ABSTRACT
An Amendment to this paper has been published and can be accessed via a link at the top of the paper.
ABSTRACT
Human immunodeficiency virus (HIV) persists indefinitely in individuals with HIV who receive antiretroviral therapy (ART) owing to a reservoir of latently infected cells that contain replication-competent virus1-4. Here, to better understand the mechanisms responsible for latency persistence and reversal, we used the interleukin-15 superagonist N-803 in conjunction with the depletion of CD8+ lymphocytes in ART-treated macaques infected with simian immunodeficiency virus (SIV). Although N-803 alone did not reactivate virus production, its administration after the depletion of CD8+ lymphocytes in conjunction with ART treatment induced robust and persistent reactivation of the virus in vivo. We found viraemia of more than 60 copies per ml in all macaques (n = 14; 100%) and in 41 out of a total of 56 samples (73.2%) that were collected each week after N-803 administration. Notably, concordant results were obtained in ART-treated HIV-infected humanized mice. In addition, we observed that co-culture with CD8+ T cells blocked the in vitro latency-reversing effect of N-803 on primary human CD4+ T cells that were latently infected with HIV. These results advance our understanding of the mechanisms responsible for latency reversal and lentivirus reactivation during ART-suppressed infection.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Interleukin-15/agonists , Simian Immunodeficiency Virus/physiology , Virus Replication , Animals , CD4-Positive T-Lymphocytes/virology , HIV Infections/immunology , HIV Infections/virology , Humans , Interleukin-15/immunology , Lymphocyte Depletion , Macaca mulatta , Mice , Simian Acquired Immunodeficiency Syndrome/immunology , Simian Acquired Immunodeficiency Syndrome/virology , Virus Latency , Virus Replication/immunologyABSTRACT
BACKGROUND: Natural killer (NK) cells are dysfunctional in chronic human immunodeficiency virus (HIV) infection as they are not able to clear virus. We hypothesized that an infusion of NK cells, supported by interleukin 2 (IL-2) or IL-15, could decrease virus-producing cells in the lymphatic tissues. METHODS: We conducted a phase 1 pilot study in 6 persons with HIV (PWH), where a single infusion of haploidentical related donor NK cells was given plus either IL-2 or N-803 (an IL-15 superagonist). RESULTS: The approach was well tolerated with no unexpected adverse events. We did not pretreat recipients with cyclophosphamide or fludarabine to "make immunologic space," reasoning that PWH on stable antiretroviral treatment remain T-cell depleted in lymphatic tissues. We found donor cells remained detectable in blood for up to 8 days (similar to what is seen in cancer pretreatment with lymphodepleting chemotherapy) and in the lymph nodes and rectum up to 28 days. There was a moderate decrease in the frequency of viral RNA-positive cells in lymph nodes. CONCLUSIONS: There was a moderate decrease in HIV-producing cells in lymph nodes. Further studies are warranted to determine the impact of healthy NK cells on HIV reservoirs and if restoring NK-cell function could be part of an HIV cure strategy. Clinical Trials Registration. NCT03346499 and NCT03899480.
Subject(s)
HIV Infections , Interleukin-15 , Interleukin-2 , Killer Cells, Natural , Humans , Killer Cells, Natural/immunology , HIV Infections/immunology , HIV Infections/virology , HIV Infections/drug therapy , Male , Middle Aged , Adult , Pilot Projects , Female , Viral Load , Lymph Nodes/immunology , HIV-1/immunologyABSTRACT
The IL-15 superagonist N-803 has been shown to enhance the function of CD8 T cells and NK cells. We previously found that in a subset of vaccinated, ART-naive, SIV+ rhesus macaques, N-803 treatment led to a rapid but transient decline in plasma viremia that positively correlated with an increase in the frequency of CD8 T cells. Here, we tested the hypothesis that prophylactic vaccination was required for the N-803 mediated suppression of SIV plasma viremia. We either vaccinated rhesus macaques with a DNA prime/Ad5 boost regimen using vectors expressing SIVmac239 gag with or without a plasmid expressing IL-12 or left them unvaccinated. The animals were then intravenously infected with SIVmac239M. 6 months after infection, the animals were treated with N-803. We found no differences in the control of plasma viremia during N-803 treatment between vaccinated and unvaccinated macaques. Interestingly, when we divided the SIV+ animals based on their plasma viral load set-points prior to the N-803 treatment, N-803 increased the frequency of SIV-specific T cells expressing ki-67+ and granzyme B+ in animals with low plasma viremia (<104 copies/mL; SIV controllers) compared to animals with high plasma viremia (>104 copies/mL; SIV noncontrollers). In addition, Gag-specific CD8 T cells from the SIV+ controllers had a greater increase in CD8+CD107a+ T cells in ex vivo functional assays than did the SIV+ noncontrollers. Overall, our results indicate that N-803 is most effective in SIV+ animals with a preexisting immunological ability to control SIV replication. IMPORTANCE N-803 is a drug that boosts the immune cells involved in combating HIV/SIV infection. Here, we found that in SIV+ rhesus macaques that were not on antiretroviral therapy, N-803 increased the proliferation and potential capacity for killing of the SIV-specific immune cells to a greater degree in animals that spontaneously controlled SIV than in animals that did not control SIV. Understanding the mechanism of how N-803 might function differently in individuals that control HIV/SIV (for example, individuals on antiretroviral therapy or spontaneous controllers) compared to settings where HIV/SIV are not controlled, could impact the efficacy of N-803 utilization in the field of HIV cure.
Subject(s)
HIV Infections , Simian Acquired Immunodeficiency Syndrome , Simian Immunodeficiency Virus , Animals , Macaca mulatta , Interleukin-15/genetics , Granzymes , Viremia , Ki-67 Antigen , CD8-Positive T-Lymphocytes , Anti-Retroviral Agents/therapeutic use , Viral Load , HIV Infections/drug therapy , Interleukin-12 , DNAABSTRACT
Vaccine strategies aimed at eliciting human immunodeficiency virus (HIV)-specific CD8+ T cells are one major target of interest in HIV functional cure strategies. We hypothesized that CD8+ T cells elicited by therapeutic vaccination during antiretroviral therapy (ART) would be recalled and boosted by treatment with the interleukin 15 (IL-15) superagonist N-803 after ART discontinuation. We intravenously immunized four simian immunodeficiency virus-positive (SIV+) Mauritian cynomolgus macaques receiving ART with vesicular stomatitis virus (VSV), modified vaccinia virus Ankara strain (MVA), and recombinant adenovirus serotype 5 (rAd-5) vectors all expressing SIVmac239 Gag. Immediately after ART cessation, these animals received three doses of N-803. Four control animals received no vaccines or N-803. The vaccine regimen generated a high-magnitude response involving Gag-specific CD8+ T cells that were proliferative and biased toward an effector memory phenotype. We then compared cells elicited by vaccination (Gag specific) to cells elicited by SIV infection and unaffected by vaccination (Nef specific). We found that N-803 treatment enhanced the frequencies of both bulk and proliferating antigen-specific CD8+ T cells elicited by vaccination and the antigen-specific CD8+ T cells elicited by SIV infection. In sum, we demonstrate that a therapeutic heterologous prime-boost-boost (HPBB) vaccine can elicit antigen-specific effector memory CD8+ T cells that are boosted by N-803. IMPORTANCE While antiretroviral therapy (ART) can suppress HIV replication, it is not a cure. It is therefore essential to develop therapeutic strategies to enhance the immune system to better become activated and recognize virus-infected cells. Here, we evaluated a novel therapeutic vaccination strategy delivered to SIV+ Mauritian cynomolgus macaques receiving ART. ART was then discontinued and we delivered an immunotherapeutic agent (N-803) after ART withdrawal with the goal of eliciting and boosting anti-SIV cellular immunity. Immunologic and virologic analysis of peripheral blood and lymph nodes collected from these animals revealed transient boosts in the frequency, activation, proliferation, and memory phenotype of CD4+ and CD8+ T cells following each intervention. Overall, these results are important in educating the field of the transient nature of the immunological responses to this particular therapeutic regimen and the similar effects of N-803 on boosting T cells elicited by vaccination or elicited naturally by infection.
