[show abstract][hide abstract] ABSTRACT: The Hedgehog (Hh) signaling network is critical for patterning and organogenesis in mammals, and has been implicated in a variety of cancers. Smoothened (Smo), the gene encoding the principal signal transducer, is overexpressed frequently in breast cancer, and constitutive activation in MMTV-SmoM2 transgenic mice caused alterations in mammary gland morphology, increased proliferation, and changes in stem/progenitor cell number. Both in transgenic mice and in clinical specimens, proliferative cells did not usually express detectable Smo, suggesting the hypothesis that Smo functioned in a non-cell autonomous manner to stimulate proliferation. Here, we employed a genetically tagged mouse model carrying a Cre-recombinase-dependent conditional allele of constitutively active Smo (SmoM2) to test this hypothesis. MMTV-Cre- or adenoviral-Cre-mediated SmoM2 expression in the luminal epithelium, but not in the myoepithelium, was required for the hyper-proliferative phenotypes. High levels of proliferation were observed in cells adjacent or in close-proximity to Smo expressing cells demonstrating that SmoM2 expressing cells were stimulating proliferation via a paracrine or juxtacrine mechanism. In contrast, Smo expression altered luminal cell differentiation in a cell-autonomous manner. SmoM2 expressing cells, purified by fluorescence activated cell sorting (FACS) via the genetic fluorescent tag, expressed high levels of Ptch2, Gli1, Gli2, Jag2 and Dll-1, and lower levels of Notch4 and Hes6, in comparison to wildtype cells. These studies provide insight into the mechanism of Smo activation in the mammary gland and its possible roles in breast tumorigenesis. In addition, these results also have potential implications for the interpretation of proliferative phenotypes commonly observed in other organs as a consequence of hedgehog signaling activation.
[show abstract][hide abstract] ABSTRACT: Estrogen signaling is required for the proliferation of normal breast epithelial cells. However, prophylactic inhibition of estrogen signaling fails to prevent 56% of human breast cancer cases. The underlying mechanism is not well understood. Aberrant activation of growth factor signaling is known to provide alternative proliferation pathways in breast cells that are fully transformed, but it is not known whether activation of growth factor signaling can substitute for estrogen signaling in causing aberrant proliferation in the normal breast epithelium. Here, we report that in a retrovirus-based somatic mouse model (replication-competent ALV-LTR splice acceptor/tumor virus A) that closely mimics the evolution of sporadic human breast cancers, mammary epithelial cells harboring PyMT or activated ErbB2 evolve into tumors independent of estrogen or other ovarian functions in contrast to previous observations of estrogen-dependent cancer formation in germ line mouse models of ErbB2 activation. Importantly, ErbB2 activation in normal mammary cells causes estrogen-independent proliferation in both estrogen receptor (ER)-negative cells as well as in normally quiescent ER-positive cells. Therefore, aberrant activation of growth factor signaling contributes to estrogen-independent proliferation of both preneoplastic and cancerous mammary cells, and prophylactic therapy against both growth factor signaling and estrogen signaling may need to be considered in women with increased risk of breast cancer.
Neoplasia (New York, N.Y.) 09/2010; 12(9):718-26. · 5.48 Impact Factor
[show abstract][hide abstract] ABSTRACT: We have reported the use of vectors based on avian leukosis virus (ALV) and ALV env pseudotyped lentivirus for stably introducing genes into somatic mammary cells in vivo for tumor modeling. These vectors require tva transgenic mice for viral infection. Here, we report, and described the method for, stably introducing genes into the mammary gland in mice using intraductal injection of a common lentiviral vector. This method does not need tva-transgenic mice for infection. This in vivo transformation method may be useful for rapid testing of the tumorigenic potential of genes potentially involved in breast carcinogenesis.
Journal of Mammary Gland Biology and Neoplasia 11/2009; 14(4):401-4. · 7.52 Impact Factor
[show abstract][hide abstract] ABSTRACT: We have adapted the avian leukosis virus RCAS (replication-competent avian sarcoma-leukosis virus LTR splice acceptor)-mediated somatic gene transfer technique to introduce oncogenes into mammary cells in mice transgenic for the avian subgroup A receptor gene, tva, under control of the mouse mammary tumor virus (MMTV) promoter. Intraductal instillation of an RCAS vector carrying the polyoma middle T antigen (PyMT) gene (RCAS-PyMT) induced multiple, oligoclonal tumors within 3 weeks in infected mammary glands of MMTV-tva transgenic mice. The rapid appearance of these tumors from a relatively small pool of infected cells (estimated to be approximately 2 x 10(3) cells per gland by infection with RCAS carrying a GFP gene; RCAS-GFP) was accompanied by a high fraction of cells positive for Ki67, Cyclin D1, and c-Myc, implying strong proliferation competence. Furthermore, the tumors displayed greater cellular heterogeneity than did tumors arising in MMTV-PyMT mice, suggesting that RCAS-PyMT transforms a relatively immature cell type. Infection of mice transgenic for both MMTV-Wnt-1 and MMTV-tva with RCAS virus carrying an activated Neu oncogene dramatically enhanced tumor formation over what is observed in uninfected bitransgenic animals. We conclude that infection of mammary glands with retrovirus vectors is an efficient means to screen candidate oncogenes for their capacity to initiate or promote mammary carcinogenesis in the mouse.
Proceedings of the National Academy of Sciences 12/2006; 103(46):17396-401. · 9.74 Impact Factor