(345c) Fluorescence Cell-Based High Throughput Assay for Screening Small-Molecule Modulators of Survivin Expression In Cancer Cells

Survivin, a member of IAP (inhibitor of apoptosis) family, is involved in the regulation of mitosis and inhibition of apoptosis. Survivin is over-expressed in embryonic and fetal tissues, malignant human cancers and some normal adult cells, including primitive hematopoietic cells, T lymphocytes, polymorphonuclear neutrophils and vascular endothelial cells. Survivin expression in most cancers is associated with patient’s low survival or disease recurrence, and resistance to chemotherapy or radiotherapy.  Therefore, anti-survivin therapy, which shows efficacy without overt toxicity, can offer a promising cancer treatment strategy. In this study, a cell-based high throughput assay was developed based on a survivin-reporter system that potentially can be used to screen drugs with cancer-specific responses. The expression of green fluorescence protein (GFP) was controlled under a survivin promoter and can be used to describe cancer specific responses for drug screening. In order to rule out false positive hits due to the cytotoxic effects of compounds, a cytomegalovirus (CMV)-reporter system was also developed. To demonstrate its potential applications in anti-cancer drug screening, the effects of Taxol and Vincristine Sulfate on survivin expression in terms of the expression of GFP were investigated. It was found that Taxol up-regulated the GFP expression, whereas Vincristine Sulfate down-regulated GFP expression, which were consistent with previous reports in the case of survivin expression. In addition, Ganoderma lucidum spore (GLS) powder, a Chinese herbal medicine with reported functions for stopping cancer cell metastasis, was evaluated for its possible effects on breast cancer cells. In general, the addition of GLS in the culture medium did not significantly affect the cell growth rate, but the expression of survivin was significantly down regulated, which was also confirmed by reverse transcriptase PCR. These results suggested that GLS itself is not cytotoxic but could reduce cancer metastasis via the down-regulation of survivin in cancer cells. Therefore, the survivin-based reporter gene system can be used to identify molecules modulating survivin expression and thus provide more reliable data in predicting in vivo interactions and efficacy of potential cancer drugs.