Human T Cell Leukemia Virus Type 1: Persistence and Pathogenesis.

Human T cell leukemia virus type 1 (HTLV-1), also known as human T lymphotropic virus type 1, was the first exogenous human retrovirus discovered. Unlike the distantly related lentivirus HIV-1, HTLV-1 causes disease in only 5-10% of infected people, depending on their ethnic origin. But whereas HIV-1 infection and the consequent diseases can be efficiently contained in most cases by antiretroviral drug treatment, there is no satisfactory treatment for the malignant or inflammatory diseases caused by HTLV-1. The purpose of the present article is to review recent advances in the understanding of the mechanisms by which the virus persists in vivo and causes disabling or fatal diseases.

Attacking Latent HIV with convertibleCAR-T Cells, a Highly Adaptable Killing Platform.

Current approaches to reducing the latent HIV reservoir entail first reactivating virus-containing cells to become visible to the immune system. A critical second step is killing these cells to reduce reservoir size. Endogenous cytotoxic T-lymphocytes (CTLs) may not be adequate because of cellular exhaustion and the evolution of CTL-resistant viruses. We have designed a universal CAR-T cell platform based on CTLs engineered to bind a variety of broadly neutralizing anti-HIV antibodies. We show that this platform, convertibleCAR-T cells, effectively kills HIV-infected, but not uninfected, CD4 T cells from blood, tonsil, or spleen and only when armed with anti-HIV antibodies. convertibleCAR-T cells also kill within 48 h more than half of the inducible reservoir found in blood of HIV-infected individuals on antiretroviral therapy. The modularity of convertibleCAR-T cell system, which allows multiplexing with several anti-HIV antibodies yielding greater breadth and control, makes it a promising tool for attacking the latent HIV reservoir.

Targeting the Latent Reservoir for HIV-1.

Antiretroviral therapy can effectively block HIV-1 replication and prevent or reverse immunodeficiency in HIV-1-infected individuals. However, viral replication resumes within weeks of treatment interruption. The major barrier to a cure is a small pool of resting memory CD4+ T cells that harbor latent HIV-1 proviruses. This latent reservoir is now the focus of an intense international research effort. We describe how the reservoir is established, challenges involved in eliminating it, and pharmacologic and immunologic strategies for targeting this reservoir. The development of a successful cure strategy will most likely require understanding the mechanisms that maintain HIV-1 proviruses in a latent state and pathways that drive the proliferation of infected cells, which slows reservoir decay. In addition, a cure will require the development of effective immunologic approaches to eliminating infected cells. There is renewed optimism about the prospect of a cure, and the interventions discussed here could pave the way.

Transcriptional Reprogramming during Effector-to-Memory Transition Renders CD4+ T Cells Permissive for Latent HIV-1 Infection.

The latent reservoir for HIV-1 in resting memory CD4+ T cells is the major barrier to curing HIV-1 infection. Studies of HIV-1 latency have focused on regulation of viral gene expression in cells in which latent infection is established. However, it remains unclear how infection initially becomes latent. Here we described a unique set of properties of CD4+ T cells undergoing effector-to-memory transition including temporary upregulation of CCR5 expression and rapid downregulation of cellular gene transcription. These cells allowed completion of steps in the HIV-1 life cycle through integration but suppressed HIV-1 gene transcription, thus allowing the establishment of latency. CD4+ T cells in this stage were substantially more permissive for HIV-1 latent infection than other CD4+ T cells. Establishment of latent HIV-1 infection in CD4+ T could be inhibited by viral-specific CD8+ T cells, a result with implications for elimination of latent HIV-1 infection by T cell-based vaccines.

Immune control of cytomegalovirus reactivation in stem cell transplantation.

