title: Characterization of human tumor cell lines transduced with the cDNA encoding either tumor necrosis factor alpha (TNF-a) or interleukin-2 (IL-2).
abstract: Tumor cell lines were generated from cancer patients, 17 with metastatic melanoma and one with colon adenocarcinoma. The lines were characterized as tumor cells by the presence of tumor associated antigens demonstrated by indirect immunofluorescence and analysing using a fluorescence activated cell sorter (FACS). The tumor cell lines were transduced using retroviruses encoding neomycin phosphotransferase and either human tumor necrosis factor alpha (TNF-alpha) or interleukin-2 (IL-2). Following transduction, cells were selected and grown in the neomycin analogue G418. Fibroblasts overgrew tumor cells in 6/18 cases following selection in G418 and 1/18 lines did not grow at all after selection. In the remaining 11 lines the expression of tumor associated antigens, growth, and susceptibility to lysis by LAK cells was similar between the selected transduced tumor cell lines and the nontransduced controls. Of the lines tested, all were positive for the presence of the cytokine gene by Southern blot or PCR analysis. In addition, no replication competent retrovirus was detected in the cell lines following transduction using an extended mink S+L- focus assay. The amount of specific cytokine produced per 10(5) transduced tumor cells in 24 h ranged from 0.2 ng to 5.8 ng of TNF-alpha for the TNF transduced lines and from 0.1 ng to 3.6 ng of IL-2 for the IL-2 transduced tumor cell lines. One transduced tumor cell line examined maintained consistent levels of cytokine production when studied at 15 different time intervals over a period of 198 days. Additionally, 40,000 rads of gamma irradiation did not stop cytokine production from two transduced tumor cell lines when studied over 6 days. This study demonstrates the feasibility of growing human tumor cell lines from surgical biopsies and genetically modifying those lines to produce a cytokine of choice for possible use as a tumor cell vaccine.
authors: Surgery Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.
authors: J R
authors: JR
authors: Yannelli
authors: C
authors: C
authors: Hyatt
authors: S
authors: S
authors: Johnson
authors: P
authors: P
authors: Hwu
authors: S A
authors: SA
authors: Rosenberg
mesh_terms: D000230:Adenocarcinoma / Q000276:immunology / Q000473:pathology*; D000951:Antigens, Neoplasm / Q000032:analysis; D000954:Antigens, Surface / Q000032:analysis; D001706:Biopsy; D015139:Blotting, Southern; D003110:Colonic Neoplasms / Q000276:immunology / Q000473:pathology*; D016207:Cytokines / Q000096:biosynthesis; D005434:Flow Cytometry; D005720:Gamma Rays; D005796:Genes; D006801:Humans; D066298:In Vitro Techniques; D007376:Interleukin-2 / Q000502:physiology*; D015979:Killer Cells, Lymphokine-Activated / Q000276:immunology; D008545:Melanoma / Q000276:immunology / Q000473:pathology*; D016133:Polymerase Chain Reaction; D013997:Time Factors; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000276:immunology* / Q000528:radiation effects; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology*
chemical_list: D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D016207:Cytokines; D007376:Interleukin-2; D014409:Tumor Necrosis Factor-alpha