Changes to the Simian Immunodeficiency Virus (SIV) Reservoir and Enhanced SIV-Specific Responses in a Rhesus Macaque Model of Functional Cure after Serial Rounds of Romidepsin Administrations.

HIV persistence requires lifelong antiretroviral therapy (ART), calling for a cure. The histone deacetylase inhibitor, romidepsin, is used in the "shock and kill" approach with the goal of reactivating virus and subsequently clearing infected cells through cell-mediated immune responses. We tested serial and double infusions of romidepsin in a rhesus macaque (RM) model of SIV functional cure, which controls virus without ART. Off ART, romidepsin reactivated SIV in all RMs. Subsequent infusions resulted in diminished reactivation, and two RMs did not reactivate the virus after the second or third infusions. Therefore, those two RMs received CD8-depleting antibody to assess the replication competence of the residual reservoir. The remaining RMs received double infusions, i.e., two doses separated by 48-h. Double infusions were well tolerated, induced immune activation, and effectively reactivated SIV. Although reactivation was gradually diminished, cell-associated viral DNA was minimally changed, and viral outgrowth occurred in 4/5 RMs. In the RM which did not reactivate after CD8 depletion, viral outgrowth was not detected in peripheral blood mononuclear cells (PBMC)-derived CD4+ cells. The frequency of SIV-specific CD8+ T cells increased after romidepsin administration, and the increased SIV-specific immune responses were associated, although not statistically, with the diminished reactivation. Thus, our data showing sequential decreases in viral reactivation with repeated romidepsin administrations with all RMs and absence of viral reactivation after CD8+ T-cell depletion in one animal suggest that, in the context of healthy immune responses, romidepsin affected the inducible viral reservoir and gradually increased immune-mediated viral control. Given the disparities between the results of romidepsin administration to ART-suppressed SIVmac239-infected RMs and HIV-infected normal progressors compared to our immune-healthy model, our data suggest that improving immune function for greater SIV-specific responses should be the starting point of HIV cure strategies. IMPORTANCE HIV cure is sought after due to the prevalence of comorbidities that occur in persons with HIV. One of the most investigated HIV cure strategies is the "shock and kill" approach. Our study investigated the use of romidepsin, a histone deacetylase (HDAC) inhibitor, in our rhesus macaque model of functional cure, which allows for better resolution of viral reactivation due to the lack of antiretroviral therapy. We found that repeated rounds of romidepsin resulted in gradually diminished viral reactivation. One animal inevitably lacked replication-competent virus in the blood. With the accompanying enhancement of the SIV-specific immune response, our data suggest that there is a reduction of the viral reservoir in one animal by the cell-mediated immune response. With the differences observed between our model and persons living with HIV (PWH) treated with romidepsin, specifically in the context of a healthy immune system in our model, our data thereby indicate the importance of restoring the immune system for cure strategies.

African green monkeys avoid SIV disease progression by preventing intestinal dysfunction and maintaining mucosal barrier integrity.

Unlike HIV infection, SIV infection is generally nonpathogenic in natural hosts, such as African green monkeys (AGMs), despite life-long high viral replication. Lack of disease progression was reportedly based on the ability of SIV-infected AGMs to prevent gut dysfunction, avoiding microbial translocation and the associated systemic immune activation and chronic inflammation. Yet, the maintenance of gut integrity has never been documented, and the mechanism(s) by which gut integrity is preserved are unknown. We sought to investigate the early events of SIV infection in AGMs, specifically examining the impact of SIVsab infection on the gut mucosa. Twenty-nine adult male AGMs were intrarectally infected with SIVsab92018 and serially sacrificed at well-defined stages of SIV infection, preramp-up (1-3 days post-infection (dpi)), ramp-up (4-6 dpi), peak viremia (9-12 dpi), and early chronic SIV infection (46-55 dpi), to assess the levels of immune activation, apoptosis, epithelial damage and microbial translocation in the GI tract and peripheral lymph nodes. Tissue viral loads, plasma cytokines and plasma markers of gut dysfunction were also measured throughout the course of early infection. While a strong, but transient, interferon-based inflammatory response was observed, the levels of plasma markers linked to enteropathy did not increase. Accordingly, no significant increases in apoptosis of either mucosal enterocytes or lymphocytes, and no damage to the mucosal epithelium were documented during early SIVsab infection of AGMs. These findings were supported by RNAseq of the gut tissue, which found no significant alterations in gene expression that would indicate microbial translocation. Thus, for the first time, we confirmed that gut epithelial integrity is preserved, with no evidence of microbial translocation, in AGMs throughout early SIVsab infection. This might protect AGMs from developing intestinal dysfunction and the subsequent chronic inflammation that drives both HIV disease progression and HIV-associated comorbidities.

