Cancer Immunol Irnmunother (1988) 26:273-279 ancer mmunology mmunotherapy © Springer-Verlag 1988 Effects of type beta transforming growth factor in combination with retinoic acid or tumor necrosis factor on proliferation of a human glioblastoma cell line and clonogenic cells from freshly resected human brain tumors Eirik Helseth 1, Geirmund Unsgaard l, Are Dalen 2, and Randi Vik 1 The Institute of Cancer Research, ~Department of Neurosurgery, 2 Department of Virology, University of Trondheim, Trondheim Regional Hospital, N-7000 Trondheim, Norway Summary. Type beta transforming growth factor ([3-TGF) is a potent regulator of cell growth and differentiation. The human glioblastoma cell line, T-MG1, was growth inhib- ited by ~-TGF under anchorage independent conditions. The antiproliferative effect of [3-TGF was potentiated to nearly total arrest by low doses of retinoic acid (RA) or tu- mor necrosis factor (TNF), while epidermal growth factor, platelet-derived growth factor, interleukin-2, and gamma interferon did not have this potentiating effect. The poten- tiation of the [3-TGF effect by RA and TNF could not be explained by modulation of the epidermal growth factor receptor, the [3-TGF receptor, or the TNF receptor. ~-TGF alone and in combination with RA or TNF were further tested on primary cultures from freshly resected human glioma biopsies (n = 13). There was great individual varia- tion in sensitivity to [3-TGF, RA, or TNF. The astrocytoma and oligodendroglioma cells were inhibited to various de- grees by [~-TGF or TNF, while most of the glioblastomas were not sensitive to these agents. Most of the biopsies were stimulated by RA. RA or TNF did not potentiate the growth inhibitory effect of [3-TGF on biopsy cells. We therefore think it unlikely that [~-TGF in combination with RA or TNF will be effective agents in the treatment of gliomas. Key words: Gliomas - Type beta transforming growth factor - Tumor necrois factor - Retionic acid Introduction The beta transforming growth factor ([3-TGF) is a potent regulator of cell growth [8, 10-12, 15] and differentiation [4, 7, 10]. It inhibits the growth of many human tumor cell lines, but only a few are totally arrested by [3-TGF [10]. Retinoic acid (RA) has been reported to potentiate the an- tiproliferative effect of [3-TGF in my¢ transfected fibro- blasts [13]. The mechanism for the antiproliferative action of [3-TGF is unknown, apart from the fact that it binds to specific cell surface high affinity receptors [3, 18]. T-MG1, a human glioblastoma cell line, was growth in- hibited by [3-TGF under anchorage independent condi- Offprint requests to: Eirik Helseth, The Institute of Cancer Re- search, University Hospital of Trondheim, N-7000 Trondheim, Norway tions. We used this cell line to search for other biological response modifiers which could potentiate the antiprolifer- ative effect of [3-TGF. Tumor necrosis factor (TNF) and RA had this potentiating effect, while epidermal growth factor (EGF), platelet-derived growth factor (PDGF), in- terleukin-2, (IL-2), and gamma interferon (G-IF) did not. Further experiments were designed to reveal a possible mechanism for the potentiation of the [3-TGF-induced in- hibition by RA and TNF. As [3-TGF is a known modulator of EGF receptor expression [1], we studied the effect of [3- TGF alone and in combination with RA or TNF on EGF receptor expression in T-MG1 cells. The effect of TNF or RA on [3-TGF receptor expression and the effect of [3-TGF on TNF receptor expression were also studied. Cell lines are widely used for screening potentially new antineoplastic agents. However, cell lines may be rather different from the primary tumor cells, because of the ex- treme selection taking place during the establishment of cell lines. Soft agar growth of primary cultures from hu- man tumor biopsies is thought to be more comparable to the in vivo situation, and the use of this technique should supplement cell line experiments in studying growth inhib- itory agents. Therefore [~-TGF in combination with RA or TNF was tested on primary cultures from freshly resected human glioma biopsies. Materials and methods Cells. The glioblastoma cell line T-MG1 [16] was passaged in 75 cm 2 tissue culture flasks (Costar, Cambridge, Mass., USA) in medium consisting of 90% RPMI 1640 with 25 mM Hepes buffer (Gibco Biocult, Glasgow, Scotland) and 10% human, heat-inactivated AB Rh + serum (Blood- bank, Trondheim, Norway). The medium was supplement- ed with 0.1 mM L-glutamine (Gibco Biocult) and 40 ~tg/ml gentamicin (Schering Co., N J, USA). Biopsies. The tumor pieces were transported with the least possible delay from the operating theatre in medium con- sisting of 30% heat-inactivated human AB Rh + serum and 70% RPMI 1640 with 25 mM Hepes buffer at a constant temperature (37 ° C). Tumor tissue was disaggregated me- chanically by mincing it into small pieces and passing it through needles of decreasing diameter. Larger clumps were removed by filtration through a prefilter. Smaller clumps were sedimented by centrifugation after which the cells were resuspended in medium consisting of 80%