Transduction of green fluorescent protein increased oxidative stress and enhanced sensitivity to cytotoxic drugs in neuroblastoma cell lines.
Green fluorescent protein (GFP) is employed as a selection marker for gene transduction and to track tumor cells. Transduction of enhanced GFP (eGFP) into human neuroblastoma cell lines via a lentiviral vector significantly sensitized CHLA-20 (wild-type and functional TP53), and to a lesser extent CHLA-90 cells (multidrug-resistant, mutant, and nonfunctional TP53) to carboplatin, doxorubicin, etoposide, or melphalan, relative to cells transduced using the cell surface antigen CD80 as a selection marker. Total glutathione (GSH) was significantly up-regulated (1.8- to 2.8-fold) after eGFP (but not CD80) transduction in cell lines with, but not in those lacking, functional p53. Cytotoxicity of GSH depletion by buthionine sulfoximine in CHLA-20 (but not in CHLA-20-eGFP) was diminished by hypoxia (2% O(2)). Thus, oxidative stress produced by GFP selects for cells with up-regulated GSH in a p53-dependent manner, and also enhanced the cytotoxicity of anticancer drugs in neuroblastoma cell lines. Our data suggest caution when employing GFP-transduced cells to assess drug sensitivity and that using a cell surface antigen as a selection marker for gene transduction may perturb cells less than GFP.
Goto
Hiroaki
H
Divisions of Hematology-Oncology and Research Immunology/Bone Marrow Transplantation, Childrens Hospital Los Angeles, CA 90027, USA.
Yang
Bo
B
Petersen
Denise
D
Pepper
Karen A
KA
Alfaro
Paul A
PA
Kohn
Donald B
DB
Reynolds
C Patrick
CP
D000970:Antineoplastic Agents / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects; D018122:B7-1 Antigen / Q000378:metabolism; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D016190:Carboplatin / Q000627:therapeutic use; D004317:Doxorubicin / Q000627:therapeutic use; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D005047:Etoposide / Q000627:therapeutic use; D005978:Glutathione / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism*; D008558:Melphalan / Q000627:therapeutic use; D009154:Mutation; D009447:Neuroblastoma / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress; D014161:Transduction, Genetic; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D000970:Antineoplastic Agents; D018122:B7-1 Antigen; D008164:Luminescent Proteins; D016159:Tumor Suppressor Protein p53; D049452:Green Fluorescent Proteins; D019328:Buthionine Sulfoximine; D005047:Etoposide; D004317:Doxorubicin; D016190:Carboplatin; D005978:Glutathione; D008558:Melphalan