Ovarian granulosa cell tumours (GCT) are hormonally active cancers characterised by indolent growth and late, invasive relapse1. We previously reported that inhibition of the X-linked inhibitor of apoptosis protein (XIAP) removes transrepression of the nuclear receptor, peroxisome proliferator-activated receptor-gamma (PPARγ) in the GCT-derived cell line, KGN. Combined XIAP inhibition and PPARγ activation results in the upregulation of PPARγ-related proteins associated with metabolism2, a significant induction in apoptosis and a reduction in cell proliferation and viability3. Given the invasive nature of GCT, our aim is to explore whether the combined treatment has an effect on invasion of the cancer.
We have used stable isotope labelling with amino acids in cell culture (SILAC), a proteomic approach to identify differentially expressed proteins in KGN cells after 24 hours of combined PPARγ activation (rosiglitazone and retinoic acid; RGZ/RA) and XIAP inhibition (smac mimetic; SM). We have used the xCELLigence RTCA system to monitor invasion of KGN cells through a Matrigel® basement membrane in real-time. KGN cells treated either with DMSO, RGZ/RA or SM alone or combined RGZ/RA/SM were plated in the upper chamber of a CIM-Plate 16 on a layer of Matrigel®. Cell invasion across the Matrigel® into the lower chamber containing serum with or without the compounds was then monitored.
We identified 52 differentially regulated proteins, including downregulation of fascin (-1.65 fold), an actin-bundling protein associated with cell motility and migration. When compared to DMSO, we observed that the RGZ/RA/SM-treated KGN cells were (i) 30% less invasive and (ii) demonstrated a delayed onset of invasion by 17 hours towards serum-containing media with and without drugs in the lower chamber, respectively.
Our findings suggest that combined targeting of PPARγ and XIAP alters the invasive properties of GCT in vitro. The effect on invasion warrants further investigation of fascin as a drug target for GCT.