title: TK.007: A novel, codon-optimized HSVtk(A168H) mutant for suicide gene therapy.
abstract: Conditional elimination of infused gene-modified alloreactive T cells, using suicide gene activation, has been shown to be an efficient strategy to abrogate severe graft-versus-host disease (GvHD) in the context of adoptive immunotherapy. To overcome shortcomings of the most widely used suicide gene, wild-type (splice-corrected) herpes simplex virus thymidine kinase (scHSVtk), we generated two new variants: the codon-optimized coHSVtk and, by introducing an additional mutation (A168H), the novel TK.007. We transduced human hematopoietic cell lines and primary T cells with retroviral "sort-suicide vectors" encoding combinations of selection markers (tCD34 and OuaSelect) with one of three HSVtk variants. In vitro we observed higher expression levels and sustained long-term expression of TK.007, indicating lower nonspecific toxicity. Also, we noted significantly improved kinetics of ganciclovir (GCV)-mediated killing for TK.007-transduced cells. In an experimental (murine) allogeneic transplantation model, TK.007-transduced T cells mediated severe GvHD, which was readily abrogated by application of GCV (10 mg/kg). Last, we established a modified allotransplantation model that allowed quantitative comparison of the in vivo activities of TK.007 versus scHSVtk. We found that TK.007 mediates both significantly faster and higher absolute killing at low GCV concentrations (10 and 25 mg/kg). In summary, we demonstrate that the novel TK.007 suicide gene combines better killing performance with reduced nonspecific toxicity (as compared with the frequently used splice-corrected wild-type scHSVtk gene), thus representing a promising alternative for suicide gene therapy.
authors.affiliation: Clinic for Stem Cell Transplantation, Research Department of Cell and Gene Therapy, University Medical Centre Hamburg-Eppendorf , 20246 Hamburg, Germany.
authors.forename: Ellen
authors.initials: E
authors.lastname: Preuss
authors.forename: Alexandra
authors.initials: A
authors.lastname: Treschow
authors.forename: Sebastian
authors.initials: S
authors.lastname: Newrzela
authors.forename: Daniela
authors.initials: D
authors.lastname: Brücher
authors.forename: Kristoffer
authors.initials: K
authors.lastname: Weber
authors.forename: Ulrika
authors.initials: U
authors.lastname: Felldin
authors.forename: Evren
authors.initials: E
authors.lastname: Alici
authors.forename: Gösta
authors.initials: G
authors.lastname: Gahrton
authors.forename: Dorothee
authors.initials: D
authors.lastname: von Laer
authors.forename: M Sirac
authors.initials: MS
authors.lastname: Dilber
authors.forename: Boris
authors.initials: B
authors.lastname: Fehse
mesh_terms: D000818:Animals; D002460:Cell Line; D003062:Codon / Q000235:genetics / Q000378:metabolism; D015774:Ganciclovir / Q000378:metabolism; D041722:Genes, Transgenic, Suicide; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006086:Graft vs Host Disease / Q000235:genetics / Q000628:therapy*; D006801:Humans; D016219:Immunotherapy, Adoptive; D051379:Mice; D008810:Mice, Inbred C57BL; D012190:Retroviridae / Q000235:genetics; D013601:T-Lymphocytes / Q000378:metabolism / Q000821:virology; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism*; D014161:Transduction, Genetic
chemical_list: D003062:Codon; D013937:Thymidine Kinase; D015774:Ganciclovir