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KMID : 0620919970290010065
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Experimental & Molecular Medicine
1997 Volume.29 No. 1 p.65 ~ p.69
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Production of IL-12 from gene modified human dermal fibroblasts:a preclinical study for IL-12 cancer gene therapy
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Chae Hwa Park/Chae Hwa Park
Won Ki Kang/Mi Sook Oh/Won Seong Kim/Jung Hyun Yang/Michael T Lotze/Sun Young Kim/Keun Chil Park/Chan H Park
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Abstract
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Cytokine has been used as an immune stimulator and administered to patients for a treatment of cancer. Interleukin-12 (IL-12) is a potent cytokine which acts through a variety of functions including interferon-γ production and cytotoxic T-cell activation. Considering the toxicity of high dose systemic IL-12 administration into human, local administration of low dose IL-12 can be a more efficient strategy. In ex vivo therapy, human dermal fibroblast has been considered as a useful vehicle for transferring genes, Here we show that human dermal fibroblast transduced with retrovirus containing IL-12 gene can be manipulated to produce reasonable amount of IL-12 protein. Human dermal fibroblast was isolated from freshly harvested skin specimens by collagenase digestion, grown in primary cultures, and transduced with a retroviral vector containing genes for human IL-12 and a selectable marker Neo(R). Following selection in G418, IL-12 producing fibroblasts were tested for secreted IL-12 level by ELISA. Six specimens of human skin were processed to obtain fibroblasts. ELISA results show that 40-150 units of IL-12 was produced for 24 h from 1x10(6) cells of transduced and selected fibroblast cultures. The primary cultures were maintained for up to nine passages about 108 days. The mean ± overall time for obtaining enough number of cells was 49 ± 2 days. The fibroblasts continued to produce IL-12 in culture for 90 days. These preliminary results can be used for the design of ex vivo gene therapy clinical trial using human dermal fibroblast.
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KEYWORD
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cancer gene therapy, human fibroblast, IL-12,
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