title: Transcriptional targeting of virus-mediated gene transfer by the human hexokinase II promoter.
abstract: The efficacy of the most commonly used form of suicide gene therapy, the HSV-TK/GCV method, utilizing herpes simplex virus thymidine kinase (HSV-TK) and antiviral drug ganciclovir (GCV) has been demonstrated in clinical trials. However, safer delivery of the therapeutic gene and more controlled regulation of the transgene expression, the essential prerequisites for successful therapeutic use, are still needed. We describe improved suicide gene therapy against cancer through transcripitional targeting by a strong and selective tumor-specific human hexokinase II promoter (hHKII). We examined the targeting properties of the human hHKII promoter in different human non-small cell lung cancer (NSCLC) and other human cancer cell lines using self-inactivating, VSV-G pseudotyped lentiviral vector. To confirm accurate transcriptional targeting of the hHKII promoter, the lack of transgene expression was verified in human primary bronchial epithelial and bronchial fibroblast cells. Furthermore, tissue-specific expression of the promoter was confirmed using transgenic mouse lines carrying the hHKII promoter driven luciferase reporter gene. We also tested the efficacy of the HSV-TK/GCV suicide gene therapy with the hHKII targeted lentiviral vector to NSCLC cells. Our results show that the hHKII promoter is strongly expressed in cancer cells. The targeted vector with the shortest hHKII promoter fragment (352 bp) appeared to have the best targeting properties because it efficiently governed the expression of the therapeutic gene in cancer cell lines, especially in certain non-small cell lung cancer cell lines, the transgene expression in human primary cells was virtually undetectable, and expression of the proximal hHKII promoter in transgenic mice was very low in most tissues. Also, the anti-cancer efficacy of HSV-TK/GCV therapy with the hHKII-targeted vector was comparable to that obtained with the control vector that utilized a commonly used constitutive promoter from the human elongation factor 1 alpha (hEF1alpha) gene. In conclusion, the transcriptionally targeted lentivirus vector with hHKII promoter can successfully direct HSV-TK/GCV suicide gene therapy to non-small cell lung cancer and other tumor cell types. These results warrant further studies with orthotopic animal tumor models and primary human cancer material.
authors.affiliation: A.I. Virtanen Institute, Department of Biotechnology and Molecular Medicine, University of Kuopio, Kuopio, Finland.
authors.forename: Ann-Marie
authors.initials: AM
authors.lastname: Määttä
authors.forename: Sanna
authors.initials: S
authors.lastname: Korja
authors.forename: Heli
authors.initials: H
authors.lastname: Venhoranta
authors.forename: Tanja
authors.initials: T
authors.lastname: Hakkarainen
authors.forename: Eija
authors.initials: E
authors.lastname: Pirinen
authors.forename: Sami
authors.initials: S
authors.lastname: Heikkinen
authors.forename: Riikka
authors.initials: R
authors.lastname: Pellinen
authors.forename: Kimmo
authors.initials: K
authors.lastname: Mäkinen
authors.forename: Jarmo
authors.initials: J
authors.lastname: Wahlfors
mesh_terms: D000818:Animals; D002289:Carcinoma, Non-Small-Cell Lung / Q000628:therapy; D045744:Cell Line, Tumor; D018014:Gene Transfer Techniques; D041722:Genes, Transgenic, Suicide; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006593:Hexokinase / Q000235:genetics*; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D008175:Lung Neoplasms / Q000628:therapy; D051379:Mice; D008822:Mice, Transgenic; D009369:Neoplasms / Q000628:therapy*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D013937:Thymidine Kinase / Q000235:genetics; D014161:Transduction, Genetic
chemical_list: D006593:Hexokinase; D013937:Thymidine Kinase