title: The murine-reduced folate carrier gene can act as a selectable marker and a suicide gene in hematopoietic cells in vivo.
abstract: Increased expression of the reduced folate carrier confers sensitivity to the antifolate drug methotrexate because it results in increased cellular uptake of this drug, and increased resistance to trimetrexate, a lipid-soluble antifolate drug, because it enables cells to take up exogenous folates that rescue cells from antifolate cytotoxicity. We therefore hypothesized that the reduced folate carrier could act as a suicide gene after treatment with methotrexate and as a selectable marker after exposure to trimetrexate. To test this hypothesis, we constructed replication-defective retroviruses containing the murine-reduced folate carrier (mRFC). Murine bone marrow cells transduced with the mRFC-containing retrovirus showed increased sensitivity to methotrexate and increased resistance to trimetrexate compared to empty vector-transduced controls in colony forming assays. Furthermore, colonies surviving trimetrexate and methotrexate treatment showed an enrichment of the mRFC gene after exposure to trimetrexate and a decrease after exposure to methotrexate. Lethally irradiated mice transplanted with bone marrow cells transduced with the mRFC-retrovirus and treated with the antifolate drugs after hematopoietic recovery demonstrated a relative increase in the number of cells containing the mRFC transgene after trimetrexate treatment and a decrease after methotrexate treatment. Therefore, these studies demonstrate the potential of the reduced folate carrier gene to play a dual role in gene therapy applications.
authors.affiliation: Department of Pediatrics, University of Kentucky, Lexington, KY 40536, USA.
authors.forename: Shuqian
authors.initials: S
authors.lastname: Liu
authors.forename: Lifu
authors.initials: L
authors.lastname: Song
authors.forename: Robert
authors.initials: R
authors.lastname: Bevins
authors.forename: Ogheneruemu
authors.initials: O
authors.lastname: Birhiray
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Moscow
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000493:pharmacokinetics* / Q000494:pharmacology; D016026:Bone Marrow Transplantation; D001943:Breast Neoplasms / Q000473:pathology; D002352:Carrier Proteins / Q000235:genetics* / Q000502:physiology; D016923:Cell Death; D003114:Colony-Forming Units Assay; D003673:Defective Viruses / Q000235:genetics*; D019008:Drug Resistance, Neoplasm; D005260:Female; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics*; D006085:Graft Survival; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D026901:Membrane Transport Proteins; D008727:Methotrexate / Q000493:pharmacokinetics* / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D016133:Polymerase Chain Reaction; D011828:Radiation Chimera; D058978:Reduced Folate Carrier Protein; D012190:Retroviridae / Q000235:genetics*; D012641:Selection, Genetic; D014161:Transduction, Genetic; D016597:Trimetrexate / Q000493:pharmacokinetics* / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000821:virology
chemical_list: D000970:Antineoplastic Agents; D002352:Carrier Proteins; D026901:Membrane Transport Proteins; D058978:Reduced Folate Carrier Protein; C548699:SLC19A1 protein, human; C548700:Slc19a1 protein, mouse; D016597:Trimetrexate; D008727:Methotrexate