Primary Adaptive Immune Responses Research Articles

Specific CD4+ T cell phenotypes associate with bacterial control in people who ‘resist’ infection with Mycobacterium tuberculosis

A subset of individuals exposed to Mycobacterium tuberculosis (Mtb) that we refer to as ‘resisters’ (RSTR) show evidence of IFN-γ− T cell responses to Mtb-specific antigens despite serially negative results on clinical testing. Here we found that Mtb-specific T cells in RSTR were clonally expanded, confirming the priming of adaptive immune responses following Mtb exposure. RSTR CD4+ T cells showed enrichment of TH17 and regulatory T cell-like functional programs compared to Mtb-specific T cells from individuals with latent Mtb infection. Using public datasets, we showed that these TH17 cell-like functional programs were associated with lack of progression to active tuberculosis among South African adolescents with latent Mtb infection and with bacterial control in nonhuman primates. Our findings suggested that RSTR may successfully control Mtb following exposure and immune priming and established a set of T cell biomarkers to facilitate further study of this clinical phenotype.

(Auto)immunity in focal epilepsy: mechanisms of (auto‑)immune-inflammatory epileptogenic neurodegeneration

While the neuronal mechanisms of epileptic hyperexcitability (HE) have been studied in detail, recent findings suggest that extraneuronal, mainly immune-mediated inflammatory and vascular mechanisms play an important role in the development and progression of HE in epilepsy and the cognitive and behavioral comorbidities. Narrative review. As in autoimmune (limbic) encephalitis (ALE/AIE) or Rasmussen's encephalitis (RE), the primary adaptive and innate immune responses and associated changes in the blood-brain barrier (BBB) and neurovascular unit (NVU) can cause acute cortical hyperexcitability (HE) and the development of hippocampal sclerosis (HS) and other structural cortical lesions with chronic HE. Cortical HE, which is associated with malformation of cortical development (MCD) and low-grade epilepsy-associated tumors (LEAT), for example, can be accompanied by secondary adaptive and innate immune responses and alterations in the BBB and NVU, potentially modulating the ictogenicity and epileptogenicity. These associations illustrate the influence of adaptive and innate immune mechanisms and associated changes in the BBB and NVU on cortical excitability and vice versa, suggesting adynamic and complex interplay of these factors in the development and progression of epilepsy in general. The described concept of a neuro-immune-vascular interaction in focal epilepsy opens up new possibilities for the pathogenetic understanding and thus also for the selective therapeutic intervention.

The effects of nanoplastics and microcystin-LR coexposure on Aristichthys nobilis at the early developmental stages

Nanoplastics (NPs) and microcystin-LR (MC-LR) are two common and harmful pollutants in water environments, especially at aquafarm where are full of plastic products and algae. It is of great significance to study the toxic effects and mechanisms of the NPs and/or MC-LR on fish at the early stage. In this study, the embryo and larvae of a filtering-feeding fish, Aristichthys nobilis, were used as the research objects. The results showed that the survival and hatching rates of the embryo were not significantly affected by the environmental concentration exposure of these two pollutants. Scanning electron microscopy (SEM) observation displayed that NPs adhered to the surface of the embryo membrane. Transcriptomic and bioinformatic analyses revealed that the NPs exposure activated neuromuscular junction development and skeletal muscle fiber in larvae, and affected C5-Branched dibasic acid metabolism. The metabolic and biosynthetic processes of zeaxanthin, xanthophyll, tetraterpenoid, and carotenoid were suppressed after the MC-LR exposure, which was harmful to the retinol metabolism of fish. Excessive production of superoxide dismutase (SOD) was detected under the MC-LR exposure. The MC-LR and NPs coexposure triggered primary immunodeficiency and adaptive immune response, leading to the possibility of reduced fitness of A.nobilis during the development. Collectively, our results indicate that environmental concentration NPs and MC-LR coexposure could cause toxic damage and enhance sick risk in A.nobilis, providing new insights into the risk of NPs and MC-LR on filtering-feeding fish.

Depleting myeloid-biased haematopoietic stem cells rejuvenates aged immunity.

