Lydie Trautmann

Upregulation of PD-1 expression on HIV-specific CD8 + T cells leads to reversible immune dysfunction

The engagement of programmed death 1 (PD-1) to its ligands, PD-L1 and PD-L2, inhibits proliferation and cytokine production mediated by antibodies to CD3 (refs. 5,6,7). Blocking the PD-1–PD-L1 pathway in mice chronically infected with lymphocytic choriomeningitis virus restores the capacity of exhausted CD8+ T cells to undergo proliferation, cytokine production and cytotoxic activity and, consequently, results in reduced viral load. During chronic HIV infection, HIV-specific CD8+ T cells are functionally impaired, showing a reduced capacity to produce cytokines and effector molecules as well as an impaired capacity to proliferate. Here, we found that PD-1 was upregulated on HIV-specific CD8+ T cells; PD-1 expression levels were significantly correlated both with viral load and with the reduced capacity for cytokine production and proliferation of HIV-specific CD8+ T cells. Notably, cytomegalovirus (CMV)-specific CD8+ T cells from the same donors did not upregulate PD-1 and maintained the production of high levels of cytokines. Blocking PD-1 engagement to its ligand (PD-L1) enhanced the capacity of HIV-specific CD8+ T cells to survive and proliferate and led to an increased production of cytokines and cytotoxic molecules in response to cognate antigen. The accumulation of HIV-specific dysfunctional CD8+ T cells in the infected host could prevent the renewal of a functionally competent HIV-specific CD8+ repertoire.

HIV reservoir size and persistence are driven by T cell survival and homeostatic proliferation

HIV persists in a reservoir of latently infected CD4+ T cells in individuals treated with highly active antiretroviral therapy (HAART). Here we identify central memory (TCM) and transitional memory (TTM) CD4+ T cells as the major cellular reservoirs for HIV and find that viral persistence is ensured by two different mechanisms. HIV primarily persists in TCM cells in subjects showing reconstitution of the CD4+ compartment upon HAART. This reservoir is maintained through T cell survival and low-level antigen-driven proliferation and is slowly depleted with time. In contrast, proviral DNA is preferentially detected in TTM cells from aviremic individuals with low CD4+ counts and higher amounts of interleukin-7–mediated homeostatic proliferation, a mechanism that ensures the persistence of these cells. Our results suggest that viral eradication might be achieved through the combined use of strategic interventions targeting viral replication and, as in cancer, drugs that interfere with the self renewal and persistence of proliferating memory T cells.

Programmed death-1–induced interleukin-10 production by monocytes impairs CD4 + T cell activation during HIV infection

Viral replication and microbial translocation from the gut to the blood during HIV infection lead to hyperimmune activation, which contributes to the decline in CD4+ T cell numbers during HIV infection. Programmed death-1 (PD-1) and interleukin-10 (IL-10) are both upregulated during HIV infection. Blocking interactions between PD-1 and programmed death ligand-1 (PD-L1) and between IL-10 and IL-10 receptor (IL-10R) results in viral clearance and improves T cell function in animal models of chronic viral infections. Here we show that high amounts of microbial products and inflammatory cytokines in the plasma of HIV-infected subjects lead to upregulation of PD-1 expression on monocytes that correlates with high plasma concentrations of IL-10. Triggering of PD-1 expressed on monocytes by PD-L1 expressed on various cell types induced IL-10 production and led to reversible CD4+ T cell dysfunction. We describe a new function for PD-1 whereby microbial products inhibit T cell expansion and function by upregulating PD-1 levels and IL-10 production by monocytes after binding of PD-1 by PD-L1.

Selection of T Cell Clones Expressing High-Affinity Public TCRs within Human Cytomegalovirus-Specific CD8 T Cell Responses

Assessment of clonal diversity of T cell responses against human CMV (HCMV), a major cause of morbidity in immunodepressed patients, provides important insights into the molecular basis of T cell immunodominance, and has also clinical implications for the immunomonitoring and immunotherapy of HCMV infections. We performed an in-depth molecular and functional characterization of CD8 T cells directed against an immunodominant HLA-A2-restricted epitope derived from HCMV protein pp65 (NLV/A2) in steady state and pathological situations associated with HCMV reactivation. NLV/A2-specific T cells in healthy HCMV-seropositive donors showed limited clonal diversity and usage of a restricted set of TCR Vβ regions. Although TCRβ-chain junctional sequences were highly diverse, a large fraction of NLV/A2-specific T cells derived from distinct individuals showed several recurrent (so-called “public”) TCR features associated in some cases with full conservation of the TCRα chain junctional region. A dramatic clonal focusing of NLV/A2-specific T cells was observed in situations of HCMV reactivation and/or chronic inflammation, which resulted in selection of a single clonotype displaying similar public TCR features in several patients. In most instances the NLV/A2-specific dominant clonotypes showed higher affinity for their Ag than subdominant ones, thus suggesting that TCR affinity/avidity is the primary driving force underlying repertoire focusing along chronic antigenic stimulation.

