Characterization of a monoclonal antibody-defined human melanoma-associated antigen susceptible to induction by immune interferon.

Abstract

To develop monoclonal antibodies (mAb) recognizing human melanoma-associated antigens (MAA) susceptible to modulation by immune interferon (IFN-gamma), hybridomas were constructed with splenocytes from a BALB/c mouse immunized with IFN-gamma-treated melanoma cells Colo 38. Screening of supernatants with control and IFN-gamma-treated melanoma cells showed that the mAb CL203 and CL207 display preferential reactivity with IFN-gamma-treated melanoma cells. The two mAb recognize the same (or spatially close) determinant on a 96,000 MAA which has a density of 0.36 X 10(6) antigenic sites/cell on untreated melanoma cells Colo 38 and of 1.39 X 10(6) and 1.54 X 10(6) on melanoma cells Colo 38 treated with IFN-gamma (final concentration, 200 U/ml) for 24 and 48 hr, respectively. The effect of IFN-gamma on the 96,000 MAA is dose- and time-dependent, reversible, and blocked by inhibitors of RNA and protein synthesis. Furthermore, the effect of IFN-gamma on the induction of the 96,000 MAA appears to be specific, inasmuch as IFN-alpha and IFN-beta do not induce the expression of the 96,000 MAA. The latter is also induced by IFN-gamma in a variety of carcinoma cell lines, but its level is markedly lower than on melanoma cells. Furthermore, the apparent m.w. of the antigen synthesized by the carcinoma cell lines in the presence of IFN-gamma ranges between 93,000 and 96,000. This molecular heterogeneity appears to reflect differences in the degree of glycosylation of the polypeptide moiety because the antigen synthesized by a variety of cell lines in the presence of tunicamycin has an apparent m.w. of 51,000.