Nonhuman Primate Testing of the Impact of Different Regulatory T Cell Depletion Strategies on Reactivation and Clearance of Latent Simian Immunodeficiency Virus.

Regulatory T cells (Tregs) may be key contributors to the HIV/SIV latent reservoir, since they harbor high levels of HIV/SIV; reverse CD4+ T cell immune activation status, increasing the pool of resting CD4+ T cells; and impair CD8+ T cell function, favoring HIV persistence. We tested the hypothesis that Treg depletion is a valid intervention toward an HIV cure by depleted Tregs in 14 rhesus macaque (RM) controllers infected with SIVsab, the virus that naturally infects sabaeus monkeys, through different strategies: administration of an anti-CCR4 immunotoxin, two doses of an anti-CD25 immunotoxin (interleukin-2 with diphtheria toxin [IL-2-DT]), or two combinations of both. All of these treatments resulted in significant depletion of the circulating Tregs (>70%) and their partial depletion in the gut (25%) and lymph nodes (>50%). The fractions of CD4+ T cells expressing Ki -67 increased up to 80% in experiments containing IL-2-DT and only 30% in anti-CCR4-treated RMs, paralleled by increases in the inflammatory cytokines. In the absence of ART, plasma virus rebounded to 103 vRNA copies/ml by day 10 after IL-2-DT administration. A large but transient boost of the SIV-specific CD8+ T cell responses occurred in IL-2-DT-treated RMs. Such increases were minimal in the RMs receiving anti-CCR4-based regimens. Five RMs received IL-2-DT on ART, but treatment was discontinued because of high toxicity and lymphopenia. As such, while all treatments depleted a significant proportion of Tregs, the side effects in the presence of ART prevent their clinical use and call for different Treg depletion approaches. Thus, based on our data, Treg targeting as a strategy for HIV cure cannot be discarded.IMPORTANCE Regulatory T cells (Tregs) can decisively contribute to the establishment and persistence of the HIV reservoir, since they harbor high levels of HIV/SIV, increase the pool of resting CD4+ T cells by reversing their immune activation status, and impair CD8+ T cell function, favoring HIV persistence. We tested multiple Treg depletion strategies and showed that all of them are at least partially successful in depleting Tregs. As such, Treg depletion appears to be a valid intervention toward an HIV cure, reducing the size of the reservoir, reactivating the virus, and boosting cell-mediated immune responses. Yet, when Treg depletion was attempted in ART-suppressed animals, the treatment had to be discontinued due to high toxicity and lymphopenia. Therefore, while Treg targeting as a strategy for HIV cure cannot be discarded, the methodology for Treg depletion has to be revisited.

Dynamics of Simian Immunodeficiency Virus Two-Long-Terminal-Repeat Circles in the Presence and Absence of CD8+ Cells.

