The vaccinia-based Sementis Copenhagen Vector coronavirus disease 2019 vaccine induces broad and durable cellular and humoral immune responses.

The ongoing coronavirus disease 2019 (COVID-19) pandemic perpetuated by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants has highlighted the continued need for broadly protective vaccines that elicit robust and durable protection. Here, the vaccinia virus-based, replication-defective Sementis Copenhagen Vector (SCV) was used to develop a first-generation COVID-19 vaccine encoding the spike glycoprotein (SCV-S). Vaccination of mice rapidly induced polyfunctional CD8 T cells with cytotoxic activity and robust type 1 T helper-biased, spike-specific antibodies, which are significantly increased following a second vaccination, and contained neutralizing activity against the alpha and beta variants of concern. Longitudinal studies indicated that neutralizing antibody activity was maintained up to 9 months after vaccination in both young and middle-aged mice, with durable immune memory evident even in the presence of pre-existing vector immunity. Therefore, SCV-S vaccination has a positive immunogenicity profile, with potential to expand protection generated by current vaccines in a heterologous boost format and presents a solid basis for second-generation SCV-based COVID-19 vaccine candidates incorporating additional SARS-CoV-2 immunogens.

Toll-Like Receptor-4 Antagonist (+)-Naltrexone Protects Against Carbamyl-Platelet Activating Factor (cPAF)-Induced Preterm Labor in Mice.

Spontaneous preterm labor is frequently caused by an inflammatory response in the gestational tissues elicited by either infectious or sterile agents. In sterile preterm labor, the key regulators of inflammation are not identified, but platelet-activating factor (PAF) is implicated as a potential rate-limiting effector agent. Since Toll-like receptor (TLR)-4 can amplify PAF signaling, we evaluated whether TLR4 contributes to inflammation and fetal loss in a mouse model of PAF-induced sterile preterm labor, and whether a small-molecule TLR4 inhibitor, (+)-naltrexone, can mitigate adverse PAF-induced effects. The administration of carbamyl (c)-PAF caused preterm labor and fetal loss in wild-type mice but not in TLR4-deficient mice. Treatment with (+)-naltrexone prevented preterm delivery and alleviated fetal demise in utero elicited after cPAF administered by i.p. or intrauterine routes. Pups born after cPAF and (+)-naltrexone treatment exhibited comparable rates of postnatal survival and growth to carrier-treated controls. (+)-Naltrexone suppressed the cPAF-induced expression of inflammatory cytokine genes Il1b, Il6, and Il10 in the decidua; Il6, Il12b, and Il10 in the myometrium; and Il1b and Il6 in the placenta. These data demonstrate that the TLR4 antagonist (+)-naltrexone inhibits the inflammatory cascade induced by cPAF, preventing preterm birth and perinatal death. The inhibition of TLR4 signaling warrants further investigation as a candidate strategy for fetal protection and delay of preterm birth elicited by sterile stimuli.

Production of a Chikungunya Vaccine Using a CHO Cell and Attenuated Viral-Based Platform Technology.

Vaccinia-based systems have been extensively explored for the development of recombinant vaccines. Herein we describe an innovative vaccinia virus (VACV)-derived vaccine platform technology termed Sementis Copenhagen Vector (SCV), which was rendered multiplication-defective by targeted deletion of the essential viral assembly gene D13L. A SCV cell substrate line was developed for SCV vaccine production by engineering CHO cells to express D13 and the VACV host-range factor CP77, because CHO cells are routinely used for manufacture of biologics. To illustrate the utility of the platform technology, a SCV vaccine against chikungunya virus (SCV-CHIK) was developed and shown to be multiplication-defective in a range of human cell lines and in immunocompromised mice. A single vaccination of mice with SCV-CHIK induced antibody responses specific for chikungunya virus (CHIKV) that were similar to those raised following vaccination with a replication-competent VACV-CHIK and able to neutralize CHIKV. Vaccination also provided protection against CHIKV challenge, preventing both viremia and arthritis. Moreover, SCV retained capacity as an effective mouse smallpox vaccine. In summary, SCV represents a new and safe vaccine platform technology that can be manufactured in modified CHO cells, with pre-clinical evaluation illustrating utility for CHIKV vaccine design and construction.

