scholarly journals Heterologous prime-boost vaccination targeting MAGE-type antigens promotes tumor T-cell infiltration and improves checkpoint blockade therapy

2021 ◽  
Vol 9 (9) ◽  
pp. e003218
Author(s):  
James McAuliffe ◽  
Hok Fung Chan ◽  
Laurine Noblecourt ◽  
Ramiro Andrei Ramirez-Valdez ◽  
Vinnycius Pereira-Almeida ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Response ◽  
Tumor Antigens ◽  
T Cell Response ◽  
Checkpoint Blockade ◽  
Cell Infiltration ◽  
Tumor Models ◽  
Murine Tumor ◽  
T Cell Infiltration

BackgroundThe clinical benefit of immune checkpoint blockade (ICB) therapy is often limited by the lack of pre-existing CD8+ T cells infiltrating the tumor. In principle, CD8+ T-cell infiltration could be promoted by therapeutic vaccination. However, this remains challenging given the paucity of vaccine platforms able to induce the strong cytotoxic CD8+ T-cell response required to reject tumors. A therapeutic cancer vaccine that induces a robust cytotoxic CD8+ T-cell response against shared tumor antigens and can be combined with ICB could improve the outcome of cancer immunotherapy.MethodsHere, we developed a heterologous prime-boost vaccine based on a chimpanzee adenovirus (ChAdOx1) and a modified vaccinia Ankara (MVA) encoding MAGE-type antigens, which are tumor-specific shared antigens expressed in different tumor types. The mouse MAGE-type antigen P1A was used as a surrogate to study the efficacy of the vaccine in combination with ICB in murine tumor models expressing the P1A antigen. To characterize the vaccine-induced immune response, we performed flow cytometry and transcriptomic analyses.ResultsThe ChAdOx1/MVA vaccine displayed strong immunogenicity with potent induction of CD8+ T cells. When combined with anti-Programmed Cell Death Protein 1 (PD-1), the vaccine induced superior tumor clearance and survival in murine tumor models expressing P1A compared with anti-PD-1 alone. Remarkably, ChAdOx1/MVA P1A vaccination promoted CD8+ T-cell infiltration in the tumors, and drove inflammation in the tumor microenvironment, turning ‘cold’ tumors into ‘hot’ tumors. Single-cell transcriptomic analysis of the P1A-specific CD8+ T cells revealed an expanded population of stem-like T cells in the spleen after the combination treatment as compared with vaccine alone, and a reduced PD-1 expression in the tumor CD8+ T cells.ConclusionsThese findings highlight the synergistic potency of ChAdOx1/MVA MAGE vaccines combined with anti-PD-1 for cancer therapy, and establish the foundation for clinical translation of this approach. A clinical trial of ChadOx1/MVA MAGE-A3/NY-ESO-1 combined with anti-PD-1 will commence shortly.

scholarly journals P03.02 Protein-based cancer vaccine combined with an oncolytic vaccine promotes potent antitumor immunity

2021 ◽  
Vol 9 (Suppl 1) ◽  
pp. A13.2-A14
Author(s):  
E Belnoue ◽  
K Das ◽  
M Rossi ◽  
T Hofer ◽  
S Danklmaier ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Microenvironment ◽  
Oncolytic Virus ◽  
Antitumor Immunity ◽  
Checkpoint Blockade ◽  
Tumor Models ◽  
Tumor Associated Antigens ◽  
T Cell Infiltration ◽  
Part Time