The reactivation of viruses from latency after allogeneic stem cell transplantation (SCT) continues to represent a major clinical challenge requiring sophisticated monitoring strategies in the context of prophylactic and/or preemptive antiviral drugs that are associated with significant expense, toxicity and rates of failure. Accumulating evidence has demonstrated the association of polyfunctional virus-specific T cells with protection from viral reactivation, affirmed by the ability of adoptively transferred virus-specific T cells to prevent and treat reactivation and disease. The roles of natural killer cells in early viral surveillance and of dendritic cells in priming of T cells have also been delineated. Most recently, a role for strain-specific humoral responses in preventing early cytomegalovirus (CMV) reactivation has been demonstrated in preclinical models. Despite these advances, many unknowns remain: what are the critical innate and adaptive responses over time; are the origin (eg, recipient vs donor) and localization (eg, in parenchymal tissue vs lymphoid organs) of these responses important; how does graft-versus-host disease and the prevention and treatment thereof (eg, high-dose steroids) affect the functionality and relevance of a particular immune axis; do the immune parameters that control latency, reactivation, and dissemination differ; and what is the impact of new antiviral drugs on the development of enduring antiviral immunity. Thus, although antiviral drugs have provided major improvements over the past two decades, understanding the immunological paradigms underpinning protective antiviral immunity after SCT offers the potential to generate nontoxic, immune-based therapeutic approaches for lasting protection from viral reactivation.

Combination therapy with anti-HIV-1 antibodies maintains viral suppression.

Individuals infected with HIV-1 require lifelong antiretroviral therapy, because interruption of treatment leads to rapid rebound viraemia. Here we report on a phase 1b clinical trial in which a combination of 3BNC117 and 10-1074, two potent monoclonal anti-HIV-1 broadly neutralizing antibodies that target independent sites on the HIV-1 envelope spike, was administered during analytical treatment interruption. Participants received three infusions of 30 mg kg-1 of each antibody at 0, 3 and 6 weeks. Infusions of the two antibodies were generally well-tolerated. The nine enrolled individuals with antibody-sensitive latent viral reservoirs maintained suppression for between 15 and more than 30 weeks (median of 21 weeks), and none developed viruses that were resistant to both antibodies. We conclude that the combination of the anti-HIV-1 monoclonal antibodies 3BNC117 and 10-1074 can maintain long-term suppression in the absence of antiretroviral therapy in individuals with antibody-sensitive viral reservoirs.

Epstein-Barr Viruses: Their Immune Evasion Strategies and Implications for Autoimmune Diseases.

Epstein-Barr virus (EBV), a member of the γ-herpesvirus family, is one of the most prevalent and persistent human viruses, infecting up to 90% of the adult population globally. EBV's life cycle includes primary infection, latency, and lytic reactivation, with the virus primarily infecting B cells and epithelial cells. This virus has evolved sophisticated strategies to evade both innate and adaptive immune responses, thereby maintaining a lifelong presence within the host. This persistence is facilitated by the expression of latent genes such as EBV nuclear antigens (EBNAs) and latent membrane proteins (LMPs), which play crucial roles in viral latency and oncogenesis. In addition to their well-known roles in several types of cancer, including nasopharyngeal carcinoma and B-cell lymphomas, recent studies have identified the pathogenic roles of EBV in autoimmune diseases such as multiple sclerosis, rheumatoid arthritis, and systemic lupus erythematosus. This review highlights the intricate interactions between EBV and the host immune system, underscoring the need for further research to develop effective therapeutic and preventive strategies against EBV-associated diseases.

Essential role of M1 macrophages in blocking cytokine storm and pathology associated with murine HSV-1 infection.