Transcription factor FOXO3a controls the persistence of memory CD4+ T cells during HIV infection

The persistence of central memory CD4+ T cells (TCM cells) is a major correlate of immunological protection in HIV/AIDS, as the rate of TCM cell decline predicts HIV disease progression. In this study, we show that TCM cells and effector memory CD4+ T cells (TEM cells) from HIV+ elite controller (EC) subjects are less susceptible to Fas-mediated apoptosis and persist longer after multiple rounds of T cell receptor triggering when compared to TCM and TEM cells from aviremic successfully treated (ST) subjects or from HIV− donors. We show that persistence of TCM cells from EC subjects is a direct consequence of inactivation of the FOXO3a pathway. Silencing the transcriptionally active form of FOXO3a by small interfering RNA or by introducing a FOXO3a dominant-negative form (FOXO3a Nt) extended the long-term survival of TCM cells from ST subjects to a length of time similar to that of TCM cells from EC subjects. The crucial role of FOXO3a in the survival of memory cells will help shed light on the underlying immunological mechanisms that control viral replication in EC subjects.

Early Interferon Therapy for Hepatitis C Virus Infection Rescues Polyfunctional, Long-Lived CD8+ Memory T Cells

ABSTRACT The majority of acute hepatitis C virus (HCV) infections progress to chronicity and progressive liver damage. Alpha interferon (IFN-α) antiviral therapy achieves the highest rate of success when IFN-α is administered early during the acute phase, but the underlying mechanisms are unknown. We used a panel of major histocompatibility complex class I tetramers to monitor the phenotypic and functional signatures of HCV-specific T cells during acute HCV infection with different infection outcomes and during early IFN therapy. We demonstrate that spontaneous resolution correlates with the early development of polyfunctional (IFN-γ- and IL-2-producing and CD107a+) virus-specific CD8+ T cells. These polyfunctional T cells are distinguished by the expression of CD127 and Bcl-2 and represent a transitional memory T-cell subset that exhibits the phenotypic and functional signatures of both central and effector memory T cells. In contrast, HCV-specific CD8+ T cells in acute infections evolving to chronicity expressed low levels of CD127 and Bcl-2, exhibited diminished proliferation and cytokine production, and eventually disappeared from the periphery. Early therapeutic intervention with pegylated IFN-α rescued polyfunctional memory T cells expressing high levels of CD127 and Bcl-2. These cells were detectable for up to 1 year following discontinuation of therapy. Our results suggest that the polyfunctionality of HCV-specific T cells can be predictive of the outcome of acute HCV infection and that early therapeutic intervention can reconstitute the pool of long-lived polyfunctional memory T cells.

Frequent Contribution of T Cell Clonotypes with Public TCR Features to the Chronic Response Against a Dominant EBV-Derived Epitope: Application to Direct Detection of Their Molecular Imprint on the Human Peripheral T Cell Repertoire

In an attempt to provide a global picture of the TCR repertoire diversity of a chronic T cell response against a common Ag, we performed an extensive TCR analysis of cells reactive against a dominant HLA-A2-restricted EBV epitope (hereafter referred to as GLC/A2), obtained after sorting PBL or synovial fluid lymphocytes from EBV-seropositive individuals using MHC/peptide multimers. Although TCR β-chain diversity of GLC/A2+ T cells was extensive and varied greatly from one donor to another, we identified in most cell lines several recurrent Vβ subsets (Vβ2, Vβ4, and Vβ16 positive) with highly conserved TCRβ complementarity-determining region 3 (CDR3) length and junctional motifs, which represented from 11 to 98% (mean, 50%) of GLC/A2-reactive cells. While TCR β-chains expressed by these subsets showed limited CDR1, CDR2, and CDR3 homology among themselves, their TCR α-chains comprised the same TCRAV region, thus suggesting hierarchical contribution of TCR α-chain vs TCR β-chain CDR to recognition of this particular MHC/peptide complex. The common occurrence of T cell clonotypes with public TCR features within GLC/A2-specific T cells allowed their direct detection within unsorted PBL using ad hoc clonotypic primers. These results, which suggest an unexpectedly high contribution of public clonotypes to the TCR repertoire against a dominant epitope, have several implications for the follow-up and modulation of T cell-mediated immunity.

