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Research Articles: Therapeutics

Construction of a novel DNA decoy that inhibits the oncogenic ß-catenin/T-cell factor pathway

¹ Department of Surgery; ² Division of Clinical Gene Therapy, Department of Geriatric Medicine; ³ Division of Gene Therapy Science, Graduate School of Medicine; ⁴ Department of Pathology, School of Allied Health Science, Faculty of Medicine, Osaka University, Osaka, Japan; and ⁵ Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, Washington, District of Columbia

Requests for reprints: Hirofumi Yamamoto, Department of Surgery and Clinical Oncology, Graduate School of Medicine, Osaka University, 2-2 Yamada-oka, Suita City, 565-0871 Osaka, Japan. Phone: 81-6-6879-3251; Fax: 81-6-6879-3259. E-mail: kobunyam{at}surg2.med.osaka-u.ac.jp

Abstract

The oncogenic ß-catenin/T-cell factor (TCF) signal is a common trigger inducing expressions of various cancer-related genes and is activated in various types of human malignancy. The aim of this study was to create an effective double-stranded DNA decoy that would interfere with endogenous TCF hyperactivity in tumor cells. We first established the TCF-activated model using nontumor human embryonic kidney 293 (HEK293) cells by introducing a ß-catenin cDNA. Based on a consensus TCF-binding sequence in the cyclin D1 and c-myc promoters, several double-stranded oligodeoxynucleotides were designed and tested for their ability to inhibit TCF activity in the HEK293 model. Among them, the 18-mer oligodeoxynucleotide stably formed double-stranded DNA and efficiently inhibited TCF activity. FITC-labeled oligodeoxynucleotide was efficiently incorporated into the nucleus at 6 hours and remained within cells for up to 72 to 96 hours. When compared with scrambled oligodeoxynucleotide, we found that the 18-mer TCF decoy significantly inhibited TCF activity and promoter activities of the downstream target genes, such as cyclin D1, c-myc, and matrix metalloproteinase 7 in HCT116 colon cancer cells. Reverse transcription-PCR assays indicated that mRNA expression of these genes decreased with treatment of the TCF decoy. Proliferation assay showed that the TCF decoy significantly inhibited growth of HCT116 tumor cells, but not of nontumor HEK293 cells. Our data provide evidence that the TCF decoy reduced both TCF activity and transcriptional activation of downstream target genes. Thus, this TCF decoy is potentially an efficient and nontoxic molecular targeting therapy for controlling malignant properties of cancer cells. [Mol Cancer Ther 2006;5(4):985–94]

Grant support: Grant-in Aid for Cancer Research from the Ministry of Education, Science, Sports, and Culture Technology, Japan (H. Yamamoto), and a grant for the third-term Comprehensive Strategy for Cancer Control from the Ministry of Health Labour and Welfare.

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.

Received 9/26/05; revised 12/30/05; accepted 1/25/06.