The murine-reduced folate carrier gene can act as a selectable marker and a suicide gene in hematopoietic cells in vivo.
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.
Liu
Shuqian
S
Department of Pediatrics, University of Kentucky, Lexington, KY 40536, USA.
Song
Lifu
L
Bevins
Robert
R
Birhiray
Ogheneruemu
O
Moscow
Jeffrey A
JA
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
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