title: Development of a therapeutic adenoviral vector for cholangiocarcinoma combining tumor-restricted gene expression and infectivity enhancement.
abstract: Cholangiocarcinoma is an invasive malignancy that is most often unresectable upon diagnosis and unresponsive to chemotherapy and radiation. While adenoviral gene therapy has shown promise in treating many tumors, systemic toxicity and low tumor transduction efficiency have hampered its application in many gastrointestinal cancers. To overcome these difficulties, we have constructed an adenoviral vector utilizing a tumor-specific promoter (TSP) for selective transgene expression and a vector with an RGD-motif in the fiber-knob region for infectivity enhancement. In seeking a TSP for cholangiocarcinoma, Secretory Leukoprotease Inhibitor, Midkine, Gastrin Releasing Peptide, VEGF, Cox-2M, and Cox-2L promoters were configures in adenoviral vectors, and evaluated in cholangiocarcinoma cells lines (Oz and SkChA-1). Luciferase assays demonstrated that Cox-2 promoters (M and L) showed the highest promoter activity, with Cox-2M appearing slightly stronger than Cox-2L. Infectivity enhanced vectors with RGD-motif in the fiber-knob region were also constructed with the luciferase transgene driven by a CMV control and the Cox-2M and Cox-2L promoters. Subsequent luciferase assays comparing the unmodified vectors to the RGD-modified versions demonstrated higher levels of luciferase activity than the RGD-infected cells. This paradigm was then applied to a therapeutic HSV-TK/GCV model by constructing RGD-enhanced HSV-TK vectors driven by Cox-2M and Cox-2L promoters. In vitro cytocidal effect analysis confirmed that the RGD-modified, cox-2 (M and L) driven vectors showed a stronger cytocidal effect upon gancyclovir administration than the vectors with wild-type fiber. The Cox-2 promoter demonstrates a favorable selectivity profile for cholangiocarcinoma, and RGD-modification further enhances transduction efficiency. This combination has potential to overcome the obstacles to clinical application of adenoviral gene therapy in cholangiocarcinoma.
authors.affiliation: Division of Human Gene Therapy, The Gene Therapy Center at UAB, Department of Medicine, University of Alabama at Birmingham, Birmingham, AL, USA.
authors.forename: Peter
authors.initials: P
authors.lastname: Nagi
authors.forename: Selwyn M
authors.initials: SM
authors.lastname: Vickers
authors.forename: Julia
authors.initials: J
authors.lastname: Davydova
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Adachi
authors.forename: Koichi
authors.initials: K
authors.lastname: Takayama
authors.forename: Shannon
authors.initials: S
authors.lastname: Barker
authors.forename: Victor
authors.initials: V
authors.lastname: Krasnykh
authors.forename: David T
authors.initials: DT
authors.lastname: Curiel
authors.forename: Masato
authors.initials: M
authors.lastname: Yamamoto
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D001650:Bile Duct Neoplasms / Q000628:therapy*; D001653:Bile Ducts, Intrahepatic; D018281:Cholangiocarcinoma / Q000628:therapy*; D015870:Gene Expression; D017930:Genes, Reporter; D015316:Genetic Therapy; D005822:Genetic Vectors; D006801:Humans; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured