IN-VITRO CHARACTERIZATION OF A RECOMBINANT P-32 PHOSPHORYLATED ANTI-(CARCINOEMBRYONIC ANTIGEN) SINGLE-CHAIN ANTIBODY

The major limitations of monoclonal antibody conjugates as therapeutic agents have been their poor tumour targeting, inadequate tumour penet ration and immunogenicity. More even and deeper tissue penetration has been demonstrated with smaller antibody fragments. The smaller size a nd absence of an Fe segment may contribute to a lowered immunogenicity with single-chain antibodies (scFv) and also permit their recombinant engineering and bacterial expression. We describe the successful engi neer ing, expression and pre-clinical characterisation of a phosphoryl atable ''kemptide'' (Leu-Arg-Arg-Ala-Ser-Gly) anticarcinoembryonic ant igen (anti-CEA) scFv (PKS-scFv), for use as a radioimmunotherapeutic a gent. Specifically, a yield of 6 mg/l induced culture was obtained. Si te-specific phosphorylation was demonstrated without loss of specifici ty. In vitro assays revealed a selective cytotoxicity of P-32-PKS-scFv for high-CEA-expressing LS-174T cells compared to the low-CEA-express ing HT-29 cells, with a rapid internalisation rate.