Isolation of the melanoma-associated antigen p23 using antibody phage display.
The general responsiveness of human melanoma to immunotherapy has been well established, but active immunotherapy of melanoma has been hampered by insufficient information on the immunogenicity of melanoma-associated Ags in patients. In this study, we isolated a recombinant phage-Fab clone (A10-5) from a phage-Fab library derived from the B cells of a melanoma patient in remission after immunotherapy. Purified A10-5 Fab bound at high levels to cultured melanoma cell lines and to tissue sections of metastatic and vertical growth phase primary melanoma, but not to radial growth phase primary melanoma, nevi, or normal skin. A10-5 Fab bound to both the surface and the cytoplasm of cultured melanoma cells, but only to the cytoplasm of cultured fibroblasts. Western blot analysis revealed A10-5 Fab reactivity with a 33- and a 23-kDa glycoprotein under nonreducing conditions, and with a 23-kDa protein only under reducing conditions. A cDNA with an open reading frame predicted to encode a 23-kDa protein was cloned by screening a melanoma cell cDNA library with A10-5 Fab. This protein (p23) is the human homologue of the murine tumor transplantation Ag P198 that interacts with the cytoplasmic domain of ErbB-3 expressed by melanoma cells. Thus, the Ab phage display method has identified a novel, stage-specific melanoma-associated Ag that may have therapeutic and diagnostic value.
Li
J
J
The Wistar Institute, Philadelphia, PA 19104.
Pereira
S
S
Van Belle
P
P
Tsui
P
P
Elder
D
D
Speicher
D
D
Deen
K
K
Linnenbach
A
A
Somasundaram
R
R
Swoboda
R
R
Herlyn
D
D
D000818:Animals; D000951:Antigens, Neoplasm / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification*; D017104:Bacteriophage M13 / Q000235:genetics / Q000378:metabolism; D001666:Binding Sites, Antibody / Q000235:genetics; D019556:COS Cells; D002460:Cell Line; D003001:Cloning, Molecular; D020650:Combinatorial Chemistry Techniques; D018076:DNA, Complementary / Q000276:immunology; D005822:Genetic Vectors / Q000276:immunology; D006649:Histocompatibility Antigens / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification*; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000235:genetics / Q000378:metabolism*; D008545:Melanoma / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D058950:Melanoma-Specific Antigens; D009363:Neoplasm Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification*; D009928:Organ Specificity / Q000235:genetics / Q000276:immunology; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D018076:DNA, Complementary; D006649:Histocompatibility Antigens; D007140:Immunoglobulin Fab Fragments; D058950:Melanoma-Specific Antigens; D009363:Neoplasm Proteins; C029237:tumor-associated transplantation antigen