Subject(s)
CD8-Positive T-Lymphocytes , SAIDS Vaccines , Simian Acquired Immunodeficiency Syndrome , Simian Immunodeficiency Virus , Animals , Cell Proliferation , Macaca mulatta/immunology , Simian Acquired Immunodeficiency Syndrome/immunology , Vaccination , Vaccinia virusABSTRACT
Mucosal associated invariant T (MAIT) cells are innate T cells that recognize bacterial metabolites and secrete cytokines and cytolytic enzymes to destroy infected target cells. This makes MAIT cells promising targets for immunotherapy to combat bacterial infections. Here, we analyzed the effects of an immunotherapeutic agent, the IL-15 superagonist N-803, on MAIT cell activation, trafficking, and cytolytic function in macaques. We found that N-803 could activate MAIT cells in vitro and increase their ability to produce IFN-γ in response to bacterial stimulation. To expand upon this, we examined the phenotypes and functions of MAIT cells present in samples collected from PBMC, airways (bronchoalveolar lavage [BAL] fluid), and lymph nodes (LN) from rhesus macaques that were treated in vivo with N-803. N-803 treatment led to a transient 6 to 7-fold decrease in the total number of MAIT cells in the peripheral blood, relative to pre N-803 time points. Concurrent with the decrease in cells in the peripheral blood, we observed a rapid decline in the frequency of CXCR3+CCR6+ MAITs. This corresponded with an increase in the frequency of CCR6+ MAITs in the BAL fluid, and higher frequencies of ki-67+ and granzyme B+ MAITs in the blood, LN, and BAL fluid. Finally, N-803 improved the ability of MAIT cells collected from PBMC and airways to produce IFN-γ in response to bacterial stimulation. Overall, N-803 shows the potential to transiently alter the phenotypes and functions of MAIT cells, which could be combined with other strategies to combat bacterial infections.
Subject(s)
Mucosal-Associated Invariant T Cells , Animals , Granzymes , Macaca mulatta , Leukocytes, Mononuclear , Ki-67 Antigen , Interferon-gammaABSTRACT
Despite the success of antiretroviral therapy (ART) to halt viral replication and slow disease progression, this treatment is not curative and there remains an urgent need to develop approaches to clear the latent HIV reservoir. The human IL-15 superagonist N-803 (formerly ALT-803) is a promising anti-cancer biologic with potent immunostimulatory properties that has been extended into the field of HIV as a potential "shock and kill" therapeutic for HIV cure. However, the ability of N-803 to reactivate latent virus and modulate anti-viral immunity in vivo under the cover of ART remains undefined. Here, we show that in ART-suppressed, simian-human immunodeficiency virus (SHIV)SF162P3-infected rhesus macaques, subcutaneous administration of N-803 activates and mobilizes both NK cells and SHIV-specific CD8+ T cells from the peripheral blood to lymph node B cell follicles, a sanctuary site for latent virus that normally excludes such effector cells. We observed minimal activation of memory CD4+ T cells and no increase in viral RNA content in lymph node resident CD4+ T cells post N-803 administration. Accordingly, we found no difference in the number or magnitude of plasma viremia timepoints between treated and untreated animals during the N-803 administration period, and no difference in the size of the viral DNA cell-associated reservoir post N-803 treatment. These results substantiate N-803 as a potent immunotherapeutic candidate capable of activating and directing effector CD8+ T and NK cells to the B cell follicle during full ART suppression, and suggest N-803 must be paired with a bona fide latency reversing agent in vivo to facilitate immune-mediated modulation of the latent viral reservoir.
Subject(s)
Anti-Retroviral Agents/administration & dosage , B-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/drug effects , HIV Infections/drug therapy , Interleukin-15/antagonists & inhibitors , Killer Cells, Natural/drug effects , Proteins/administration & dosage , Animals , B-Lymphocytes/cytology , B-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/immunology , Cell Movement/drug effects , Disease Models, Animal , HIV Infections/genetics , HIV Infections/immunology , HIV Infections/physiopathology , HIV-1/drug effects , HIV-1/physiology , Humans , Interleukin-15/genetics , Interleukin-15/immunology , Killer Cells, Natural/cytology , Killer Cells, Natural/immunology , Lymph Nodes/drug effects , Lymph Nodes/immunology , Macaca mulatta , Recombinant Fusion Proteins , Simian Acquired Immunodeficiency Syndrome/drug therapy , Simian Acquired Immunodeficiency Syndrome/immunology , Simian Acquired Immunodeficiency Syndrome/physiopathology , Simian Acquired Immunodeficiency Syndrome/virology , Simian Immunodeficiency Virus/drug effects , Simian Immunodeficiency Virus/physiology , Virus Latency/drug effectsABSTRACT
