Prostaglandin E2 induction by cytosolic Listeria monocytogenes in phagocytes is necessary for optimal T-cell priming

Listeria monocytogenes is an intracellular bacterium that elicits robust CD8 + T-cell responses. Despite the ongoing development of L. monocytogenes -based platforms as cancer vaccines, our understanding of how L. monocytogenes drives robust CD8 + T-cell responses remains incomplete. One overarching hypothesis is that activation of cytosolic innate pathways is critical for immunity, as strains of L. monocytogenes that are unable to access the cytosol fail to elicit robust CD8 + T-cell responses and in fact inhibit optimal T-cell priming. Counterintuitively, however, activation of known cytosolic pathways, such as the inflammasome and type I IFN, lead to impaired immunity. Here, we describe a cytosol-dependent response that is critical for immunity to L. monocytogenes , namely production of prostaglandin E 2 (PGE 2 ) downstream of cyclooxygenase-2 (COX-2). Vacuole-constrained L. monocytogenes elicit reduced PGE 2 production compared to wild-type strains in macrophages and dendritic cells ex vivo . In vivo, infection with wild-type L. monocytogenes leads to 10-fold increases in PGE 2 production early during infection whereas vacuole-constrained strains fail to induce PGE 2 over mock-immunized controls. Mice deficient in COX-2 specifically in Lyz2 + or CD11c + cells produce less PGE 2 , suggesting these cell subsets contribute to PGE 2 levels in vivo, while depletion of phagocytes with clodronate abolishes PGE 2 production completely . Taken together, this work identifies the first known cytosol-dependent innate immune response critical for generating CD8 + T-cell responses to L. monocytogenes, suggesting that one reason cytosolic access is required to prime CD8 + T-cell responses may be due to induction of PGE 2 .

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Phagocytes produce prostaglandin E2 in response to cytosolic Listeria monocytogenes

PLoS Pathogens ◽

10.1371/journal.ppat.1009493 ◽

2021 ◽

Vol 17 (9) ◽

pp. e1009493

Author(s):

Courtney E. McDougal ◽

Zachary T. Morrow ◽

Tighe Christopher ◽

Seonyoung Kim ◽

Drake Carter ◽

...

Keyword(s):

Listeria Monocytogenes ◽

T Cell ◽

T Cell Responses ◽

Prostaglandin E ◽

Type I ◽

Wild Type ◽

Cell Responses ◽

T Cell Priming ◽

Cox 2

Listeria monocytogenes is an intracellular bacterium that elicits robust CD8+ T-cell responses. Despite the ongoing development of L. monocytogenes-based platforms as cancer vaccines, our understanding of how L. monocytogenes drives robust CD8+ T-cell responses remains incomplete. One overarching hypothesis is that activation of cytosolic innate pathways is critical for immunity, as strains of L. monocytogenes that are unable to access the cytosol fail to elicit robust CD8+ T-cell responses and in fact inhibit optimal T-cell priming. Counterintuitively, however, activation of known cytosolic pathways, such as the inflammasome and type I IFN, lead to impaired immunity. Conversely, production of prostaglandin E2 (PGE2) downstream of cyclooxygenase-2 (COX-2) is essential for optimal L. monocytogenes T-cell priming. Here, we demonstrate that vacuole-constrained L. monocytogenes elicit reduced PGE2 production compared to wild-type strains in macrophages and dendritic cells ex vivo. In vivo, infection with wild-type L. monocytogenes leads to 10-fold increases in PGE2 production early during infection whereas vacuole-constrained strains fail to induce PGE2 over mock-immunized controls. Mice deficient in COX-2 specifically in Lyz2+ or CD11c+ cells produce less PGE2, suggesting these cell subsets contribute to PGE2 levels in vivo, while depletion of phagocytes with clodronate abolishes PGE2 production completely. Taken together, this work demonstrates that optimal PGE2 production by phagocytes depends on L. monocytogenes access to the cytosol, suggesting that one reason cytosolic access is required to prime CD8+ T-cell responses may be to facilitate production of PGE2.