An immunogenic phenotype in paternal antigen-specific CD8+ T cells at embryo implantation elicits later fetal loss in mice.

Central to pregnancy success is a state of T cell tolerance to paternal antigens, which is initiated at conception. The role and regulation of specific phenotypes of CD8+ T cells in mediating pregnancy tolerance is not clear. This study aimed to investigate the impact on pregnancy outcome of altering the cytokine environment during maternal CD8+ T cell priming in early pregnancy. Transgenic Act-mOVA male mice were mated to C57BL/6 (B6) females to generate fetuses expressing ovalbumin (OVA) as a model paternal antigen. OVA-reactive CD8+ OT-I T cells were activated in vitro with OVA in the presence of either transforming growth factor-ß1 (TGFB1) plus interleukin-10 (IL10), or IL2, to mimic normal or dysregulated uterine conditions, respectively, and transferred into pregnant mice on gestational day 3.5. OT-I T cells activated with TGFB1 and IL10, like naive OT-I T cells, did not alter embryo implantation or fetal viability. In contrast, OT-I T cells activated with IL2 caused extensive fetal loss manifesting in mid-gestation. IL2-activated OT-I T cells expressed less FOXP3 and higher interferon-γ (IFNG) than cells activated with TGFB1 and IL10. Fetal loss did not occur in females mated with B6 males, demonstrating the antigen specificity of fetal loss, and was not abrogated by maternal genetic C1q deficiency indicating a mechanism independent of antibody-mediated cytotoxicity. These data indicate that alternative phenotypes generated in maternal CD8+ T cells at the time of priming with paternal antigens can impact pregnancy outcome, such that inappropriate activation of CD8+ T cells before implantation is capable of causing antigen-specific fetal loss later in pregnancy.

Evaluation of trained immunity by ß-1, 3 (d)-glucan on murine monocytes in vitro and duration of response in vivo.

The ß-1, 3 (d)-glucan (ß-glucan) present in the cell wall of Candida albicans induces epigenetic changes in human monocytes resulting in primed macrophages exhibiting increased cytokine responsiveness to reinfection. This phenomenon is referred to as trained immunity or innate immune memory. However, whether ß-glucan can reprogramme murine monocytes in vitro or induce lasting effects in vivo has yet to be elucidated. Thus, purified murine spleen-derived monocytes were primed with ß-glucan in vitro and assessed for markers of differentiation and survival. Important macrophage cell markers during monocyte-to-macrophage differentiation were downregulated and survival enhanced due to partial inhibition of apoptosis. Increased survival and not the ß-glucan training effect explained the elevated production of tumour necrosis factor-α (TNFα) and interleukin-6 (IL-6) induced by subsequent lipopolysaccharide (LPS) challenge. In vivo, 4 days after systemic administration of ß-glucan, mice were more responsive to LPS challenge as shown by the increased serum levels of TNFα, IL-6 and IL-10, an effect shown to be short lived as enhanced cytokine production was lost by day 20. Here, we have characterised murine macrophages derived from ß-glucan-primed monocytes based on their surface marker expression and for the first time provide evidence that the training effect of ß-glucan in vivo declines within a 3-week period.

The majority of murine γδ T cells at the maternal-fetal interface in pregnancy produce IL-17.