BackgroundKISIMATM platform allows the development of protein-based cancer vaccines able to induce a potent, tumor-specific CD8 and CD4 T cells response. While the cell penetrating peptide and peptide agonist for Toll like receptor (TLR)-2 and TLR-4 confer, respectively, the cell delivery and self-adjuvanticity properties, the multiantigenic domain allows the targeting of different cancer antigens, resulting in anti-tumoral efficacy in different murine models. Oncolytic viruses exert their therapeutic effects by a prolonged oncolytic action and the associated intratumoral inflammation as well as general immune activation. Arming oncolytic virus with tumor associated antigens can additionally enhance the tumor-specific T cell portion and therefore positively affect the balance of antitumor versus antiviral immune responses. The protein vaccine KISIMATM and the recombinant oncolytic virus VSV-GP-TAA (vesicular stomatitis virus pseudotyped with LCMV GP expressing tumor-associated antigens) are both promising vaccine candidates that offer a new cancer vaccination opportunity when combined in heterologous prime-boost regimen.Materials and MethodsMice were vaccinated with subcutaneous (s.c.) injection of KISIMA-TAA vaccine and/or with intravenous injection of VSV-GP-TAA in different settings. Immunogenicity was assessed by measuring the peripheral antigen-specific response. Anti-tumoral efficacy as well as in depth monitoring of TILs and tumor microenvironment modulation were assessed following therapeutic vaccination in different tumor models. Additionally, transcriptome and immunohistochemistry analyses of the TC-1 tumor have been performed. Combination of heterologous prime-boost with checkpoint blockade PD-1 therapy has been assessed.ResultsPriming with KISIMA-TAA followed by VSV-GP-TAA boost induced a large pool of polyfunctional and persistent antigen-specific cytotoxic T cells in the periphery as well as within the tumor in several tumor models. Frequencies of antigen specific T cells are significantly higher than the respective homologous vaccinations. Additionally, transcriptome analysis of a cold tumor model revealed profound changes in the tumor microenvironment upon heterologous vaccination, including a strong upregulation of gene signatures of several pro-inflammatory cytokines and chemokines required for antitumor immunity along with dendritic and T cell trafficking and activation. This was corroborated by flow-cytometric analysis of tumor-infiltrating leukocytes showing massive CD8+ and CD4+ T cell infiltration as well as repolarization of M2-like macrophages towards M1-phenotype. The presence of the CD8+ T cells within the tumor core was confirmed by immunohistochemistry analysis. Moreover, combining heterologous vaccination with checkpoint blockade further improved its therapeutic efficacy and the number of long-term survivors.ConclusionsThe KISIMA/VSV-GP heterologous prime-boost approach holds great promise for patients with primary or acquired resistance to checkpoint blockade due to its ability to induce tumor-specific T cell, improve T cell infiltration and increase tumor inflammation, even in tumors with limited permissivity for the oncolytic virus.Disclosure InformationE. Belnoue: A. Employment (full or part-time); Significant; AMAL Therapeutics SA. K. Das: None. M. Rossi: A. Employment (full or part-time); Significant; AMAL Therapeutics SA. T. Hofer: None. S. Danklmaier: None. T. Nolden: A. Employment (full or part-time); Significant; Viratherapeutics GmbH. L. Schreiber: None. K. Angerer: None. J. Kimpel: None. S. Hoegler: None. L. Kenner: None. D. von Laer: None. K. Elbers: A. Employment (full or part-time); Significant; Viratherapeutics GmbH. G. Wollmann: None. M. Derouazi: A. Employment (full or part-time); Significant; AMAL Therapeutics SA.

Azacitidine Induces Expansion of Regulatory T Cells and Tumour Specific CD8+ T Lymphocytes After Allogeneic Stem Cell Transplantation: A Strategy for Epigenetic Manipulation of a Graft-Versus-Leukemia Response

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 324-324 ◽  
Author(s):  
Mike Dennis ◽  
Oliver Goodyear ◽  
Justin Loke ◽  
Nadiria Jilani ◽  
Shamyla Siddique ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Response ◽  
T Regulatory Cells ◽  
Tumor Antigens ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
Regulatory Cells ◽  
Post Transplant ◽  
Graft Versus Leukemia