The "shock and kill" strategy predicates that virus reactivation in latently infected cells is required to eliminate the human immunodeficiency virus (HIV) reservoir. In a recent study, we showed robust and persistent induction of plasma viremia in antiretroviral therapy (ART)-treated simian immunodeficiency virus-infected rhesus macaques (RMs) undergoing CD8α depletion and treated with the interleukin-15 (IL-15) superagonist N-803 (J. B. McBrien et al., Nature 578:154-159, 2020, https://doi.org/10.1038/s41586-020-1946-0). Of note, in that study we used an antibody targeting CD8α, thereby depleting NK cells, NKT cells, and γδ T cells, in addition to CD8+ T cells. In the current proof-of-concept study, we tested whether virus reactivation can be induced by administration of N-803 to simian-human chimeric immunodeficiency virus-infected, ART-treated RMs that are selectively depleted of CD8+ T cells via the CD8ß-targeting antibody CD8b255R1. CD8ß depletion was performed in five SHIVSF162P3-infected RMs treated with ART for 12 months and with plasma viremia consistently below 3 copies/ml. All animals received four weekly doses of N-803 starting at the time of CD8b255R1 administration. The induction of detectable plasma viremia was observed in three out of five RMs, with the level of virus reactivation seemingly correlated with the frequency of CD8+ T cells following CD8ß depletion as well as the level of virus reactivation observed when the same animals underwent CD8α depletion and N-803 administration after 24 weeks of ART. These data indicate that CD8ß depletion and N-803 administration can induce virus reactivation in SHIVSF162P3-infected RMs despite suboptimal depletion of CD8+ T cells and profound ART-induced suppression of virus replication, confirming a critical role for these cells in suppressing virus production and/or reactivation in vivo under ART.IMPORTANCE The "shock and kill" HIV cure strategy attempts to reverse and eliminate the latent viral infection that prevents eradication of the virus. Latency-reversing agents tested in clinical trials to date have failed to affect the HIV viral reservoir. IL-15 superagonist N-803, currently involved in a clinical trial for HIV cure, was recently shown by our laboratory to induce robust and persistent induction of plasma viremia during ART in three in vivo animal models of HIV infection. These results suggest a substantial role for CD8+ lymphocytes in suppressing the latency reversal effect of N-803 by promoting the maintenance of viral latency. In this study, we tested whether the use of a CD8ß-targeting antibody, which would specifically deplete CD8+ T cells, would yield similar levels of virus reactivation. We observed the induction of plasma viremia, which correlated with the efficacy of the CD8 depletion strategy.
Subject(s)
Anti-Retroviral Agents/pharmacology , CD8 Antigens/immunology , HIV Infections/immunology , Interleukin-15/immunology , Simian Acquired Immunodeficiency Syndrome/immunology , Simian Immunodeficiency Virus/drug effects , Simian Immunodeficiency Virus/immunology , Animals , Anti-Retroviral Agents/therapeutic use , CD8-Positive T-Lymphocytes/immunology , Disease Models, Animal , HIV/drug effects , Killer Cells, Natural/drug effects , Lymphocyte Depletion , Macaca mulatta , Viral Load , Viremia/virology , Virus Latency/drug effects , Virus Replication/drug effectsABSTRACT
Clinical trials investigating histone deacetylase inhibitors (HDACi) to reverse HIV-1 latency aim to expose reservoirs in antiretroviral (ARV)-treated individuals to clearance by immune effectors, yet have not driven measurable reductions in the frequencies of infected cells. We therefore investigated the effects of the class I-selective HDACi nanatinostat and romidepsin on various blocks to latency reversal and elimination, including viral splicing, antigen presentation, and CD8+ T cell function. In ex vivo CD4+ T cells from ARV-suppressed individuals, both HDACi significantly induced viral transcription, but not splicing nor supernatant HIV-1 RNA. In an HIV-1 latency model using autologous CD8+ T cell clones as biosensors of antigen presentation, neither HDACi-treated CD4+ T cell condition induced clone degranulation. Both HDACi also impaired the function of primary CD8+ T cells in viral inhibition assays, with nanatinostat causing less impairment. These findings suggest that spliced or cell-free HIV-1 RNAs are more indicative of antigen expression than unspliced HIV-RNAs and may help to explain the limited abilities of HDACi to generate CD8+ T cell targets in vivoIMPORTANCE Antiretroviral (ARV) drug regimens suppress HIV-1 replication but are unable to cure infection. This leaves people living with HIV-1 burdened by a lifelong commitment to expensive daily medication. Furthermore, it has become clear that ARV therapy does not fully restore health, leaving individuals at elevated risk for cardiovascular disease, certain types of cancers, and neurocognitive disorders, as well as leaving them exposed to stigma. Efforts are therefore under way to develop therapies capable of curing infection. A key focus of these efforts has been on a class of drugs called histone deacetylase inhibitors (HDACi), which have the potential of exposing hidden reservoirs of HIV-1 to elimination by the immune system. Unfortunately, clinical trial results with HDACi have thus far been disappointing. In the current study, we integrate a number of experimental approaches to build a model that provides insights into the limited activity of HDACi in clinical trials and offers direction for future approaches.