Ocular HSV-1 infection is a major cause of eye disease and innate and adaptive immunity both play a role in protection and pathology associated with ocular infection. Previously we have shown that M1-type macrophages are the major and earliest infiltrates into the cornea of infected mice. We also showed that HSV-1 infectivity in the presence and absence of M2-macrophages was similar to wild-type (WT) control mice. However, it is not clear whether the absence of M1 macrophages plays a role in protection and disease in HSV-1 infected mice. To explore the role of M1 macrophages in HSV-1 infection, we used mice lacking M1 activation (M1-/- mice). Our results showed that macrophages from M1-/- mice were more susceptible to HSV-1 infection in vitro than were macrophages from WT mice. M1-/- mice were highly susceptible to ocular infection with virulent HSV-1 strain McKrae, while WT mice were refractory to infection. In addition, M1-/- mice had higher virus titers in the eyes than did WT mice. Adoptive transfer of M1 macrophages from WT mice to M1-/- mice reduced death and rescued virus replication in the eyes of infected mice. Infection of M1-/- mice with avirulent HSV-1 strain KOS also increased ocular virus replication and eye disease but did not affect latency-reactivation seen in WT control mice. Severity of virus replication and eye disease correlated with significantly higher inflammatory responses leading to a cytokine storm in the eyes of M1-/- infected mice that was not seen in WT mice. Thus, for the first time, our study illustrates the importance of M1 macrophages specifically in primary HSV-1 infection, eye disease, and survival but not in latency-reactivation.

Cytotoxic CD4+ T-cells specific for EBV capsid antigen BORF1 are maintained in long-term latently infected healthy donors.

Epstein Barr Virus (EBV) infects more than 95% of the population whereupon it establishes a latent infection of B-cells that persists for life under immune control. Primary EBV infection can cause infectious mononucleosis (IM) and long-term viral carriage is associated with several malignancies and certain autoimmune diseases. Current efforts developing EBV prophylactic vaccination have focussed on neutralising antibodies. An alternative strategy, that could enhance the efficacy of such vaccines or be used alone, is to generate T-cell responses capable of recognising and eliminating newly EBV-infected cells before the virus initiates its growth transformation program. T-cell responses against the EBV structural proteins, brought into the newly infected cell by the incoming virion, are prime candidates for such responses. Here we show the structural EBV capsid proteins BcLF1, BDLF1 and BORF1 are frequent targets of T-cell responses in EBV infected people, identify new CD8+ and CD4+ T-cell epitopes and map their HLA restricting alleles. Using T-cell clones we demonstrate that CD4+ but not CD8+ T-cell clones specific for the capsid proteins can recognise newly EBV-infected B-cells and control B-cell outgrowth via cytotoxicity. Using MHC-II tetramers we show a CD4+ T-cell response to an epitope within the BORF1 capsid protein epitope is present during acute EBV infection and in long-term viral carriage. In common with other EBV-specific CD4+ T-cell responses the BORF1-specific CD4+ T-cells in IM patients expressed perforin and granzyme-B. Unexpectedly, perforin and granzyme-B expression was sustained over time even when the donor had entered the long-term infected state. These data further our understanding of EBV structural proteins as targets of T-cell responses and how CD4+ T-cell responses to EBV change from acute disease into convalescence. They also identify new targets for prophylactic EBV vaccine development.

'Rinse and Replace': Boosting T Cell Turnover To Reduce HIV-1 Reservoirs.

Latent HIV-1 persists indefinitely during antiretroviral therapy (ART) as an integrated silent genome in long-lived memory CD4+ T cells. In untreated infections, immune activation increases the turnover of intrinsically long-lived provirus-containing CD4+ T cells. Those are 'washed out' as a result of their activation, which when coupled to viral protein expression can facilitate local inflammation and recruitment of uninfected cells to activation sites, causing latently infected cells to compete for survival. De novo infection can counter this washout. During ART, inflammation and CD4+ T cell activation wane, resulting in reduced cell turnover and a persistent reservoir. We propose accelerating reservoir washout during ART by triggering sequential waves of polyclonal CD4+ T cell activation while simultaneously enhancing virus protein expression. Reservoir reduction as an adjunct to other therapies might achieve lifelong viral control.

CD32a is a marker of a CD4 T-cell HIV reservoir harbouring replication-competent proviruses.