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779 Inhibiting type-I interferon signaling promotes memory T-cell formation following immunization with Listeria anti-cancer vaccines

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2021-sitc2021.779 ◽

2021 ◽

Vol 9 (Suppl 3) ◽

pp. A814-A814

Author(s):

Zachary Morrow ◽

John-Demian Sauer

Keyword(s):

T Cells ◽

T Cell ◽

Type I Interferon ◽

T Cell Responses ◽

Cd8 T Cell ◽

T Cell Response ◽

Type I ◽

Interferon Signaling ◽

Wild Type ◽

Cell Responses

BackgroundThe aspiration of cancer immunotherapy is to generate large numbers of highly functional anti-tumor CD8+ T-cells. We and others have optimized Listeria monocytogenes as a powerful anti-cancer vaccine platform to drive such T-cell responses. Early clinical trial data suggest the number of T-cells generated correlates with efficacy, demanding an understanding of the factors that dictate vaccine-induced T-cell responses. The CD8+ T-cell response is intimately linked to magnitude and quality of the innate immune response triggered by vaccines. Listeria-based vaccines activate numerous innate pathways and can be engineered to hyper- or hypo-induce these pathways. We sought to understand how modulating innate immunity would impact vaccine efficacy.MethodsTo dissect the impact of type I interferon signaling and the inflammasomes on L. monocytogenes induced T-cell responses, we immunized IFNAR-/-, Caspase1/11-/-, and novel IFNAR-/-Caspase1/11-/- double knockouts mice we generated for this study. CD8+ T-cell responses were assessed at the peak T-cell response, after contraction and memory formation, and after rechallenge. The phenotype and magnitude of CD8+ T-cells was assessed at each stage, and functional outcomes were assessed by measuring protection from reinfection by wild-type Listeria.ResultsIFNAR-/- mice developed the largest number of CD8+ T-cells during the peak primary response contradicting the dogma that Type-I Interferon promotes robust CD8+ T-cell responses. Caspase1/11-/- mice were not significantly different from wild-type mice. The frequency of short-lived effector cells (assessed by expression of CD127 and KLRG1) was no different between wild-type and IFNAR-/- mice, however we observed more than twice as many memory precursor cells at the peak CD8+ T-cell response. These findings extend to the memory and recall stage with more antigen-specific T-cells observed after contraction and upon rechallenge. Finally, IFNAR-/- mice are remarkably more protected from wild-type Listeria rechallenge than their counterparts after immunization demonstrating the efficacy of the increased memory T-cell pool. Data are representative of at least two independent replicates with at least 5 mice per group and significance was assessed by one-way ANOVA with *p<0.05.ConclusionsWe demonstrated that type-I interferon signaling deficiency leads to enhanced prophylactic vaccine efficacy through increased memory T-cell formation. Ultimately, for patients with slow growing tumors or with high-risk mutations, prophylactic tumor vaccines could elicit life-long protection from disease. Importantly, increased memory precursor T-cell abundance did not come at the expense of short-lived effectors leaving open the possibility that blocking Type-I IFN could potentiate lasting immunological memory in both the therapeutic and prophylactic setting.

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608 Immunodominant listeria epitopes compete with vaccine-directed cd8+ t-cell responses rescued by peptide-MHC stabilizing modifications

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-sitc2020.0608 ◽

2020 ◽

Vol 8 (Suppl 3) ◽

pp. A644-A644

Author(s):

John Flickinger ◽

Jagmohan Singh ◽

Yanki Yarman ◽

Robert Carlson ◽

Scott Waldman ◽

...

Keyword(s):