Subject(s)
Histone Deacetylase Inhibitors/pharmacology , Virus Latency/drug effects , Adult , CD4-Positive T-Lymphocytes/drug effects , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/immunology , Depsipeptides/pharmacology , Female , HIV Infections/immunology , HIV Seropositivity/drug therapy , HIV-1/metabolism , HIV-1/pathogenicity , HIV-1/physiology , Histone Deacetylases/metabolism , Humans , Male , Middle Aged , Primary Cell Culture , Virus Latency/physiology , Virus Replication/drug effectsABSTRACT
BACKGROUND: Although mother-to-child human immunodeficiency virus (HIV) transmission has dramatically decreased with maternal antiretroviral therapy, breast milk transmission accounts for most of the 180 000 new infant HIV infections annually. Broadly neutralizing antibodies (bNAb) may further reduce transmission. METHODS: A Phase 1 safety and pharmacokinetic study was conducted: a single subcutaneous (SC) dose of 20 or 40 mg/kg (Dose Groups 1 and 2, respectively) of the bNAb VRC01 was administered to HIV-exposed infants soon after birth. Breastfeeding infants (Dose Group 3) received 40 mg/kg SC VRC01 after birth and then 20 mg/kg/dose SC monthly. All infants received appropriate antiretroviral prophylaxis. RESULTS: Forty infants were enrolled (21 in the United States, 19 in Africa). Subcutaneous VRC01 was safe and well tolerated with only mild-to-moderate local reactions, primarily erythema, which rapidly resolved. For multiple-dose infants, local reactions decreased with subsequent injections. VRC01 was rapidly absorbed after administration, with peak concentrations 1-6 days postdose. The 40 mg/kg dose resulted in 13 of 14 infants achieving the serum 50 micrograms (mcg)/mL target at day 28. Dose Group 3 infants maintained concentrations greater than 50 mcg/mL throughout breastfeeding. CONCLUSIONS: Subcutaneous VRC01 as single or multiple doses is safe and well tolerated in very young infants and is suitable for further study to prevent HIV transmission in infants.
Subject(s)
Antibodies, Monoclonal/administration & dosage , Broadly Neutralizing Antibodies/administration & dosage , HIV Antibodies/administration & dosage , HIV Infections/drug therapy , HIV-1/drug effects , Infectious Disease Transmission, Vertical/prevention & control , Africa , Antibodies, Monoclonal/adverse effects , Antibodies, Monoclonal/pharmacokinetics , Broadly Neutralizing Antibodies/adverse effects , Female , HIV Antibodies/adverse effects , HIV Infections/blood , Humans , Infant, Newborn , Injections, Subcutaneous , Linear Models , Male , United StatesABSTRACT
Efforts to cure human immunodeficiency virus (HIV) infection are obstructed by reservoirs of latently infected CD4+ T cells that can reestablish viremia. HIV-specific broadly neutralizing antibodies (bNAbs), defined by unusually wide neutralization breadths against globally diverse viruses, may contribute to the elimination of these reservoirs by binding to reactivated cells, thus targeting them for immune clearance. However, the relationship between neutralization of reservoir isolates and binding to corresponding infected primary CD4+ T cells has not been determined. Thus, the extent to which neutralization breadths and potencies can be used to infer the corresponding parameters of infected cell binding is currently unknown. We assessed the breadths and potencies of bNAbs against 36 viruses reactivated from peripheral blood CD4+ T cells from antiretroviral (ARV)-treated HIV-infected individuals by using paired neutralization and infected cell binding assays. Single-antibody breadths ranged from 0 to 64% for neutralization (80% inhibitory concentration [IC80] of ≤10 µg/ml) and from 0 to 89% for binding, with two-antibody combinations (results for antibody combinations are theoretical/predicted) reaching levels of 0 to 83% and 50 to 100%, respectively. Infected cell binding correlated with virus neutralization for 10 of 14 antibodies (e.g., for 3BNC117, r = 0.82 and P < 0.0001). Heterogeneity was observed, however, with a lack of significant correlation for 2G12, CAP256.VRC26.25, 2F5, and 4E10. Our results provide guidance on the selection of bNAbs for interventional cure studies, both by providing a direct assessment of intra- and interindividual variabilities in neutralization and infected cell binding in a novel cohort and by defining the relationships between these parameters for a panel of bNAbs.IMPORTANCE Although antiretroviral therapies have improved the lives of people who are living with HIV, they do not cure infection. Efforts are being directed towards harnessing the immune system to eliminate the virus that persists, potentially resulting in virus-free remission without medication. HIV-specific antibodies hold promise for such therapies owing to their ability to both prevent the infection of new cells (neutralization) and direct the killing of infected cells. We isolated 36 HIV strains from individuals whose virus was suppressed by medication and tested 14 different antibodies for neutralization of these viruses and for binding to cells infected with the same viruses (critical for engaging natural killer cells). For both neutralization and infected cell binding, we observed variation both between individuals and amongst different viruses within an individual. For most antibodies, neutralization activity correlated with infected cell binding. These data provide guidance on the selection of antibodies for clinical trials.
Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , HIV Antibodies/immunology , HIV Infections/immunology , HIV Infections/virology , HIV-1/immunology , Viremia/immunology , Adult , Antibody-Dependent Cell Cytotoxicity , HIV Envelope Protein gp120/immunology , Humans , Male , Middle Aged , Neutralization Tests , Viremia/virologyABSTRACT
Yegor Voronin and colleagues explore how monoclonal antibodies against HIV could provide a new opportunity to further reduce mother-to-child transmission of HIV and propose that new interventions should consider issues related to implementation, feasibility, and access. Please see later in the article for the Editors' Summary.
Subject(s)
Antibodies, Monoclonal/blood , HIV Antibodies/blood , HIV Infections/prevention & control , HIV Infections/transmission , HIV/immunology , Adult , Child , Female , HIV Infections/virology , Humans , Infant , Infant, Newborn , Male , MothersABSTRACT
HIV vaccine development has been hindered by significant challenges over four decades. Despite persistent efforts, all efficacy trials to date have yielded disappointing results. This has pushed the field back to the discovery phase and created uncertainty about the future involvement of large pharmaceutical companies. Currently, the HIV vaccine landscape is dominated by startup biotech firms, which face a complex array of obstacles. These include evolving HIV prevention methods, waning interest in vaccine research, and difficulties securing sustainable funding. This viewpoint explores the challenges faced by these biotech companies and the support mechanisms necessary for their continued involvement in HIV vaccine development. By leveraging insights from both pharmaceutical and biotech sectors, we propose a multi-faceted approach that includes enhanced communication, fostering innovation, and implementing strategic funding models.
Subject(s)
AIDS Vaccines , Biotechnology , HIV Infections , Vaccine Development , AIDS Vaccines/immunology , AIDS Vaccines/administration & dosage , Humans , HIV Infections/prevention & control , Drug Industry/economics , Drug Development/trends , Drug Development/economicsABSTRACT
The eradication of the viral reservoir represents the major obstacle to the development of a clinical cure for established HIV-1 infection. Here, we demonstrate that the administration of N-803 (brand name Anktiva) and broadly neutralizing antibodies (bNAbs) results in sustained viral control after discontinuation of antiretroviral therapy (ART) in simian-human AD8 (SHIV-AD8)-infected, ART-suppressed rhesus macaques. N-803+bNAbs treatment induced immune activation and transient viremia but only limited reductions in the SHIV reservoir. Upon ART discontinuation, viral rebound occurred in all animals, which was followed by durable control in approximately 70% of all N-803+bNAb-treated macaques. Viral control was correlated with the reprogramming of CD8+ T cells by N-803+bNAb synergy. Thus, complete eradication of the replication-competent viral reservoir is likely not a prerequisite for the induction of sustained remission after discontinuation of ART.
Subject(s)
Anti-Retroviral Agents , Recombinant Fusion Proteins , Simian Acquired Immunodeficiency Syndrome , Simian Immunodeficiency Virus , Animals , Humans , Anti-Retroviral Agents/therapeutic use , Anti-Retroviral Agents/pharmacology , Broadly Neutralizing Antibodies/administration & dosage , CD8-Positive T-Lymphocytes/virology , Immunotherapy , Macaca mulatta , Simian Acquired Immunodeficiency Syndrome/drug therapy , Simian Acquired Immunodeficiency Syndrome/therapy , Viral Load , Recombinant Fusion Proteins/administration & dosage , Recombinant Fusion Proteins/pharmacology , Remission Induction , Drug Therapy, CombinationABSTRACT
The main barrier to HIV cure is a persistent reservoir of latently infected CD4+ T cells harboring replication-competent provirus that fuels rebound viremia upon antiretroviral therapy (ART) interruption. A leading approach to target this reservoir involves agents that reactivate latent HIV proviruses followed by direct clearance of cells expressing induced viral antigens by immune effector cells and immunotherapeutics. We previously showed that AZD5582, an antagonist of inhibitor of apoptosis proteins and mimetic of the second mitochondrial-derived activator of caspases (IAPi/SMACm), induces systemic reversal of HIV/SIV latency but with no reduction in size of the viral reservoir. In this study, we investigated the effects of AZD5582 in combination with four SIV Env-specific Rhesus monoclonal antibodies (RhmAbs) ± N-803 (an IL-15 superagonist) in SIV-infected, ART-suppressed rhesus macaques. Here we confirm the efficacy of AZD5582 in inducing SIV reactivation, demonstrate enhancement of latency reversal when AZD5582 is used in combination with N-803 and show a reduction in total and replication-competent SIV-DNA in lymph-node-derived CD4+ T cells in macaques treated with AZD5582 + RhmAbs. Further exploration of this therapeutic approach may contribute to the goal of achieving an HIV cure.
