Abstract
Recombinant immunotoxins (RITs) are genetically engineered proteins being developed to treat cancer. They are composed of an Fv that targets a cancer antigen and a portion of a protein toxin. Their clinical success is limited by their immunogenicity. Our goal is to produce a new RIT that targets mesothelin and is non-immunogenic by combining mutations that decrease B- and T-cell epitopes. Starting with an immunotoxin that has B-cell epitopes suppressed, we added mutations step-wise that suppress T-cell epitopes. The final protein (LMB-T14) has greatly reduced antigenicity as assessed by binding to human anti-sera and a greatly decreased ability to activate helper T-cells evaluated in a T-cell activation assay. It is very cytotoxic to mesothelioma cells from patients, and to cancer cell lines. LMB-T14 produces complete remissions of a mesothelin expressing cancer (A431/H9) xenograft. The approach used here can be used to de-immunize other therapeutic foreign proteins.
Keywords:
epitope; immunogenicity; mesothelioma; pancreatic cancer; rational design.
MeSH terms
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Adaptive Immunity
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Animals
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Antibodies, Monoclonal / immunology
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Bacterial Toxins / genetics
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Bacterial Toxins / immunology
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Cell Line, Tumor
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Epitopes, B-Lymphocyte / genetics
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Epitopes, T-Lymphocyte / genetics
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Exotoxins / genetics
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Exotoxins / immunology
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Female
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GPI-Linked Proteins / antagonists & inhibitors*
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Humans
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Immunoglobulin Variable Region / immunology
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Immunotherapy / methods
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Immunotoxins / genetics
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Immunotoxins / immunology*
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Immunotoxins / therapeutic use
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Lung Neoplasms / immunology
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Lung Neoplasms / therapy*
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Mesothelin
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Mesothelioma / immunology
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Mesothelioma / therapy*
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Mesothelioma, Malignant
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Mice
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Mice, Nude
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Molecular Targeted Therapy / methods
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Mutation
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Protein Engineering
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / immunology*
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Recombinant Fusion Proteins / therapeutic use
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T-Lymphocytes, Helper-Inducer
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Xenograft Model Antitumor Assays
Substances
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Antibodies, Monoclonal
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Bacterial Toxins
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Epitopes, B-Lymphocyte
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Epitopes, T-Lymphocyte
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Exotoxins
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GPI-Linked Proteins
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Immunoglobulin Variable Region
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Immunotoxins
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Msln protein, mouse
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Recombinant Fusion Proteins
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Mesothelin