Project Details

Activating mutations in the BRAF oncogene are found in approximately 10% of patients with metastatic colon cancers. These patients tend to have a poor prognosis and have limited treatment options once they fail chemotherapy. The development of new treatments for these patients is therefore urgently needed. Drugs which target and inhibit mutant BRAF have recently been developed, and have shown remarkable clinical activity in BRAF mutant melanoma. Clinical activity of these drugs in colon cancer has also been observed, however response rates are significantly less than those observed in melanoma. One proposed explanation for this difference is the presence of concurrent mutations in the PI3K/PTEN pathway in colon tumours, which reduces the dependence of the tumour on BRAF, and thus sensitivity to BRAF inhibitors. In this project a panel of BRAF mutant colon cancer cell lines will be screened for response to BRAF inhibitors using the MTS and FACS-based assays, to identify colon tumours sensitive and resistance to these drugs. The impact of concurrent mutations in the PI3K/PTEN pathway on response to BRAF inhibitors will then be determined. Should mutations in PIK3CA/PTEN be found to impact response, the effect of dual targeting of BRAF and PI3K signalling on tumour growth will be determined by combination treatment with BRAF and PI3K-inhibitors. Should PIK3CA/PTEN mutations not impact response to BRAF inhibitors, alternative mechanisms of response will be examined by gene expression profiling of sensitive and resistant cell lines treated with these drugs. This thesis is designed to provide the student with insight into the genetic basis of cancer, an understanding of the treatment strategies used in cancer and exposure to translational cancer research.

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Project Details

We have recently discovered a subset of colon cancers in which the cytoskeletal protein, villin, is completely absent. Remarkably, loss of villin occurs selectively in colon cancers with a characteristic known as “microsatellite instability” or MSI. Cancers with this characteristic are less likely to metastasize and have a better outcome. This honours project will examine the transcriptional mechanism by which loss of villin expression occurs in colon cancer. Specifically, the role of the Cdx-1 and HNF1 transcription factors in regulating villin expression will be explored. First, whether Cdx-1 and HNF1 expression correlates with villin expression will be examined by immunohistochemistry in a panel of colon cancer cell lines and primary colon tumors by real time PCR and immunohistochemistry. Second, the effect of over and underexpression of Cdx-1 and HNF1 on villin promoter activity and expression will be tested by transient transfection of these genes into colon cancer cells. Third, the impact of mutating Cdx-1 and HNF1 binding sites in the villin promoter by site-directed mutagenesis will also be tested, and binding of these factors to the villin promoter examined by ChIP analysis. The project will provide the student with a sound introduction to fundamental concepts in cancer biology, and experience in cell and molecular biology. This Hons project will be conducted in the Oncogenic Transcription Laboratory, Ludwig Institute for Cancer Research, Austin Hospital, Heidelberg.

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