Abstract
Activation of murine glucocorticoid-induced tumor necrosis factor-related receptor (mGITR) by its natural ligand (GITRL) or antiGITR agonist mAb enhances T-cell responses, inhibits regulatory T-cell (Treg)-mediated suppression and induces tumor immunity in a variety of murine tumor models. However, systemic administration of these costimulatory agents can lead to global T-cell activation and autoimmunity. To specifically manipulate the T-cell compartment in the tumor microenvironment we propose to target the tumor infiltrating T cells with a bispecific mGITRL fusion protein. For that purpose, mGITRL is linked to a single-chain antibody targeting fibroblast activation protein (FAP) as FAP expression is restricted to cancer-associated fibroblasts (CAFs) found in the stroma of epithelial cancers. AntiFAP-mGITRL fusion protein forms dimers and binds to murine GITR with 1.2 μM affinity and to murine FAP with 4.5 nM. The construct is able to costimulate CD8+ and CD4+ effector T cells resulting in increased proliferation, IFN-γ and IL-2 production. This costimulatory effect is enhanced when the fusion protein is bound to a FAP-positive cell line mimicking FAP CAFs. In suppression assays, membrane-bound antiFAP-mGITRL is 100-fold more effective in overcoming Treg-mediated suppression than unbound fusion protein. These studies suggest that targeted tumor therapy with antiFAP-mGITRL fusion protein could induce tumor rejection while minimizing autoimmune side effects.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Antigens, Neoplasm / immunology
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Antigens, Neoplasm / metabolism*
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Cell Line, Tumor
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Cell Proliferation / drug effects
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Cytokines / genetics
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Cytokines / metabolism
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Endopeptidases
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Gelatinases / genetics
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Gelatinases / immunology
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Gelatinases / metabolism
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Glucocorticoid-Induced TNFR-Related Protein
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Immunotherapy*
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Lymphocyte Activation / drug effects
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Lymphocytes, Tumor-Infiltrating / immunology
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Membrane Proteins / genetics
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Membrane Proteins / immunology
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Membrane Proteins / metabolism
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Mice
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Mice, Inbred BALB C
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Neoplasms, Experimental / drug therapy
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Neoplasms, Experimental / immunology*
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Protein Structure, Tertiary / genetics
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Receptors, Nerve Growth Factor / immunology
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Receptors, Nerve Growth Factor / metabolism*
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Receptors, Tumor Necrosis Factor / immunology
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Receptors, Tumor Necrosis Factor / metabolism*
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / immunology
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Recombinant Fusion Proteins / metabolism
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Serine Endopeptidases / genetics
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Serine Endopeptidases / immunology
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Serine Endopeptidases / metabolism
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Single-Chain Antibodies / genetics
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Single-Chain Antibodies / immunology
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Single-Chain Antibodies / metabolism
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T-Lymphocyte Subsets / drug effects
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T-Lymphocyte Subsets / immunology
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T-Lymphocyte Subsets / metabolism
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T-Lymphocyte Subsets / pathology
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T-Lymphocytes, Regulatory / drug effects*
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T-Lymphocytes, Regulatory / immunology
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T-Lymphocytes, Regulatory / metabolism
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T-Lymphocytes, Regulatory / pathology
Substances
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Antigens, Neoplasm
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Cytokines
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Glucocorticoid-Induced TNFR-Related Protein
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Membrane Proteins
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Receptors, Nerve Growth Factor
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Receptors, Tumor Necrosis Factor
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Recombinant Fusion Proteins
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Single-Chain Antibodies
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Tnfrsf18 protein, mouse
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Endopeptidases
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Serine Endopeptidases
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fibroblast activation protein alpha
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Gelatinases