Proteomic Study of Novel Platinum Compounds in Glioma

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DESCRIPTION (provided by applicant): Gliomas remain a serious public health problem with no effective treatment, making the identification of new chemotherapeutics a priority. Equally important is the timely development of molecular tools allow the stratification of patients into likely responders and non-responders, so that therapies can be targeted effectively. A new family of platinum compounds, founded by the trinuclear BBR3464, is much more potent than established platinum agents in the killing of glioma cells in vitro. The most potent compound, BBR3610 has an IC90 in a glioma cell clonogenic assay of 8 nM, as compared to 2 mM for cisplatin. We are attempting to establish molecular profiles of responding tumors, and investigating the mechanism of action of these promising drugs by various means, and in this proposal seek support for the newest approach, which is based on displaying the proteome using two-dimensional liquid chromatography. Proteins are separated in the first dimension by a chromatofocusing column and in the second dimension on a reverse phase HPLC column. Absorbance data at 214 nM is then displayed using specialized software allowing the rapid identification of expression differences. In addition to being turnkey and suited to high throughput, this approach leaves proteins entirely in the liquid phase making recovery for downstream analysis, such as mass spectrometry, easy. We propose to apply this technology to two model systems with the aim of testing the hypothesis that cell death in response to platinum compounds is associated with characteristic changes in the proteome and that the proteins involved can be used to profile patients. In the first model, we will treat glioma cell lines with isodoses of cisplatin and BBR compounds in vitro and in vivo and assess the impact on their proteomes. Sensitive and resistant isolates of the same cell lines will be compared. In the second model, we will compare the impact of p53 mutation, which can sensitize or protect cells, on the response of cells to platinum compounds at the protein level, by using transformed p53-null astrocytes engineered to express human p53 mutants. We hope to identify changes in the proteome that are characteristic of effective glioma cell killing by platinum compounds, as such information may facilitate identifying patients likely to respond to a given drug, as well as aid in understanding their mechanism of action.