P06.05 Endogenous T-cell responses to ten major cancer testis antigens are frequent in esophago-gastric adenocarcinoma and antigen-specific T cells can be expanded using CD40-activated B cells

BackgroundTumor-associated antigens (TAAs) and especially cancer testis antigens (CTAs) are classical tumor-specific targets for immunotherapies. As TAAs are shared between patients, strategies aiming to exploit these targets are scalable and potentially applicable across different types of cancer. Loss of target antigens and other mechanisms of immune escape have limited the success of CTA-directed immunotherapy. CAR T cells and other highly effective cellular therapies have renewed the interest in TAAs. Especially combined targeting of multiple antigens appears highly promising as recently shown in lymphoma. In our study, we aimed to characterize CTA-expression patterns and their impact on endogenous T-cell responses, T-cell abundance and antigen-presentation in esophago-gastric adenocarcinoma (EGA).Materials and Methods41 treatment-naïve EGA patients were included in our study. RNA of tumor and patient-matched healthy tissue was isolated and used for NanoString based RNA expression analysis of 26 CTAs and 25 genes associated with antigen-presentation. Based on CTA expression, 10 peptide pools were selected and co-cultured with peripheral blood mononuclear cells (PBMCs, n=21) to determine cellular anti-tumor immune responses in a FluoroSpot assay. T-cell abundance was assessed using immunohistochemistry (CD3, CD8) and digital image analyses of tumor area and invasive margin. Autologous CD40 activated B cells were used to expand antigen-specific T cells using peptide pools of CTAs.ResultsNanoString analysis revealed pronounced differences regarding CTA expression, with CEP55 and MAGEA3/6 showing strong expression, while NY-ESO-1 or MAGEA1 were only weakly expressed. 68.3% (28/41) of the patients showed expression of ≥ 5/10 analyzed TAAs simultaneously. In line with the frequent expression, 75.0% of the patients showed a cellular response against at least one of the TAAs. T-cell responses were most frequently detected to Survivin and NY-ESO-1 (65.0% and 52.6% of patients, respectively), while only 20.0% responded to CEP55 or TTK peptide pools. Overall, 6/20 patients showed cellular responses against ≥5 TAAs simultaneously. We found a strong correlation of T-cell abundance and antigen-presentation. In addition, patients with a high Immune-Score showed increased TAA expression. Finally, we demonstrate feasibility of TAA-specific T-cell expansion using CD40 activated B cells as potential strategy to induce or enhance TAA immune responses in EGA.ConclusionsOur study highlights the importance of TAAs in EGA. The identified antigens are highly relevant for immunomonitoring of clinical trials and as targets for immunotherapy. Personalized immunotherapeutic strategies targeting EGA-specific or even patient specific TAAs appear highly promising in this challenging disease.Disclosure InformationM. Thelen: None. M. Garcia-Marquez: None. J. Lehmann: None. D. Keller: None. E. Preugszat: None. M. von Bergwelt-Baildon: B. Research Grant (principal investigator, collaborator or consultant and pending grants as well as grants already received); Modest; Astellas, Roche, MSD. D. Speakers Bureau/Honoraria (speakers bureau, symposia, and expert witness); Modest; BMS. F. Consultant/Advisory Board; Modest; BMS. H.A. Schlößer: B. Research Grant (principal investigator, collaborator or consultant and pending grants as well as grants already received); Significant; Astra Zeneca.

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Epigenetic Manipulation of Cancer Testis Antigen (CTA) Expression: A Strategy for Manipulating the Graft-Versus Leukaemia Response in Patients Allografted for Haematological Malignancies

Blood ◽

10.1182/blood.v112.11.600.600 ◽

2008 ◽

Vol 112 (11) ◽

pp. 600-600 ◽

Cited By ~ 2

Author(s):

Tatjana Stankovic ◽

Andrew McLarnon ◽

Angelo Agathanggelou ◽

Oliver Goodyear ◽

Charles Craddock ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Cell Lines ◽