The persistence of the HIV reservoir in infected individuals is a major obstacle to the development of a cure for HIV. Here, using an in vitro model of HIV-infected quiescent CD4 T cells, we reveal a gene expression signature of 103 upregulated genes that are specific for latently infected cells, including genes for 16 transmembrane proteins. In vitro screening for surface expression in HIV-infected quiescent CD4 T cells shows that the low-affinity receptor for the immunoglobulin G Fc fragment, CD32a, is the most highly induced, with no detectable expression in bystander cells. Notably, productive HIV-1 infection of T-cell-receptor-stimulated CD4 T cells is not associated with CD32a expression, suggesting that a quiescence-dependent mechanism is required for its induction. Using blood samples from HIV-1-positive participants receiving suppressive antiretroviral therapy, we identify a subpopulation of 0.012% of CD4 T cells that express CD32a and host up to three copies of HIV DNA per cell. This CD32a+ reservoir was highly enriched in inducible replication-competent proviruses and can be predominant in some participants. Our discovery that CD32a+ lymphocytes represent the elusive HIV-1 reservoir may lead to insights that will facilitate the specific targeting and elimination of this reservoir.

Autologous IgG antibodies block outgrowth of a substantial but variable fraction of viruses in the latent reservoir for HIV-1.

In untreated HIV-1 infection, rapid viral evolution allows escape from immune responses. Viral replication can be blocked by antiretroviral therapy. However, HIV-1 persists in a latent reservoir in resting CD4+ T cells, and rebound viremia occurs following treatment interruption. The reservoir, which is maintained in part by clonal expansion, can be measured using quantitative viral outgrowth assays (QVOAs) in which latency is reversed with T cell activation to allow viral outgrowth. Recent studies have shown that viruses detected in QVOAs prior to treatment interruption often differ from rebound viruses. We hypothesized that autologous neutralizing antibodies directed at the HIV-1 envelope (Env) protein might block outgrowth of some reservoir viruses. We modified the QVOA to reflect pressure from low concentrations of autologous antibodies and showed that outgrowth of a substantial but variable fraction of reservoir viruses is blocked by autologous contemporaneous immunoglobulin G (IgG). A reduction in outgrowth of >80% was seen in 6 of 15 individuals. This effect was due to direct neutralization. We established a phylogenetic relationship between rebound viruses and viruses growing out in vitro in the presence of autologous antibodies. Some large infected cell clones detected by QVOA carried neutralization-sensitive viruses, providing a cogent explanation for differences between rebound virus and viruses detected in standard QVOAs. Measurement of the frequency of reservoir viruses capable of outgrowth in the presence of autologous IgG might allow more accurate prediction of time to viral rebound. Ultimately, therapeutic immunization targeting the subset of variants resistant to autologous IgG might contribute to a functional cure.

Interferon Regulatory Factor 3 Supports the Establishment of Chronic Gammaherpesvirus Infection in a Route- and Dose-Dependent Manner.

Gammaherpesviruses are ubiquitous pathogens that establish lifelong infections and are associated with several malignancies, including B cell lymphomas. Uniquely, these viruses manipulate B cell differentiation to establish long-term latency in memory B cells. This study focuses on the interaction between gammaherpesviruses and interferon regulatory factor 3 (IRF-3), a ubiquitously expressed transcription factor with multiple direct target genes, including beta interferon (IFN-ß), a type I IFN. IRF-3 attenuates acute replication of a plethora of viruses, including gammaherpesvirus. Furthermore, IRF-3-driven IFN-ß expression is antagonized by the conserved gammaherpesvirus protein kinase during lytic virus replication in vitro In this study, we have uncovered an unexpected proviral role of IRF-3 during chronic gammaherpesvirus infection. In contrast to the antiviral activity of IRF-3 during acute infection, IRF-3 facilitated establishment of latent gammaherpesvirus infection in B cells, particularly, germinal center and activated B cells, the cell types critical for both natural infection and viral lymphomagenesis. This proviral role of IRF-3 was further modified by the route of infection and viral dose. Furthermore, using a combination of viral and host genetics, we show that IRF-3 deficiency does not rescue attenuated chronic infection of a protein kinase null gammaherpesvirus mutant, highlighting the multifunctional nature of the conserved gammaherpesvirus protein kinases in vivo In summary, this study unveils an unexpected proviral nature of the classical innate immune factor, IRF-3, during chronic virus infection.IMPORTANCE Interferon regulatory factor 3 (IRF-3) is a critical component of the innate immune response, in part due to its transactivation of beta interferon (IFN-ß) expression. Similar to that observed in all acute virus infections examined to date, IRF-3 suppresses lytic viral replication during acute gammaherpesvirus infection. Because gammaherpesviruses establish lifelong infection, this study aimed to define the antiviral activity of IRF-3 during chronic infection. Surprisingly, we found that, in contrast to acute infection, IRF-3 supported the establishment of gammaherpesvirus latency in splenic B cells, revealing an unexpected proviral nature of this classical innate immune host factor.

