SITE-SPECIFIC MODIFICATIONS OF LIGHT-CHAIN GLYCOSYLATED ANTILYMPHOMA (LL2) AND ANTICARCINOEMBRYONIC ANTIGEN (HIMMU-14-N) ANTIBODY DIVALENT FRAGMENTS

Site-specific introduction of metal-chelating groups into F(ab')(2) fr agments of an antilymphoma antibody (LL2) possessing a natural Asn-lin ked light chain carbohydrate and an anti-carcinoembryonic antigen anti body (hImmu-14-N) grafted with a light chain carbohydrate site is desc ribed. For this purpose, four yttrium- (and indium)-chelating agents w ere used, containing a primary amino group for antibody binding and 1- (4-substituted benzyl)diethylenetriaminepentaacetic acid as the metal- chelator, separated by structurally different additional linkers, Conj ugates were prepared by reacting excess chelator with oxidized carbohy drate of F(ab')(2) fragments, with or without a subsequent reduction s tep. The conjugates, with up to an average of 5.5 chelating groups att ached to a F(ab')(2) fragment, were readily labeled with Y-90 and In-1 11 and were found to retain antigen-binding ability in in vitro assays . Tumor targeting was demonstrated using a Y-88-labeled hImmu-14-N F(a b')(2) carbohydrate-modified conjugate, 2-Pyridyldithiopropionic hydra zide was conjugated to the carbohydrate region, and the disulfide was selectively deprotected to the thiol group, which is reactive with red uced Tc-99m. These initial experiments establish that light chain carb ohydrate modification of F(ab')(2) is as facile as with the Fc-region carbohydrate of intact IgG, and thereby offer the possibility of desig ning site-specifically substituted F(ab')(2) fragments with favorable pharmacokinetic properties.