title: Recombinant immunotoxin for cancer treatment with low immunogenicity by identification and silencing of human T-cell epitopes.
abstract: Nonhuman proteins have valuable therapeutic properties, but their efficacy is limited by neutralizing antibodies. Recombinant immunotoxins (RITs) are potent anticancer agents that have produced many complete remissions in leukemia, but immunogenicity limits the number of doses that can be given to patients with normal immune systems. Using human cells, we identified eight helper T-cell epitopes in PE38, a portion of the bacterial protein Pseudomonas exotoxin A which consists of the toxin moiety of the RIT, and used this information to make LMB-T18 in which three epitopes were deleted and five others diminished by point mutations in key residues. LMB-T18 has high cytotoxic and antitumor activity and is very resistant to thermal denaturation. The new immunotoxin has a 93% decrease in T-cell epitopes and should have improved efficacy in patients because more treatment cycles can be given. Furthermore, the deimmunized toxin can be used to make RITs targeting other antigens, and the approach we describe can be used to deimmunize other therapeutically useful nonhuman proteins.
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;Department of Molecular Microbiology and Biotechnology, The George S. Wise Faculty of Life Sciences, Tel Aviv University, Ramat Aviv 6998, Israel;
authors.forename: Ronit
authors.initials: R
authors.lastname: Mazor
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
authors.forename: Jaime A
authors.initials: JA
authors.lastname: Eberle
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
authors.forename: Xiaobo
authors.initials: X
authors.lastname: Hu
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
authors.forename: Aaron N
authors.initials: AN
authors.lastname: Vassall
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
authors.forename: Masanori
authors.initials: M
authors.lastname: Onda
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
authors.forename: Richard
authors.initials: R
authors.lastname: Beers
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
authors.forename: Elizabeth C
authors.initials: EC
authors.lastname: Lee
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Kreitman
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
authors.forename: Byungkook
authors.initials: B
authors.lastname: Lee
authors.affiliation: Institute for Protein Design, Department of Biochemistry, University of Washington, Seattle, WA 98122; and.
authors.forename: David
authors.initials: D
authors.lastname: Baker
authors.affiliation: Institute for Protein Design, Department of Biochemistry, University of Washington, Seattle, WA 98122; and.
authors.forename: Chris
authors.initials: C
authors.lastname: King
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;Thoracic and Gastrointestinal Oncology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.
authors.forename: Raffit
authors.initials: R
authors.lastname: Hassan
authors.affiliation: Department of Molecular Microbiology and Biotechnology, The George S. Wise Faculty of Life Sciences, Tel Aviv University, Ramat Aviv 6998, Israel;
authors.forename: Itai
authors.initials: I
authors.lastname: Benhar
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892; pastani@mail.nih.gov.
authors.forename: Ira
authors.initials: I
authors.lastname: Pastan
mesh_terms: D036002:ADP Ribose Transferases / Q000235:genetics / Q000276:immunology; D000596:Amino Acids / Q000235:genetics / Q000276:immunology; D000818:Animals; D000917:Antibody Formation / Q000276:immunology; D001427:Bacterial Toxins / Q000235:genetics / Q000276:immunology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D018604:Epitope Mapping; D018984:Epitopes, T-Lymphocyte / Q000276:immunology*; D005098:Exotoxins / Q000235:genetics / Q000276:immunology; D005260:Female; D006801:Humans; D007167:Immunotherapy / Q000379:methods; D000922:Immunotoxins / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology; D051379:Mice; D016513:Mice, SCID; D008958:Models, Molecular; D009369:Neoplasms / Q000276:immunology* / Q000473:pathology / Q000628:therapy; D010455:Peptides / Q000235:genetics / Q000276:immunology; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology* / Q000627:therapeutic use; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D037521:Virulence Factors / Q000235:genetics / Q000276:immunology; D023041:Xenograft Model Antitumor Assays; D000097668:Pseudomonas aeruginosa Exotoxin A
chemical_list: D036002:ADP Ribose Transferases; D000596:Amino Acids; D001427:Bacterial Toxins; D018984:Epitopes, T-Lymphocyte; D005098:Exotoxins; D000922:Immunotoxins; D010455:Peptides; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A
keywords: deimmunization; protein engineering