TGF-Beta Suppresses VEGFA-Mediated Angiogenesis in Colon Cancer Metastasis

The FET cell line, derived from an early stage colon carcinoma, is non-tumorigenic in athymic nude mice. Engineered FET cells that express TGF-alpha (FET alpha) display constitutively active EGFR/ErbB signaling. These cells readily formed xenograft tumors in athymic nude mice. Importantly, FET alpha cells retained their response to TGF-beta-mediated growth inhibition, and, like the parental FET cells, expression of a dominant negative TGF-beta type II receptor (DNRII) in FET alpha cells (FET alpha/DNRII) abrogated responsiveness to TGF-beta-induced growth inhibition and apoptosis under stress conditions in vitro and increased metastatic potential in an orthotopic model in vivo, which indicates metastasis suppressor activity of TGF-beta signaling in this model. Cancer angiogenesis is widely regarded as a key attribute for tumor formation and progression. Here we show that TGF-beta signaling inhibits expression of vascular endothelial growth factor A (VEGFA) and that loss of autocrine TGF-beta in FET alpha/DNRII cells resulted in increased expression of VEGFA. Regulation of VEGFA expression by TGF-beta is not at the transcriptional level but at the post-transcriptional level. Our results indicate that TGF-beta decreases VEGFA protein stability through ubiquitination and degradation in a PKA-and Smad3-dependent and Smad2-independent pathway. Immunohistochemical (IHC) analyses of orthotopic tumors showed significantly reduced TGF-beta signaling, increased CD31 and VEGFA staining in tumors of FET alpha/DNRII cells as compared to those of vector control cells. These results indicate that inhibition of TGF-beta signaling increases VEGFA expression and angiogenesis, which could potentially contribute to enhanced metastasis of those cells in vivo. IHC studies performed on human colon adenocarcinoma specimens showed that TGF-beta signaling is inversely correlated with VEGFA expression, indicating that TGF-beta-mediated suppression of VEGFA expression exists in colon cancer patients.