Subject(s)
HIV Infections , HIV-1 , Simian Acquired Immunodeficiency Syndrome , Simian Immunodeficiency Virus , Animals , Simian Immunodeficiency Virus/physiology , Macaca mulatta , Anti-Retroviral Agents/pharmacology , Anti-Retroviral Agents/therapeutic use , Virus Latency , Virus Replication , Antibodies/therapeutic use , Lymph Nodes , CD4-Positive T-Lymphocytes , Viral LoadABSTRACT
We assessed if immune responses are enhanced in CD-1 mice by heterologous vaccination with two different nucleic acid-based COVID-19 vaccines: a next-generation human adenovirus serotype 5 (hAd5)-vectored dual-antigen spike (S) and nucleocapsid (N) vaccine (AdS+N) and a self-amplifying and -adjuvanted S RNA vaccine (AAHI-SC2) delivered by a nanostructured lipid carrier. The AdS+N vaccine encodes S modified with a fusion motif to increase cell-surface expression and an N antigen modified with an Enhanced T-cell Stimulation Domain (N-ETSD) to direct N to the endosomal/lysosomal compartment and increase MHC class I and II stimulation potential. The S sequence in the AAHI-SC2 vaccine comprises the D614G mutation, two prolines to stabilize S in the prefusion conformation, and 3 glutamines in the furin cleavage region to confer protease resistance. CD-1 mice received vaccination by homologous and heterologous prime > boost combinations. Humoral responses to S were the highest with any regimen that included the AAHI-SC2 vaccine, and IgG bound to wild type and Delta (B.1.617.2) variant S1 at similar levels. An AAHI-SC2 prime followed by an AdS+N boost particularly enhanced CD4+ and CD8+ T-cell responses to both wild type and Delta S peptides relative to all other vaccine regimens. Sera from mice receiving AAHI-SC2 homologous or heterologous vaccination were found to be highly neutralizing for all pseudovirus strains tested: Wuhan, Beta, Delta, and Omicron strains. The findings here, taken in consideration with the availability of both vaccines in thermostable formulations, support the testing of heterologous vaccination by an AAHI-SC2 > AdS+N regimen in animal models of SARS-CoV-2 infection to assess its potential to provide increased protection against emerging SARS-CoV-2 variants particularly in regions of the world where the need for cold-chain storage has limited the distribution of other vaccines.
Subject(s)
COVID-19 , Viral Vaccines , Animals , Antibodies, Neutralizing , Antigens, Heterophile , COVID-19/prevention & control , COVID-19 Vaccines , DNA , Humans , Mice , SARS-CoV-2 , Vaccination , Vaccines, Synthetic , mRNA VaccinesABSTRACT
There is no cure for HIV infection, and lifelong antiretroviral therapy (ART) is required. N-803 is an IL-15 superagonist comprised of an N72D mutant IL-15 molecule attached to its alpha receptor and a human IgG1 fragment designed to increase IL-15 activity. Preclinical studies with both HIV and SIV suggest that the drug has potential to reduce virus reservoirs by activating virus from latency and enhancing effector function. We conducted a phase 1 study of N-803 ( NCT02191098 ) in people living with HIV, the primary objective of which was to assess the safety and tolerability of the drug, with an exploratory objective of assessing the impact on peripheral virus reservoirs. ART-suppressed individuals were enrolled into a dose-escalation study of N-803 in four different cohorts (0.3, 1.0, 3.0 and 6.0 mcg kg-1). Each cohort received three doses total, separated by at least 1 week. We enrolled 16 individuals, of whom 11 completed all three doses. The maximum tolerated dose was 6.0 mcg kg-1. The primary clinical adverse events (AEs) reported were injection site rash and adenopathy, and four participants experienced a grade 1 or grade 2 QTc prolongation. No significant laboratory AEs attributable to N-803 were observed. In exploratory analyses, N-803 was associated with proliferation and/or activation of CD4+ and CD8+ T cells and natural killer cells that peaked at 4 d after dosing. IFN-γ, IP-10, MCP-1 and IL-15 increased during treatment. HIV transcription in memory CD4 T cells and intact proviral DNA initially increased after N-803 treatment; however, there was a small but significant decrease in the frequency of peripheral blood mononuclear cells with an inducible HIV provirus that persisted for up to 6 months after therapy. These data suggest that N-803 administration in ART-suppressed people living with HIV is safe and that larger clinical trials are needed to further investigate the effects of N-803 on HIV reservoirs.