Immune activation alters cellular and humoral responses to yellow fever 17D vaccine

BACKGROUND Defining the parameters that modulate vaccine responses in African populations will be imperative to design effective vaccines for protection against HIV, malaria, tuberculosis, and dengue virus infections. This study aimed to evaluate the contribution of the patient-specific immune microenvironment to the response to the licensed yellow fever vaccine 17D (YF-17D) in an African cohort. METHODS We compared responses to YF-17D in 50 volunteers in Entebbe, Uganda, and 50 volunteers in Lausanne, Switzerland. We measured the CD8+ T cell and B cell responses induced by YF-17D and correlated them with immune parameters analyzed by flow cytometry prior to vaccination. RESULTS We showed that YF-17D-induced CD8+ T cell and B cell responses were substantially lower in immunized individuals from Entebbe compared with immunized individuals from Lausanne. The impaired vaccine response in the Entebbe cohort associated with reduced YF-17D replication. Prior to vaccination, we observed higher frequencies of exhausted and activated NK cells, differentiated T and B cell subsets and proinflammatory monocytes, suggesting an activated immune microenvironment in the Entebbe volunteers. Interestingly, activation of CD8+ T cells and B cells as well as proinflammatory monocytes at baseline negatively correlated with YF-17D-neutralizing antibody titers after vaccination. Additionally, memory T and B cell responses in preimmunized volunteers exhibited reduced persistence in the Entebbe cohort but were boosted by a second vaccination. CONCLUSION Together, these results demonstrate that an activated immune microenvironment prior to vaccination impedes efficacy of the YF-17D vaccine in an African cohort and suggest that vaccine regimens may need to be boosted in African populations to achieve efficient immunity. TRIAL REGISTRATION Registration is not required for observational studies. FUNDING This study was funded by Canadas Global Health Research Initiative, Defense Threat Reduction Agency, National Institute of Allergy and Infectious Diseases, Bill & Melinda Gates Foundation, and United States Agency for International Development.

The Tat Inhibitor Didehydro-Cortistatin A Prevents HIV-1 Reactivation from Latency

ABSTRACT Antiretroviral therapy (ART) inhibits HIV-1 replication, but the virus persists in latently infected resting memory CD4+ T cells susceptible to viral reactivation. The virus-encoded early gene product Tat activates transcription of the viral genome and promotes exponential viral production. Here we show that the Tat inhibitor didehydro-cortistatin A (dCA), unlike other antiretrovirals, reduces residual levels of viral transcription in several models of HIV latency, breaks the Tat-mediated transcriptional feedback loop, and establishes a nearly permanent state of latency, which greatly diminishes the capacity for virus reactivation. Importantly, treatment with dCA induces inactivation of viral transcription even after its removal, suggesting that the HIV promoter is epigenetically repressed. Critically, dCA inhibits viral reactivation upon CD3/CD28 or prostratin stimulation of latently infected CD4+ T cells from HIV-infected subjects receiving suppressive ART. Our results suggest that inclusion of a Tat inhibitor in current ART regimens may contribute to a functional HIV-1 cure by reducing low-level viremia and preventing viral reactivation from latent reservoirs. IMPORTANCE Antiretroviral therapy (ART) reduces HIV-1 replication to very low levels, but the virus persists in latently infected memory CD4+ T cells, representing a long-lasting source of resurgent virus upon ART interruption. Based on the mode of action of didehydro-cortistatin A (dCA), a Tat-dependent transcription inhibitor, our work highlights an alternative approach to current HIV-1 eradication strategies to decrease the latent reservoir. In our model, dCA blocks the Tat feedback loop initiated after low-level basal reactivation, blocking transcriptional elongation and hence viral production from latently infected cells. Therefore, dCA combined with ART would be aimed at delaying or halting ongoing viral replication, reactivation, and replenishment of the latent viral reservoir. Thus, the latent pool of cells in an infected individual would be stabilized, and death of the long-lived infected memory T cells would result in a continuous decay of this pool over time, possibly culminating in the long-awaited sterilizing cure. Antiretroviral therapy (ART) reduces HIV-1 replication to very low levels, but the virus persists in latently infected memory CD4+ T cells, representing a long-lasting source of resurgent virus upon ART interruption. Based on the mode of action of didehydro-cortistatin A (dCA), a Tat-dependent transcription inhibitor, our work highlights an alternative approach to current HIV-1 eradication strategies to decrease the latent reservoir. In our model, dCA blocks the Tat feedback loop initiated after low-level basal reactivation, blocking transcriptional elongation and hence viral production from latently infected cells. Therefore, dCA combined with ART would be aimed at delaying or halting ongoing viral replication, reactivation, and replenishment of the latent viral reservoir. Thus, the latent pool of cells in an infected individual would be stabilized, and death of the long-lived infected memory T cells would result in a continuous decay of this pool over time, possibly culminating in the long-awaited sterilizing cure.

Dominant TCR Vα usage by virus and tumor‐reactive T cells with wide affinity ranges for their specific antigens

We have studied the TCR features and functional responses of three sets of human cytolytic T cell (CTL) clones, recognizing antigenic peptides presented by HLA‐A2 and derived from the Epstein‐Barr virus proteins BMLF1 and BRLF1 and from the melanoma protein Melan‐A/MART‐1. Within each set, a majority of clones used a recurrent Vα region, even though they expressed highly diverse TCR β chains and V(D)J junctional sequences. Functional assays and peptide/MHC multimer binding studies indicated that this restricted Vα usage was not associated with the affinity/avidity of the CTL clones. The Vα dominance, which may be a frequent feature of antigen‐specific T cells, likely reflects a restricted geometry of TCR/peptide/MHC complexes, primarily determined by Vα CDR.