Ageing of the immune system is characterized by decreased lymphopoiesis and adaptive immunity, and increased inflammation and myeloid pathologies1,2. Age-related changes in populations of self-renewing haematopoietic stem cells (HSCs) are thought to underlie these phenomena3. During youth, HSCs with balanced output of lymphoid and myeloid cells (bal-HSCs) predominate over HSCs with myeloid-biased output (my-HSCs), thereby promoting the lymphopoiesis required for initiating adaptive immune responses, while limiting the production of myeloid cells, which can be pro-inflammatory4. Ageing is associated with increased proportions of my-HSCs, resulting in decreased lymphopoiesis and increased myelopoiesis3,5,6. Transfer of bal-HSCs results in abundant lymphoid and myeloid cells, a stable phenotype that is retained after secondary transfer; my-HSCs also retain their patterns of production after secondary transfer5. The origin and potential interconversion of these two subsets is still unclear. If they are separate subsets postnatally, it might be possible to reverse the ageing phenotype by eliminating my-HSCs in aged mice. Here we demonstrate that antibody-mediated depletion of my-HSCs in aged mice restores characteristic features of a more youthful immune system, including increasing common lymphocyte progenitors, naive T cells and B cells, while decreasing age-related markers of immune decline. Depletion of my-HSCs in aged mice improves primary and secondary adaptive immune responses to viral infection. These findings may have relevance to the understanding and intervention of diseases exacerbated or caused by dominance of the haematopoietic system by my-HSCs.

IL-10 Negatively Controls the Primary T Cell Response of Tilapia by Triggering the JAK1/STAT3/SOCS3 Axis That Suppresses NF-κB and MAPK/ERK Signaling.

The braking mechanisms to protect the host from tissue damage and inflammatory disease caused by an overexuberant immune response are common in many T cell subsets. However, the negative regulation of T cell responses and detailed mechanisms are not well understood in early vertebrates. In the current study, using a Nile tilapia (Oreochromis niloticus) model, we investigated the suppression of T cell immunity by IL-10. Tilapia encodes an evolutionarily conserved IL-10, whose expression in lymphocytes is markedly induced during the primary adaptive immune response against Aeromonas hydrophila infection. Activated T cells of tilapia produce IL-10, which in turn inhibits proinflammatory cytokine expression and suppresses PHA-induced T cell activation. Moreover, administration of IL-10 impairs the proliferation of tilapia T cells, reduces their potential to differentiate into Th subsets, and cripples the cytotoxic function, rendering the animals more vulnerable to pathogen attack. After binding to its receptor IL-10Ra, IL-10 activates the JAK1/STAT3 axis by phosphorylation and enhances the expression of the suppressor of cytokine signaling 3 (SOCS3), which in turn attenuates the activation of the NF-κB and MAPK/ERK signaling pathways, thus suppressing the T cell response of tilapia. Our findings elucidate a negative regulatory mechanism of T cell immunity in a fish species and support the notion that the braking mechanism of T cells executed through IL-10 existed prior to the divergence of the tetrapod lineage from teleosts. Therefore, this study, to our knowledge, provides a novel perspective on the evolution of the adaptive immune system.

Comparative RNA-Sequencing Analysis Reveals High Complexity and Heterogeneity of Transcriptomic and Immune Profiles in Hepatocellular Carcinoma Tumors of Viral (HBV, HCV) and Non-Viral Etiology.

