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Section of Hematology/Oncology, Department of Pediatrics, Herman B Wells Center for Pediatric Research, Indianapolis, Indiana; and Departments of Biochemistry & Molecular Biology and Pharmacology & Toxicology, Indiana University School of Medicine, Indianapolis, Indiana

Requests for reprints: Mark R. Kelley, Department of Pediatrics, Herman B Wells Center for Pediatric Research, 1044 W. Walnut, R4-302C, Indianapolis, IN 46202. Phone: 317-274-2755; Fax: 317-278-9298. E-mail: mkelley{at}iupui.edu

Osteosarcoma is the most common highly malignant bone tumor with primary appearance during the second and third decade of life. It is associated with a high risk of relapse, possibly resulting from a developed resistance to chemotherapy agents. As a means to overcome osteosarcoma tumor cell resistance and/or to sensitize tumor cells to currently used chemotherapeutic treatments, we examined the role of human apurinic endonuclease 1 (APE1) in osteosarcoma tumor cell resistance and prognosis. Sixty human samples of archived conventional (intramedullary) osteosarcoma were analyzed. APE1 protein was elevated in 72% of these tissues and among those with a known clinical outcome, there was a significant correlation between high APE1 expression levels and reduced survival times. The remaining 28% of samples showed low expression of APE1. Given that APE1 was overexpressed in osteosarcoma, we decreased APE1 levels using silencing RNA (siRNA) targeting technology in the osteosarcoma cell line, human osteogenic sarcoma (HOS), to enhance chemo- and radiation sensitivity. Using siRNA targeted technology of APE1, protein levels were reduced by more than 90% within 24 hours, remained low for 72 hours, and returned to normal levels at 96 hours. There was also a clear loss of APE1 endonuclease activity following APE1-siRNA treatment. A decrease in APE1 levels in siRNA-treated human osteogenic sarcoma cells led to enhanced cell sensitization to the DNA damaging agents: methyl methanesulfonate, H₂O₂, ionizing radiation, and chemotherapeutic agents. The findings presented here have both prognostic and therapeutic implications for treating osteosarcoma. The APE1-siRNA results demonstrate the feasibility for the therapeutic modulation of APE1 using a variety of molecules and approaches.

Key Words: APE1/ref-1 • siRNA • base excision repair • translational • osteosarcoma

Grant support: NIH CA094025, CA106298, NS38506, ES05865, ES03456, and P30 DK49218; CDMRP OC00113 grant (M. Kelley); and Riley Children's Foundation postdoctoral fellowship (M. Luo).

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

² M. Lou, A. Reed, Y. He, and M. R. Kelley, unpublished data.

³ Data in preparation.

Received 2/19/04; revised 4/ 8/04; accepted 4/13/04.