Abstract
Here we constructed and produced a recombinant human 4-1BB ligand (4-1BBL)/anti-CD20 fusion protein and examined its antitumor activity, alone and in combination with an anti-CD3/anti-CD20 bispecific diabody. The 4-1BBL/anti-CD20 fusion protein retained both the costimulatory activity of 4-1BBL on T cells and the tumor targeting ability of CD20 antibody on B cells. The fusion protein bound as efficiently to 4-1BB- and CD20-positive cells as its respective parental antibodies, and was capable of cross-linking human T lymphocytes and CD20-positive tumor cells. Combination treatment with 4-1BBL/anti-CD20 fusion protein and anti-CD3/anti-CD20 diabody led to significantly increased T-cell cytotoxicity to human B-lymphoma cells in vitro and drastically more potent tumor inhibitory activity in vivo in xenografted B-cell lymphoma in severe combined immunodeficiency disease mice. Mechanistic studies revealed that the combination treatment remarkably inhibited apoptosis of human peripheral blood lymphocytes, accompanied by upregulation of Bcl-XL and Bf1-1, perforin and granzyme B mRNA, and increased interleukin-2 production. Taken together, these results suggest that targeted delivery of 4-1BBL to the tumor site, when combined with anti-CD3/anti-CD20 diabody, could strongly potentiate the antitumor activity of the diabody, thus may have significant clinical application in the treatment of human CD20-positive B-cell malignancies.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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4-1BB Ligand / administration & dosage
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4-1BB Ligand / genetics
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4-1BB Ligand / metabolism
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Animals
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Antibodies, Bispecific / administration & dosage
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Antibodies, Bispecific / genetics
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Antibodies, Bispecific / metabolism
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Antibodies, Monoclonal / administration & dosage
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Antibodies, Monoclonal / genetics
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Antibodies, Monoclonal / metabolism
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Antibodies, Monoclonal, Murine-Derived
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Antineoplastic Combined Chemotherapy Protocols / therapeutic use*
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CD3 Complex / administration & dosage
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CD3 Complex / genetics
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CD3 Complex / immunology
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CD3 Complex / metabolism
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Cytotoxicity, Immunologic / drug effects
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Drug Synergism
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Female
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Humans
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Interleukin-2 / biosynthesis*
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Interleukin-2 / genetics
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Interleukin-2 / metabolism
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Jurkat Cells
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Lymphoma, B-Cell / drug therapy*
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Lymphoma, B-Cell / immunology
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Lymphoma, B-Cell / pathology
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Lymphoma, B-Cell / physiopathology
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Mice
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Mice, Inbred BALB C
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Mice, SCID
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Protein Engineering
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Recombinant Fusion Proteins / administration & dosage
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Rituximab
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T-Lymphocytes / drug effects
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T-Lymphocytes / immunology
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T-Lymphocytes / metabolism*
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T-Lymphocytes / pathology
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bcl-X Protein / biosynthesis*
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bcl-X Protein / genetics
Substances
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4-1BB Ligand
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Antibodies, Bispecific
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Antibodies, Monoclonal
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Antibodies, Monoclonal, Murine-Derived
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CD3 Complex
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Interleukin-2
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Recombinant Fusion Proteins
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bcl-X Protein
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Rituximab