T Cell Responses ◽

Interferon Γ ◽

Haematological Malignancies ◽

Cell Responses ◽

Cancer Testis Antigens ◽

Reduced Intensity ◽

Cancer Testis

Abstract An immunologically mediated graft-versus-tumor (GVT) underlies the curative effect of reduced intensity allografts in acute myeloid leukaemia (AML) and other haematologic malignancies. Cancer testis antigens (CTA) represent a family of immunodominant proteins that are variably expressed in haematological malignancies and represent a potential target of a GVT response. Importantly a number of CTA genes demonstrate promoter hypermethylation in solid tumours which can be reversed using demethylating agents such as azacitidine. We have therefore examined whether donor T cell responses to CTAs are observed in patients allografted for AML and multiple myeloma (MM) and whether epigenetic therapies can be used to manipulate T cell mediating killing of haemopoietic targets. We screened 37 patients with AML and 8 patients with MM, who had not received demethylating agent treatment, for T cell responses to 25 peptides derived from 10 CTA genes, including BAGE, LAGE, MAGE-A1, MAGE-A2, MAGE-A3, MAGE-A4, MAGE-C2, RAGE-1, by interferon-γ cytokine secretion assay (IFN-γ CSA). We found CTA specific T cells in 11.1% of patients (5 of 45) with frequencies ranging from 0.0005% to 0.2% (median 0.024%). Subsequently, we studied expression of 15 cancer-testis antigens (CTA) at the RNA and protein levels in AML, MM and Hodgkins’ lymphoma cell lines before and after exposure to demethylating agent 5-aza-2′-deoxycytidine (Aza) and histone deacetylase inhibitor sodium valproate (VPA), as single agents and in combination. We found that expression of CTAs HAGE, SACA3, SPANXB, LAGE, XAGE, MAGEA3 could be induced in a dose dependent manner by Aza alone but not with VPA alone. We also observed that expression induced by Aza treatment was increased further by combination treatment with VPA. Induction of CTAs was confirmed in vitro in primary AML cells and in vivo in 2/6 patients on VPA/Aza trial. Furthermore, using interferon-γ ELISA assay we observed that Aza-induced expression of the CTA MAGEA3 in MM cell lines was accompanied by increased recognition by MAGEA3-specific T cells. These studies confirm the importance of members of the CTA family as targets of a T cell mediated immune response. Our data demonstrate that the expression of these putative immunodominant antigens in haematological malignancies in disease such as AML and myeloma, can be significantly up-regulated by epigenetic therapies with functional increases in target cell killing and support the use of adjunctive epigenetic therapies after allogeneic transplantation with the aim of augmenting a GVL response. A Phase II clinical trial combining post transplant azacitidine with a reduced intensity allograft is currently ongoing in patients with AML.

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B cell depletion impairs vaccination-induced CD8+ T cell responses in a type I interferon-dependent manner

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2021-220435 ◽

2021 ◽

pp. annrheumdis-2021-220435

Author(s):

Theresa Graalmann ◽

Katharina Borst ◽

Himanshu Manchanda ◽

Lea Vaas ◽

Matthias Bruhn ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

B Cell ◽

Cell Expansion ◽

De Novo ◽

T Cell Responses ◽

Type I ◽

Cell Responses ◽

T Cell Expansion

ObjectivesThe monoclonal anti-CD20 antibody rituximab is frequently applied in the treatment of lymphoma as well as autoimmune diseases and confers efficient depletion of recirculating B cells. Correspondingly, B cell-depleted patients barely mount de novo antibody responses during infections or vaccinations. Therefore, efficient immune responses of B cell-depleted patients largely depend on protective T cell responses.MethodsCD8+ T cell expansion was studied in rituximab-treated rheumatoid arthritis (RA) patients and B cell-deficient mice on vaccination/infection with different vaccines/pathogens.ResultsRituximab-treated RA patients vaccinated with Influvac showed reduced expansion of influenza-specific CD8+ T cells when compared with healthy controls. Moreover, B cell-deficient JHT mice infected with mouse-adapted Influenza or modified vaccinia virus Ankara showed less vigorous expansion of virus-specific CD8+ T cells than wild type mice. Of note, JHT mice do not have an intrinsic impairment of CD8+ T cell expansion, since infection with vaccinia virus induced similar T cell expansion in JHT and wild type mice. Direct type I interferon receptor signalling of B cells was necessary to induce several chemokines in B cells and to support T cell help by enhancing the expression of MHC-I.ConclusionsDepending on the stimulus, B cells can modulate CD8+ T cell responses. Thus, B cell depletion causes a deficiency of de novo antibody responses and affects the efficacy of cellular response including cytotoxic T cells. The choice of the appropriate vaccine to vaccinate B cell-depleted patients has to be re-evaluated in order to efficiently induce protective CD8+ T cell responses.

