CONSTRUCTION AND EXPRESSION OF 2 MOUSE-HUMAN CHIMERIC ANTIBODIES WITHHIGH SPECIFICITY AND AFFINITY FOR CARCINOEMBRYONIC ANTIGEN

We have previously reported that a group of monoclonal antibodies (MAb s) to carcinoembryonic antigen (CEA), designated Group F MAbs, are abl e to discriminate CEA in tumor tissues from the CEA-related normal ant igens and that CEA assay systems utilizing at least one Group F MAb sh ow the improved cancer diagnosis. In this study, we cloned the genes c oding for two Group F MAbs (F11-35 and F11-39) and deduced the amino a cid sequences of the variable regions for their heavy and light chains . The variable region for the heavy chain of F11-35 contained a possib le N-glycosylation site (Asn/Asp/Thr) at amino acid positions 89-91. T hen, we constructed two mouse-human chimeric antibodies by using the F 11-35 and F11-39 variable region genes of heavy and light chains (V(H) and Vkappa) and human heavy and light chain constant region genes (ga mma1 and kappa) derived from a human plasma cell leukemia line (ARH77) . The chimeric gene constructs were sequentially co-transfected into m urine non-Ig-producing myeloma (P3-U1) or hybridoma (Sp2/0) cells by e lectroporation. The resulting chimeric heavy chain of F11-35 showed a slightly but significantly higher molecular weight than that of F11-39 , but the molecular weights of their unglycosylated peptides synthesiz ed in the presence of tunicamycin were similar, indicating the glycosy lation at the possible N-glycosylation site in the variable region of the Ch F11-35 heavy chain. Both chimeric antibodies exhibited the same specificity and affinity for CEA as those of the parental murine hybr idoma antibodies, respectively. Ascites production of Sp2/0 transfecto mas is sufficiently high (600-900 mug/ml) for initial clinical studies with the chimeric antibodies.