CD8+ cells play a key role in human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) infection, but their specific mechanism(s) of action in controlling the virus is unclear. Two-long-terminal-repeat (2-LTR) circles are extrachromosomal products generated upon failed integration of HIV/SIV. To understand the specific effects of CD8+ cells on infected cells, we analyzed the dynamics of 2-LTR circles in SIVmac251-infected rhesus macaques (RMs) treated with an integrase inhibitor (INT). Twenty RMs underwent CD8+ cell depletion and received raltegravir (RAL) monotherapy or a combination of both. Blood, lymph nodes (LNs), and gut biopsy specimens were routinely sampled. Plasma viral loads (pVLs) and 2-LTR circles from peripheral blood mononuclear cells (PBMCs) and LN lymphocytes were measured with quantitative reverse transcription-PCR (qRT-PCR). In the CD8 depletion group, an ∼1-log increase in pVLs and a slow increase in PBMC 2-LTRs occurred following depletion. In the INT group, a strong decline in pVLs upon treatment initiation and no change in 2-LTR levels were observed. In the INT and CD8+ cell depletion group, an increase in pVLs following CD8 depletion similar to that in the CD8 depletion group was observed, with a modest decline following INT initiation, and 2-LTR circles significantly increased in PBMCs and LNs. Analyzing the 2-LTR data across all treatment groups with a mathematical model indicates that the data best support an effect of CD8+ cells in killing cells prior to viral integration. Sensitivity analyses of these results confirm that effect but also allow for additional effects, which the data do not discriminate well. Overall, we show that INT does not significantly increase the levels of 2-LTR circles. However, CD8+ cell depletion increases the 2-LTR levels, which are enhanced in the presence of an INT.IMPORTANCE CD8+ T cells play an essential role in controlling HIV and SIV infection, but the specific mechanisms involved remain poorly understood. Due to failed viral infection, HIV and SIV can form 2-LTR extrachromosomal circles that can be quantified. We present novel data on the dynamics of these 2-LTR forms in a SIV-infected macaque model under three different treatment conditions: depletion of CD8+ cells, administration of the integrase inhibitor in a monotherapy, which favors the formation of 2-LTR circles, and a combination of the two treatments. We used a new mathematical model to help interpret the data, and the results suggest that CD8+ cells exert a killing effect on infected cells prior to virus integration. These results provide new insights into the mechanisms of action of CD8+ cells in SIV infection. Confirmation of our results would be an important step in understanding immune control of HIV.

Marginal Effects of Systemic CCR5 Blockade with Maraviroc on Oral Simian Immunodeficiency Virus Transmission to Infant Macaques.

Current approaches do not eliminate all human immunodeficiency virus type 1 (HIV-1) maternal-to-infant transmissions (MTIT); new prevention paradigms might help avert new infections. We administered maraviroc (MVC) to rhesus macaques (RMs) to block CCR5-mediated entry, followed by repeated oral exposure of a CCR5-dependent clone of simian immunodeficiency virus (SIV) mac251 (SIVmac766). MVC significantly blocked the CCR5 coreceptor in peripheral blood mononuclear cells and tissue cells. All control animals and 60% of MVC-treated infant RMs became infected by the 6th challenge, with no significant difference between the number of exposures (P = 0.15). At the time of viral exposures, MVC plasma and tissue (including tonsil) concentrations were within the range seen in humans receiving MVC as a therapeutic. Both treated and control RMs were infected with only a single transmitted/founder variant, consistent with the dose of virus typical of HIV-1 infection. The uninfected RMs expressed the lowest levels of CCR5 on the CD4+ T cells. Ramp-up viremia was significantly delayed (P = 0.05) in the MVC-treated RMs, yet peak and postpeak viral loads were similar in treated and control RMs. In conclusion, in spite of apparent effective CCR5 blockade in infant RMs, MVC had a marginal impact on acquisition and only a minimal impact on the postinfection delay of viremia following oral SIV infection. Newly developed, more effective CCR5 blockers may have a more dramatic impact on oral SIV transmission than MVC.IMPORTANCE We have previously suggested that the very low levels of simian immunodeficiency virus (SIV) maternal-to-infant transmissions (MTIT) in African nonhuman primates that are natural hosts of SIVs are due to a low availability of target cells (CCR5+ CD4+ T cells) in the oral mucosa of the infants, rather than maternal and milk factors. To confirm this new MTIT paradigm, we performed a proof-of-concept study in which we therapeutically blocked CCR5 with maraviroc (MVC) and orally exposed MVC-treated and naive infant rhesus macaques to SIV. MVC had only a marginal effect on oral SIV transmission. However, the observation that the infant RMs that remained uninfected at the completion of the study, after 6 repeated viral challenges, had the lowest CCR5 expression on the CD4+ T cells prior to the MVC treatment appears to confirm our hypothesis, also suggesting that the partial effect of MVC is due to a limited efficacy of the drug. New, more effective CCR5 inhibitors may have a better effect in preventing SIV and HIV transmission.