Background and Objectives: Hepatocellular carcinoma (HCC), the most common type of primary liver cancer, is the leading cause of cancer-related mortality. It arises and progresses against fibrotic or cirrhotic backgrounds mainly due to infection with hepatitis viruses B (HBV) or C (HCV) or non-viral causes that lead to chronic inflammation and genomic changes. A better understanding of molecular and immune mechanisms in HCC subtypes is needed. Materials and Methods: To identify transcriptional changes in primary HCC tumors with or without hepatitis viral etiology, we analyzed the transcriptomes of 24 patients by next-generation sequencing. Results: We identified common and unique differentially expressed genes for each etiological tumor group and analyzed the expression of SLC, ATP binding cassette, cytochrome 450, cancer testis, and heat shock protein genes. Metascape functional enrichment analysis showed mainly upregulated cell-cycle pathways in HBV and HCV and upregulated cell response to stress in non-viral infection. GeneWalk analysis identified regulator, hub, and moonlighting genes and highlighted CCNB1, ACTN2, BRCA1, IGF1, CDK1, AURKA, AURKB, and TOP2A in the HCV group and HSF1, HSPA1A, HSP90AA1, HSPB1, HSPA5, PTK2, and AURKB in the group without viral infection as hub genes. Immune infiltrate analysis showed that T cell, cytotoxic, and natural killer cell markers were significantly more highly expressed in HCV than in non-viral tumors. Genes associated with monocyte activation had the highest expression levels in HBV, while high expression of genes involved in primary adaptive immune response and complement receptor activity characterized tumors without viral infection. Conclusions: Our comprehensive study underlines the high degree of complexity of immune profiles in the analyzed groups, which adds to the heterogeneous HCC genomic landscape. The biomarkers identified in each HCC group might serve as therapeutic targets.

Interleukin-2 inducible T cell kinase (ITK) may participate in the anti-bacterial immune response of Nile tilapia via regulating T-cell activation

Interleukin-2 inducible T cell kinase (ITK) plays a predominant role in the T-cell receptor (TCR) signaling cascade to ensure valid T-cell activation and function. Nevertheless, whether it regulates T-cell response of early vertebrates remains unknown. Herein, we investigated the involvement of ITK in the lymphocyte-mediated adaptive immune response, and its regulation to T-cell activation in the Nile tilapia Oreochromis niloticus. Both sequence and structure of O. niloticus ITK (OnITK) were remarkably conserved with its homologues from other vertebrates, implying its potential conserved function. OnITK mRNA was extensively expressed in lymphoid-related tissues, and with the relative highest level in peripheral blood. Once Nile tilapia was infected by Edwardsiella piscicida, OnITK in splenic lymphocytes was significantly up-regulated on 7-day post infection at both transcription and translation levels, suggesting that OnITK might involve in the primary adaptive immune response of teleost. Furthermore, upon splenic lymphocytes were stimulated by T-cell specific mitogen PHA, OnITK mRNA and protein levels were dramatically elevated. More importantly, treatment of splenic lymphocytes with specific inhibitor significantly crippled OnITK expression, which in turn impaired the inducible expression of T-cell activation markers IFN-γ, IL-2 and CD122, indicating the critical roles of ITK in regulating T-cell activation of Nile tilapia. Taken together, our results suggest that ITK takes part in the lymphocyte-mediated adaptive immunity of tilapia, and is indispensable for T-cell activation of teleost. Our findings thus provide novel evidences for understanding the mechanism regulating T-cell immunity of early vertebrates, as well as the evolution of adaptive immune system.

Preliminary evaluation of selected inflammatory cytokine gene expression in lymphocytes isolated from whole human blood infected with trans-anethole-treated Staphylococcus aureus Newman strain.

In our previous study based on a whole-blood model of sepsis infected with trans-anethole (TA)-treated Staphylococcus aureus, we have found that innate immune response was more effective in comparison to non-treated cells. Due to the previous observation, in the current preliminary study, a primary adaptive immune response was analysed. This study was conducted to evaluate the expression of selected cytokine (IL1B, IL2, IL6, IL10, TNF, TGFB1, IFNG) and Toll-like receptor (TLR2) genes in lymphocytes isolated from whole human blood infected with S. aureus Newman strain treated with TA. The lymphocytes were isolated by density gradient centrifugation from blood samples infected with S. aureus, as well as from non-infected samples. Gene expression was measured using quantitative real-time PCR. The lymphocytes isolated from the blood infected with TA-treated staphylococcal cells demonstrated significantly greater IL10, IL1B, IL6, TNF and TLR2 expression. Hence, it is possible that the previously observed changes in the surface structure of TA-treated S. aureus Newman strain may significantly increase the relative expression of IL10, IL1B, IL6, TNF and TLR2 genes in lymphocytes; however, further studies are needed.

