Tissue distribution of an immunogenic melanoma associated antigen, D-1

A human melanoma-associated antigen immunogenic in patients was recently identified by screening an expression cDNA library constructed from cultured human melanoma cell line with sera from patients with melanoma. The nucleic acid sequence of the cloned D-1 cDNA has no significant homology with previously reported mammalian genes. The cDNA D-1 encodes a peptide of about 37 kDa, which showed fivefold higher reactivity with sera from patients with melanoma than with sera from normal donors. In order to detect D-1 gene expression in vivo, in-situ hybridization and immunostaining with cRNA probe and murine anti-D-1 sera were carried out on surgically removed tissues. Digoxigenin-labeled cRNA D-1 was exclusively hybridized with mRNA in the cytoplasm of melanoma cells but not with keratinocytes and fibroblasts adjacent to melanoma nests. Polyclonal anti-D-1 antibodies were obtained by immunization of Balb/c mice with recombinant D-1 peptide and clearly reacted with melanoma cells but not with keratinocytes and fibroblasts, similar to the results of in-situ hybridization. The above information will help to assess the suitability of recombinant D-1 peptide to implement active specific immunotherapy in patients with melanoma.