Abstract
Abstract Background: We hypothesize that numerous molecular differences exist between ovarian cancer stem cells (OCSC) and non-ovarian cancer stem cells (non-OCSC). Identifying these differences and describing their cellular interactions could be utilized as sites of targeted therapies for more effective treatment strategies. Methods: Fluorescence Activated Cell Sorting (FACS) was utilized to sort 2 papillary serous ovarian cancer cell lines (OV90 & SKOV3) into 2 distinct populations: OCSC (CD44+/CD24-) and non-OCSC (all other cells). RNA was isolated from each sorted cell subtype, converted to cDNA, and analyzed via PCR array designed to evaluate numerous molecular pathways described in cancer. This array profiles the expression of 84 genes representative of 6 biological functional pathways involved in tumorigenesis. Molecular pathway mapping identified a common molecular pathway, the Hedgehog (Hh) pathway. Expression of Hh pathway proteins (SMOH, PTCH, GLI1) was determined by Western blot. Localization of intranuclear GLI1, which signifies activated Hh pathway, was detected via immunofluorescent staining of GLI1. Cancer cell growth was evaluated after treatment with Hh agonists (SHH and IHH) as well as treatment with an Hh inhibitor (cyclopamine) with and without standard chemotherapy. Results: Genes with ≥ 2-fold upregulation in OCSC compared to non-OCSC where considered significant. Genes grouped by function are as follows: Apoptosis: BCL2 (+2.5), CFLAR (+2.6), and MDM2 (+3.4); Angiogenesis: PIK3R1 (+3.4), VEGFA (+5.1), FGFR2 (+5.1), ANGPT2 (+6.0), and THBS1 (+6.1); Invasion: MMP1 (+2.3), SNCG (+2.7), PLAU (+3.0), and PLAUR (+5.8); and Adhesion: ITGA2 (+9.4). Molecular pathway mapping from these upregulated genes identified the Hh pathway as a common pathway. OV90 and SKOV3 demonstrated increased intranuclear GLI1 expression indicating an activated Hh pathway. Immunofluorescent-staining of GLI1 confirmed these innate findings, however near complete exclusion of intranuclear GLI1 was seen with anti-SMOH treatment with cyclopamine. Both Hh agonists demonstrated significant increases in cancer growth of at least 42-fold for SHH-treated cells and 46-fold for IHH-treated cells (p< 0.001). Inhibition of the Hh pathway with cyclopamine demonstrated a 5-fold decrease in cancer cell growth when used in combination with paclitaxel and carboplatin (p < 0.001). Conclusions: Based on the numerous molecular differences between OCSC and non-OCSC, the Hedgehog pathway appears to be important in OCSC regulation. As such, the inhibition of Hh pathway in combination with standard chemotherapy for ovarian cancer warrants further evaluation of the potential targeting of this pathway in ovarian cancer treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3406. doi:10.1158/1538-7445.AM2011-3406
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