Compared with lymphoid tissues, the immune cell compartment at mucosal sites is enriched with T cells bearing the γδ T-cell receptor (TCR). The female reproductive tract, along with the placenta and uterine decidua during pregnancy, are populated by γδ T cells predominantly expressing the invariant Vγ6(+)Vδ1(+) receptor. Surprisingly little is understood about the function of these cells. We found that the majority of γδ T cells in the non-pregnant uterus, pregnant uterus, decidua and placenta of mice express the transcription factor RORγt and produce interleukin-17 (IL-17). In contrast, IFNγ-producing γδ T cells were markedly reduced in gestational tissues compared with uterine-draining lymph nodes and spleen. Both uterine-resident invariant Vγ6(+) and Vγ4(+) γδ T cells which are more typically found in lymphoid tissues and circulating blood, were found to express IL-17. Vγ4(+) γδ T cells were particularly enriched in the placenta, suggesting a pregnancy-specific recruitment or expansion of these cells. A small increase in IL-17-producing γδ T cells was observed in allogeneic compared with syngeneic pregnancy, suggesting a contribution to regulating the maternal response to paternally-derived alloantigens. However, their high proportions also in non-pregnant uteri and gestational tissues of syngeneic pregnancy imply a role in the prevention of intrauterine infection or quality control of fetal development. These data suggest the need for a more rigorous evaluation of the role of IL-17 in sustaining normal pregnancy, particularly as emerging data points to a pathogenic role for IL-17 in pre-eclampsia, pre-term birth, miscarriage and maternal immune activation-induced behavioral abnormalities in offspring.

The functional contribution of calcium ion flux heterogeneity in T cells.

The role of intracellular calcium ion oscillations in T-cell physiology is being increasingly appreciated by studies that describe how unique temporal and spatial calcium ion signatures can control different signalling pathways. Within this review, we provide detailed mechanisms of calcium ion oscillations, and emphasise the pivotal role that calcium signalling plays in directing crucial events pertaining to T-cell functionality. We also describe methods of calcium ion quantification, and take the opportunity to discuss how a deeper understanding of calcium signalling combined with new detection and quantification methodologies can be used to better design immunotherapies targeting T-cell responses.

Alcohol-induced sedation and synergistic interactions between alcohol and morphine: a key mechanistic role for Toll-like receptors and MyD88-dependent signaling.

Increasing evidence demonstrates induction of proinflammatory Toll-like receptor (TLR) 2 and TLR4 signaling by morphine and, TLR4 signaling by alcohol; thus indicating a common site of drug action and a potential novel innate immune-dependent hypothesis for opioid and alcohol drug interactions. Hence, the current study aimed to assess the role of TLR2, TLR4, MyD88 (as a critical TLR-signaling participant), NF-κB, Interleukin-1ß (IL-1ß; as a downstream proinflammatory effector molecule) and the µ opioid receptor (MOR; as a classical site for morphine action) in acute alcohol-induced sedation (4.5g/kg) and alcohol (2.5g/kg) interaction with morphine (5mg/kg) by assessing the loss of righting reflex (LORR) as a measure of sedation. Wild-type male Balb/c mice and matched genetically-deficient TLR2, TLR4, and MyD88 strains were utilized, together with pharmacological manipulation of MOR, NF-κB, TLR4 and Interleukin-1ß. Alcohol induced significant LORR in wild-type mice; this was halved by MyD88 and TLR4 deficiency, and surprisingly nearly completely eliminated by TLR2 deficiency. In contrast, the interaction between morphine and alcohol was found to be MOR-, NF-κB-, TLR2- and MyD88-dependent, but did not involve TLR4 or Interleukin-1ß. Morphine-alcohol interactions caused acute elevations in microglial cell counts and NF-κB-p65 positive cells in the motor cortex in concordance with wild-type and TLR2 deficient mouse behavioral data, implicating neuroimmunopharmacological signaling as a pivotal mechanism in this clinically problematic drug-drug interaction.

Toll-like receptor 7 gene deficiency and early-life Pneumovirus infection interact to predispose toward the development of asthma-like pathology in mice.