Abstract Abstract 324 Disease relapse and graft-versus-host disease (GVHD) remain the commonest causes of treatment failure after allogeneic stem cell transplantation (SCT) for Acute Myeloid Leukemia (AML). Azacitidine (AZA) possesses inherent anti-leukemic activity but also expands immunomodulatory T regulatory cells in animal models and up-regulates the expression of tumor antigens in vitro. Reasoning that AZA might selectively augment a graft-versus-leukemia (GVL) without a concomitant increase in GVHD we have studied the tolerability and immunological sequelae of AZA administration in patients who have undergone a reduced intensity allogeneic SCT for AML. All patients were transplanted using a conditioning regimen consisting of fludarabine (30 mg/m2 IV × 5 days), melphalan (140 mg/m2IV), and alemtuzumab (10 mg IV × 5 days) with cyclosporine GVHD prophylaxis. Patients received AZA (36 mg/m2) × 5 days every 28 days commencing after sustained neutrophil and platelet engraftment at day +42 with the aim of administering monthly cycles of AZA until 12 months post-transplant. Numbers of CD4+CD25+CD127loFoxP3+ T regulatory cells were measured by flow cytometry. Tumor specific cytotoxic T lymphocytes (CTL) recognising members of the cancer testis antigen (CTA) family and WT1 were quantitated using a CD137 expression and enrichment assay. We report results on 27 patients (median age 59 years) who commenced treatment with AZA (follow-up 3–21 months). 11 patients were transplanted from an HLA identical related donor and 16 from a volunteer unrelated donor. Disease status at the time of transplant was: CR1 n=18; CR2 n=7; first relapse n=2. Post-transplant AZA was well tolerated and 24 patients tolerated at least three cycles of AZA. Haematological toxicity was modest with only two patients experiencing treatment delay for neutropenia, or thrombocytopenia. Three patients developed Grade 2 acute GVHD and no patient developed >Grade 2 acute GVHD. Two patients developed limited chronic GVHD. To date seven patients have relapsed at a median time of 6 months (4–15 months) post-transplant. Administration of AZA had no impact on absolute lymphocyte counts or CD8+ and CD4+ cell numbers compared with a control population of 17 patients transplanted using an identical conditioning regimen. However AZA administration was noted to increase the number of CD4+CD25+CD127loFoxP3+ regulatory T cells within the first 3 months post transplant compared with a time-matched control population (p=0.017) although there was no difference detectable at 6 or 12 months. AZA administration also induced a cytotoxic CD8+ T cell response to candidate tumor antigens MAGE, GAGE, RAGE and WT-1 in the peripheral blood. A CD8+ T cell response to these candidate tumor antigens was only detectable in 1/22 patients pre-transplant but circulating CTA- or WT1 specific CD8+ T cells were detected in 14/16 patients who had received at least six cycles of AZA with a frequency between 0.001–1.4% (mean 0.25%). In four patients with paired peripheral blood and bone marrow samples the size of the CD8+ T cell response was noted to be up to 100 fold greater in the bone marrow. Ex vivo characterisation of the CTA response using dextramers demonstrated the CD8+ T cell response to be effector memory (CD45RA-CCR7-) and functional assays confirmed by mobilization of CD107a and secretion of interferon-g, TNF-a and IL-2 in response to peptide. These data confirm the tolerability of adjunctive AZA post-transplant and its administration appears to be associated with a low incidence of both acute and chronic GVHD. The demonstration that AZA induces both early expansion of T regulatory cells and a tumor specific CD8+ CTL response highlight its potential use as a strategy to epigenetically manipulate a graft-versus-leukemia reaction after allogeneic SCT. Disclosures: Dennis: Celgene: Honoraria, Research Funding. Craddock:Celgene: Honoraria, Research Funding.

scholarly journals Clonal replacement of tumor-specific T cells following PD-1 blockade

2019 ◽  
Author(s):  
Kathryn E. Yost ◽  
Ansuman T. Satpathy ◽  
Daniel K. Wells ◽  
Yanyan Qi ◽  
Chunlin Wang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Carcinoma ◽  
Single Cell ◽  
Cell Response ◽  
T Cell Response ◽  
Checkpoint Blockade ◽  
T Cell Clones ◽  
Cell Clones ◽  
Clonal Replacement

