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 ins ufficient information on the immunogenicity of melanoma-associated Ags in p atients. In this study, we isolated a recombinant phage-Fab clone (A10-5) f rom a phage-Fab library derived from the B cells of a melanoma patient in r emission after immunotherapy, Purified A10-5 Fab bound at high levels to cu ltured melanoma cell lines and to tissue sections of metastatic and vertica l growth phase primary melanoma, but not to radial growth phase primary mel anoma, nevi, or normal skin. A10-5 Fab bound to both the surface and the cy toplasm of cultured melanoma cells, but only to the cytoplasm of cultured f ibroblasts. Western blot analysis revealed A10-5 Fab reactivity with a 33- and a 23-kDa glycoprotein under nonreducing conditions, and with a 23-kDa p rotein only under reducing conditions, A cDNA with an open reading frame pr edicted to encode a 23-kDa protein was cloned by screening a melanoma cell cDNA library with A10-5 Feb. This protein (p23) is the human homologue of t he murine tumor transplantation Ag P198 that interacts with the cytoplasmic domain of ErbB-3 expressed by melanoma cells. Thus, the Ab phage display m ethod has identified a novel, stage-specific melanoma-associated Ag that ma y have therapeutic and diagnostic value.