BACKGROUND: Respiratory tract viruses are a major environmental risk factor for both the inception and exacerbations of asthma. Genetic defects in Toll-like receptor (TLR) 7-mediated signaling, impaired type I interferon responses, or both have been reported in asthmatic patients, although their contribution to the onset and exacerbation of asthma remains poorly understood. OBJECTIVE: We sought to determine whether Pneumovirus infection in the absence of TLR7 predisposes to bronchiolitis and the inception of asthma. METHODS: Wild-type and TLR7-deficient (TLR7(-/-)) mice were inoculated with the rodent-specific pathogen pneumonia virus of mice at 1 (primary), 7 (secondary), and 13 (tertiary) weeks of age, and pathologic features of bronchiolitis or asthma were assessed. In some experiments infected mice were exposed to low-dose cockroach antigen. RESULTS: TLR7 deficiency increased viral load in the airway epithelium, which became sloughed and necrotic, and promoted an IFN-α/ß(low), IL-12p70(low), IL-1ß(high), IL-25(high), and IL-33(high) cytokine microenvironment that was associated with the recruitment of type 2 innate lymphoid cells/nuocytes and increased TH2-type cytokine production. Viral challenge of TLR7(-/-) mice induced all of the cardinal pathophysiologic features of asthma, including tissue eosinophilia, mast cell hyperplasia, IgE production, airway smooth muscle alterations, and airways hyperreactivity in a memory CD4(+) T cell-dependent manner. Importantly, infections with pneumonia virus of mice promoted allergic sensitization to inhaled cockroach antigen in the absence but not the presence of TLR7. CONCLUSION: TLR7 gene defects and Pneumovirus infection interact to establish an aberrant adaptive response that might underlie virus-induced asthma exacerbations in later life.

The multifunctional alarmin HMGB1 with roles in the pathophysiology of sepsis and cancer.

Although originally described as a highly conserved nuclear protein involved in DNA replication, transcription and repair, high-mobility group box-1 protein (HMGB1) has emerged as a key mediator in the regulation of immune responses to infection and sterile injury by exhibiting all the properties of a prototypic 'alarmin'. These include rapid passive release in response to pathogenic infection and/or traumatic injury, active secretion providing for chemotactic and cytokine-like function and an ability to resolve inflammation, including tissue repair and remodelling. In this review, we will give an overview of the post-translational modifications necessary for such diversity in biological activity, concentrating particularly on how differences in oxidation of highly conserved redox-sensitive cysteine residues can potentiate inflammatory responses and dictate cellular fate. We will also review the most recent literature on HMGB1 and its involvement in the pathophysiology of sepsis and cancer, as well as cancer therapy-induced mucositis.

Antigen-specific T-cell responses to a recombinant fowlpox virus are dependent on MyD88 and interleukin-18 and independent of Toll-like receptor 7 (TLR7)- and TLR9-mediated innate immune recognition.

Fowlpox virus (FWPV) is a double-stranded DNA virus long used as a live-attenuated vaccine against poultry diseases, but more recent interest has focused on its use as a mammalian vaccine vector. Here, in a mouse model system using FWPV encoding the nominal target antigen chicken ovalbumin (OVA) (FWPV(OVA)), we describe for the first time some of the fundamental processes by which FWPV engages both the innate and adaptive immune systems. We show that Toll-like receptor 7 (TLR7) and TLR9 are important for type I interferon secretion by dendritic cells, while TLR9 is solely required for proinflammatory cytokine secretion. Despite this functional role for TLR7 and TLR9 in vitro, only the adapter protein myeloid differentiation primary response gene 88 (MyD88) was shown to be essential for the formation of adaptive immunity to FWPV(OVA) in vivo. The dependence on MyD88 was confined only to the T-cell compartment and was not related to its contribution to TLR signaling, dendritic cell maturation, or the capture and presentation of FWPV-derived OVA antigen. We demonstrate that this is not by means of mediating T-cell-dependent interleukin-1 (IL-1) signaling, but rather, we suggest that MyD88 functions to support T-cell-specific IL-18 receptor signaling, which in turn is essential for the formation of adaptive immunity to FWPV-encoded OVA.

