Antitumor Effects of Systemically Delivered Adenovirus Harboring Trans-Splicing Ribozyme in Intrahepatic Colon Cancer Mouse Model Jin-Sook Jeong, 1 Seong-Wook Lee, 2 Seung-Hee Hong, 3 Yoon-Jong Lee, 3 Haeng-Im Jung, 3 Kyung-Sook Cho, 3 Hye-Hyun Seo, 3 Sang-Jin Lee, 3 Sohee Park, 3 Min-Sun Song, 2 Chang-Min Kim, 3 and In-Hoo Kim 3 Abstract Purpose: Our previous studies suggested that human telomerase reverse transcriptase (hTERT) RNA-targeting trans-splicing ribozyme could be a useful tool for cancer gene therapy. Here, we investigated whether adenoviruses harboring this ribozyme can be systemically delivered to mice, and whether they selectively mark tumors expressing hTERT and sensitize them to ganciclovir treatments. Experimental Design: We constructed adenoviral vectors containing modified hTERT-targeting trans-splicing ribozyme with downstream reporter gene (Ad-Ribo-LacZ) or suicide gene (Ad- Ribo-HSVtk) driven by a cytomegalovirus promoter. The tumor-specific trans-splicing reaction and the tumor-killing effect of adenoviruses harboring ribozyme were investigated both in vitro and in vivo using mice with intrahepatic colon cancer metastasis via systemic administration. The safety of systemic administration of the viruses was also evaluated. Results: We showed that Ad-Ribo-LacZ, when injected i.v., performs a highly specific trans- splicing reaction on hTERT mRNA and that it selectively marks tumors expressing hTERT in mice. More importantly, i.v. injection of Ad-Ribo-HSVtk plus ganciclovir significantly reduced tumor burden, with minimal liver toxicity, in mice with metastatic liver cancer, compared with the untreated group (P = 0.0009). Moreover, animals receiving Ad-Ribo-HSVtk showed improved survival compared with controls (P < 0.0001). Conclusions: This study shows that systemically delivered adenovirus harboring trans-splicing ribozyme can recognize cancer-specific transcripts and reprogram them to combat the cancer cells. Use of trans-splicing ribozymes seems to be a potentially useful gene therapy for cancer. Colorectal cancer is one of the most common cancers worldwide, both in terms of incidence and mortality (1). Despite progress in technologies for diagnosis and screening, liver metastasis occurs in 20% to 70% of patients with colorectal cancer and is the leading cause of death. Surgical resection or surgery coupled with systemic chemotherapy of liver metastasis is the treatment currently available for these patients (2). However, only 25% of liver metastases are amenable to surgery and f60% of those patients relapse (3). Therefore, there remains a continued need for a universally effective treatment for cancer and for new therapeutic approaches. Ribozymes are potential tools for gene therapy because of their ability to discriminate targets that differ by a single nucleotide, and splice and edit RNA transcripts (4, 5). The self- splicing group I intron from Tetrahymena thermophila was shown to catalyze its own excision without the aid of proteins and also perform a trans -splicing reaction that specifically cleaves target RNAs, leading to revision of gene information (6–8). In performing this trans -splicing reaction, group I ribozyme recognizes and binds a substrate RNA, cleaves the targeted RNA, and splices a 3¶ exon sequence that is initially attached to the 3¶ end of the ribozyme (9 – 11). The trans - splicing ribozyme has therefore been used to repair defective RNA messages such as mutant transcripts associated with genetic diseases (12 – 14) and cancer (15, 16). A previous study suggested that group I trans -splicing ribozyme can be used to splice therapeutic RNA sequences Cancer Therapy: Preclinical Authors’Affiliations: 1 Department of Pathology and Medical Research Center for Cancer Molecular Therapy, Dong-A University College of Medicine, Busan, Korea; 2 Department of Molecular Biology, Institute of Nanosensor and Biotechnology, Dankook University, Yongin, Korea; and 3 Research Institute and Hospital, National Cancer Center, Goyang, Korea Received 6/22/07; revised 9/26/07; accepted 10/9/07. Grant support: National Cancer Center grant 04100203, and KOSEF grants for the Medical Research Center at Dong-A University (J-S. Jeong) and M10534000004- 06N3400-00410 (S-W. Lee). 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. Note: J-S. Jeong and S-W. Lee contributed equally to this work. Present address for S-H. Hong and Y-J. Lee: Korea Human Resource Development Institute for Health and Welfare, Seoul 122-701, Korea. Requests for reprints: In-Hoo Kim, Molecular Imaging and Therapy Branch, Research Institute, National Cancer Center, 809 Madu-dong, Ilsan-gu, Goyang-si, Gyeonggi 410-769, Korea. Phone: 31-920-2580; Fax: 31-920-2542; E-mail: ihkim@ncc.re.kr. F 2008 American Association for Cancer Research. doi:10.1158/1078-0432.CCR-07-1524 www.aacrjournals.org Clin Cancer Res 2008;14(1) January 1, 2008 281 Downloaded from http://aacrjournals.org/clincancerres/article-pdf/14/1/281/1973837/281.pdf by guest on 17 June 2022