ZAP70 activation is an early event of T cell immunity that involved in the anti-bacterial adaptive immune response of Nile tilapia

ZAP70 is essential for initiating the early events of T-cell antigen receptor (TCR) signaling cascade to ensure proper T cell activation and function. However, whether this molecule takes part in the T cell immune response of early vertebrates remains unclear. In the present study, using a teleost model Nile tilapia (Oreochromis niloticus), we investigated the potential involvement of ZAP70 in the T cell activation and adaptive immunity of fish species. Both primary and tertiary structures of O. niloticus ZAP70 (On-ZAP70) are highly conserved with those from other vertebrates. On-ZAP70 protein was widely expressed in lymphoid tissues, and with the highest level in thymus. Once Nile tilapia was infected by Aeromonas hydrophila, mRNA of On-ZAP70 in spleen lymphocytes was induced on day 5 and 8 after infection; meanwhile, phosphorylation of On-ZAP70 was also enhanced, suggesting that On-ZAP70 potentially participated in primary adaptive immune response of Nile tilapia. Furthermore, the frequency of ZAP70 positive lymphocytes was increased during the anti-bacterial adaptive immune response. More importantly, when spleen lymphocytes were activated by T cell specific mitogen PHA, a dramatical augment of On-ZAP70 could be observed at transcription, phosphorylation and cellular level, indicating the involvement of this molecule in T cells activation of Nile tilapia. Altogether, our results demonstrated that ZAP70 activation is an early event of T cell immunity that involved in the anti-bacterial adaptive immune response of Nile tilapia, and thus provided a new evidence to understand the evolution of the lymphocyte-mediated adaptive immunity.

Macrophages Harbor Transcriptionally Active Defective HIV Provirus Which Contribute to Immune Activation

Abstract People living with HIV suffer from a variety of co-morbidities including neurological deficits and cardiovascular disease even while on antiretroviral therapy (ART). Chronic inflammation has been implicated in these co-morbidities and has long been described in HIV patients. Characterization of the HIV DNA (provirus) reservoir in CD4 T cells from HIV patients on ART has revealed that more than 90% are defective and unable to complete the viral life cycle. Macrophages are susceptible to HIV infection and are positioned at the nexus of pathogen detection, inflammation, and priming of adaptive immune responses. We hypothesized that HIV-infected macrophages harbor defective HIV proviruses which remain transcriptionally competent and affect adaptive immune responses. To test this, we examined the status of HIV proviruses in primary human monocyte-derive macrophages (MDMs) and their expression of HIV transcripts. MDMs were infected with HIV in vitro and using an intact provirus detection assay we observed that roughly half of HIV proviruses were defective. Reverse transcription droplet-digital PCR (RT-ddPCR) detected a distribution of HIV transcripts, many of which lack the full 5′ LTR sequence consistent with transcription starting from an intragenic site. When CD8 T cells were co-cultured with autologous HIV-infected MDMs, they increased their expression of CD107a, a degranulation marker, indicating that infected macrophages could trigger CD8+ activation. Together, these results support that alternative transcriptional mechanisms, which persist in the absence of a fully functional LTR, are active during HIV infection in macrophages, contributing to macrophage dysfunction and persistent HIV-associated diseases.

Berberine Delays Onset of Collagen-Induced Arthritis through T Cell Suppression.

