BarTeL, a Genetically Versatile, Bioluminescent and Granule Neuron Precursor-Targeted Mouse Model for Medulloblastoma.

Medulloblastomas are the most common malignant pediatric brain tumor and have been divided into four major molecular subgroups. Animal models that mimic the principal molecular aberrations of these subgroups will be important tools for preclinical studies and allow greater understanding of medulloblastoma biology. We report a new transgenic model of medulloblastoma that possesses a unique combination of desirable characteristics including, among others, the ability to incorporate multiple and variable genes of choice and to produce bioluminescent tumors from a limited number of somatic cells within a normal cellular environment. This model, termed BarTeL, utilizes a Barhl1 homeobox gene promoter to target expression of a bicistronic transgene encoding both the avian retroviral receptor TVA and an eGFP-Luciferase fusion protein to neonatal cerebellar granule neuron precursor (cGNP) cells, which are cells of origin for the sonic hedgehog (SHH) subgroup of human medulloblastomas. The Barhl1 promoter-driven transgene is expressed strongly in mammalian cGNPs and weakly or not at all in mature granule neurons. We efficiently induced bioluminescent medulloblastomas expressing eGFP-luciferase in BarTeL mice by infection of a limited number of somatic cGNPs with avian retroviral vectors encoding the active N-terminal fragment of SHH and a stabilized MYCN mutant. Detection and quantification of the increasing bioluminescence of growing tumors in young BarTeL mice was facilitated by the declining bioluminescence of their uninfected maturing cGNPs. Inclusion of eGFP in the transgene allowed enriched sorting of cGNPs from neonatal cerebella. Use of a single bicistronic avian vector simultaneously expressing both Shh and Mycn oncogenes increased the medulloblastoma incidence and aggressiveness compared to mixed virus infections. Bioluminescent tumors could also be produced by ex vivo transduction of neonatal BarTeL cerebellar cells by avian retroviruses and subsequent implantation into nontransgenic cerebella. Thus, BarTeL mice provide a versatile model with opportunities for use in medulloblastoma biology and therapeutics.

Stromelysin-1 (MMP-3) is a target and a regulator of Wnt1-induced epithelial-mesenchymal transition (EMT).

Matrix metalloproteinases (MMPs) play a well-defined role in later stages of tumor progression. However, there has been evidence that they also contribute to earlier stages of malignant transformation. The Wnt signaling transduction pathway plays a critical role in development and in the pathogenesis of many epithelial cancers. Here we have used Wnt1-induced epithelial-mesenchymal transition (EMT) in C57MG murine mammary epithelial cells to study the role of MMPs in this early step of malignant progression. Overexpression of Wnt1 in C57MG cells promoted EMT, the translocation of ß-catenin from the cell membrane to the nucleus and its transcriptional activity, cell proliferation and cell motility. Simultaneously, we observed an increased expression of stromelysin-1 (MMP-3) and a 5.5-fold increase in MMP-3 promoter activity in C57MG cells expressing Wnt1 compared with C57MG cells. Treatment of Wnt-overexpressing cells with MMP inhibitor AG3340 decreased MMP-3 expression. We also found evidence that MMP-3 and Wnt3a cooperate in enhancing the transcriptional activity of ß-catenin in C57MG cells. Consistently, the effects of Wnt1 on EMT, proliferation and migration were inhibited by MMP inhibitors, or upon downregulation of MMP-3 by siRNA. These results suggest that MMP-3 is both a direct transcriptional target and a necessary contributor of the Wnt/ß-catenin signaling pathway.

Androgen inducibility of Fgf8 in Shionogi carcinoma 115 cells correlates with an adjacent t(5;19) translocation.

Fgf8 (fibroblast growth factor 8) was initially cloned from a mouse mammary tumor cell line derived from the androgen-dependent Shionogi carcinoma 115. The androgen-inducible expression of Fgf8 in this tumor controls its androgen-dependent phenotype, thus stimulating interest in this gene as a possible factor in human prostate cancer and other androgen-sensitive cancers. However, apart from Shionogi carcinoma 115, the androgen inducibility of Fgf8 is controversial. In the present study, having not detected androgen-inducible expression of Fgf8 in other mouse mammary cell lines or mouse prostate, we examined the Shionogi carcinoma 115-derived S115 cell line for mouse mammary tumor virus (MMTV) insertions or other nearby DNA rearrangements that might explain the androgen inducibility of Fgf8 in these cells. Southern blotting did not detect MMTV insertions near Fgf8 but did reveal a specific DNA rearrangement 3.7 kb upstream of Fgf8 in S115 cells and in other cells (SC115) independently derived from Shionogi carcinoma 115. Spectral karyotyping of S115 cells and sequencing of the cloned rearrangement junctions indicate that Fgf8 is involved in a t(5;19) translocation. The chromosome 5 sequence joined to Fgf8 is immediately adjacent to Smr2 (submaxillary gland androgen-regulated protein 2) and includes Muc10 (mucin 10), two genes that we show are testosterone inducible in S115 cells, suggesting that the androgen-dependent expression of Fgf8 in Shionogi carcinoma 115 and derivative cells results from this translocation. Together, these results suggest that androgen inducibility is not an inherent property of the Fgf8 gene, which has implications regarding this gene's proposed role in the etiology of hormone-responsive cancers.