CROI 2019 Abstract eBook

Abstract eBook

Oral Abstracts

of replication-competent, transcriptionally-silent proviruses in a latent state. Latently-infected cells, mainly resting CD4+ T cells but possibly other infected cell types, are insensitive to cART and can evade the patient immune system. However, latency is a reversible state and reactivation of HIV-1 gene expression from latently-infected cells constitutes a permanent source for virus production in cART-treated patients. One of the most explored therapeutic approach aiming at purging HIV-1 reservoirs, the shock and kill strategy, consists in reactivating HIV-1 gene expression from the latently-infected cellular reservoirs, followed by killing of the virus-producing infected cells. Several classes of latency reversing agents (LRAs), including epigenetic modifying agents, have been studied to reactivate viral gene expression, based on the understanding of the molecular mechanisms involved in HIV-1 latency. Due to the small numbers of latently- infected cells found in vivo, these molecular mechanisms have been mainly studied in in vitro cell line and primary cell models for HIV-1 latency and in ex vivo models obtained with patient-derived latently-infected cells. However, many of these studies have highlighted the major contribution of epigenetic and transcriptional mechanisms to HIV-1 silencing. Clinical trials using individual LRAs have yielded variable, but sometimes encouraging results concerning their ability to induce HIV-1 transcription. However, none of these trials have caused significant and persistent reduction in the HIV-1 reservoir size. The multiplicity of the silencing mechanisms involved in HIV-1 latency, the intrinsically dynamic and heterogeneous nature of the latent HIV-1 cellular reservoirs, the variations in patient clinical history and the lack of selectivity of LRAs constitute causes of the LRA ineffectiveness in clinical trials. These causes will need to be understood in order to rationally improve the “shock” strategy so that it could reach clinical success. Over the past decade, our studies found that HIV-1 latency is driven by stochastic fluctuations in Tat transcription (Weinberger et al. Cell 2005), which provided the first evidence for a classic theory that transcriptional fluctuations (a.k.a., ‘noise’) are harnessed for cell-fate decisions. We subsequently characterized a ‘hardwired’ virus-intrinsic HIV circuit that regulates latency and appears optimized by evolution (Razooky et al. Cell 2015; Rouzine et al. Cell 2015). Our studies also found FDA-approved compounds that act as noise-enhancer molecules and potentiate current LRAs, dramatically increasing their potency (Dar et al. PNAS 2012; Dar et al. Science 2014). Recently, we discovered that HIV alternative splicing is post transcriptional, thereby generating a noise- attenuating circuit that regulates HIV’s latency decision (Hansen et al. Cell 2018). Perturbing the HIV latency circuit, for example with noise-enhancer or suppressor molecules, may represent a novel strategy for HIV cure, and functional cure, approaches. 110 SILENCING OF RETROVIRAL GENE EXPRESSION BY THE HUSH COMPLEX Stephen P. Goff , Columbia University Medical Center, New York, NY, USA Retroviral DNAs are transcriptionally silenced in a number of settings and in specific cell types, an important mechanism for the inhibition of virus replication. The HUSH complex (containing the three subunits TASOR/ FAM208A, MPP8, and periphilin) was originally identified as required for the maintenance of silencing of transfected DNAs integrated into heterochromatic regions of vertebrate genomes. HUSH binds to histone H3K9me3 marks, and brings the histone methyltransferase SETDB1/ESET to sustain and likely spread this H3K9me3 silencing modification along chromatin. HUSH was found to be important for the silencing of those retroviral DNAs that have integrated into heterochromatin. This position-dependent silencing by HUSH is distinct from the more universal silencing of retroviral DNAs in embryonic stem cells, mediated by zinc finger proteins tethering TRIM28/Kap1 to specific sequence elements of the viral DNA. We have recently found that the HUSH complex is also involved in the silencing of unintegrated viral DNAs of many retroviruses in virtually all cell types. HUSH is recruited to unintegrated DNA of the mouse leukemia viruses by NP220, a large DNA-binding protein with preference for oligo(C) sequences. This silencing, involving both SETDB1/ESET and histone deacetylases (HDACs), is relieved upon integration of the DNA into euchromatic regions of permissive cell types. NP220 and HUSH thereby act to inhibit early viral gene expression and the overall rate of virus replication. The Vpx and Vpr proteins of HIV-2 and various strains of the simian immunodeficiency viruses bind HUSH and induce its proteasomal degradation, stimulating virus 109 DISCOVERY AND CHARACTERIZATION OF A VIRUS-INTRINSIC HIV LATENCY CIRCUIT Leor Weinberger , Gladstone Institutes, San Francisco, CA, USA