Listeria Monocytogenes ◽

T Cell ◽

Cell Line ◽

Mhc Class I ◽

T Cell Responses ◽

Cd8 T Cell ◽

Class I ◽

Peptide Ligand ◽

Altered Peptide Ligand ◽

Cell Responses

BackgroundThe Gram-positive bacterium Listeria monocytogenes (Lm) is a promising vector for cancer immunotherapy due to its ability to directly infect antigen-presenting cells, induce potent CD8+ T-cell immunity, and remodel immunosuppressive tumor microenvironments.1 Recent clinical trials have demonstrated safety and immunogenicity of Lm-based cancer vaccines in lung, cervical, pancreatic, and other cancers. In colorectal cancer, the transmembrane receptor guanylyl cyclase C (GUCY2C) is an emerging target for immunotherapy.2 Here, we examined the immunogenicity of a recombinant strain of Listeria monocytogenes secreting GUCY2C (Lm-GUCY2C). Surprisingly, Lm-GUCY2C vaccination induced robust Lm-specific CD8+ T-cell immunity but failed to prime GUCY2C-specific CD8+ T-cell responses. These studies explore the hypothesis that immunodominant Lm antigens suppress primary immunity to subdominant GUCY2C epitopes in Lm-GUCY2CMethodsLm-GUCY2C expresses the extracellular domain of mouse GUCY2C23-429 downstream of an ActA promoter integrated into the genome of the live, attenuated delta actA delta inlB Lm strain. Altered peptide ligands were designed based on NetMHCpan 4.0 peptide-MHC binding algorithms and similarly cloned into Lm. Peptide-MHC class I complex stability was quantified by FACS-based surface peptide-MHC dissociation on the TAP-deficient cell line, RMA-S H-2Kd.3In vivo efficacy studies employed IFNγ-ELISpot quantification of T-cell responses and tumor challenge studies with the CT26 colorectal cancer cell line. Adenovirus expressing GUCY2C was used as a positive control.2 4ResultsLm-GUCY2C vaccination of BALB/c mice generated Lm-specific CD8+ T-cell responses but an absence of GUCY2C-specific immunity. Peptide-MHC stability studies revealed poor stability of the dominant GUCY2C254-262 epitope complexed with H-2Kd compared to H-2Kd-restricted Lm epitopes derived from the LLO and p60 Lm antigens. Mutation of the GUCY2C254-262 peptide at critical anchoring residues for binding H-2Kd revealed that the altered peptide ligand with an F255Y mutation significantly improved the stability of the GUCY2C254-262-H-2Kd complex. Similarly, vaccination of mice with recombinant Lm-GUCY2C expressing the altered peptide ligand (Lm-GUCY2CF255Y) restored GUCY2C immunogenicity and antitumor immunity.ConclusionsImmunodominant Lm antigens may interfere with immune responses directed to the vaccine target antigen GUCY2C by competing with GUCY2C epitope for MHC class I binding and presentation. Moreover, use of a substituted GUCY2C -peptide ligand with enhanced peptide-MHC class I stability restored GUCY2C-specific immunity in the context of Lm-GUCY2C, an approach that can be translated to patients. Importantly, these studies also suggest that ongoing Lm-based vaccine development programs targeting a variety of antigens in other cancer types may be similarly limited by the immunodominance of Lm epitopes.AcknowledgementsThe authors thank Dr. Peter Lauer for providing the pPL2 integration vector used in cloning Lm-GUCY2C and Dr. Sean Murphy for providing the RMA-S H-2Kd cell line.Ethics ApprovalStudies were approved by the Thomas Jefferson University IACUC (Protocol # 01956).ReferencesFlickinger JC, Rodeck U, Snook AE. Listeria monocytogenes as a Vector for Cancer Immunotherapy: Current Understanding and Progress. Vaccines (Basel) 2018;6. doi:10.3390/vaccines6030048.Snook AE, Baybutt TR, Xiang B, Abraham TS, Flickinger JC, Hyslop T, et al. Split tolerance permits safe Ad5-GUCY2C-PADRE vaccine-induced T-cell responses in colon cancer patients. J Immunother Cancer 2019;7:104. doi:10.1186/s40425-019-0576-2.Müllbacher A, Lobigs M, Kos FJ, Langman R. Alloreactive cytotoxic T-cell function, peptide nonspecific. Scand J Immunol 1999;49:563–9.Flickinger J. JC, Singh J, Carlson R, Leong E, Baybutt T, Barton J, et al. Chimeric Ad5.F35 vector evades anti-adenovirus serotype 5 neutralization opposing GUCY2C-targeted antitumor immunity. J Immunother Cancer 2020.

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Neutrophil subtypes shape HIV-specific CD8 T-cell responses after vaccinia virus infection

npj Vaccines ◽

10.1038/s41541-021-00314-7 ◽

2021 ◽

Vol 6 (1) ◽

Author(s):

Mauro Di Pilato ◽

Miguel Palomino-Segura ◽

Ernesto Mejías-Pérez ◽

Carmen E. Gómez ◽

Andrea Rubio-Ponce ◽

...

Keyword(s):

T Cell ◽

Immune Responses ◽

Vaccinia Virus ◽

Adaptive Immune System ◽

T Cell Responses ◽

Cd8 T Cell ◽

Cell Responses ◽

Adaptive Immune

AbstractNeutrophils are innate immune cells involved in the elimination of pathogens and can also induce adaptive immune responses. Nα and Nβ neutrophils have been described with distinct in vitro capacity to generate antigen-specific CD8 T-cell responses. However, how these cell types exert their role in vivo and how manipulation of Nβ/Nα ratio influences vaccine-mediated immune responses are not known. In this study, we find that these neutrophil subtypes show distinct migratory and motility patterns and different ability to interact with CD8 T cells in the spleen following vaccinia virus (VACV) infection. Moreover, after analysis of adhesion, inflammatory, and migration markers, we observe that Nβ neutrophils overexpress the α4β1 integrin compared to Nα. Finally, by inhibiting α4β1 integrin, we increase the Nβ/Nα ratio and enhance CD8 T-cell responses to HIV VACV-delivered antigens. These findings provide significant advancements in the comprehension of neutrophil-based control of adaptive immune system and their relevance in vaccine design.

