Expression of two non-mutated genetic elements is sufficient to stimulate oncogenic transformation of human mammary epithelial cells

Trefoil factor 3 (TFF3) expression is positively associated with advanced clinicopathological features of mammary carcinoma (MC). Herein, we provide evidence for a functional role of TFF3 in oncogenic transformation of immortalized, but otherwise normal human mammary epithelial cells (HMECs), namely, HMEC-hTERT, MCF10A, and MCF12A. Forced expression of TFF3 in immortalized-HMECs enhanced cell proliferation, cell survival, anchorage-independent growth, produced highly disorganised three-dimensional (3D) acinar structures and generated tumours in immunocompromised mice. Forced expression of TFF3 in immortalized-HMECs stimulated STAT3 activity that was required for TFF3-stimulated cell proliferation, survival, and anchorage-independent growth. TFF3 specifically utilised STAT3 activity to govern a transcriptional program, which was required for TFF3-stimulated oncogenic transformation of immortalized-HMECs, including transcriptional upregulation of CCND1 and BCL2. siRNA-mediated depletion or functional inhibition of STAT3 significantly inhibited the TFF3-stimulated transcription of CCND1 and BCL2 and oncogenicity in immortalized-HMECs. Furthermore, DOX-inducible expression of TFF3 in HMEC-hTERT cells also permitted anchorage-independent growth and produced disorganized acinar structures in 3D Matrigel culture. Removal of DOX-induced expression of TFF3 in HMEC-hTERT cells, previously grown with DOX, resulted in efficient normalisation of the disorganized acinar architecture and attenuated cell viability in Matrigel culture. Cumulatively, these findings suggest that TFF3 is a potent oncogene and its increased expression along with hTERT in HMECs is sufficient to produce oncogenic transformation.