There is evidence that berberine (BBR), a clinically relevant plant compound, ameliorates clinically apparent collagen-induced arthritis (CIA) in vivo. However, to date, there are no studies involving the use of BBR which explore its prophylactic potential in this model of rheumatoid arthritis (RA). The aim of this study was to determine if prophylactic BBR use during the preclinical phase of collagen-induced arthritis would delay arthritic symptom onset, and to characterize the cellular mechanism underlying such an effect. DBA/1J mice were injected with an emulsion of bovine type II collagen (CII) and complete Freund’s adjuvant (day 0) and a booster injection of CII in incomplete Freund’s adjuvant (day 18) to induce arthritis. Mice were then given i.p. injections of 1 mg/kg/day of BBR or PBS (vehicle with 0.01% DMSO) from days 0 to 28, were left untreated (CIA control), or were in a non-arthritic control group (n = 15 per group). Incidence of arthritis in BBR-treated mice was 50%, compared to 90% in both the CIA and PBS controls. Populations of B and T cells from the spleens and draining lymph nodes of mice were examined on day 14 (n = 5 per group) and day 28 (n = 10 per group). BBR-treated mice had significantly reduced populations of CD4+Th and CD4+CXCR5+ Tfh cells, and an increased proportion of Foxp3+ Treg at days 14 and 28, as well as reduced expression of co-stimulatory molecules CD28 and CD154 at both endpoints. The effect seen on T cell populations and co-stimulatory molecule expression in BBR-treated mice was not mirrored in CD19+ B cells. Additionally, BBR-treated mice experienced reduced anti-CII IgG2a and anti-CII total IgG serum concentrations. These results indicate a potential role for BBR as a prophylactic supplement for RA, and that its effect may be mediated specifically through T cell suppression. However, the cellular effector involved raises concern for BBR prophylactic use in the context of vaccine efficacy and other primary adaptive immune responses.

Fish NF‐κB couples TCR and IL‐17 signals to regulate ancestral T‐cell immune response against bacterial infection

As fish constitute the first evolutionary group with primordial T cells, they are of importance for understanding the origin and evolution of adaptive immunity. Yet, the knowledge about how ancestral T cells function remains limited. Therefore, the teleost model Nile tilapia (Oreochromis niloticus) was used in this study to investigate the regulatory mechanisms of T-cell immunity in fish. We identified an evolutionarily conserved canonical NF-κB signaling pathway in Nile tilapia, which participates in primary adaptive immune response during Streptococcus agalactiae infection. Blockade of NF-κB activity severely impairs T-cell activation and expansion, rendering the animals more vulnerable to pathogen attack. Meanwhile, NF-κB signaling is indispensable for fish T cells to produce IL-17A during the antibacterial immune response. Moreover, IL-17A binds its receptor IL-17RA, initiates the ACT1-TRAF6-TAK1 axis, and triggers NF-κB-dependent T-cell activation, thus forming a positive feedback loop of T-cell immunity in Nile tilapia. Furthermore, IL-17A seems to promote innate immunity by regulating pro-inflammatory cytokines via TRAF6-NF-κB axis, indicating the presence of an NF-κB-dependent IL-17A signaling pathway for coordinating adaptive and innate immunity in fish. Our results suggest that fish NF-κB couples TCR and IL-17 signals to modulate ancestral T-cell immunity against bacterial infection, and the regulation of T-cell immunity by NF-κB and IL-17 is a strategy that existed prior to the divergence of the tetrapod lineage from teleost fish. This study, therefore, provides a new perspective on the evolution of adaptive immunity.

Lymph node stromal cell subsets—Emerging specialists for tailored tissue-specific immune responses

The effective priming of adaptive immune responses depends on the precise dispatching of lymphocytes and antigens into and within lymph nodes (LNs), which are strategically dispersed throughout the body. Over the past decade, a growing body of evidence has advanced our understanding of lymph node stromal cells (LNSCs) from viewing them as mere accessory cells to seeing them as critical cellular players for the modulation of adaptive immune responses. In this review, we summarize current advances on the pivotal roles that LNSCs play in orchestrating adaptive immune responses during homeostasis and infection, and highlight the imprinting of location-specific information by micro-environmental cues into LNSCs, thereby tailoring tissue-specific immune responses.