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COVID-19 convalescents exhibit deficient humoral and T cell responses to variant of concern Spike antigens at 12 month post-infection

10.1101/2021.11.08.21266035 ◽

2021 ◽

Author(s):

Pablo Garcia-Valtanen ◽

Christopher Martin Hope ◽

Makutiro Ghislain Masavuli ◽

Arthur Eng Lip Yeow ◽

Harikrishnan Balachandran ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

Post Infection ◽

T Cell Responses ◽

Memory B Cells ◽

Pseudotyped Virus ◽

Cell Responses ◽

Specific Memory ◽

Cell Functionality

Background The duration and magnitude of SARS-CoV-2 immunity after infection, especially with regard to the emergence of new variants of concern (VoC), remains unclear. Here, immune memory to primary infection and immunity to VoC was assessed in mild-COVID-19 convalescents one year after infection and in the absence of viral re-exposure or COVID-19 vaccination. Methods Serum and PBMC were collected from mild-COVID-19 convalescents at ~6 and 12 months after a COVID-19 positive PCR (n=43) and from healthy SARS-CoV-2-seronegative controls (n=15-40). Serum titers of RBD and Spike-specific Ig were quantified by ELISA. Virus neutralisation was assessed against homologous, pseudotyped virus and homologous and VoC live viruses. Frequencies of Spike and RBD-specific memory B cells were quantified by flow cytometry. Magnitude of memory T cell responses was quantified and phenotyped by activation-induced marker assay, while T cell functionality was assessed by intracellular cytokine staining using peptides specific to homologous Spike virus antigen and four VoC Spike antigens. Findings At 12 months after mild-COVID-19, >90% of convalescents remained seropositive for RBD-IgG and 88.9% had circulating RBD-specific memory B cells. Despite this, only 51.2% convalescents had serum neutralising activity against homologous live-SARS-CoV-2 virus, which decreased to 44.2% when tested against live B.1.1.7, 4.6% against B.1.351, 11.6% against P.1 and 16.2%, against B.1.617.2 VoC. Spike and non-Spike-specific T cells were detected in >50% of convalescents with frequency values higher for Spike antigen (95% CI, 0.29-0.68% in CD4+ and 0.11-0.35% in CD8+ T cells), compared to non-Spike antigens. Despite the high prevalence and maintenance of Spike-specific T cells in Spike 'high-responder' convalescents at 12 months, T cell functionality, measured by cytokine expression after stimulation with Spike epitopes corresponding to VoC was severely affected. Interpretations SARS-CoV-2 immunity is retained in a significant proportion of mild COVID-19 convalescents 12 months post-infection in the absence of re-exposure to the virus. Despite this, changes in the amino acid sequence of the Spike antigen that are present in current VoC result in virus evasion of neutralising antibodies, as well as evasion of functional T cell responses.

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B cell depletion with anti-CD20 mAb exacerbates anti-donor CD4+ T cell responses in highly sensitized transplant recipients

Scientific Reports ◽

10.1038/s41598-021-97748-9 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Asuka Tanaka ◽

Kentaro Ide ◽

Yuka Tanaka ◽

Masahiro Ohira ◽

Hiroyuki Tahara ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

B Cell ◽

T Cell Responses ◽

Cell Depletion ◽

B Cell Depletion ◽

Transplant Recipients ◽

Cell Responses ◽

Anti Cd20

AbstractPretransplant desensitization with rituximab has been applied to preformed donor-specific anti-human leukocyte antigen antibody (DSA)-positive recipients for elimination of preformed DSA. We investigated the impact of pretransplant desensitization with rituximab on anti-donor T cell responses in DSA-positive transplant recipients. To monitor the patients’ immune status, mixed lymphocyte reaction (MLR) assays were performed before and after desensitization with rituximab. Two weeks after rituximab administration, the stimulation index (SI) of anti-donor CD4+ T cells was significantly higher in the DSA-positive recipients than in the DSA-negative recipients. To investigate the mechanisms of anti-donor hyper responses of CD4+ T cells after B cell depletion, highly sensitized mice models were injected with anti-CD20 mAb to eliminate B cells. Consistent with clinical observations, the SI values of anti-donor CD4+ T cells were significantly increased after anti-CD20 mAb injection in the sensitized mice models. Adding B cells isolated from untreated sensitized mice to MLR significantly inhibited the enhancement of anti-donor CD4+ T cell response. The depletion of the CD5+ B cell subset, which exclusively included IL-10-positive cells, from the additive B cells abrogated such inhibitory effects. These findings demonstrate that IL-10+ CD5+ B cells suppress the excessive response of anti-donor CD4+ T cells responses in sensitized recipients.

