Expression of Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) Receptor DR5 in Ewing Sarcoma Family Tumors (ESFT)
S Galli, G Li, Y Ren, M Tsokos. NCI/NIH, Bethesda, MD
Background: TRAIL induces apoptosis in a wide range of tumor cells but not in normal tissues and is a promising candidate for novel anticancer therapies. The efficacy of a TRAIL agonistic antibody in pediatric sarcomas is currently studied in phase I clinical trials. TRAIL induces apoptosis through binding with two cell surface death receptors (DR), DR4 and DR5. Each one of the TRAIL death receptors is independently sufficient to induce apoptosis, but DR5 is more frequently expressed in tumors and can be induced by chemotherapeutic agents. We have previously shown that ESFT, which are common bone and soft tissue sarcomas in children and young adults, express TRAIL receptors in vitro and in vivo. However, immunohistochemical studies in paraffin tissues do not provide information of receptor expression on the tumor cell surface, which is necessary for the functionality of the receptors. Also, studies on established cell lines may not be indicative of the expression status of the receptors in vivo. To address these questions, we studied DR5 expression in early passage primary cell cultures and corresponding tumor tissues from ESFT patients. We also studied DR5 inducibility by doxorubicin.
Design: Eight early passage ESFT cell cultures were established in our laboratory (TC-324, TC-389, TC-390, TC-392, TC-393, TC-394, TC-399 and TC-400) and studied for expression of DR5 by Western blot and flow cytometry. Six available tumor specimens corresponding to the primary cultures were studied by immunohistochemistry for DR5 expression. Five of these cultures were treated with low to high concentrations of doxorubicin (0.1, 0.5 and 1.0ug/ml) for 16 and 24 hours and DR5 protein expression was studied by Western blot before and after treatment.
Results: All primary cultures expressed moderate to high levels of DR5 by Western blot. Flow cytometric analysis showed high DR5 surface expression in 5 and low in 3 cases. Immunohistochemical staining correlated with the flow cytometric data, as it was high in the 3 studied tumors with high flow cytometric expression and low or absent in the 3 tumors with low DR5 expression by flow cytometry. DR5 expression was induced in all five studied lines even by low concentration of doxorubicin (0.1ug/ml).
Conclusions: DR5 is expressed on the surface of ESFT cells and is induced by doxorubicin. These data suggest that ESFT are amenable to treatment with TRAIL alone or in combination with chemotherapeutic agents, which can increase TRAIL sensitivity at non toxic concentrations.
Monday, March 9, 2009 1:00 PM
Poster Session II # 208, Monday Afternoon