title: Cancer cell specific cytotoxic gene expression mediated by ARF tumor suppressor promoter constructs.
abstract: In current cancer treatment protocols, such as radiation and chemotherapy, side effects on normal cells are major obstacles to radical therapy. To avoid these side effects, a cancer cell-specific approach is needed. One way to specifically target cancer cells is to utilize a cancer specific promoter to express a cytotoxic gene (suicide gene therapy) or a viral gene required for viral replication (oncolytic virotherapy). For this purpose, the selected promoter should have minimal activity in normal cells to avoid side effects, and high activity in a wide variety of cancers to obtain optimal therapeutic efficacy. In contrast to the AFP, CEA and PSA promoters, which have high activity only in a limited spectrum of tumors, the E2F1 promoter exhibits high activity in wide variety of cancers. This is based on the mechanism of carcinogenesis. Defects in the RB pathway and activation of the transcription factor E2F, the main target of the RB pathway, are observed in almost all cancers. Consequently, the E2F1 promoter, which is mainly regulated by E2F, has high activity in wide variety of cancers. However, E2F is also activated by growth stimulation in normal growing cells, suggesting that the E2F1 promoter may also be highly active in normal growing cells. In contrast, we found that the tumor suppressor ARF promoter is activated by deregulated E2F activity, induced by forced inactivation of pRB, but does not respond to physiological E2F activity induced by growth stimulation. We also found that the deregulated E2F activity, which activates the ARF promoter, is detected only in cancer cell lines. These observations suggest that ARF promoter is activated by E2F only in cancer cells and therefore may be more cancer cell-specific than E2F1 promoter to drive gene expression. We show here that the ARF promoter has lower activity in normal growing fibroblasts and shows higher cancer cell-specificity compared to the E2F1 promoter. We also demonstrate that adenovirus expressing HSV-TK under the control of the ARF promoter shows lower cytotoxicity than that of the E2F1 promoter, in normal growing fibroblasts but has equivalent cytotoxicity in cancer cell lines. These results suggest that the ARF promoter, which is specifically activated by deregulated E2F activity, is an excellent candidate to drive therapeutic cytotoxic gene expression, specifically in cancer cells.
authors.affiliation: Department of Bioscience, School of Science and Technology, Kwansei Gakuin University, 2-1 Gakuen, Sanda, Hyogo 669-1337, Japan.
authors.forename: Kenta
authors.initials: K
authors.lastname: Kurayoshi
authors.affiliation: Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary, University of London, John Vane Science Centre, Charterhouse Square, London EC1M 6BQ, UK.
authors.forename: Eiko
authors.initials: E
authors.lastname: Ozono
authors.affiliation: Department of Obstetrics and Gynecology, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO 80045, USA.
authors.forename: Ritsuko
authors.initials: R
authors.lastname: Iwanaga
authors.affiliation: Department of Obstetrics and Gynecology, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO 80045, USA.
authors.forename: Andrew P
authors.initials: AP
authors.lastname: Bradford
authors.affiliation: Center for Stem Cell Biology, Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109, USA.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Komori
authors.affiliation: Department of Bioscience, School of Science and Technology, Kwansei Gakuin University, 2-1 Gakuen, Sanda, Hyogo 669-1337, Japan. Electronic address: btm88939@kwansei.ac.jp.
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Ohtani
mesh_terms: D017209:Apoptosis / Q000235:genetics; D045744:Cell Line, Tumor; D050687:E2F1 Transcription Factor / Q000235:genetics*; D041722:Genes, Transgenic, Suicide / Q000235:genetics; D016147:Genes, Tumor Suppressor; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D009374:Neoplasms, Experimental / Q000235:genetics* / Q000628:therapy*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D025803:Tumor Suppressor Protein p14ARF / Q000235:genetics*; D025521:Tumor Suppressor Proteins / Q000235:genetics*
chemical_list: D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D025803:Tumor Suppressor Protein p14ARF; D025521:Tumor Suppressor Proteins
keywords: ARF promoter; Cancer-specific; E2F; E2F1 promoter; Normal growing cells; pRB