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RNA-loaded CD40-activated B cells stimulate antigen-specific T cell responses in dogs with spontaneous lymphoma

Veterinary Immunology and Immunopathology ◽

10.1016/j.vetimm.2008.10.278 ◽

2009 ◽

Vol 128 (1-3) ◽

pp. 341-342

Author(s):

Nicola J. Mason ◽

Christina M. Coughlin ◽

Jarish N. Cohen ◽

Theresa A. Colligon ◽

Craig A. Clifford ◽

...

Keyword(s):

T Cell ◽

B Cells ◽

T Cell Responses ◽

Cell Responses ◽

Stimulate Antigen ◽

Activated B Cells

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Ectopic Lymphoid Follicles in Multiple Sclerosis: Centers for Disease Control?

Frontiers in Neurology ◽

10.3389/fneur.2020.607766 ◽

2020 ◽

Vol 11 ◽

Author(s):

Austin Negron ◽

Olaf Stüve ◽

Thomas G. Forsthuber

Keyword(s):

Multiple Sclerosis ◽

T Cells ◽

T Cell ◽

B Cells ◽

B Cell ◽

T Cell Responses ◽

Memory B Cells ◽

Lymphoid Follicles ◽

Cell Responses ◽

B Cell Responses

While the contribution of autoreactive CD4+ T cells to the pathogenesis of Multiple Sclerosis (MS) is widely accepted, the advent of B cell-depleting monoclonal antibody (mAb) therapies has shed new light on the complex cellular mechanisms underlying MS pathogenesis. Evidence supports the involvement of B cells in both antibody-dependent and -independent capacities. T cell-dependent B cell responses originate and take shape in germinal centers (GCs), specialized microenvironments that regulate B cell activation and subsequent differentiation into antibody-secreting cells (ASCs) or memory B cells, a process for which CD4+ T cells, namely follicular T helper (TFH) cells, are indispensable. ASCs carry out their effector function primarily via secreted Ig but also through the secretion of both pro- and anti-inflammatory cytokines. Memory B cells, in addition to being capable of rapidly differentiating into ASCs, can function as potent antigen-presenting cells (APCs) to cognate memory CD4+ T cells. Aberrant B cell responses are prevented, at least in part, by follicular regulatory T (TFR) cells, which are key suppressors of GC-derived autoreactive B cell responses through the expression of inhibitory receptors and cytokines, such as CTLA4 and IL-10, respectively. Therefore, GCs represent a critical site of peripheral B cell tolerance, and their dysregulation has been implicated in the pathogenesis of several autoimmune diseases. In MS patients, the presence of GC-like leptomeningeal ectopic lymphoid follicles (eLFs) has prompted their investigation as potential sources of pathogenic B and T cell responses. This hypothesis is supported by elevated levels of CXCL13 and circulating TFH cells in the cerebrospinal fluid (CSF) of MS patients, both of which are required to initiate and maintain GC reactions. Additionally, eLFs in post-mortem MS patient samples are notably devoid of TFR cells. The ability of GCs to generate and perpetuate, but also regulate autoreactive B and T cell responses driving MS pathology makes them an attractive target for therapeutic intervention. In this review, we will summarize the evidence from both humans and animal models supporting B cells as drivers of MS, the role of GC-like eLFs in the pathogenesis of MS, and mechanisms controlling GC-derived autoreactive B cell responses in MS.

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Multiple Epitopes in the Murine Cytomegalovirus Early Gene Product M84 Are Efficiently Presented in Infected Primary Macrophages and Contribute to Strong CD8+-T-Lymphocyte Responses and Protection following DNA Immunization

Journal of Virology ◽

10.1128/jvi.78.20.11233-11245.2004 ◽

2004 ◽

Vol 78 (20) ◽

pp. 11233-11245 ◽

Cited By ~ 13

Author(s):

Ming Ye ◽

Christopher S. Morello ◽

Deborah H. Spector

Keyword(s):

