a long-lasting source of resurgent virus upon ART interruption. Based on the mode of action of didehydro-cortistatin A (dCA), a Tat-dependent transcription inhibitor, our work highlights an alternative approach to current HIV-1 eradication strategies to decrease the latent reservoir. In our model, dCA blocks the Tat feedback loop initiated after low-level basal reactivation, blocking transcriptional elongation and hence viral production from latently infected cells. Therefore, dCA combined with ART would be aimed at delaying or halting ongoing viral replication, reactivation, and replenishment of the latent viral reservoir. Thus, the latent pool of cells in an infected individual would be stabilized, and death of the long-lived infected memory T cells would result in a continuous decay of this pool over time, possibly culminating in the long-awaited sterilizing cure.
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Project description:Summary Noise-modulating chemicals can synergize with transcriptional activators in reactivating latent HIV to eliminate latent HIV reservoirs. To understand the underlying biomolecular mechanism, we investigate a previous two-gene-state model and identify two necessary conditions for the synergy: an assumption of the inhibition effect of transcription activators on noise enhancers; and frequent transitions to the gene non-transcription-permissive state. We then develop a loop-four-gene-state model with Tat transcription/translation and find that drug synergy is mainly determined by the magnitude and direction of energy input into the genetic regulatory kinetics of the HIV promoter. The inhibition effect of transcription activators is actually a phenomenon of energy dissipation in the nonequilibrium gene transition system. Overall, the loop-four-state model demonstrates that energy dissipation plays a crucial role in HIV latency reactivation, which might be useful for improving drug effects and identifying other synergies on lentivirus latency reactivation. Graphical abstract Highlights • The inhibition of Activator on Noise enhancer is necessary for their synergy in reactivating HIV• The drug synergy is a nonequilibrium phenomenon in the gene regulatory system• The magnitude and direction of energy input determine the drug synergy• This nonequilibrium mechanism is general without regarding molecular details Drugs; Virology; Mathematical biosciences
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Project description:Highly active antiretroviral therapy (HAART) has improved the outlook for the HIV epidemic, but does not provide a cure. The proposed "shock-and-kill" strategy is directed at inducing latent HIV reservoirs, which may then be purged via boosted immune response or targeting infected cells. We describe five novel compounds that are capable of reversing HIV latency without affecting the general T-cell activation state. The new compounds exhibit synergy for reactivation of latent provirus with other latency-reversing agents (LRAs), in particular ingenol-3-angelate/PEP005. One compound, designated PH02, was efficient at reactivating viral transcription in several cell lines bearing reporter HIV-1 at different integration sites. Furthermore, it was capable of reversing latency in resting CD4+ T lymphocytes from latently infected aviremic patient cells on HAART, while producing minimal cellular toxicity. The combination of PH02 and PEP005 produces a strong synergistic effect for reactivation, as demonstrated through a quantitative viral outgrowth assay (qVOA), on CD4+ T lymphocytes from HIV-1-infected individuals. We propose that the PH02/PEP005 combination may represent an effective novel treatment for abrogating persistent HIV-1 infection.
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