Wnt-1 and int-2 mammary oncogene effects on the beta-catenin pathway in immortalized mouse mammary epithelial cells are not sufficient for tumorigenesis

Development of strategies for prevention of breast cancer development requi res an understanding of the effects of mammary oncogenes on mammary cells a t early stages in neoplastic transformation. As mammary oncogenes wnt-1 and int-2 affect different signal transduction pathways, we investigated their effects on established mouse mammary epithelial cell lines (MMECLs) reflec ting early stages in tumorigenesis. Normal interactions between beta -caten in and E-cadherin were abrogated in all three immortalized MMECLs and the c ells lacked beta -catenin-mediated transactivation activity, detectable usi ng a reporter assay, suggesting that alterations in cell adhesion may be ve ry early events in mammary tumorigenesis. Immortalized FSK4 and EL12 cells and hyperplastic TM3 cells were stably transfected with expression vectors encoding wnt-1 or int-2 or the control vector, and drug-selected pooled cel ls from each tine were confirmed by reverse transcription-polymerase chain reaction to express the transfected oncogene; this expression persisted in the cells analysed in vitro and in vivo. Resultant phenotypic changes depen ded both on the oncogene and the target mammary cell line. In FSK4 cells, e xpression of wnt-1 or int-2 resulted in proliferative changes in vitro, inc luding reduced contact inhibition, increased beta -catenin expression, and decreased p53 transcriptional activity, but neither oncogene conferred upon those cells the ability to produce tumors in vivo. EL12 cells were highly refractory to the effects of both oncogenes, with the only measurable chang es being increased E-cadherin levels induced by both oncogenes and increase d proliferation of the int-2-transfected cells in the absence of serum. Par ental TM3 cells were phenotypically similar to wnt-1- or int-2-transfected FSK4 cells and displayed an increased rate of proliferation in vitro and ma rkedly increased tumorigenicity in vivo following transfection with int-2 b ut not with wnt-1. These results suggest that wnt-1 signaling is redundant in the hyperplastic TM3 cells and indicate that wnt-1-induced effects in th e immortalized FSK4 and EL12 cells were not sufficient to mediate a tumorig enic phenotype. This study showed that the wnt-1 and int-2 oncogenes have s imilar but distinguishable effects on immortalized MMECLs and that the gene tic background of the mammary cells greatly influences the consequences of oncogene expression at early stages of cell transformation.