Antibiotic and Antiinflammatory Therapy Transiently Reduces Inflammation and Hypercoagulation in Acutely SIV-Infected Pigtailed Macaques.

Increased chronic immune activation and inflammation are hallmarks of HIV/SIV infection and are highly correlated with progression to AIDS and development of non-AIDS comorbidities, such as hypercoagulability and cardiovascular disease. Intestinal dysfunction resulting in microbial translocation has been proposed as a lead cause of systemic immune activation and hypercoagulability in HIV/SIV infection. Our goal was to assess the biological and clinical impact of a therapeutic strategy designed to reduce microbial translocation through reduction of the microbial content of the intestine (Rifaximin-RFX) and of gut inflammation (Sulfasalazine-SFZ). RFX is an intraluminal antibiotic that was successfully used in patients with hepatic encephalopathy. SFZ is an antiinflammatory drug successfully used in patients with mild to moderate inflammatory bowel disease. Both these clinical conditions are associated with increased microbial translocation, similar to HIV-infected patients. Treatment was administered for 90 days to five acutely SIV-infected pigtailed macaques (PTMs) starting at the time of infection; seven untreated SIVsab-infected PTMs were used as controls. RFX+SFZ were also administered for 90 days to three chronically SIVsab-infected PTMs. RFX+SFZ administration during acute SIVsab infection of PTMs resulted in: significantly lower microbial translocation, lower systemic immune activation, lower viral replication, better preservation of mucosal CD4+ T cells and significantly lower levels of hypercoagulation biomarkers. This effect was clear during the first 40 days of treatment and was lost during the last stages of treatment. Administration of RFX+SFZ to chronically SIVsab-infected PTMs had no discernible effect on infection. Our data thus indicate that early RFX+SFZ administration transiently improves the natural history of acute and postacute SIV infection, but has no effect during chronic infection.

Multi-dose Romidepsin Reactivates Replication Competent SIV in Post-antiretroviral Rhesus Macaque Controllers.

Viruses that persist despite seemingly effective antiretroviral treatment (ART) and can reinitiate infection if treatment is stopped preclude definitive treatment of HIV-1 infected individuals, requiring lifelong ART. Among strategies proposed for targeting these viral reservoirs, the premise of the "shock and kill" strategy is to induce expression of latent proviruses [for example with histone deacetylase inhibitors (HDACis)] resulting in elimination of the affected cells through viral cytolysis or immune clearance mechanisms. Yet, ex vivo studies reported that HDACis have variable efficacy for reactivating latent proviruses, and hinder immune functions. We developed a nonhuman primate model of post-treatment control of SIV through early and prolonged administration of ART and performed in vivo reactivation experiments in controller RMs, evaluating the ability of the HDACi romidepsin (RMD) to reactivate SIV and the impact of RMD treatment on SIV-specific T cell responses. Ten RMs were IV-infected with a SIVsmmFTq transmitted-founder infectious molecular clone. Four RMs received conventional ART for >9 months, starting from 65 days post-infection. SIVsmmFTq plasma viremia was robustly controlled to <10 SIV RNA copies/mL with ART, without viral blips. At ART cessation, initial rebound viremia to ~106 copies/mL was followed by a decline to < 10 copies/mL, suggesting effective immune control. Three post-treatment controller RMs received three doses of RMD every 35-50 days, followed by in vivo experimental depletion of CD8+ cells using monoclonal antibody M-T807R1. RMD was well-tolerated and resulted in a rapid and massive surge in T cell activation, as well as significant virus rebounds (~104 copies/ml) peaking at 5-12 days post-treatment. CD8+ cell depletion resulted in a more robust viral rebound (107 copies/ml) that was controlled upon CD8+ T cell recovery. Our results show that RMD can reactivate SIV in vivo in the setting of post-ART viral control. Comparison of the patterns of virus rebound after RMD administration and CD8+ cell depletion suggested that RMD impact on T cells is only transient and does not irreversibly alter the ability of SIV-specific T cells to control the reactivated virus.

