Colon carcinoma cells switch their response to transforming growth factor β₁ with tumor progression

Transforming growth factor β₁ (TGF-β₁) switches from an inhibitor of tumor cell growth to a stimulator of growth and invasion during human colon carcinoma progression. We originally observed that metastatic colon carcinoma cells in primary culture responded to TGF-β₁ by proliferation, whereas moderate to well-differentiated primary site colon carcinomas were growth inhibited by TGF-β₁ (P. Schroy et al., Cancer Res., 50: 261-265, 1990). We then cloned several colon carcinoma cell lines which modeled these responses to TGF-β₁ and expressed TGF-β₁ (M. M. Hafez et al., Cell Growth and Differ., 1: 617-626, 1990; 3: 753-762, 1992). Two of these colon carcinoma cell lines, U9 and HD3, which activate approximately equal amounts of TGF- β₁ and express equal amounts of TGF-β receptors, are now used to compare the effects of TGF-β₁ in modulating invasive behavior. The U9 cell line exhibits autocrine-positive growth regulation in vitro by TGF-β₁, whereas the HD3 cell line shows the opposite response, autocrine-negative regulation. Blocking endogenous TGF-β₁ with isotype-specific antibody inhibited U9 cell growth because autocrine TGF-β₁ acts as a mitogen for U9 cells. In contrast, antibody to TGF-β₁ stimulated HD3 cell proliferation because autocrine TGF-β₁ inhibits growth of these cells. U9 cells were 13-fold more invasive in vitro through a collagen I layer than HD3 cells. In vitro invasion by U9 cells but not by HD3 cells was stimulated 2-fold by TGF-β₁, and U9 invasion was inhibited by antibody to TGF-β₁. When injected into athymic mice, U9 cells induced tumors 6-fold the size of those induced by HD3 cells. Tumors induced by each cell line retained expression of TGF-β₁ in vivo, as shown by immunohistochemistry using isotype-specific antibody. Antibody to TGF-β₁ inhibited the growth of U9 cells in athymic mice 4-fold compared to control antibody, demonstrating that autocrine TGF-β₁ stimulated U9 cell growth in vivo. Thus, the U9 cells with autocrine-positive growth regulation by TGF-β₁ were more tumorigenic in vivo and more invasive in vitro than HD3 cells with autocrine-negative regulation by TGF-β₁. Differences in the induction of three immediate early genes were observed between TGF-β₁ growth-stimulated U9 cells and TGF-β₁ growth-inhibited HD3 cells. Thirty min of TGF-β₁ treatment stimulated expression of c-fos approximately 50-fold in TGF-β₁ growth-inhibited HD3 cells, whereas only a 2-fold induction was observed in U9 cells. A small increase (50%) in expression of c-jun was seen with TGF-β₁ treatment in HD3 cells, whereas expression of c-jun was inhibited in U9. In contrast, c-myc was inhibited 3- 5-fold by exogenous TGF-β₁ in both HD3 and U9 cells, suggesting a common role for c-myc in both responses to TGF-β₁.