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Listeria Monocytogenes: A Model Pathogen Continues to Refine Our Knowledge of the CD8 T Cell Response

Pathogens ◽

10.3390/pathogens7020055 ◽

2018 ◽

Vol 7 (2) ◽

pp. 55 ◽

Cited By ~ 11

Author(s):

Zhijuan Qiu ◽

Camille Khairallah ◽

Brian Sheridan

Keyword(s):

Listeria Monocytogenes ◽

T Cell ◽

Cell Response ◽

Protective Immunity ◽

T Cell Responses ◽

Cd8 T Cell ◽

Oral Infection ◽

T Cell Response ◽

Infection Model ◽

Cell Responses

Listeria monocytogenes (Lm) infection induces robust CD8 T cell responses, which play a critical role in resolving Lm during primary infection and provide protective immunity to re-infections. Comprehensive studies have been conducted to delineate the CD8 T cell response after Lm infection. In this review, the generation of the CD8 T cell response to Lm infection will be discussed. The role of dendritic cell subsets in acquiring and presenting Lm antigens to CD8 T cells and the events that occur during T cell priming and activation will be addressed. CD8 T cell expansion, differentiation and contraction as well as the signals that regulate these processes during Lm infection will be explored. Finally, the formation of memory CD8 T cell subsets in the circulation and in the intestine will be analyzed. Recently, the study of CD8 T cell responses to Lm infection has begun to shift focus from the intravenous infection model to a natural oral infection model as the humanized mouse and murinized Lm have become readily available. Recent findings in the generation of CD8 T cell responses to oral infection using murinized Lm will be explored throughout the review. Finally, CD8 T cell-mediated protective immunity against Lm infection and the use of Lm as a vaccine vector for cancer immunotherapy will be highlighted. Overall, this review will provide detailed knowledge on the biology of CD8 T cell responses after Lm infection that may shed light on improving rational vaccine design.

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Imatinib Mesylate Inhibits Antigen-Specific Memory CD8 T Cell Responses In Vivo

The Journal of Immunology ◽

10.4049/jimmunol.178.4.2028 ◽

2007 ◽

Vol 178 (4) ◽

pp. 2028-2037 ◽

Cited By ~ 24

Author(s):

Parisa Sinai ◽

Rance E. Berg ◽

J. Marshall Haynie ◽

Merrill J. Egorin ◽

Robert L. Ilaria ◽

...

Keyword(s):

T Cell ◽

Imatinib Mesylate ◽

T Cell Responses ◽

Cd8 T Cell ◽

Cell Responses ◽

Specific Memory ◽

Memory Cd8 T Cell

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TRANCE, a Tumor Necrosis Factor Family Member Critical for CD40 Ligand–independent T Helper Cell Activation

Journal of Experimental Medicine ◽

10.1084/jem.189.7.1025 ◽

1999 ◽

Vol 189 (7) ◽

pp. 1025-1031 ◽

Cited By ~ 191

Author(s):

Martin F. Bachmann ◽

Brian R. Wong ◽

Régis Josien ◽

Ralph M. Steinman ◽

Annette Oxenius ◽

...

Keyword(s):

T Cell ◽

Family Member ◽

T Cell Responses ◽

Cd40 Ligand ◽

Cd4 T Cell ◽

Cell Responses ◽

T Cell Priming ◽

Deficient Mice ◽

Necrosis Factor

CD40 ligand (CD40L), a tumor necrosis factor (TNF) family member, plays a critical role in antigen-specific T cell responses in vivo. CD40L expressed on activated CD4+ T cells stimulates antigen-presenting cells such as dendritic cells, resulting in the upregulation of costimulatory molecules and the production of various inflammatory cytokines required for CD4+ T cell priming in vivo. However, CD40L- or CD40-deficient mice challenged with viruses mount protective CD4+ T cell responses that produce normal levels of interferon γ, suggesting a CD40L/CD40-independent mechanism of CD4+ T cell priming that to date has not been elucidated. Here we show that CD4+ T cell responses to viral infection were greatly diminished in CD40-deficient mice by administration of a soluble form of TNF-related activation-induced cytokine receptor (TRANCE-R) to inhibit the function of another TNF family member, TRANCE. Thus, the TRANCE/TRANCE-R interaction provides costimulation required for efficient CD4+ T cell priming during viral infection in the absence of CD40L/CD40. These results also indicate that not even the potent inflammatory microenvironment induced by viral infections is sufficient to elicit efficient CD4+ T cell priming without proper costimulation provided by the TNF family (CD40L or TRANCE). Moreover, the data suggest that TRANCE/TRANCE-R may be a novel and important target for immune intervention.

