title: A bidirectional Tet-dependent promotor construct regulating the expression of E1A for tight control of oncolytic adenovirus replication.
abstract: Tight regulation of oncolytic adenoviruses (oAdV) represents an important requirement for their safe application. Here we describe a new doxycycline (Dox)-dependent oAdV with a bidirectional expression cassette, which drives the expression of the reverse tetracycline-controlled transactivator (rtTA(s)-M2) from a lung tumor-specific promoter and, in the opposite direction, the expression of the adenoviral E1A gene from a second generation TetO(7) sequence linked to an isolated TATA box. In H441 lung cancer cells, this oAdV showed a strictly Dox-dependent E1A expression, adenoviral replication, cell killing activity and a 450-fold induction of progeny virus production. The virus could be shut off again by withdrawal of Dox and, in contrast to a control oAdV expressing E1A directly from the SP-B promoter, did not replicate in non-target cells. However, the absolute values of virus production and the cell killing activity in the presence of the inducer were still reduced as compared to the control oAdV. The results demonstrate, for the first time, Dox-dependent oAdV replication from a single adenoviral vector genome. Future improvement of the Dox-dependent E1A regulation cassette should lead to the generation of an oAdV well suited to meet the demands for a highly regulated and efficient oncolytic virus for in vivo applications.
authors.affiliation: Department of Cardiology and Pulmology, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Hindenburgdamm 30, 12200 Berlin, Germany. henry.fechner@charite.de
authors.forename: Henry
authors.initials: H
authors.lastname: Fechner
authors.forename: Xiaomin
authors.initials: X
authors.lastname: Wang
authors.forename: Almudena Hurtado
authors.initials: AH
authors.lastname: Picó
authors.forename: Judith
authors.initials: J
authors.lastname: Wildner
authors.forename: Lennart
authors.initials: L
authors.lastname: Suckau
authors.forename: Sandra
authors.initials: S
authors.lastname: Pinkert
authors.forename: Isaac
authors.initials: I
authors.lastname: Sipo
authors.forename: Stefan
authors.initials: S
authors.lastname: Weger
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Poller
mesh_terms: D000256:Adenoviridae / Q000235:genetics* / Q000502:physiology*; D017500:Adenovirus E1A Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D004259:DNA-Directed DNA Polymerase / Q000187:drug effects / Q000235:genetics*; D004318:Doxycycline / Q000494:pharmacology; D005818:Genetic Engineering; D005822:Genetic Vectors / Q000235:genetics / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D050504:Oncolytic Viruses / Q000235:genetics / Q000502:physiology*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D014779:Virus Replication
chemical_list: D017500:Adenovirus E1A Proteins; C062255:Tet DNA polymerase; D004259:DNA-Directed DNA Polymerase; D004318:Doxycycline