Pharmacokinetics and biodistribution of a light-chain-shuffled CC49 single-chain Fv antibody construct

Murine monoclonal antibodies to tumor-associated glycoprotein 72 (anti-TAG- 72 mAb B72.3 and CC49) are among the most extensively studied mAb for immun otherapy of adenocarcinomas. They have been used clinically to localize pri mary and metastatic tumor sites; however, murine mAb generally induce poten t human anti-(mouse antibody) responses. The immunogenicity of murine mAb c an be minimized by genetic humanization of these antibodies, where non-huma n regions are replaced by the corresponding human sequences or complementar y determining regions are grafted into the human framework regions. We have developed a humanized CC49 single-chain antibody construct (hu/muCC49 scFv ) by replacing the murine CC49 variable light chain with the human subgroup IV germline variable light chain (Hum4 V-L) The major advantages of scFv m olecules are their excellent penetration into the tumor tissue, rapid clear ance rate, and much lower exposure to normal organs, especially bone marrow , than occur with intact antibody. The biochemical properties of hu/muCC49 scFv were compared to those of the murine CC49 scFv (muCC49 scFv). The asso ciation constants (K-a) for hu/muCC49 and muCC49 constructs were 1.1 x 10(6 ) M-1 and 1.4 x 10(6) M-1 respectively. Pharmacokinetic studies in mice sho wed similar rapid blood and whole-body clearance with a half-life of 6 min for both scFv. The biodistribution studies demonstrated equivalent tumor ta rgeting to human colon carcinoma xenografts for muCC49 and hu/muCC49 scFv. These results indicate that the human variable light-chain subgroup IV can be used for the development of humanized or human immunoglobulin molecules potentially useful in both diagnostic and therapeutic applications with TAG -72-positive tumors.