OVER-EXPRESSION OF TRANSFORMING GROWTH-FACTOR-ALPHA ANTAGONIZES THE ANTI-TUMORIGENIC BUT NOT THE DIFFERENTIATION ACTIONS OF RETINOIC ACID IN A HUMAN TERATOCARCINOMA CELL

All-trans retinoic acid (RA) treatment of the multipotent human terato carcinoma (TC) cell line NTERA-2 clone D1 (abbreviated NT2/D1) induces a neuronal phenotype and other cell lineages. NT2/D1 cells basally ex press transforming growth factor alpha (TGF-alpha) mRNA and secreted p rotein. After RA-treatment TGF-alpha expression is markedly reduced. T his decline in TGF-alpha expression accompanies the induction of the n euronal phenotype and a marked reduction of tumorigenicity in athymic mice. This suggested a causal link between reduced TGF-alpha expressio n and the induced differentiation or loss of tumorigenicity of these R A-treated TC cells. To evaluate this possibility, an RA-refractory NT2 /D1 subclone was analysed. This subclone, designated NT2/D1-R1, failed to induce differentiation or to decrease TGF-alpha expression despite RA treatment. To further explore the relationship between TGF-alpha e xpression and RA actions in this human TC cell, a TGF-alpha cDNA was s tably transfected and expressed in NT2/D1 cells. RA-treatment of indep endently obtained TGF-alpha over-expressing clones and a representativ e control transfectant only expressing the neomycin resistance gene pr oduced a neuronal phenotype similar to parental NT2/D1 cells as assess ed by morphologic, immunophenotypic, and gene expression markers of di fferentiation. RA-treatment of these clones also induced a G1 arrest s imilar to parental cells. However, only the TGF-alpha over-expressing clones that secreted high levels of TGF-alpha protein into the conditi oned media before and after RA treatment still developed tumors in ath ymic mice despite prior exposure of these cells to RA. This finding de monstrates that TGF-alpha can inhibit the anti-tumorigenic effects of RA in human TCs. Thus, over-expression of a single growth factor that normally declines with RA treatment antagonizes the antitumorigenic bu t not the differentiation actions of RA in this human tumor cell.