Human brain tumor cell culture characterization after immunostimulatory gene transfer

Ian F. Parney, Maxine A. Farr-Jones, Anu Koshal, Lung Ji Chang, Kenneth C. Petruk, Charles Y. Liu, Berislav V. Zlokovic, James T. Rutka

Research output: Contribution to journalArticlepeer-review

6 Scopus citations


OBJECTIVE: Immunogene therapy is a novel cancer treatment strategy based on vaccination with irradiated autologous tumor cells transduced with immunostimulatory genes. To characterize such cells before clinical applications, we studied a human glioma cell line (D54 MG) and early passage human glioma (Ed147.BT, Ed149.BT) and melanoma (Ed141.MEL) cultures after immunostimulatory gene transfer. METHODS: Granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-12 (IL-12), and B7-2 genes were retrovirally transferred to tumor cells. Gene expression before and after irradiation (200 Gy) was assessed by enzyme-linked immunosorbent assay (GM-CSF, IL-12) and flow cytometry (B7-2). Viability and clonogenicity were determined via trypan blue staining before and after irradiation. Growth rates were determined by serial cell counts. RESULTS: GM-CSF expression was high in GM-CSF-transduced (10.36-162.10 ng/106 cells/d preirradiation and 10.22-122.02 ng/106 cells/d postirradiation) but lower in B7-2/GM-CSF-transduced cultures (1.41-2.90 ng/106 cells/d preirradiation, 1.96-5.02 ng/106 cells/d postirradiation). IL-12 expression also was lower (1.30-2.10 ng/106 cells/d preirradiation, 0.47-1.70 ng/106 cells/d postirradiation). B7-2 expression was high (one- to two-logarithm increase in fluorescence) and unaffected by radiation. Postirradiation viability was initially high (94.20 ± 8.46%, Day 1) but decreased rapidly (28.13 ± 4.64%, Day 10). No cultures demonstrated evidence of clonogenicity (i.e., cell division) after 200-Gy irradiation. Growth rates were similar in wild-type and gene-transduced Ed141.MEL, Ed147.BT, and Ed149.BT. However, D54MG-IL-12 growth was slower than that of wild-type D54MG. CONCLUSION: GM-CSF, IL-12, and B7-2 genes can be transferred to human glioma and melanoma cell cultures efficiently by use of our retroviral vectors. Irradiation (200 Gy) does not significantly alter therapeutic gene expression. Irradiated cells remain viable for several days but cannot undergo further cell division. Early passage culture growth rates are not altered by therapeutic gene expression but are decreased by IL-12 in an immortalized cell line (D54MG). These results suggest that it is feasible to create vaccines with irradiated, autologous, genetically modified brain tumor cells.

Original languageEnglish (US)
Pages (from-to)1094-1102
Number of pages9
Issue number5
StatePublished - May 1 2002


  • Gene therapy
  • Glioma
  • Immunogene therapy
  • Immunology
  • Melanoma

ASJC Scopus subject areas

  • Surgery
  • Clinical Neurology


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