In Vivo Tumor Delivery of a Recombinant Single-Chain Fv::Tumor Necrosis Factor: A Fusion Protein

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Abstract
Locoregional and intratumoral administration of tumor necrosis factor α (TNFα) has been successful in obtaining inhibition or regression of tumor growth in the clinic. This potent antitumor activity of TNFα has not yet been exploited as a systemic agent in cancer therapy, mainly due to high levels of toxicity to normal tissues before a therapeutic dose of TNFα in the tumor has been achieved. To address this, we have targeted TNFα using antitumor antibodies. We have used a genetic fusion of human recombinant TNFα with MFE-23, a single-chain Fv antibody fragment directed against carcinoembryonic antigen. MFE-23::TNFα fusion protein is isolated in high yields (28 mg/L) from bacterial inclusion bodies and purified to homogeneity by affinity chromatography. It is a 144 kDa trimer in native form and possesses the antigen-binding activity of the sFv and the cytotoxicity to both WEHI 164 and a human adenocarcinoma cell line (LoVo) of rhTNFα. Radiolabeled MFE-23::TNFα binds both human and mouse TNF receptor 1 in vitro and is able to localize effectively in nude (nu/nu) mice bearing human LS174T xenografts; tumor/tissue ratios of 21:1 and 60:1 are achieved 24 and 48 h after intravenous injection. These studies indicate that MFE-23::TNFα will provide an effective means for systemically administered cancer therapy with TNFα.