Immobilized streptavidin gradients as bioconjugation platforms.

Surface density gradients of streptavidin (SAV) were created on solid surfaces and demonstrated functionality as a bioconjugation platform. The surface density of immobilized streptavidin steadily increased in one dimension from 0 to 235 ng cm(-2) over a distance of 10 mm. The density of coupled protein was controlled by its immobilization onto a polymer surface bearing a gradient of aldehyde group density, onto which SAV was covalently linked using spontaneous imine bond formation between surface aldehyde functional groups and primary amine groups on the protein. As a control, human serum albumin was immobilized in the same manner. The gradient density of aldehyde groups was created using a method of simultaneous plasma copolymerization of ethanol and propionaldehyde. Control over the surface density of aldehyde groups was achieved by manipulating the flow rates of these vapors while moving a mask across substrates during plasma discharge. Immobilized SAV was able to bind biotinylated probes, indicating that the protein retained its functionality after being immobilized. This plasma polymerization technique conveniently allows virtually any substrate to be equipped with tunable protein gradients and provides a widely applicable method for bioconjugation to study effects arising from controllable surface densities of proteins.

Type I interferons mediate the innate cytokine response to recombinant fowlpox virus but not the induction of plasmacytoid dendritic cell-dependent adaptive immunity.

Type I interferons (IFNs) are considered to be important mediators of innate immunity due to their inherent antiviral activity, ability to drive the transcription of a number of genes involved in viral clearance, and their role in the initiation of innate and adaptive immune responses. Due to the central role of type I IFNs, we sought to determine their importance in the generation of immunity to a recombinant vaccine vector fowlpox virus (FPV). In analyzing the role of type I IFNs in immunity to FPV, we show that they are critical to the secretion of a number of innate and proinflammatory cytokines, including type I IFNs themselves as well as interleukin-12 (IL-12), tumor necrosis factor-alpha (TNF-alpha), IL-6, and IL-1beta, and that deficiency leads to enhanced virus-mediated antigen expression. Interestingly, however, type I IFNs were not required for adaptive immune responses to recombinant FPV even though plasmacytoid dendritic cells (pDCs), the primary producers of type I IFNs, have been shown to be requisite for this to occur. Furthermore, we provide evidence that the importance of pDCs may lie in their ability to capture and present virally derived antigen to T cells rather than in their capacity as professional type I IFN-producing cells.

Cross-presentation of male seminal fluid antigens elicits T cell activation to initiate the female immune response to pregnancy.

The events that generate T cell-mediated immune tolerance in early pregnancy are ill-defined. To investigate the significance of seminal fluid Ags in activating maternal T cells, and define the underlying Ag presentation pathways, OVA-specific T cells were adoptively transferred to female mice inseminated by males ubiquitously expressing membrane-bound OVA. OVA-reactive CD8(+) OT-I and CD4(+) OT-II T cells transferred to mated recipients expressed activation markers CD25 and CD69 and proliferated vigorously in the para-aortic lymph nodes, but not in distal lymph nodes or spleen, and OT-I T cells expressed IFN-gamma and IL-2. In contrast, OT-I T cells transferred later in pregnancy or up to 10 days postpartum expressed CD25 and CD69 and proliferated in all peripheral lymphoid tissues examined. OVA Ag was present predominantly in the plasma fraction of seminal fluid, and seminal plasma, but not sperm, was necessary for T cell proliferation. Female H-2K(b) bone marrow-derived cells expressing TAP were essential for OT-I T cell proliferation, but responses were not elicited by OVA Ag presented by paternal MHC in seminal fluid or associated with placental cells. This study shows that at conception, seminal fluid drives activation and expansion of paternal Ag-reactive CD4(+) and CD8(+) T cell populations, and female APCs have an essential role in cross-presenting Ag to CD8(+) T cells via a TAP-dependent pathway. Delivery of paternal Ags and immune-deviating cytokines by seminal fluid at conception may activate Ag-dependent CD4(+) and CD8(+) regulatory T cells mediating tolerance of pregnancy.