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Adrenergic signaling controls early transcriptional programs during CD8+ T cell responses to viral infection

10.1101/2021.10.20.465178 ◽

2021 ◽

Author(s):

Leonardo Estrada ◽

Didem Agac Cobanoglu ◽

Aaron Wise ◽

Robert Maples ◽

Murat Can Cobanoglu ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Cd8 T Cells ◽

T Cell Responses ◽

Cd8 T Cell ◽

Receptor Activation ◽

Cell Development ◽

Primary Response ◽

Cell Responses

Viral infections drive the expansion and differentiation of responding CD8+ T cells into variegated populations of cytolytic effector and memory cells. While pro-inflammatory cytokines and cell surface immune receptors play a key role in guiding T cell responses to infection, T cells are also markedly influenced by neurotransmitters. Norepinephrine is a key sympathetic neurotransmitter, which acts to suppress CD8 + T cell cytokine secretion and lytic activity by signaling through the beta2-adrenergic receptor (ADRB2). Although ADRB2 signaling is considered generally immunosuppressive, its role in regulating differentiation of effector T cells in response to infection has not been investigated. Using an adoptive transfer approach, we compared the expansion and differentiation of wild type (WT) to Adrb2-/- CD8 + T cells throughout the primary response to vesicular stomatitis virus (VSV) infection in vivo. We measured the dynamic changes in transcriptome profiles of antigen-specific CD8 + T cells as they responded to VSV. Within the first 7 days of infection, WT cells out-paced the expansion of Adrb2-/- cells, which correlated with reduced expression of IL-2 and the IL-2Ralpha; in the absence of ADRB2. RNASeq analysis identified over 300 differentially expressed genes that were both temporally regulated following infection and selectively regulated in WT vs Adrb2-/- cells. These genes contributed to major transcriptional pathways including cytokine receptor activation, signaling in cancer, immune deficiency, and neurotransmitter pathways. By parsing genes within groups that were either induced or repressed over time in response to infection, we identified three main branches of genes that were differentially regulated by the ADRB2. These gene sets were predicted to be regulated by specific transcription factors involved in effector T cell development, such as Tbx21 and Eomes. Collectively, these data demonstrate a significant role for ADRB2 signaling in regulating key transcriptional pathways during CD8 + T cells responses to infection that may dramatically impact their functional capabilities and downstream memory cell development.

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DC subset–specific induction of T cell responses upon antigen uptake via Fcγ receptors in vivo

Journal of Experimental Medicine ◽

10.1084/jem.20160951 ◽

2017 ◽

Vol 214 (5) ◽

pp. 1509-1528 ◽

Cited By ~ 21

Author(s):

Christian H.K. Lehmann ◽

Anna Baranska ◽

Gordon F. Heidkamp ◽

Lukas Heger ◽

Kirsten Neubert ◽

...

Keyword(s):

T Cell ◽

T Cell Responses ◽

Cd8 T Cell ◽

Fcγ Receptors ◽

Specific Expression ◽

Antigen Uptake ◽

Surface Receptors ◽

Cell Responses ◽

Inflammatory Conditions

Dendritic cells (DCs) are efficient antigen-presenting cells equipped with various cell surface receptors for the direct or indirect recognition of pathogenic microorganisms. Interestingly, not much is known about the specific expression pattern and function of the individual activating and inhibitory Fcγ receptors (FcγRs) on splenic DC subsets in vivo and how they contribute to the initiation of T cell responses. By targeting antigens to select activating and the inhibitory FcγR in vivo, we show that antigen uptake under steady-state conditions results in a short-term expansion of antigen-specific T cells, whereas under inflammatory conditions especially, the activating FcγRIV is able to induce superior CD4+ and CD8+ T cell responses. Of note, this effect was independent of FcγR intrinsic activating signaling pathways. Moreover, despite the expression of FcγRIV on both conventional splenic DC subsets, the induction of CD8+ T cell responses was largely dependent on CD11c+CD8+ DCs, whereas CD11c+CD8− DCs were critical for priming CD4+ T cell responses.

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