Essential role of 4E-BP1 for lymphocyte activation and proliferation in the adaptive immune response of Nile tilapia

Eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) is a translation repressor downstream of mTORC1 pathway, and plays a pivotal role in the adaptive immune response. However, whether 4E-BP1 participates in the regulation of adaptive immunity of early vertebrates is still unclear. In present study, using Nile tilapia Oreochromis niloticus as a model, we investigated the regulatory roles of 4E-BP1 (On-4E-BP1) on lymphocyte-mediated adaptive immune response of fish species. On-4E-BP1 is highly conserved compared with the homologues from other vertebrates, and it is widely distributed in the immune-related tissues of Nile tilapia with the highest expression level in the gill. Once the animals were infected by Aeromonas hydrophila, transcription level of On-4E-BP1 in spleen lymphocytes was significantly induced on day 5 after infection. Meanwhile, phosphorylation of On-4E-BP1 in lymphocytes was also enhanced during the primary adaptive immune response, suggesting that 4E-BP1 is involved in the adaptive immunity of Nile tilapia. Furthermore, when lymphocytes were activated by agonist PMA or T cell specific mitogen PHA in vitro, phosphorylation of On-4E-BP1 was obviously up-regulated. More importantly, once mTORC1/4E-BP1 activity was blocked by specific inhibitor, the inducible expression of T cell activation markers IFN-γ and CD122 was severely impaired during PHA-induced T cell activation, suggesting this signaling regulates T lymphocyte activation of Nile tilapia. In addition, blockade of mTORC1/4E-BP1 axis also resulted in a crippled proliferation of lymphocyte during the primary response of anti-bacterial adaptive immunity. Collectively, we found that mTORC1/4E-BP1 signaling participates in the adaptive immune response by regulating lymphocyte activation and proliferation in Nile tilapia. This study enriches our current knowledge on teleost adaptive immunity, and provides novel perspective to understand the evolution of adaptive immune system.

Akt1/mTORC1 signaling modulates adaptive immune response of Nile tilapia by promoting lymphocyte activation and proliferation

Serving as a significant signaling molecule, RAC-alpha serine/threonine-protein kinase (Akt1) plays indispensable roles in cell cycle, growth, survival, metabolism, as well as immune response. However, how Akt1 regulates adaptive immune response in early vertebrate, especially the teleost, is largely unknown. Here, using a Nile tilapia Oreochromis niloticus model, we investigated the regulatory role of Akt1 in adaptive immunity of teleost. Both sequence and structure of the O. niloticus Akt1 (OnAkt1), were evolutionarily conserved comparing with the counterparts from other vertebrates. mRNA of OnAkt1 was widely expressed in lymphoid organs/tissues of Nile tilapia, with relative higher level in PBL. After Nile tilapia was infected by Aeromonas hydrophila, both transcription and phosphorylation levels of OnAkt1 were obviously elevated in spleen lymphocytes at the adaptive immune stage, suggesting Akt1 participated in primary adaptive immune response of Nile tilapia. Furthermore, OnAkt1 transcript or phosphorylation was dramatically augmented after spleen lymphocytes were activated by T cell specific mitogen PHA or lymphocyte agonist PMA. More critically, inhibition of Akt1 by specific inhibitor crippled the activation of downstream mTORC1 signaling, and impaired the up-regulation of T cell activation markers CD44, IFN-γ and CD122 in spleen lymphocytes upon PHA-induced T cell activation. Meanwhile, blockade of Akt1-activated mTORC1 signaling also decreased the frequency of BrdU+ lymphocytes during A. hydrophila infection, indicating the critical role of Akt1 in regulating lymphocyte proliferation of Nile tilapia. Together, our results demonstrated that Akt1 modulated adaptive immune response of Nile tilapia by promoting lymphocyte activation and proliferation via mTORC1 signaling. Our study enriched the regulatory mechanism of lymphocyte-mediated adaptive immunity in teleost, and thus provided novel insights into the evolution of adaptive immune system.

A New Bioassay Platform Design for the Discovery of Small Molecules with Anticancer Immunotherapeutic Activity.

Immunotherapy takes advantage of the immune system to prevent, control, and eliminate neoplastic cells. The research in the field has already led to major breakthroughs to treat cancer. In this work, we describe a platform that integrates in vitro bioassays to test the immune response and direct antitumor effects for the preclinical discovery of anticancer candidates. The platform relies on the use of dendritic cells that are professional antigen-presenting cells (APC) able to activate T cells and trigger a primary adaptive immune response. The experimental procedure is based on two phenotypic assays for the selection of chemical leads by both a panel of nine tumor cell lines and growth factor-dependent immature mouse dendritic cells (D1). The positive hits are then validated by a secondary test on human monocyte-derived dendritic cells (MoDCs). The aim of this approach is the selection of potential immunotherapeutic small molecules from natural extracts or chemical libraries.