AbstractImmunotherapies that block inhibitory checkpoint receptors on T cells have transformed the clinical care of cancer patients. However, which tumor-specific T cells are mobilized following checkpoint blockade remains unclear. Here, we performed paired single-cell RNA- and T cell receptor (TCR)-sequencing on 79,046 cells from site-matched tumors from patients with basal cell carcinoma (BCC) or squamous cell carcinoma (SCC) pre- and post-anti-PD-1 therapy. Tracking TCR clones and transcriptional phenotypes revealed a coupling of tumor-recognition, clonal expansion, and T cell dysfunction: the T cell response to treatment was accompanied by clonal expansions of CD8+CD39+T cells, which co-expressed markers of chronic T cell activation and exhaustion. However, this expansion did not derive from pre-existing tumor infiltrating T cell clones; rather, it comprised novel clonotypes, which were not previously observed in the same tumor. Clonal replacement of T cells was preferentially observed in exhausted CD8+T cells, compared to other distinct T cell phenotypes, and was evident in BCC and SCC patients. These results, enabled by single-cell multi-omic profiling of clinical samples, demonstrate that pre-existing tumor-specific T cells may be limited in their capacity for re-invigoration, and that the T cell response to checkpoint blockade relies on the expansion of a distinct repertoire of T cell clones that may have just recently entered the tumor.

scholarly journals Metformin reduces PD-L1 on tumor cells and enhances the anti-tumor immune response generated by vaccine immunotherapy

2021 ◽  
Vol 9 (11) ◽  
pp. e002614
Author(s):  
Luis Enrique Munoz ◽  
Lei Huang ◽  
Ramireddy Bommireddy ◽  
Richa Sharma ◽  
Lenore Monterroza ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Cd8 T Cells ◽  
Checkpoint Blockade ◽  
Cell Infiltration ◽  
In Vitro Assays ◽  
Immune Checkpoints ◽  
Cell Phenotype ◽  
Tumor Models

BackgroundPD-L1 is one of the major immune checkpoints which limits the effectiveness of antitumor immunity. Blockade of PD-L1/PD-1 has been a major improvement in the treatment of certain cancers, however, the response rate to checkpoint blockade remains low suggesting a need for new therapies. Metformin has emerged as a potential new drug for the treatment of cancer due to its effects on PD-L1 expression, T cell responses, and the immunosuppressive environment within tumors. While the benefits of metformin in combination with checkpoint blockade have been reported in animal models, little remains known about its effect on other types of immunotherapy.MethodsVaccine immunotherapy and metformin were administered to mice inoculated with tumors to investigate the effect of metformin and TMV vaccine on tumor growth, metastasis, PD-L1 expression, immune cell infiltration, and CD8 T cell phenotype. The effect of metformin on IFN-γ induced PD-L1 expression in tumor cells was assessed by flow cytometry, western blot, and RT-qPCR.ResultsWe observed that tumors that respond to metformin and vaccine immunotherapy combination show a reduction in surface PD-L1 expression compared with tumor models that do not respond to metformin. In vitro assays showed that the effect of metformin on tumor cell PD-L1 expression was mediated in part by AMP-activated protein kinase signaling. Vaccination results in increased T cell infiltration in all tumor models, and this was not further enhanced by metformin. However, we observed an increased number of CD8 T cells expressing PD-1, Ki-67, Tim-3, and CD62L as well as increased effector cytokine production after treatment with metformin and tumor membrane vesicle vaccine.ConclusionsOur data suggest that metformin can synergize with vaccine immunotherapy to augment the antitumor response through tumor-intrinsic mechanisms and also alter the phenotype and function of CD8 T cells within the tumor, which could provide insights for its use in the clinic.