Cutting Edge: T Regulatory Cell Depletion Reactivates Latent Simian Immunodeficiency Virus (SIV) in Controller Macaques While Boosting SIV-Specific T Lymphocytes.

T regulatory cells (Tregs) are critical in shaping the latent HIV/SIV reservoir, as they are preferentially infected, reverse CD4+ T cell activation status, and suppress CTL responses. To reactivate latent virus and boost cell-mediated immune responses, we performed in vivo Treg depletion with Ontak (denileukin diftitox) in two SIVsab-infected controller macaques. Ontak induced significant (>75%) Treg depletion and major CD4+ T cell activation, and only minimally depleted CD8+ T cells. The overall ability of Tregs to control immune responses was significantly impaired despite their incomplete depletion, resulting in both reactivation of latent virus (virus rebound to 103 viral RNA copies/ml plasma in the absence of antiretroviral therapy) and a significant boost of SIV-specific CD8+ T cell frequency, with rapid clearance of reactivated virus. As none of the latency-reversing agents in development have such dual activity, our strategy holds great promise for cure research.

Population Bottlenecks and Pathogen Extinction: "Make This Everyone's Mission to Mars, Including Yours".

Kapusinszky et al. (J Virol 89:8152-8161, 2015, http://dx.doi.org/10.1128/JVI.00671-15) report that host population bottlenecks may result in pathogen extinction, which provides a compelling argument for an alternative approach to vaccination for the control of virus spread. By comparing the prevalence levels of three viral pathogens in two populations of African green monkeys (AGMs) (Chlorocebus sabaeus) from Africa and two Caribbean Islands, they convincingly show that a major host bottleneck resulted in the eradication of select pathogens from a given host.

Critical Role for the Adenosine Pathway in Controlling Simian Immunodeficiency Virus-Related Immune Activation and Inflammation in Gut Mucosal Tissues.

UNLABELLED: The role of the adenosine (ADO) pathway in human immunodeficiency virus type 1/simian immunodeficiency virus (HIV-1/SIV) infection remains unclear. We compared SIVsab-induced changes of markers related to ADO production (CD39 and CD73) and breakdown (CD26 and adenosine deaminase) on T cells from blood, lymph nodes, and intestine collected from pigtailed macaques (PTMs) and African green monkeys (AGMs) that experience different SIVsab infection outcomes. We also measured ADO and inosine (INO) levels in tissues by mass spectrometry. Finally, we assessed the suppressive effect of ADO on proinflammatory cytokine production after T cell receptor stimulation. The baseline level of both CD39 and CD73 coexpression on regulatory T cells and ADO levels were higher in AGMs than in PTMs. Conversely, high INO levels associated with dramatic increases in CD26 expression and adenosine deaminase activity were observed in PTMs during chronic SIV infection. Immune activation and inflammation markers in the gut and periphery inversely correlated with ADO and directly correlated with INO. Ex vivo administration of ADO significantly suppressed proinflammatory cytokine production by T cells in both species. In conclusion, the opposite dynamics of ADO pathway-related markers and contrasting ADO/INO levels in species with divergent proinflammatory responses to SIV infection support a key role of ADO in controlling immune activation/inflammation in nonprogressive SIV infections. Changes in ADO levels predominately occurred in the gut, suggesting that the ADO pathway may be involved in sparing natural hosts of SIVs from developing SIV-related gut dysfunction. Focusing studies of the ADO pathway on mucosal sites of viral replication is warranted. IMPORTANCE: The mechanisms responsible for the severe gut dysfunction characteristic of progressive HIV and SIV infection in humans and macaques are not completely elucidated. We report that ADO may play a key role in controlling immune activation/inflammation in nonprogressive SIV infections by limiting SIV-related gut inflammation. Conversely, in progressive SIV infection, significant degradation of ADO occurs, possibly due to an early increase of ADO deaminase complexing protein 2 (CD26) and adenosine deaminase. Our study supports therapeutic interventions to offset alterations of this pathway during progressive HIV/SIV infections. These potential approaches to control chronic immune activation and inflammation during pathogenic SIV infection may prevent HIV disease progression.

