PROCESSING OF CARCINOEMBRYONIC ANTIGEN BY KUPFFER CELLS - RECOGNITIONOF A PENTA-PEPTIDE SEQUENCE

Carcinoembryonic antigen (CEA) binds to an 80-kDa cell surface recepto r on Kupffer cells via the peptide sequence PELPK (residues 108-112) l ocated at the hinge region between the N and A1 immunoglobulinlike dom ains. This study is aimed at analyzing the specificity of the peptide binding, determining biodistribution of 80-kDa receptor, and processin g of CEA by this receptor. We synthesized a number of bovine serum alb umin (BSA) derivatives carrying PELPK and related sequences. A series of peptides (YPELPK, YPDLPK, YPDLPR, and YPELGK) were conjugated to bo vine serum albumin using N-hydroxysuccinimidyl-4-azidobenzoate. When I -125 peptide conjugates, CEA, and BSA were injected intravenously into rats CEA and the PELPK-albumin conjugate were cleared rapidly. The ot her peptide conjugates and BSA cleared at a much slower rate. Activity of I-125-labeled CEA and PELPK-albumin conjugate per gram of tissue w as highest for the liver and spleen. Clearance of I-125-CEA was inhibi ted by the presence of higher concentrations of the PELPK-albumin conj ugate. With isolated rat Kupffer cells, only CEA and the PELPK-albumin conjugate were bound and internalized in vitro and CEA binding was in hibited by higher concentrations of the PELPK-albumin conjugate. Simil arly, binding of the PELPK-albumin conjugate was inhibited by the pres ence of unlabeled CEA. Use of a heterobifunctional cross linking agent demonstrated reaction of the PELPK-albumin with an 80-kDa protein on the Kupffer cell surface by SDS-polyacrylamide gel electrophoresis (SD S-PAGE). This semisynthetic ligand (PELPK-albumin) allows us to examin e the function of the 80-kDa receptor without interference due to othe r properties of CEA including its ability to bind lectins and to cause homotypic aggregation of cells. The consequences of CEA binding to th e 80-kDa receptor may have implications in the development of hepatic metastasis from colorectal cancer. (C) 1996 Academic Press, Inc.