Dynamics of T-cell infiltration during the course of ovarian cancer: The gradual shift from a Th17 effector cell response to a predominant infiltration by regulatory T-cells

2012 ◽  
Vol 132 (5) ◽  
pp. 1070-1079 ◽  
Author(s):  
Anna Fialová ◽  
Simona Partlová ◽  
Luděk Sojka ◽  
Hana Hromádková ◽  
Tomáš Brtnický ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Cell Response ◽  
Effector Cell ◽  
Cell Infiltration ◽  
T Cell Infiltration ◽  
Gradual Shift

scholarly journals 49 In situ phenotypic analysis of T cells in the tumor microenvironment of a pre-clinical model of non-small cell lung cancer (NSCLC) by tissue sectioning and whole-mount immunofluorescence imaging

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A52-A52
Author(s):  
Elen Torres ◽  
Stefani Spranger
Keyword(s):  
T Cells ◽  
Spatial Distribution ◽  
T Cell ◽  
Spatial Location ◽  
Cell Infiltration ◽  
Cell Populations ◽  
Tumor Bearing ◽  
T Cell Infiltration ◽  
Volume Imaging ◽  
Whole Mount

BackgroundUnderstanding the interactions between tumor and immune cells is critical for improving current immunotherapies. Pre-clinical and clinical evidence has shown that failed T cell infiltration into lung cancer lesions might be associated with low responsiveness towards checkpoint blockade.1 For this reason, it is necessary to characterize not only the phenotype of T cells in tumor-bearing lungs but also their spatial location in the tumor microenvironment (TME). Multiplex immunofluorescence staining allows the simultaneous use of several cell markers to study the state and the spatial location of cell populations in the tissue of interest. Although this technique is usually applied to thin tissue sections (5 to 12 µm), the analysis of large tissue volumes may provide a better understanding of the spatial distribution of cells in relation to the TME. Here, we analyzed the number and spatial distribution of cytotoxic T cells and other immune cells in the TME of tumor-bearing lungs, using both 12 µm sections and whole-mount preparations imaged by confocal microscopy.MethodsLung tumors were induced in C57BL/6 mice by tail vein injection of a cancer cell line derived from KrasG12D/+ and Tp53-/- mice. Lung tissue with a diverse degree of T cell infiltration was collected after 21 days post tumor induction. Tissue was fixed in 4% PFA, followed by snap-frozen for sectioning. Whole-mount preparations were processed according to Weizhe Li et al. (2019) 2 for tissue clearing and multiplex volume imaging. T cells were labeled with CD8 and FOXP3 antibodies to identify cytotoxic or regulatory T cells, respectively. Tumor cells were labeled with a pan-Keratin antibody. Images were acquired using a Leica SP8 confocal microscope. FIJI3 and IMARIS were used for image processing.ResultsWe identified both cytotoxic and regulatory T cell populations in the TME using thin sections and whole-mount. However, using whole-mount after tissue clearing allowed us to better evaluate the spatial distribution of the T cell populations in relation to the tumor structure. Furthermore, tissue clearance facilitates the imaging of larger volumes using multiplex immunofluorescence.ConclusionsAnalysis of large lung tissue volumes provides a better understanding of the location of immune cell populations in relation to the TME and allows to study heterogeneous immune infiltration on a per-lesion base. This valuable information will improve the characterization of the TME and the definition of cancer-immune phenotypes in NSCLC.ReferencesTeng MW, et al., Classifying cancers based on T-cell infiltration and PD-L1. Cancer Res 2015;75(11): p. 2139–45.Li W, Germain RN, and Gerner MY. High-dimensional cell-level analysis of tissues with Ce3D multiplex volume imaging. Nat Protoc 2019;14(6): p. 1708–1733.Schindelin J, et al, Fiji: an open-source platform for biological-image analysis. Nat Methods 2012;9(7): p. 676–82.

scholarly journals Mechanisms that allow vaccination against an oncolytic vesicular stomatitis virus-encoded transgene to enhance safety without abrogating oncolysis

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Amanda W. K. AuYeung ◽  
Robert C. Mould ◽  
Ashley A. Stegelmeier ◽  
Jacob P. van Vloten ◽  
Khalil Karimi ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Vesicular Stomatitis Virus ◽  
Cell Response ◽  
Viral Infections ◽  
T Cell Response ◽  
Oncolytic Viruses ◽  
Oncolytic Virotherapy ◽  
Cell Immunity ◽  
Vesicular Stomatitis

