A New Epitope on Human Melanoma-Associated Antigen CD63/ME491 Expressed by Both Primary and Metastatic Melanoma

Abstract

A monoclonal antibody (MAb) designated FC-5.01 (IgG2a) was generated that binds to several human carcinomas and malignant melanoma. It has revealed no or very low reactivity with most human normal tissues, except for the fact that FC-5.01 binds to some cells from the neuroendocrine system, macrophages, and some renal proximal convolute tubules with an intracellular pattern. Biochemical studies indicate that FC-5.01 recognizes a heterogeneous glycoprotein (broadband between 30-60 kDa) in melanoma tumors. The epitopes reside in the protein core and are presumably conformational, with disulphide bonds implicated in MAb recognition. The current study presents evidence that MAb FC-5.01 reacts with CD63 antigen (Ag), which has been initially described as a melanoma associated Ag, and is a member of the tetraspan family. Reactivity of MAb FC-5.01 with CD63 was demonstrated by Western blot, immunodepletion assay, and FACS analysis of the CD63-negative melanoma cells (KM3) after transfection with the genomic copy of CD63. The epitope recognized by MAb FC-5.01 was shown to be different from the epitope recognized by another anti-CD63 MAb, ME491, by an inhibition radioimmunoassay. Opposite to what has been stated for MAb ME491, no significant differences were found in CD63 expression between primary and metastatic melanoma using MAb FC-5.01.