Regulatory B Cells: Dark Horse in Pregnancy Immunotherapy?

There are many unanswered questions surrounding the function of immune cells and how they interact with the reproductive system to support successful pregnancy or contribute to pregnancy pathologies. While the role of immune cells such as uterine natural killer and dendritic cells, and more recently regulatory T cells has been established, the role of another major immune cell population, the B cell, and particularly the regulatory B cells, is relatively poorly understood. This review outlines what is known about B-cell subsets in the context of pregnancy, what constitutes a regulatory B cell and what role they may play, particularly during early pregnancy. Lastly, we discuss why immunotherapies for the treatment of pregnancy disorders is not widely progressed clinically and speculate on the potential of functional regulatory B cells as the basis of novel immunotherapeutic approaches for the treatment of immune-based pregnancy pathologies.

Macrophages exert homeostatic actions in pregnancy to protect against preterm birth and fetal inflammatory injury.

Macrophages are commonly thought to contribute to the pathophysiology of preterm labor by amplifying inflammation - but a protective role has not previously been considered to our knowledge. We hypothesized that given their antiinflammatory capability in early pregnancy, macrophages exert essential roles in maintenance of late gestation and that insufficient macrophages may predispose individuals to spontaneous preterm labor and adverse neonatal outcomes. Here, we showed that women with spontaneous preterm birth had reduced CD209+CD206+ expression in alternatively activated CD45+CD14+ICAM3- macrophages and increased TNF expression in proinflammatory CD45+CD14+CD80+HLA-DR+ macrophages in the uterine decidua at the materno-fetal interface. In Cd11bDTR/DTR mice, depletion of maternal CD11b+ myeloid cells caused preterm birth, neonatal death, and postnatal growth impairment, accompanied by uterine cytokine and leukocyte changes indicative of a proinflammatory response, while adoptive transfer of WT macrophages prevented preterm birth and partially rescued neonatal loss. In a model of intra-amniotic inflammation-induced preterm birth, macrophages polarized in vitro to an M2 phenotype showed superior capacity over nonpolarized macrophages to reduce uterine and fetal inflammation, prevent preterm birth, and improve neonatal survival. We conclude that macrophages exert a critical homeostatic regulatory role in late gestation and are implicated as a determinant of susceptibility to spontaneous preterm birth and fetal inflammatory injury.

Polyinosinic: Polycytidylic Acid and Murine Cytomegalovirus Modulate Expression of Murine IL-10 and IL-21 in White Adipose Tissue.

White adipose tissue (WAT) produces interleukin-10 and other immune suppressors in response to pathogen-associated molecular patterns (PAMPs). It also homes a subset of B-cells specialized in the production of IL-10, referred to as regulatory B-cells. We investigated whether viral stimuli, polyinosinic: polycytidylic acid (poly(I:C)) or whole replicative murine cytomegalovirus (MCMV), could stimulate the expression of IL-10 in murine WAT using in vivo and ex vivo approaches. Our results showed that in vivo responses to systemic administration of poly(I:C) resulted in high levels of endogenously-produced IL-10 and IL-21 in WAT. In ex vivo WAT explants, a subset of B-cells increased their endogenous IL-10 expression in response to poly(I:C). Finally, MCMV replication in WAT explants resulted in decreased IL-10 levels, opposite to the effect seen with poly(I:C). Moreover, downregulation of IL-10 correlated with relatively lower number of Bregs. To our knowledge, this is the first report of IL-10 expression by WAT and WAT-associated B-cells in response to viral stimuli.

Maternal host responses to poly(I:C) during pregnancy leads to both dysfunctional immune profiles and altered behaviour in the offspring.