AbstractVaccination can prevent viral infections via virus-specific T cells, among other mechanisms. A goal of oncolytic virotherapy is replication of oncolytic viruses (OVs) in tumors, so pre-existing T cell immunity against an OV-encoded transgene would seem counterproductive. We developed a treatment for melanomas by pre-vaccinating against an oncolytic vesicular stomatitis virus (VSV)-encoded tumor antigen. Surprisingly, when the VSV-vectored booster vaccine was administered at the peak of the primary effector T cell response, oncolysis was not abrogated. We sought to determine how oncolysis was retained during a robust T cell response against the VSV-encoded transgene product. A murine melanoma model was used to identify two mechanisms that enable this phenomenon. First, tumor-infiltrating T cells had reduced cytopathic potential due to immunosuppression. Second, virus-induced lymphopenia acutely removed virus-specific T cells from tumors. These mechanisms provide a window of opportunity for replication of oncolytic VSV and rationale for a paradigm change in oncolytic virotherapy, whereby immune responses could be intentionally induced against a VSV-encoded melanoma-associated antigen to improve safety without abrogating oncolysis.

scholarly journals Checkpoint blockade accelerates a novel switch from an NKT-driven TNFα response toward a T cell driven IFN-γ response within the tumor microenvironment

2021 ◽  
Vol 9 (6) ◽  
pp. e002269
Author(s):  
Shota Aoyama ◽  
Ryosuke Nakagawa ◽  
Satoshi Nemoto ◽  
Patricio Perez-Villarroel ◽  
James J Mulé ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Tumor Growth ◽  
Ex Vivo ◽  
T Cell Response ◽  
Effector Function ◽  
Checkpoint Blockade ◽  
Necrosis Factor Alpha ◽  
Ifn Γ

BackgroundThe temporal response to checkpoint blockade (CB) is incompletely understood. Here, we profiled the tumor infiltrating lymphocyte (TIL) landscape in response to combination checkpoint blockade at two distinct timepoints of solid tumor growth.MethodsC57BL/6 mice bearing subcutaneous MC38 tumors were treated with anti-PD-1 and/or anti-CTLA-4 antibodies. At 11 or 21 days, TIL phenotype and effector function were analyzed in excised tumor digests using high parameter flow cytometry. The contributions of major TIL populations toward overall response were then assessed using ex vivo cytotoxicity and in vivo tumor growth assays.ResultsThe distribution and effector function among 37 distinct TIL populations shifted dramatically between early and late MC38 growth. At 11 days, the immune response was dominated by Tumor necrosis factor alpha (TNFα)-producing NKT, representing over half of all TIL. These were accompanied by modest frequencies of natural killer (NK), CD4+, or CD8+ T cells, producing low levels of IFN-γ. At 21 days, NKT populations were reduced to a combined 20% of TIL, giving way to increased NK, CD4+, and CD8+ T cells, with increased IFN-γ production. Treatment with CB accelerated this switch. At day 11, CB reduced NKT to less than 20% of all TIL, downregulated TNFα across NKT and CD4+ T cell populations, increased CD4+ and CD8+ TIL frequencies, and significantly upregulated IFN-γ production. Degranulation was largely associated with NK and NKT TIL. Blockade of H-2kb and/or CD1d during ex vivo cytotoxicity assays revealed NKT has limited direct cytotoxicity against parent MC38. However, forced CD1d overexpression in MC38 cells significantly diminished tumor growth, suggesting NKT TIL exerts indirect control over MC38 growth.ConclusionsDespite an indirect benefit of early NKT activity, CB accelerates a switch from TNFα, NKT-driven immune response toward an IFN-γ driven CD4+/CD8+ T cell response in MC38 tumors. These results uncover a novel NKT/T cell switch that may be a key feature of CB response in CD1d+ tumors.

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