bserved across the rows. Following normalisation the average activity for each sequence-specific siRNA over the three screens was calculated and used for subsequent statistical analysis. Due to the three-fold redundancy of the siRNA library, three activity values per gene were used for the statistical analysis. Statistical analysis. Statistical analysis was carried out using redundant siRNA activity analysis as described in. Two different analyses were carried out: one for identification of positive regulators and one for identification of negative regulators. ~~ During the last decade, it has become obvious that progression and severity of malignant diseases is often not caused by a single genetic aberration or deregulation of a single signaling pathway, but actually requires the cooperation of oncogenic-signaling pathways in cancer cells. For instance, Hedgehog /GLI and EGF-driven signaling can synergize and promote events, such as neural stem cell proliferation, as well as tumor initiation and progression . Deregulation of at least one of the two pathways has been implicated in about one-third of all cancers, and frequently, both pathways are found aberrantly activated in the same tumor. This understanding has helped to develop the hypothesis that a simultaneous activation of both pathways can drive tumor development. Although HH- as well as EGF-mediated signaling have been intensely studied, the details of how signals derived from HH or EGF are integrated at the molecular level still needs to be clarified for distinct cell types, and in different cancer entities . The first insights into HH/ GLI and EGF crosstalk in cancer was provided by Kasper et al. and Schnidar, et al., who pointed out that co-activation of both pathways results in the induction of a specific gene expression pattern, which induces malignant transformation of human Synergism between Hedgehog-GLI and EGFR Signaling keratinocytes . The hypothesis that both pathways 
10608278 merge at the level of transcriptional regulation was also supported by other studies, which showed that several different genes indeed possess binding sites for GLI and EGF-regulated transcription factors, such as c-JUN/AP-1 . Evidence for cooperative effects was also obtained at the level of protein activation, by demonstrating that GLI1 transcription factors need to be stabilized by MAPK and PI3K/AKT-signaling , which also presented a prerequisite for cytoplasmic/nuclear shuttling of GLI proteins. Finally, Whisenant et al. revealed a direct phosphorylation of GLI1 10914735 by ERK, which was anticipated as the explanation for the modified transcriptional activity of GLI proteins upon activation of MAPK and PI3K/AKT signaling . The interaction of HH/GLI with EGF-induced signaling has been described in a number of tumor types such as skin, prostate, and pancreas , while other kinases such as PKC and mTOR/S6K also positively regulate GLI activity. The lack of human cancer cell lines clearly responsive to HH stimulation frequently complicates an unambiguous Halofuginone interpretation at the molecular level. The fact that many studies have been carried out in murine fibroblast cell lines, and have been based on overexpression of HH pathway components raises the question about the possible physiological relevance for the analysis of human cancer. Given this, we screened human cell lines for their SMO expression level and identified high level SMO expression in the Daoy human medulloblastoma cell line. This cell line revealed in f
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