replication. Vpx can also induce expression of HIV-1 proviruses in various models of latency, suggesting that HUSH may help maintain repression of silent proviruses in the latent reservoir. 111 THE POWER OF THE HIV PROMOTER: IMPACT OF HIV-DRIVEN VIRAL AND HOST GENE EXPRESSION Ya-Chi Ho , Yale University, New Haven, CT, USA Antiretroviral therapy (ART) inhibits the enzymatic function of viral proteins or blocks viral entry but does not inhibit HIV-1 LTR promoter activity. The HIV-1 LTR remains functional and active despite effective ART. Around 1–64 per million CD4 + T cells contain replication-competent HIV-1 proviruses. HIV-1 LTR drives the expression of intact HIV-1 proviruses, leading to viral rebound upon treatment interruptions. Around 100–1000 per million CD4 + T cells contain replication-defective HIV-1 proviruses. These replication-defective HIV-1 proviruses can be transcribed and translated, leading to antigen presentation to CD8 + T cells and immune distraction. Meanwhile, cells harboring intact and defective HIV-1 proviruses can both undergo clonal expansion, and the clonally expanded cells increase over time. HIV-1 integration site analysis revealed striking enrichment of HIV-1 integration at specific sites of proliferation-related genes in the clonally expanded cells, but the mechanisms remain unknown. The rarity and the lack of cellular markers of HIV-1-infected cells makes mechanistic studies challenging. Using single-cell RNAseq and high-resolution RNA landscape mapping on CD4 + T cells from virally suppressed, HIV-1-infected individuals, we show that HIV-1 LTR dominates over the host promoter and drives the host gene expression at the integration sites in genes associated with in vivo clonal expansion. HIV-1 activates cryptic host gene splice sites and induces aberrant host gene transcription, while HIV-1 viral genome transcription remains intact. Overall, HIV-1 LTR drives viral and host gene expression at the same time. Strategies targeting HIV-1 LTR should be explored to disrupt HIV-1-induced immune activation and HIV-1-driven aberrant proliferation gene expression. The Global Health Sector Strategy GHSS calls for the elimination of viral hepatitis as a public health threat by 2030 (reducing new infections by 90% and mortality by 65%). Indeed, with the advent of highly successful and well tolerated direct acting antiviral combinations, allowing HCV cure after short durations of treatment within 8-12 weeks in more than 95% of all treated patients, HCV elimination appears to be a reachable goal. Nevertheless, the WHO report from 2016 clearly describes some significant obstacles which need to be overcome in order to approach HCV elimination. The first major obstacle clearly is underdiagnoses with only 20% of people with HCV worldwide having been diagnosed so far. Equally disturbing is that while approximately 71 million people were thought to be infected with HCV in 2015, only 1.76 million people received HCV treatment in 2016 despite all praised advances in HCV therapy. Microelimination in well targeted patient groups with regular monitoring such as hemophiliacs, dialysis patients and also HIV/HCV coinfected patients therefore, appear low-hanging fruits on the pathway to global HCV elimination. First national studies from Netherlands and Switzerland suggest that indeed by increasing treatment uptake in all HIV/HCV coinfected men who have sex with men (MSM) the incidence of newly acquired acute HCV infections has been successfully reduced by over 50%. Nevertheless, increased HCV outbreaks among HIV negative MSM using PrEP as well as the high risk of HCV reinfection in MSM in general jeopardize these first encouraging reports. Clearly, earlier HCV treatment initiation and use of HCV-RNA and HCV-antigen testing rather than HCV-serology, allowing earlier acute HCV diagnosis, will be needed to impact HCV dynamics in the long-term. Under consideration of the still significantly increasing PrEP user number in Western Europe a call for action is needed to prevent a further spread of HCV into the MSM community. The World Health Organization set goals in 2016 for reductions in prevalent and incident hepatitis C infection necessary to achieve elimination of HCV as a public health problem by 2030. Modeling demonstrates that global HCV control will require annual rates of cure that are consistently and significantly higher than new HCV infection rates. However, a recent study showed that nearly 60% of surveyed countries had the opposite—more HCV infections 112 DYNAMICS OF ACUTE HCV IN WESTERN EUROPE Jürgen K. Rockstroh , University of Bonn, Bonn, Germany 113 A VACCINE TO PREVENT HCV: ARE WE GETTING THERE? Andrea L. Cox , Johns Hopkins University, Baltimore, MD, USA

Oral Abstracts

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CROI 2019