Characterization of the B cell epitopes associated with a truncated form of Pseudomonas exotoxin (PE38) used to make immunotoxins for the treatment of cancer patients.
Recombinant immunotoxins composed of an Ab Fv fragment joined to a truncated portion of Pseudomonas exotoxin A (termed PE38) have been evaluated in clinical trials for the treatment of various human cancers. Immunotoxin therapy is very effective in hairy cell leukemia and also has activity in other hemological malignancies; however, a neutralizing Ab response to PE38 in patients with solid tumors prevents repeated treatments to maximize the benefit. In this study, we analyze the murine Ab response as a model to study the B cell epitopes associated with PE38. Sixty distinct mAbs to PE38 were characterized. Mutual competitive binding of the mAbs indicated the presence of 7 major epitope groups and 13 subgroups. The competition pattern indicated that the epitopes are discrete and could not be reproduced using a computer simulation program that created epitopes out of random surface residues on PE38. Using sera from immunotoxin-treated patients, the formation of human Abs to each of the topographical epitopes was demonstrated. One epitope subgroup, E1a, was identified as the principal neutralizing epitope. The location of each epitope on PE38 was determined by preparing 41 mutants of PE38 in which bulky surface residues were mutated to either alanine or glycine. All 7 major epitope groups and 9 of 13 epitope subgroups were identified by 14 different mutants and these retained high cytotoxic activity. Our results indicate that a relatively small number of discrete immunogenic sites are associated with PE38, most of which can be eliminated by point mutations.
Onda
Masanori
M
Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, 37 Convent Drive, Bethesda, MD 20892, USA.
Nagata
Satoshi
S
FitzGerald
David J
DJ
Beers
Richard
R
Fisher
Robert J
RJ
Vincent
James J
JJ
Lee
Byungkook
B
Nakamura
Michihiro
M
Hwang
Jaulang
J
Kreitman
Robert J
RJ
Hassan
Raffit
R
Pastan
Ira
I
D036002:ADP Ribose Transferases / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use; D000818:Animals; D000911:Antibodies, Monoclonal; D000917:Antibody Formation; D001427:Bacterial Toxins / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D018985:Epitopes, B-Lymphocyte / Q000276:immunology*; D005098:Exotoxins / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use; D006801:Humans; D007128:Immunoglobulin Fragments; D000922:Immunotoxins; D051379:Mice; D009369:Neoplasms / Q000188:drug therapy*; D011549:Pseudomonas; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D000911:Antibodies, Monoclonal; D001427:Bacterial Toxins; D018985:Epitopes, B-Lymphocyte; D005098:Exotoxins; D007128:Immunoglobulin Fragments; D000922:Immunotoxins; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A; C055966:immunoglobulin Fv