A recombinant immunotoxin targeting CD22 with low immunogenicity, low nonspecific toxicity, and high antitumor activity in mice.
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.
Hansen
Johanna K
JK
Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
Weldon
John E
JE
Xiang
Laiman
L
Beers
Richard
R
Onda
Masanori
M
Pastan
Ira
I
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
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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