Kinetics of myeloid dendritic cell trafficking and activation: impact on progressive, nonprogressive and controlled SIV infections.

We assessed the role of myeloid dendritic cells (mDCs) in the outcome of SIV infection by comparing and contrasting their frequency, mobilization, phenotype, cytokine production and apoptosis in pathogenic (pigtailed macaques, PTMs), nonpathogenic (African green monkeys, AGMs) and controlled (rhesus macaques, RMs) SIVagmSab infection. Through the identification of recently replicating cells, we demonstrated that mDC mobilization from the bone marrow occurred in all species postinfection, being most prominent in RMs. Circulating mDCs were depleted with disease progression in PTMs, recovered to baseline values after the viral peak in AGMs, and significantly increased at the time of virus control in RMs. Rapid disease progression in PTMs was associated with low baseline levels and incomplete recovery of circulating mDCs during chronic infection. mDC recruitment to the intestine occurred in all pathogenic scenarios, but loss of mucosal mDCs was associated only with progressive infection. Sustained mDC immune activation occurred throughout infection in PTMs and was associated with increased bystander apoptosis in blood and intestine. Conversely, mDC activation occurred only during acute infection in nonprogressive and controlled infections. Postinfection, circulating mDCs rapidly became unresponsive to TLR7/8 stimulation in all species. Yet, stimulation with LPS, a bacterial product translocated in circulation only in SIV-infected PTMs, induced mDC hyperactivation, apoptosis and excessive production of proinflammatory cytokines. After infection, spontaneous production of proinflammatory cytokines by mucosal mDCs increased only in progressor PTMs. We thus propose that mDCs promote tolerance to SIV in the biological systems that lack intestinal dysfunction. In progressive infections, mDC loss and excessive activation of residual mDCs by SIV and additional stimuli, such as translocated microbial products, enhance generalized immune activation and inflammation. Our results thus provide a mechanistic basis for the role of mDCs in the pathogenesis of AIDS and elucidate the causes of mDC loss during progressive HIV/SIV infections.

CD8+ T cells control SIV infection using both cytolytic effects and non-cytolytic suppression of virus production.

Whether CD8+ T lymphocytes control human immunodeficiency virus infection by cytopathic or non-cytopathic mechanisms is not fully understood. Multiple studies highlighted non-cytopathic effects, but one hypothesis is that cytopathic effects of CD8+ T cells occur before viral production. Here, to examine the role of CD8+ T cells prior to virus production, we treated SIVmac251-infected macaques with an integrase inhibitor combined with a CD8-depleting antibody, or with either reagent alone. We analyzed the ensuing viral dynamics using a mathematical model that included infected cells pre- and post- viral DNA integration to compare different immune effector mechanisms. Macaques receiving the integrase inhibitor alone experienced greater viral load decays, reaching lower nadirs on treatment, than those treated also with the CD8-depleting antibody. Models including CD8+ cell-mediated reduction of viral production (non-cytolytic) were found to best explain the viral profiles across all macaques, in addition an effect in killing infected cells pre-integration (cytolytic) was supported in some of the best models. Our results suggest that CD8+ T cells have both a cytolytic effect on infected cells before viral integration, and a direct, non-cytolytic effect by suppressing viral production.

High-fat diet exacerbates SIV pathogenesis and accelerates disease progression.

