CONSTRUCTION AND CHARACTERIZATION OF A BISPECIFIC DIABODY FOR RETARGETING T-CELLS TO HUMAN CARCINOMAS

We describe the construction of a recombinant bispecific antibody frag ment in the diabody format with specificity for both the well-establis hed human pancarcinoma associated target antigen EGP2 (epithelial glyc oprotein 2, also known as the CO 17-1A antigen or KSA) and the CD3 eps ilon chain of human TCR/CD3 complex. The murine anti-ECP2 (MOC31) sing le chain variable fragment (scFv) and the humanized anti-CD3 (Ucht1v9) scFv were cast into a diabody format (designated Dia5v9) using a shor t 5 amino acid Gly-Ser linker between immunoglobulin heavy-chain and l ight-chain variable domains. Purification of the poly-histidine tagged Dia5v9 was achieved from extracts of protease deficient Escerichia co li by IMAC chromatography. The Dia5v9 diabody showed strong binding to both EGP2 and CD3 in transfected cells. The in vitro efficacy of Dia5 v9 in mediating tumor cell lysis by interleukin-2 activated human T ce lls appeared to be similar to that of the hybrid-hybridoma-derived BsF (ab')(2) Bis1 (anti-EGP2/anti-CD3) in a standard 4-hr Cr-51-release as say. This small and partially humanized recombinant bispecific antibod y fragment may be valuable for T-cell-based immuno-therapeutical treat ment protocols, retargeting activated peripheral blood T lymphocytes t o lyse various human carcinomas in vivo. (C) 1998 Wiley-Liss, Inc.