T Cells ◽

T Cell ◽

Antigen Presentation ◽

T Cell Responses ◽

Dna Vaccination ◽

Early Gene ◽

Murine Cytomegalovirus ◽

Specific Response ◽

Mcmv Infection ◽

Cell Responses

ABSTRACT We previously demonstrated that after vaccination of BALB/c mice with DNA encoding murine cytomegalovirus (MCMV) IE1 or M84, a similar level of protection against MCMV infection was achieved. However, the percentage of antigen-specific CD8+ T cells elicited by IE1 was higher than that by M84 as measured by intracellular cytokine staining when splenocytes were stimulated with an epitope peptide (M. Ye at al., J. Virol. 76:2100-2112, 2002). We show here that after DNA vaccination with M84, a higher percentage of M84-specific CD8+ T cells was detected when splenocytes were stimulated with J774 cells expressing full-length M84. When the defined M84 epitope 297-305 was deleted, the mutant DNA vaccine was still protective against MCMV replication and induced strong M84-specific CD8+-T-cell responses. The M84 gene was subsequently subcloned into three fragments encoding overlapping protein fragments. When mice were immunized with each of the M84 subfragment DNAs, at least two additional protective CD8+-T-cell epitopes were detected. In contrast to strong responses after DNA vaccination, M84-specific CD8+-T-cell responses were poorly induced during MCMV infection. The weak M84-specific response after MCMV infection was not due to poor antigen presentation in antigen-presenting cells, since both J774 macrophages and primary peritoneal macrophages infected with MCMV in vitro were able to efficiently and constitutively present M84-specific epitopes starting at the early phase of infection. These results indicate that antigen presentation by macrophages is not sufficient for M84-specific CD8+-T-cell responses during MCMV infection.

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HIV-1 Antibody Neutralization Breadth Is Associated with Enhanced HIV-Specific CD4+T Cell Responses

Journal of Virology ◽

10.1128/jvi.02278-15 ◽

2015 ◽

Vol 90 (5) ◽

pp. 2208-2220 ◽

Cited By ~ 22

Author(s):

Srinika Ranasinghe ◽

Damien Z. Soghoian ◽

Madelene Lindqvist ◽

Musie Ghebremichael ◽

Faith Donaghey ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

Hiv Infection ◽

Neutralizing Antibodies ◽

T Cell Responses ◽

Neutralizing Antibody ◽

Antibody Activity ◽

Cell Responses

ABSTRACTAntigen-specific CD4+T helper cell responses have long been recognized to be a critical component of effective vaccine immunity. CD4+T cells are necessary to generate and maintain humoral immune responses by providing help to antigen-specific B cells for the production of antibodies. In HIV infection, CD4+T cells are thought to be necessary for the induction of Env-specific broadly neutralizing antibodies. However, few studies have investigated the role of HIV-specific CD4+T cells in association with HIV neutralizing antibody activity in vaccination or natural infection settings. Here, we conducted a comprehensive analysis of HIV-specific CD4+T cell responses in a cohort of 34 untreated HIV-infected controllers matched for viral load, with and without neutralizing antibody breadth to a panel of viral strains. Our results show that the breadth and magnitude of Gag-specific CD4+T cell responses were significantly higher in individuals with neutralizing antibodies than in those without neutralizing antibodies. The breadth of Gag-specific CD4+T cell responses was positively correlated with the breadth of neutralizing antibody activity. Furthermore, the breadth and magnitude of gp41-specific, but not gp120-specific, CD4+T cell responses were significantly elevated in individuals with neutralizing antibodies. Together, these data suggest that robust Gag-specific CD4+T cells and, to a lesser extent, gp41-specific CD4+T cells may provide important intermolecular help to Env-specific B cells that promote the generation or maintenance of Env-specific neutralizing antibodies.IMPORTANCEOne of the earliest discoveries related to CD4+T cell function was their provision of help to B cells in the development of antibody responses. Yet little is known about the role of CD4+T helper responses in the setting of HIV infection, and no studies to date have evaluated the impact of HIV-specific CD4+T cells on the generation of antibodies that can neutralize multiple different strains of HIV. Here, we addressed this question by analyzing HIV-specific CD4+T cell responses in untreated HIV-infected persons with and without neutralizing antibodies. Our results indicate that HIV-infected persons with neutralizing antibodies have significantly more robust CD4+T cell responses targeting Gag and gp41 proteins than individuals who lack neutralizing antibodies. These associations suggest that Gag- and gp41-specific CD4+T cell responses may provide robust help to B cells for the generation or maintenance of neutralizing antibodies in natural HIV-infection.

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