Consuming a high-fat diet (HFD) is a risk factor for obesity and diabetes; both of these diseases are also associated with systemic inflammation, similar to HIV infection. A HFD induces intestinal dysbiosis and impairs liver function and coagulation, with a potential negative impact on HIV/SIV pathogenesis. We administered a HFD rich in saturated fats and cholesterol to nonpathogenic (African green monkeys) and pathogenic (pigtailed macaques) SIV hosts. The HFD had a negative impact on SIV disease progression in both species. Thus, increased cell-associated SIV DNA and RNA occurred in the HFD-receiving nonhuman primates, indicating a potential reservoir expansion. The HFD induced prominent immune cell infiltration in the adipose tissue, an important SIV reservoir, and heightened systemic immune activation and inflammation, altering the intestinal immune environment and triggering gut damage and microbial translocation. Furthermore, HFD altered lipid metabolism and HDL oxidation and also induced liver steatosis and fibrosis. These metabolic disturbances triggered incipient atherosclerosis and heightened cardiovascular risk in the SIV-infected HFD-receiving nonhuman primates. Our study demonstrates that dietary intake has a discernable impact on the natural history of HIV/SIV infections and suggests that dietary changes can be used as adjuvant approaches for HIV-infected subjects, to reduce inflammation and the risk of non-AIDS comorbidities and possibly other infectious diseases.

Inflammatory monocytes expressing tissue factor drive SIV and HIV coagulopathy.

In HIV infection, persistent inflammation despite effective antiretroviral therapy is linked to increased risk of noninfectious chronic complications such as cardiovascular and thromboembolic disease. A better understanding of inflammatory and coagulation pathways in HIV infection is needed to optimize clinical care. Markers of monocyte activation and coagulation independently predict morbidity and mortality associated with non-AIDS events. We identified a specific subset of monocytes that express tissue factor (TF), persist after virological suppression, and trigger the coagulation cascade by activating factor X. This subset of monocytes expressing TF had a distinct gene signature with up-regulated innate immune markers and evidence of robust production of multiple proinflammatory cytokines, including interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), and IL-6, ex vivo and in vitro upon lipopolysaccharide stimulation. We validated our findings in a nonhuman primate model, showing that TF-expressing inflammatory monocytes were associated with simian immunodeficiency virus (SIV)-related coagulopathy in the progressive [pigtail macaques (PTMs)] but not in the nonpathogenic (African green monkeys) SIV infection model. Last, Ixolaris, an anticoagulant that inhibits the TF pathway, was tested and potently blocked functional TF activity in vitro in HIV and SIV infection without affecting monocyte responses to Toll-like receptor stimulation. Strikingly, in vivo treatment of SIV-infected PTMs with Ixolaris was associated with significant decreases in D-dimer and immune activation. These data suggest that TF-expressing monocytes are at the epicenter of inflammation and coagulation in chronic HIV and SIV infection and may represent a potential therapeutic target.

Animal Models for HIV Cure Research.

The HIV-1/AIDS pandemic continues to spread unabated worldwide, and no vaccine exists within our grasp. Effective antiretroviral therapy (ART) has been developed, but ART cannot clear the virus from the infected patient. A cure for HIV-1 is badly needed to stop both the spread of the virus in human populations and disease progression in infected individuals. A safe and effective cure strategy for human immunodeficiency virus (HIV) infection will require multiple tools, and appropriate animal models are tools that are central to cure research. An ideal animal model should recapitulate the essential aspects of HIV pathogenesis and associated immune responses, while permitting invasive studies, thus allowing a thorough evaluation of strategies aimed at reducing the size of the reservoir (functional cure) or eliminating the reservoir altogether (sterilizing cure). Since there is no perfect animal model for cure research, multiple models have been tailored and tested to address specific quintessential questions of virus persistence and eradication. The development of new non-human primate and mouse models, along with a certain interest in the feline model, has the potential to fuel cure research. In this review, we highlight the major animal models currently utilized for cure research and the contributions of each model to this goal.