A New Experimental Approach to the Specific Immunotherapy for Malignant Brain Tumor

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Abstract
The in vivo efficacy of T-cell growth factor (TCGF)-dependent tumor-specific cytotoxic T lymphocyte (CTL) clone against a syngeneic murine malignant glioma (a 20-methylcholanthrene-induced ependymoblastoma of C57BL/6 mouse origin, 203-glioma) was investigated. Prior to this goal, the immunobiological properties of TCGF were first characterized in culture supernatants of concanavalin A (Con A)-activated murine (C57BL/6) or rat (Wistar) lymphocytes, respectively. 5×106/ml lymphocytes were cultured with 2 μg/ml Con A in RPMI-1640 medium supplemented with 2% fetal calf serum for 48 hours. The culture supernatants (Con A sup) were concentrated by ammonium sulphate precipitation and partially purified by successive diethylaminoethyl (DEAE)-cellulose ion exchange chromatography and Sephadex G-100 gel filtration. TCGF was assayed for a quantitative analysis using a TCGF-dependent murine glioma-specific CTL clone (G-CTLL). The major portion of TCGF activity in murine and rat Con A sup was recovered in the 50 to 80%, and 40 to 80% (NH4)2SO4 fraction, respectively. These activities were found in proteins of (3.0 ?? 3.6) × 104, (1.2 ?? 1.8) × 104 daltons in size as estimated by gel filtration, respectively and eluted from ion exchange chromatography in a NaCl gradient on 150, and 50 mM, respectively. It was also found that rat Con A sup contained TCGF activity approximately 8 times as much as murine Con A sup and that the CTL activity of sensitized T lymphocytes was fully enhanced in a short-term culture with TCGF. The substantial in vivo anti-tumor activity of G-CTLL cell line was observed from the intracranial Winn-type neutralization assay and the systemic adoptive transfer assay. Accordingly, this new experimental approach to the specific immunotherapy with TCGF and the cloned CTL line such as G-CTLL may be particularly useful for malignant gliomas.

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