Identifying hedgehog target genes in basal cell carcinoma

Research project P 14227 Hedgehog target genes in Basal Cell Carcinoma Anna-Maria FRISCHAUF 06.03.2000 The aim of this project is the analysis of the molecular events leading to the formation of basal cell carcinoma (BCC), the most common human cancer. The primary event in tumorigenesis of basal keratinocytes is the uncontrolled activation of the sonic hedgehog (SHH) pathway in epidermal cells of the skin. This has been shown by the identification of inactivating mutations in Patched, a repressor of SHH signalling, in patients with the hereditary disease predisposing to BCCs, Nevoid Basal Cell Carcinoma (Hahn et al., 1996. Cell 85, 841-851; Johnson et al., 1996. Science 272, 1668-1671). To elucidate the next steps in tumorigenesis it is necessary to identify SHH target genes in human epidermis. To do so we will establish an in vitro model for gene expression in BCCs by controlled turn-on of Hh-signalling in a human keratinocyte cell line. This approach is necessary for the identification of early target genes. Two strategies will be employed: hedgehog itself or a constitutively active positive regulator of the pathway, Smoothened (SMO), will be expressed from a tetracycline-regulated construct. Alternatively, a hormone-regulated construct of the transcription factor Gli1, a Hh target that is likely to mediate BCC formation, will be expressed in keratinocytes. Changes in gene expression will be analysed by differential hybridisation of high-density cDNA arrays, which allows the simultaneous measurement of the concentrations of thousands of different mRNA species. This method has been successfully used in our laboratory (see Appendix 1). Newly identified SHH target genes will be further characterised and the role of a small number of genes likely to be involved in tumorigensis will be explored.

Basal Cell Carcinoma (BCC) is the most common tumour in the Western world. It occurs mainly in sun exposed skin and, in most cases can easily be removed my standard procedures. BCC does not metastasise but can be invasive and there is also a tendency for BCCs to reccur. BCCs are caused by inappropriate activation of the hegehog signalling pathway, a cascade of cellular events that was thought to be normally active only during development, but has now also been implicated in stem cell proliferation and repair of injury. Recently the hedgehog signalling pathway has also been shown to be active in common lethal tumours like carcinoma of the oesophagus, the stomach and the pancreas. The project has focused on one particular aspect of tumorigenesis in basal cell carcinoma: which genes are specifically expressed or not expressed when hedgehog signalling is active in the skin cells that can give rise to tumours. Since the pattern of gene expression determines what a cell looks like and what it does, some of the genes whose expression is changed by the hedgehog cascade must be responsible for the cell becoming a tumour cell. Transcription factors are proteins that control gene expression and, if it has been shown that the action of such a transcription factor promotes cancer, then some genes that are controlled by this protein also play a role. We have identified more than 500 genes that are expressed higher or lower than in normal cells in response to two transcription factors, GLI1 and GLI2, which are part of hedgehog signalling and can cause basal cell carcinoma in mice. Within this project we have shown how those two transcription factors influence each other, where they do similar things, and how only one but not the other can turn genes off. Among those 500 genes controlled by GLI1 and GLI2 are some that are known to cause cancer in different tissues but have not previously been known to play a role in basal cell carcinoma. They may turn out to be useful in designing new treatments for this cancer.