title: A recombinant immunotoxin targeting CD22 with low immunogenicity, low nonspecific toxicity, and high antitumor activity in mice.
abstract: Recombinant immunotoxins (RITs) are genetically engineered proteins designed to kill cancer cells. The RIT HA22 contains the Fv portion of an anti-CD22 antibody fused to a 38 kDa fragment of Pseudomonas exotoxin A (PE38). As PE38 is a bacterial protein, patients frequently produce antibodies that neutralize its activity, preventing retreatment. We have earlier shown in mice that PE38 contains 7 major B-cell epitopes located in domains II and III of the protein. Here we present a new mutant RIT, HA22-LR-6X, in which we removed most B-cell epitopes by deleting domain II and mutating 6 residues in domain III. HA22-LR-6X is cytotoxic to several lymphoma cell lines, has very low nonspecific toxicity, and retains potent antitumor activity in mice with CA46 lymphomas. To assess its immunogenicity, we immunized 3 MHC-divergent strains of mice with 5 microg doses of HA22-LR-6X, and found that HA22-LR-6X elicited significantly lower antibody responses than HA22 or other mutant RITs with fewer epitopes removed. Furthermore, large (50 microg) doses of HA22-LR-6X induced markedly lower antibody responses than 5 microg of HA22, indicating that high doses can be administered with low immunogenicity. Our experiments show that we have correctly identified and removed B-cell epitopes from PE38, producing a highly active immunotoxin with low immunogenicity and low animal toxicity. Future studies will determine if these properties carry over to humans with cancer.
authors.affiliation: Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Johanna K
authors.initials: JK
authors.lastname: Hansen
authors.forename: John E
authors.initials: JE
authors.lastname: Weldon
authors.forename: Laiman
authors.initials: L
authors.lastname: Xiang
authors.forename: Richard
authors.initials: R
authors.lastname: Beers
authors.forename: Masanori
authors.initials: M
authors.lastname: Onda
authors.forename: Ira
authors.initials: I
authors.lastname: Pastan
mesh_terms: D036002:ADP Ribose Transferases / Q000235:genetics / Q000276:immunology; D000818:Animals; D000906:Antibodies / Q000097:blood / Q000276:immunology; D000907:Antibodies, Bacterial / Q000097:blood / Q000276:immunology; D001427:Bacterial Toxins / Q000235:genetics / Q000276:immunology; D002051:Burkitt Lymphoma / Q000276:immunology* / Q000473:pathology / Q000517:prevention & control; D045744:Cell Line, Tumor; D004797:Enzyme-Linked Immunosorbent Assay; D005098:Exotoxins / Q000235:genetics / Q000276:immunology; D005260:Female; D006801:Humans; D007114:Immunization / Q000379:methods; D000922:Immunotoxins / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D016688:Mice, Inbred NOD; D008815:Mice, Inbred Strains; D016513:Mice, SCID; D011994:Recombinant Proteins / Q000276:immunology; D051918:Sialic Acid Binding Ig-like Lectin 2 / Q000276:immunology*; D013045:Species Specificity; D037521:Virulence Factors / Q000235:genetics / Q000276:immunology; D023041:Xenograft Model Antitumor Assays; D000097668:Pseudomonas aeruginosa Exotoxin A
chemical_list: D036002:ADP Ribose Transferases; D000906:Antibodies; D000907:Antibodies, Bacterial; D001427:Bacterial Toxins; D005098:Exotoxins; D000922:Immunotoxins; D011994:Recombinant Proteins; D051918:Sialic Acid Binding Ig-like Lectin 2; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A
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