Distribution and immunochemical characterization of a keratin-like antigen in epithelial tumors using mouse and human monoclonal antibodies.
Using conventional murine hybridoma technology, we have produced a monoclonal antibody (MAb), 89E5, which recognizes two keratin-like polypeptides (Mr 53,000 and 45,000), which are preferentially expressed by epithelial tumors. In addition to detection of tumor cells by immunohistochemistry, MAb 89E5 was able to localize to tumor xenografts in nude mice after iodination of its F(ab')2 fragments. To develop potentially less immunogenic antibodies to antigens defined by MAb 89E5, studies were performed to produce a human counterpart to the mouse MAb. The mouse 89E5 MAb was used to purify the 89E5 polypeptides from tumor cell lines. The partially purified 89E5 antigen was then used to sensitize human splenic lymphocytes in vitro. Immortalization of the sensitized cells by cell fusion resulted in a human IgM MAb, PA1, which showed the same reactivity pattern on a panel of cell lines as did the mouse MAb 89E5. Immunofluorescent studies showed that both 89E5 and PA1 had staining patterns on epithelial cells indicative of antibodies to cytokeratin. Furthermore, PA1 immunoprecipitated two polypeptides (Mr 53,000 and 45,000) which comigrated with the 89E5 polypeptides. Competitive binding assays showed that the PA1 MAb and 89E5 MAb recognized closely associated epitopes. As with the 89E5 MAb, PA1 was reactive with tumor tissues in immunohistochemical studies. These studies indicate that the PA1 MAb is a human counterpart of the mouse 89E5 MAb. Direct comparison of human MAb and mouse MAb against the same antigen could yield valuable information on the efficacy of using human MAb in vivo.
Ho
M K
MK
Medical Products Department, E.I. Du Pont de Nemours & Co., North Billerica, Massachusetts 01862.
Geist
C
C
Murray
J
J
Kato
K
K
DeLellis
R
R
Wolfe
H
H
Carney
W
W
Rabin
H
H
Durda
P
P
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000951:Antigens, Neoplasm / Q000032:analysis*; D002277:Carcinoma / Q000276:immunology*; D006801:Humans; D006825:Hybridomas; D007150:Immunohistochemistry; D007633:Keratins / Q000032:analysis* / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008970:Molecular Weight
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D007633:Keratins