The Role of Insulin Regulated Aminopeptidase in Endocytic Trafficking and Receptor Signaling in Immune Cells.

Insulin regulated aminopeptidase (IRAP) is a type II transmembrane protein with broad tissue distribution initially identified as a major component of Glut4 storage vesicles (GSV) in adipocytes. Despite its almost ubiquitous expression, IRAP had been extensively studied mainly in insulin responsive cells, such as adipocytes and muscle cells. In these cells, the enzyme displays a complex intracellular trafficking pattern regulated by insulin. Early studies using fusion proteins joining the IRAP cytosolic domain to various reporter proteins, such as GFP or the transferrin receptor (TfR), showed that the complex and regulated trafficking of the protein depends on its cytosolic domain. This domain contains several motifs involved in IRAP trafficking, as demonstrated by mutagenesis studies. Also, proteomic studies and yeast two-hybrid experiments showed that the IRAP cytosolic domain engages in multiple protein interactions with cytoskeleton components and vesicular trafficking adaptors. These findings led to the hypothesis that IRAP is not only a cargo of GSV but might be a part of the sorting machinery that controls GSV dynamics. Recent work in adipocytes, immune cells, and neurons confirmed this hypothesis and demonstrated that IRAP has a dual function. Its carboxy-terminal domain located inside endosomes is responsible for the aminopeptidase activity of the enzyme, while its amino-terminal domain located in the cytosol functions as an endosomal trafficking adaptor. In this review, we recapitulate the published protein interactions of IRAP and summarize the increasing body of evidence indicating that IRAP plays a role in intracellular trafficking of several proteins. We describe the impact of IRAP deletion or depletion on endocytic trafficking and the consequences on immune cell functions. These include the ability of dendritic cells to cross-present antigens and prime adaptive immune responses, as well as the control of innate and adaptive immune receptor signaling and modulation of inflammatory responses.

Targeting Inflammation and Immunosenescence to Improve Vaccine Responses in the Elderly

One of the most appreciated consequences of immunosenescence is an impaired response to vaccines with advanced age. While most studies report impaired antibody responses in older adults as a correlate of vaccine efficacy, it is now widely appreciated that this may fail to identify important changes occurring in the immune system with age that may affect vaccine efficacy. The impact of immunosenescence on vaccination goes beyond the defects on antibody responses as T cell-mediated responses are reshaped during aging and certainly affect vaccination. Likewise, age-related changes in the innate immune system may have important consequences on antigen presentation and priming of adaptive immune responses. Importantly, a low-level chronic inflammatory status known as inflammaging has been shown to inhibit immune responses to vaccination and pharmacological strategies aiming at blocking baseline inflammation can be potentially used to boost vaccine responses. Yet current strategies aiming at improving immunogenicity in the elderly have mainly focused on the use of adjuvants to promote local inflammation. More research is needed to understand the role of inflammation in vaccine responses and to reconcile these seemingly paradoxical observations. Alternative approaches to improve vaccine responses in the elderly include the use of higher vaccine doses or alternative routes of vaccination showing only limited benefits. This review will explore novel targets and potential new strategies for enhancing vaccine responses in older adults, including the use of anti-inflammatory drugs and immunomodulators.

Abstract 5602: BT-001, an oncolytic vaccinia virus armed with a Treg-depletion-optimized recombinant human anti-CTLA4 antibody and GM-CSF to target the tumor microenvironment