T Cell-Intrinsic Interferon Regulatory Factor 1 Expression Suppresses Differentiation of CD4+ T Cell Populations That Support Chronic Gammaherpesvirus Infection.

Gammaherpesviruses are ubiquitous pathogens that establish lifelong infection and are associated with B cell lymphomas. To establish chronic infection, these viruses usurp B cell differentiation and drive a robust germinal center response to expand the latent viral reservoir and gain access to memory B cells. Germinal center B cells, while important for the establishment of latent infection, are also thought to be the target of viral transformation. The host and viral factors that impact the gammaherpesvirus-driven germinal center response are not clearly defined. We show that the global expression of the antiviral and tumor suppressor interferon regulatory factor 1 (IRF-1) selectively attenuates the murine gammaherpesvirus 68 (MHV68)-driven germinal center response and restricts the expansion of the latent viral reservoir. In this study, we found that T cell-intrinsic IRF-1 expression recapitulates some aspects of the antiviral state imposed by IRF-1 during chronic MHV68 infection, including the attenuation of the germinal center response and viral latency in the spleen. We also discovered that global and T cell-intrinsic IRF-1 deficiency leads to an unhindered rise of interleukin-17A (IL-17A)-expressing and follicular helper T cell populations, two CD4+ T cell subsets that support chronic MHV68 infection. Thus, this study unveils a novel aspect of the antiviral activity of IRF-1 by demonstrating IRF-1-mediated suppression of specific CD4+ T cell subsets that support chronic gammaherpesvirus infection. IMPORTANCE Gammaherpesviruses infect over 95% of the adult population, last the lifetime of the host, and are associated with multiple cancers. These viruses usurp the germinal center response to establish lifelong infection in memory B cells. This manipulation of B cell differentiation by the virus is thought to contribute to lymphomagenesis, although exactly how the virus precipitates malignant transformation in vivo is unclear. IRF-1, a host transcription factor and a known tumor suppressor, restricts the MHV68-driven germinal center response in a B cell-extrinsic manner. We found that T cell-intrinsic IRF-1 expression attenuates the MHV68-driven germinal center response by restricting the CD4+ T follicular helper population. Furthermore, our study identified IRF-1 as a novel negative regulator of IL-17-driven immune responses, highlighting the multifaceted role of IRF-1 in gammaherpesvirus infection.

An HSV-2 nucleoside-modified mRNA genital herpes vaccine containing glycoproteins gC, gD, and gE protects mice against HSV-1 genital lesions and latent infection.

HSV-1 causes 50% of first-time genital herpes infections in resource-rich countries and affects 190 million people worldwide. A prophylactic herpes vaccine is needed to protect against genital infections by both HSV-1 and HSV-2. Previously our laboratory developed a trivalent vaccine that targets glycoproteins C, D, and E present on the HSV-2 virion. We reported that this vaccine protects animals from genital disease and recurrent virus shedding following lethal HSV-2 challenge. Importantly the vaccine also generates cross-reactive antibodies that neutralize HSV-1, suggesting it may provide protection against HSV-1 infection. Here we compared the efficacy of this vaccine delivered as protein or nucleoside-modified mRNA immunogens against vaginal HSV-1 infection in mice. Both the protein and mRNA vaccines protected mice from HSV-1 disease; however, the mRNA vaccine provided better protection as measured by lower vaginal virus titers post-infection. In a second experiment, we compared protection provided by the mRNA vaccine against intravaginal challenge with HSV-1 or HSV-2. Vaccinated mice were totally protected against death, genital disease and infection of dorsal root ganglia caused by both viruses, but somewhat better protected against vaginal titers after HSV-2 infection. Overall, in the two experiments, the mRNA vaccine prevented death and genital disease in 54/54 (100%) mice infected with HSV-1 and 20/20 (100%) with HSV-2, and prevented HSV DNA from reaching the dorsal root ganglia, the site of virus latency, in 29/30 (97%) mice infected with HSV-1 and 10/10 (100%) with HSV-2. We consider the HSV-2 trivalent mRNA vaccine to be a promising candidate for clinical trials for prevention of both HSV-1 and HSV-2 genital herpes.

