Monoclonal antibodies for characterization of a tumor-associated antigen from a mouse lung carcinoma

Binding specifities of putative tumor-specific antibodies intended for use in radio imaging of tumors are routinely characterized by in vitro binding assays to tissue cultured cells, frozen tis sue sections and tissue lysates. 135-13C MoAb was characterized in vitro as specific to line 1 alveolar carcinoma cells and other lung carcinomas from mice. 135-13C MoAb precipitates a protein of 180,000 M.W., tumor-surface protein-lBO, from solubilized surface-labeled line 1 cells. I analyzed the distribution of radioiodinated 135-13C MoAb and 135-14 MoAb in tumor-bearing mice. Localization indexes for lung, spleen and kidney indicated that 135-13C MoAb might be binding to normal cells of these organs. Results from the analysis of clearance of radiolabeled 135-13C MoAb and control MoAb, 135-14, from the blood of normal and tumor-bearing mice are consistent with this hypothesis since 135-13C MoAb is cleared from the blood faster than the control MoAb, 135-14, in normal as well as tumor-bearing mice. Results show that putative imaging antibodies should be analyzed for clearance kinetics in vivoin normal subjects prior to imaging trials to determine if there is significant binding to normal cell sites.

135-13C MoAb was not expected to bind to normal cell sites. Characterization of TSP-l80-like antigens from normal cells requires a more sensitive assay for the detection and isolation of proteins than the assays used in the characterization of 135-13C MoAb. A MoAb (346-11) to a second site on TSP-180 was developed for future use in a two-site immunoassay. Two site assays increase the sensitivity of detection by concentrating the antigen with one MoAb prior to detection of antigen with a second MoAb. 346-11 binds to the surface of line 1 cells. It specifically precipitates TSP-180 from surface labeled line 1 cells as identified by SDS-PAGE of immunoprecipitates of solubilized line 1 cells. Reciprocal competition binding analysis of 135-13C MoAb and 346-11 MoAb demonstrated that 346-11 binds to a different site than 135-13C MoAb on the TSP-I80 molecule.