PROBLEM: Autism spectrum disorder (ASD)-like phenotypes in murine models are linked to elevated pro-inflammatory cytokine profiles caused by maternal immune activation (MIA), but whether MIA alters the immune response in the offspring remains unclear. METHOD OF STUDY: Polyinosinic:polycytidylic acid (poly:[IC]) was used to induce MIA in immunocompetent and control TLR3-deficient pregnant mice, and cytokine levels were measured in maternal and foetal organs. Furthermore, cytokines and behaviour responses were tested after challenge with lipopolysaccharide in 7-day-old and adult mice. RESULTS: MIA induced on E12 resulted in changes in the cytokine expression profile in maternal and foetal organs and correlated with TNFα and IL-18 dysregulation in immune organs and brains from neonatal mice born to MIA-induced dams. Such changes further correlated with altered behavioural responses in adulthood. CONCLUSION: MIA induced by pathogens during pregnancy can interfere with the development of the foetal immune and nervous systems leading to dysfunctional immune responses and behaviour in offspring.

The vaccinia virus based Sementis Copenhagen Vector vaccine against Zika and chikungunya is immunogenic in non-human primates.

The Sementis Copenhagen Vector (SCV) is a new vaccinia virus-derived, multiplication-defective, vaccine technology assessed herein in non-human primates. Indian rhesus macaques (Macaca mulatta) were vaccinated with a multi-pathogen recombinant SCV vaccine encoding the structural polyproteins of both Zika virus (ZIKV) and chikungunya virus (CHIKV). After one vaccination, neutralising antibody responses to ZIKV and four strains of CHIKV, representative of distinct viral genotypes, were generated. A second vaccination resulted in significant boosting of neutralising antibody responses to ZIKV and CHIKV. Following challenge with ZIKV, SCV-ZIKA/CHIK-vaccinated animals showed significant reductions in viremias compared with animals that had received a control SCV vaccine. Two SCV vaccinations also generated neutralising and IgG ELISA antibody responses to vaccinia virus. These results demonstrate effective induction of immunity in non-human primates by a recombinant SCV vaccine and illustrates the utility of SCV as a multi-disease vaccine platform capable of delivering multiple large immunogens.

Transforming growth factor-beta-mediated signaling in T lymphocytes impacts on prostate-specific immunity and early prostate tumor progression.

T cells are in general tolerant of prostate-specific tumor antigens. That prostate tumor tissue makes transforming growth factor-beta (TGFbeta) is thought to play a role in the induction of T-cell tolerance within the host and to contribute to tumor progression itself. Here we sought to investigate the influence of TGFbeta signaling on prostate antigen-specific T-cell responses as well as prostate tumorogenesis in an autochthonous murine model of the disease. The response of naive and activated ovalbumin (OVA) antigen-specific T cells, which had been rendered incapable of responding to TGFbeta through T-cell-specific transgenic expression of a dominant-negative variant of the TGFbeta receptor II (dnTGFRII), was analyzed after adoptive transfer into prostate OVA-expressing transgenic (POET) mice. The role of TGFbeta signaling in endogenous T cells in mice, which spontaneously form tumors, was also assessed by monitoring prostate tumor formation and progression in F1 progeny of productive matings between transgenic adenocarcinoma of the mouse prostate (TRAMP) and dnTGFRII mice. TGFbeta-resistant CD8(+) T cells proliferated more and produced IFNgamma more readily after OVA stimulation in vitro. OVA-specific T cells did not damage the prostate gland of POET mice irrespective of TGFbeta responsiveness. However, ex vivo activation facilitated entry of TGFbeta-insensitive T cells into the prostate and was associated with prostate tissue damage. Early tumor progression was delayed in TRAMP mice that carried endogenous TGFbeta-insensitive T cells. Together, these results suggest that TGFbeta-signaling represses CD8(+) T-cell responses to a prostate-specific antigen. TGFbeta-mediated repression of T-cell function may include production of IFNgamma, which is known to contribute to tumor immunosurveillance.