Diverse effects of interferon alpha on the establishment and reversal of HIV latency.

HIV latency is the major barrier to a cure for people living with HIV (PLWH) on antiretroviral therapy (ART) because the virus persists in long-lived non-proliferating and proliferating latently infected CD4+ T cells. Latently infected CD4+ T cells do not express viral proteins and are therefore not visible to immune mediated clearance. Therefore, identifying interventions that can reverse latency and also enhance immune mediated clearance is of high interest. Interferons (IFNs) have multiple immune enhancing effects and can inhibit HIV replication in activated CD4+ T cells. However, the effects of IFNs on the establishment and reversal of HIV latency is not understood. Using an in vitro model of latency, we demonstrated that plasmacytoid dendritic cells (pDC) inhibit the establishment of HIV latency through secretion of type I IFNα, IFNß and IFNω but not IFNε or type III IFNλ1 and IFNλ3. However, once latency was established, IFNα but no other IFNs were able to efficiently reverse latency in both an in vitro model of latency and CD4+ T cells collected from PLWH on suppressive ART. Binding of IFNα to its receptor expressed on primary CD4+ T cells did not induce activation of the canonical or non-canonical NFκB pathway but did induce phosphorylation of STAT1, 3 and 5 proteins. STAT5 has been previously demonstrated to bind to the HIV long terminal repeat and activate HIV transcription. We demonstrate diverse effects of interferons on HIV latency with type I IFNα; inhibiting the establishment of latency but also reversing HIV latency once latency is established.

Latency reversal agents modulate HIV antigen processing and presentation to CD8 T cells.

Latency reversal agents (LRA) variably induce HIV re-expression in CD4 T cells but reservoirs are not cleared. Whether HIV epitope presentation is similar between latency reversal and initial infection of CD4 T cells is unknown yet crucial to define immune responses able to detect HIV-infected CD4 T cells after latency reversal. HIV peptides displayed by MHC comes from the intracellular degradation of proteins by proteasomes and post-proteasomal peptidases but the impact of LRAs on antigen processing is not known. Here we show that HDAC inhibitors (HDCAi) reduced cytosolic proteolytic activities while PKC agonists (PKCa) increased them to a lesser extent than that induced by TCR activation. During the cytosolic degradation of long HIV peptides in LRA-treated CD4 T cells extracts, HDACi and PKCa modulated degradation patterns of peptides and altered the production of HIV epitopes in often opposite ways. Beyond known HIV epitopes, HDACi narrowed the coverage of HIV antigenic fragments by 8-11aa degradation peptides while PKCa broadened it. LRAs altered HIV infection kinetics and modulated CD8 T cell activation in an epitope- and time-dependent manner. Interestingly the efficiency of endogenous epitope processing and presentation to CD8 T cells was increased by PKCa Ingenol at early time points despite low levels of antigens. LRA-induced modulations of antigen processing should be considered and exploited to enhance and broaden HIV peptide presentation by CD4 T cells and to improve immune recognition after latency reversal. This property of LRAs, if confirmed with other antigens, might be exploited to improve immune detection of diseased cells beyond HIV.

Mining for humoral correlates of HIV control and latent reservoir size.