Abstract Oncolytic virotherapy is a clinically validated class of immune modulatory drugs with activity in solid cancer. The mode of action of armed oncolytic Vaccinia viruses (oVV) includes cell death induction following selective replication in tumor cells, local inflammation and priming of adaptive immune responses, and intra-tumoral delivery of therapeutic modalities e.g. antibodies, cytokines, enzymes and engineered receptor ligands. BioInvent and Transgene are collaborating in the development of a next generation armed oncolytic viruses, bringing respectively their n-CoDeR F.I.R.S.T antibody (mAb) discovery and Invir.IO oVV platforms. The first product, named BT-001, consists of the Copenhagen oVV strain deleted in J2R and I4L viral genes allowing restricted replication in proliferating cells, and the human IgG1 4-E03 recognizing human CTLA4. 4-E03 shows improved Treg-depleting activity compared with ipilimumab. BT-001 also encodes GM-CSF, the cytokine expressed in clinically advanced products like T-Vec (Imlygic) or Pexa-Vec. A surrogate mAb with similar pronounced Treg-depleting properties to 4-E03 was vectorized into the same oVV (mBT-1) allowing for functional and mechanistic in vivo studies. Since virus replication and expression of recombinant transgenes are tightly linked, the armed oVV is an interesting design for intra-tumoral delivery of therapeutic proteins exhibiting toxicological limitation (e.g. anti-CTLA4) and/or pharmacokinetic issues (e.g. GM-CSF). Oncoviral tumor-localized delivery of anti-CTLA4 may also allow better tolerated and more effective combination therapy with approved antibodies targeting the PD-1/PDL1 axis. Our studies demonstrate that 4-E03 and GM-CSF were expressed at high level as functional molecules after infection by BT-001 of several human tumoral cell lines in vitro. Moreover, following intratumoral administration in immune competent and immune deficient mice transplanted with mouse or human tumors, transgene expression was sustained at levels associated with receptor saturation for days to weeks. In contrast, and supporting the tumor-selective nature of oVV, blood concentrations of anti-CTLA4 mAb were lower compared to those observed following I.V. administration of therapeutic doses (3 mg/kg) of mAb. The in vivo anti-tumor activity of mBT-1 was assessed in multiple syngeneic mouse tumor models including CT26, EMT6, A20, and C38. mBT-1 conferred cures in the majority of challenged mice irrespective of tumor origin. The excellent anti-tumoral profile depends on anti-CTLA4 expression and could be boosted by co-administration of anti-PD-1 mAb. A clinical batch of BT-001 has been produced and toxicological evaluation is ongoing. Transgene and BioInvent are preparing CTA and IND filings for a multicentre clinical trial targeting injectable superficial tumors. Citation Format: Jean-Baptiste Marchand, Monika Semmrich, Laetitia Fend, Matilda Rehn, Nathalie Silvestre, Ingrid Teige, Johann Foloppe, Linda Mårtensson, Eric Quéméneur, Björn Frendeus. BT-001, an oncolytic vaccinia virus armed with a Treg-depletion-optimized recombinant human anti-CTLA4 antibody and GM-CSF to target the tumor microenvironment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5602.

Early IFN-I limits virus-specific immunity

Abstract Type I interferons (IFN-I) play a major role in the innate immune response. These cytokines are induced within hours of infection, conferring rapid protection before the onset of adaptive immunity. IFN-I are also critical for the priming of adaptive immune responses, but it is not clear if early antiviral restriction by IFN-I can also limit the amount of antigen available to prime the adaptive immune system. Here we demonstrate that reinforcement of IFN-I signaling early after viral immunization limits the priming of adaptive immunity, whereas blocking IFN-I signaling improves adaptive immunity and enhances subsequent host protection following viral re-infection. Paradoxically, short-term blockade of IFN-I improved the efficacy of clinically approved vaccines, such as the yellow fever virus vaccine (YFV-17D) and the smallpox virus vaccine, as well as experimental HIV vaccines in mice. Short-term blockade of IFN-I also improved immune responses following acute viral infections, such as those with Zika virus, lymphocytic choriomeningitis virus (LCMV), and vesicular stomatitis virus (VSV). This unexpected adjuvant effect of blocking early IFN-I responses was associated with a hyperacute increase in antigen availability, which could not be recapitulated by increasing the dose of the virus inoculum. The improvement of virus-specific immunity following acute IFN-I blockade was mechanistically due to an increase in antigen presentation and costimulation. These findings warrant a re-evaluation of the immunoregulatory roles of IFN-I during immune memory differentiation, which may provide a framework for rational vaccine design.