Cloning and in vitro expression of a melanoma-associated antigen immunogenic in patients with melanoma

The purpose of this study was to identify human melanoma-associated Ag (MAA) that are immunogenic in patients, because these molecules may be useful immunogens to implement active specific immunotherapy. To this end, an expression cDNA library constructed from the human melanoma cell line A375 was screened with sera from patients with melanoma. A 1029-bp cDNA (designated D-1) was isolated. Its nucleotide sequence showed no significant homology with viral and mammalian sequences stored in GE-NETYX. cDNA D-1 hybridized to a 2.0-kb mRNA species from human melanoma, neuroblastoma, erythroleukemia, B lymphoid, and T lymphoid cell lines but not from a renal carcinoma cell line, PBL, and cultured skin fibroblasts. The D-1 clone produced a fusion protein that displayed a significantly higher reactivity with sera from patients with melanoma than from healthy controls. Furthermore, D-1 fusion protein induced in mice antibodies that immunoprecipitated a 50-kDa component from cultured human melanoma cells. The structural properties of D-1 MAA are different from those of previously described MAA. These results suggest that the approach we have applied may be useful to identify novel MAA expressed by melanoma cells. Furthermore, the immunogenicity of recombinant D-1 protein suggests that it may be a valuable immunogen to implement active specific immunotherapy in patients with melanoma, if additional experiments show that it has the appropriate tissue distribution.