title: Lentiviral vector conferring resistance to mycophenolate mofetil and sensitivity to ganciclovir for in vivo T-cell selection.
abstract: Several clinical studies of gene-modified T cells have shown limited in vivo function of the cells, immunogenicity of the transgene, and lack of a selective advantage for gene-modified T cells. To address these problems, we developed a lentiviral vector (LV) that provides a selectable, proliferative advantage and potentially decreases immunogenicity for transduced T cells. The bicistronic vector expressed two genes linked with an internal ribosomal entry site. One gene is a variant of the inosine monophosphate dehydrogenase 2, inosine monophosphate dehydrogenase (IMPDH(IY)), conferring resistance to the immunosuppressive drug mycophenolate mofetil (MMF). The other is a suicide gene, herpes simplex virus thymidine kinase (HSV-TK), rendering proliferating cells sensitive to ablation with ganciclovir, fused to the selectable transmembrane marker DeltaCD34 (DeltaCD34/TK). Cells transduced with LV-DeltaCD34/TK.IMPDH(IY) were efficiently enriched by immunomagnetic selection for CD34, proliferated in 0.5-5 microM MMF, and were killed by 0.5-25 microg ml(-1) ganciclovir. We demonstrate efficient selection and killing of gene-modified cells and suggest LV-DeltaCD34/TK.IMPDH(IY)-transduced T cells could be used to facilitate allogeneic hematopoietic cell engraftment. The expression of IMPDH(IY) would allow in vivo selection with MMF, and DeltaCD34/TK expression would allow rapid and safe elimination of transduced T cells if graft-versus-host disease developed.
authors.affiliation: Transplantation Biology, Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.
authors.forename: D
authors.initials: D
authors.lastname: Sangiolo
authors.forename: M
authors.initials: M
authors.lastname: Lesnikova
authors.forename: R A
authors.initials: RA
authors.lastname: Nash
authors.forename: M C
authors.initials: MC
authors.lastname: Jensen
authors.forename: A
authors.initials: A
authors.lastname: Nikitine
authors.forename: H-P
authors.initials: HP
authors.lastname: Kiem
authors.forename: G E
authors.initials: GE
authors.lastname: Georges
mesh_terms: D000998:Antiviral Agents / Q000627:therapeutic use; D049109:Cell Proliferation; D003001:Cloning, Molecular; D004351:Drug Resistance; D015774:Ganciclovir / Q000627:therapeutic use; D041722:Genes, Transgenic, Suicide; D005818:Genetic Engineering; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage / Q000235:genetics*; D006086:Graft vs Host Disease / Q000517:prevention & control; D018380:Hematopoietic Stem Cell Transplantation; D018259:Herpesvirus 1, Human / Q000201:enzymology*; D006801:Humans; D007168:IMP Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D018189:Immunomagnetic Separation / Q000379:methods; D007166:Immunosuppressive Agents / Q000494:pharmacology; D020014:K562 Cells; D009173:Mycophenolic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000821:virology; D013937:Thymidine Kinase / Q000235:genetics*; D014161:Transduction, Genetic / Q000379:methods; D014184:Transplantation, Homologous