Eradication of chemotherapy‐resistant CD44+ human ovarian cancer stem cells in mice by intraperitoneal administration of clostridium perfringens enterotoxin

Abstract

BACKGROUND: Emerging evidence has suggested that the capability to sustain tumor formation, growth, and chemotherapy resistance in ovarian as well as other human malignancies exclusively resides in a small proportion of tumor cells termed cancer stem cells. During the characterization of CD44⁺ ovarian cancer stem cells, we found a high expression of the genes encoding for claudin‐4. Because this tight junction protein is the natural high‐affinity receptor for Clostridium perfringens enterotoxin (CPE), we have extensively investigated the sensitivity of ovarian cancer stem cells to CPE treatment in vitro and in vivo. METHODS: Real‐time polymerase chain reaction and flow cytometry were used to evaluate claudin‐3/‐4 expression in ovarian cancer stem cells. Small interfering RNA knockdown experiments and MTS assays were used to evaluate CPE‐induced cytotoxicity against ovarian cancer stem cell lines in vitro. C.B‐17/SCID mice harboring ovarian cancer stem cell xenografts were used to evaluate CPE therapeutic activity in vivo. RESULTS: CD44⁺ ovarian cancer stem cells expressed claudin‐4 gene at significantly higher levels than matched autologous CD44⁻ ovarian cancer cells, and regardless of their higher resistance to chemotherapeutic agents died within 1 hour after exposure to 1.0 μg/mL of CPE in vitro. Conversely, small‐interfering RNA‐mediated knockdown of claudin‐3/‐4 expression in CD44⁺ cancer stem cells significantly protected cancer stem cells from CPE‐induced cytotoxicity. Importantly, multiple intraperitoneal administrations of sublethal doses of CPE in mice harboring xenografts of chemotherapy‐resistant CD44⁺ ovarian cancer stem cells had a significant inhibitory effect on tumor progression leading to the cure and/or long‐term survival of all treated animals (ie, 100% reduction in tumor burden in 50% of treated mice; P < .0001). CONCLUSIONS: CPE may represent an unconventional, potentially highly effective strategy to eradicate chemotherapy‐resistant cancer stem cells. Cancer 2011;. © 2011 American Cancer Society.