Epithelial ovarian cancer (EOC) strikes more than 25,000 American women annually, and >50% of these women will die from their disease, especially if diagnosed, as is common, with late-stage disease. To develop new therapies for this disease, it will be necessary to better understand the molecular events that are associated with the neoplastic transformation. Alternative splicing (AS), by allowing production of multiple protein isoforms from a single gene, plays a major role in expanding proteomic complexity in metazoans and adaptations to internal and external environmental changes. In general, more genes in tumors, especially those associated with tumorigenesis, exhibit alternative splice forms compared to normal tissues. There is mounting evidence that AS is associated with and possibly involved in tumor progression and/or metastasis. At present, the mechanisms leading to splicing alterations in cancer are poorly understood, but they are likely due either to mutations in cis-elements or, more likely, changes in trans-acting factors, such as splicing factors. Indeed, several splicing factors, including PTB and SRp20, are overexpressed in tumors and one, SF2/ASF, was shown to be an oncogene. PTB and SRp20 are overexpressed in human ovarian tumors, compared to normal ovarian epithelium, where PTB is either not expressed or expressed at very low levels, and expression of these SFs is associated with malignancy of these tumors. Knockdown of PTB in ovarian tumor cells in vitro by siRNA substantially suppressed tumor cell growth, colony formation, and invasiveness and led to slower tumor growth in mice bearing ovarian tumors. Our results provide proof-of-principle that splicing factors may be a class of novel and unexplored therapeutic targets for cancer treatment, at least PTB in EOC, a disease that has a dismal outcome and for which there are no good therapeutic options, especially for advanced-stage disease. In this proposal, we focus on one splicing factor, PTB, and one tumor type, EOC, and hypothesize that PTB is a druggable therapeutic target in this disease. To test our hypothesis, we propose to develop a cell- based high-throughput screen of small molecule libraries to identify small molecules that interfere with PTB activity and then study whether these identified small molecules are able to inhibit ovarian tumor growth. To this end, we offer three specific aims: (1) Develop and optimize a proprietary cell-based reporter system to monitor the activity of PTB in the cell;(2) Develop and optimize a high throughput screen (HTS) of small molecule libraries, including those containing FDA-approved small molecules as well as extracts from natural sources or compounds generated by chemical synthesis to identify PTB inhibitors;and (3) Validation of effective small molecules on ovarian tumor cell growth in vitro and in vivo. If successful, our approach will validate PTB as a novel therapeutic target and will identify a novel class of small molecule drugs based on the manipulation of alternative splicing for the treatment of ovarian cancer, and possibly other epithelial cancers.
Epithelial ovarian cancer (EOC) strikes more than 25,000 American women annually, and >50% of these women will die from their disease, especially if diagnosed, as is common, with late-stage disease. If successful, the research proposed here will validate targeting of splicing factors and alternative splicing as a novel therapeutic approach to the treatment of ovarian and other cancers. At the end of the project period we would have identified small molecules leading to new drugs for the treatment of these cancers, and this will clearly have a positive impact on public health.
|Arslan, Ahmet Dirim; He, Xiaolong; Wang, Minxiu et al. (2013) A high-throughput assay to identify small-molecule modulators of alternative pre-mRNA splicing. J Biomol Screen 18:180-90|
|He, X; Arslan, A D; Pool, M D et al. (2011) Knockdown of splicing factor SRp20 causes apoptosis in ovarian cancer cells and its expression is associated with malignancy of epithelial ovarian cancer. Oncogene 30:356-65|