Title: a Claudin-targeting Molecule as an Inhibitor of Tumor Metastasis A) a Running Title: Claudin-targeting and Metastasis

This article has not been copyedited and formatted. The final version may differ from this version. Abstract Tumor metastasis of epithelium-derived tumors is the major cause of death from malignant tumors. Overexpression of claudin is frequently observed in malignant tumors. However, claudin-targeting anti-metastasis therapy has never been investigated. We previously prepared a claudin-4-targeting anti-tumor molecule that consisted of the C-terminal fragment of Clostridium perfringens enterotoxin (C-CPE) fused to protein synthesis inhibitory factor (PSIF) derived from Pseudomonas exotoxin. In the present study, we investigated whether claudin CPE receptors can be a target for tumor metastasis by using the C-CPE-fused PSIF as a claudin-targeting agent. One of the most popular murine metastasis models is the lung metastasis of intravenously injected B16 cells. Therefore, we first investigated the effects of the C-CPE-fused PSIF on lung metastasis of claudin-4-expressing B16 cells (CL4-B16 cells). Intravenous administration of the C-CPE-fused PSIF suppressed lung metastasis of CL4-B16 cells but not B16 cells. Injection of C-CPE-fused PSIF also inhibited tumor growth and spontaneous lung metastasis of murine breast cancer 4T1 cells inoculated into the subcutis. Treatment with C-CPE-fused PSIF did not show apparent side effects in mice. These findings indicate that claudin-targeting may be a novel strategy for inhibiting some tumor metastases. This article has not been copyedited and formatted. The final version may differ from this version.