While antiretroviral therapy (ART) has effectively revolutionized HIV care, the virus is never fully eliminated. Instead, immune dysfunction, driven by persistent non-specific immune activation, ensues and progressively leads to premature immunologic aging. Current biomarkers monitoring immunologic changes encompass generic inflammatory biomarkers, that may also change with other infections or disease states, precluding the antigen-specific monitoring of HIV-infection associated changes in disease. Given our growing appreciation of the significant changes in qualitative and quantitative properties of disease-specific antibodies in HIV infection, we used a systems approach to explore humoral profiles associated with HIV control. We found that HIV-specific antibody profiles diverge by spontaneous control of HIV, treatment status, viral load and reservoir size. Specifically, HIV-specific antibody profiles representative of changes in viral load were largely quantitative, reflected by differential HIV-specific antibody levels and Fc-receptor binding. Conversely, HIV-specific antibody features that tracked with reservoir size exhibited a combination of quantitative and qualitative changes marked by more distinct subclass selection profiles and unique HIV-specific Fc-glycans. Our analyses suggest that HIV-specific antibody Fc-profiles provide antigen-specific resolution on both cell free and cell-associated viral loads, pointing to potentially novel biomarkers to monitor reservoir activity.

CTL Clonotypes with Higher TCR Affinity Have Better Ability to Reduce the HIV Latent Reservoir.

The success of the shock and kill strategy for the HIV cure depends both on the reactivation of the latent reservoir and on the ability of the immune system to eliminate infected cells. As latency reversal alone has not shown any impact in the size of the latent reservoir, ensuring that effector CTLs are able to recognize and kill HIV-infected cells could contribute to reservoir reduction. In this study, we investigated which functional aspects of human CTLs are associated with a better capacity to kill HIV-infected CD4+ T cells. We isolated Gag- and Nef-specific CTL clones with different TCR sequences from the PBMC of donors in acute and chronic infection. High-affinity clonotypes that showed IFN-γ production preserved even when the CD8 coreceptor was blocked, and clones with high Ag sensitivity exhibited higher efficiency at reducing the latent reservoir. Although intrinsic cytotoxic capacity did not differ according to TCR affinity, clonotypes with high TCR affinity showed a better ability to kill HIV-infected CD4+ T cells obtained from in vivo-infected PBMC and subjected to viral reactivation. Strategies aiming to specifically boost and maintain long-living memory CTLs with high TCR affinity in vivo prior to latency-reversing treatment might improve the efficacy of the shock and kill approach to reduce the latent reservoir.

Combination Immune Checkpoint Blockade to Reverse HIV Latency.

In people living with HIV on antiretroviral therapy, HIV latency is the major barrier to a cure. HIV persists preferentially in CD4+ T cells expressing multiple immune checkpoint (IC) molecules, including programmed death (PD)-1, T cell Ig and mucin domain-containing protein 3 (TIM-3), lymphocyte associated gene 3 (LAG-3), and T cell immunoreceptor with Ig and ITIM domains (TIGIT). We aimed to determine whether these and other IC molecules have a functional role in maintaining HIV latency and whether blocking IC molecules with Abs reverses HIV latency. Using an in vitro model that establishes latency in both nonproliferating and proliferating human CD4+ T cells, we show that proliferating cells express multiple IC molecules at high levels. Latent infection was enriched in proliferating cells expressing PD-1. In contrast, nonproliferating cells expressed IC molecules at significantly lower levels, but latent infection was enriched in cells expressing PD-1, TIM-3, CTL-associated protein 4 (CTLA-4), or B and T lymphocyte attenuator (BTLA). In the presence of an additional T cell-activating stimulus, staphylococcal enterotoxin B, Abs to CTLA-4 and PD-1 reversed HIV latency in proliferating and nonproliferating CD4+ T cells, respectively. In the absence of staphylococcal enterotoxin B, only the combination of Abs to PD-1, CTLA-4, TIM-3, and TIGIT reversed latency. The potency of latency reversal was significantly higher following combination IC blockade compared with other latency-reversing agents, including vorinostat and bryostatin. Combination IC blockade should be further explored as a strategy to reverse HIV latency.