Subject(s)
HIV Infections , HIV-1 , CD4-Positive T-Lymphocytes , CD8-Positive T-Lymphocytes , HIV Infections/drug therapy , Humans , Interleukin-15/genetics , Leukocytes, Mononuclear , Recombinant Fusion Proteins , Viral LoadABSTRACT
We have developed a non-radioactive flow-cytometry assay to monitor and quantify the target-cell killing activities mediated by cytotoxic T lymphocytes (CTLs). This flow-cytometry CTL (FCC) assay is predicated on measurement of CTL-induced caspase activation in target cells through detection of the specific cleavage of fluorogenic caspase substrates. Here we show that this assay reliably detects antigen-specific CTL killing of target cells, and demonstrate that it provides a more sensitive, more informative and safer alternative to the standard 51Cr-release assay most often used to quantify CTL responses. The FCC assay can be used to study CTL-mediated killing of primary host target cells of different cell lineages, and enables the study of antigen-specific cellular immune responses in real time at the single-cell level. As such, the FCC assay can provide a valuable tool for studies of infectious disease pathogenesis and development of new vaccines and immunotherapies.
Subject(s)
Caspases/metabolism , Cytotoxicity, Immunologic/physiology , T-Lymphocytes, Cytotoxic/immunology , Animals , Chromium Radioisotopes , Female , Flow Cytometry/methods , Mice , Mice, Inbred C57BL , Substrate SpecificityABSTRACT
BACKGROUND: Children with recurrent and/or metastatic osteosarcoma (OS), neuroblastoma (NB) and glioblastoma multiforme (GBM) have a dismal event-free survival (<25%). The majority of these solid tumors highly express GD2. Dinutuximab, an anti-GD2 monoclonal antibody, significantly improved event-free survival in children with GD2+ NB post autologous stem cell transplantation and enhanced natural killer (NK) cell-mediated antibody-dependent cell cytotoxicity. Thus, approaches to increase NK cell number and activity, improve persistence and trafficking, and enhance tumor targeting may further improve the clinical benefit of dinutuximab. N-803 is a superagonist of an interleukin-15 (IL-15) variant bound to an IL-15 receptor alpha Su-Fc fusion with enhanced biological activity. METHODS: The anti-tumor combinatorial effects of N-803, dinutuximab and ex vivo expanded peripheral blood NK cells (exPBNK) were performed in vitro using cytoxicity assays against GD2+ OS, NB and GBM cells. Perforin and interferon (IFN)-γ levels were measured by ELISA assays. Multiple cytokines/chemokines/growth factors released were measured by multiplex assays. Human OS, GBM or NB xenografted NOD/SCID/IL2rγnull (NSG) mice were used to investigate the anti-tumor combinatorial effects in vivo. RESULTS: N-803 increased the viability and proliferation of exPBNK. The increased viability and proliferation are associated with increased phosphorylation of Stat3, Stat5, AKT, p38MAPK and the expression of NK activating receptors. The combination of dinutuximab and N-803 significantly enhanced in vitro cytotoxicity of exPBNK with enhanced perforin and IFN-γ release against OS, GBM and NB. The combination of exPBNK+N-803+dinutuximab significantly reduced the secretion of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), platelet-derived growth factor-BB (PDGF-BB), and stem cell growth factor beta (SCGF-ß) from OS or GBM tumor cells. Furthermore, OS or GBM significantly inhibited the secretion of regulated on activation, normal T cell expressed and presumably secreted (RANTES) and stromal cell-derived factor-1 alpha (SDF-1α) from exPBNK cells (p<0.001) but significantly enhanced monokine induced by gamma interferon (MIG) secretion from exPBNK cells (p<0.001). N-803 combined with dinutuximab and exPBNK cells significantly extended the survival of OS, GBM or NB xenografted NSG mice. CONCLUSIONS: Our results provide the rationale for the development of a clinical trial of N-803 in combination with dinutuximab and ex vivo exPBNK cells in patients with recurrent or metastatic GD2+ solid tumors.