Expression of ABCA transporters was associated with poor outcome in serous ovarian cancer, implicating lipid trafficking as a potentially important process in EOC.

BackgroundAcquired resistance to Tamoxifen remains a critical problem in breast cancer patient treatment, yet the underlying causes of resistance have not been fully elucidated. Abberations in the Wnt signalling pathway have been linked to many human cancers, including breast cancer, and appear to be associated with more metastatic and aggressive types of cancer. Here, our aim was to investigate if this key pathway was involved in acquired Tamoxifen resistance, and could be targeted therapeutically.MethodsAn in vitro model of acquired Tamoxifen resistance (named TamR) was generated by growing the estrogen receptor alpha (ER) positive MCF7 breast cancer cell line in increasing concentrations of Tamoxifen (up to 5 uM). Alterations in the Wnt signalling pathway and epithelial to mesenchymal transition (EMT) in response to Tamoxifen and treatment with the Wnt inhibitor, IWP-2 were measured via quantitative RT-PCR (qPCR) and TOP/FOP Wnt reporter assays. Resistance to Tamoxifen, and effects of IWP-2 treatment were determined by MTT proliferation assays.ResultsTamR cells exhibited increased Wnt signalling as measured via the TOP/FOP Wnt luciferase reporter assays. Genes associated with both the β-catenin dependent (AXIN2, MYC, CSNK1A1) and independent arms (ROR2, JUN), as well as general Wnt secretion (PORCN) of the Wnt signalling pathway were upregulated in the TamR cells compared to the parental MCF7 cell line. Treatment of the TamR cell line with human recombinant Wnt3a (rWnt3a) further increased the resistance of both MCF7 and TamR cells to the anti-proliferative effects of Tamoxifen treatment. TamR cells demonstrated increased expression of EMT markers (VIM, TWIST1, SNAI2) and decreased CDH1, which may contribute to their resistance to Tamoxifen. Treatment with the Wnt inhibitor, IWP-2 inhibited cell proliferation and markers of EMT.ConclusionsThese data support the role of the Wnt signalling pathway in acquired resistance to Tamoxifen. Further research into the mechanism by which activated Wnt signalling inhibits the effects of Tamoxifen should be undertaken. As a number of small molecules targeting the Wnt pathway are currently in pre-clinical development, combinatorial treatment with endocrine agents and Wnt pathway inhibitors may be a useful therapeutic option in the future for a subset of breast cancer patients.

Objective ABCB1 encodes the multi-drug efflux pump P-glycoprotein (P-gp) and has been implicated in multi-drug resistance. We comprehensively evaluated this gene and flanking regions for an association with clinical outcome in epithelial ovarian cancer (EOC). Methods The best candidates from fine-mapping analysis of 21 ABCB1 SNPs tagging C1236T (rs1128503), G2677T/A (rs2032582), and C3435T (rs1045642) were analysed in 4,616 European invasive EOC patients from thirteen Ovarian Cancer Association Consortium (OCAC) studies and The Cancer Genome Atlas (TCGA). Additionally we analysed 1,562 imputed SNPs around ABCB1 in patients receiving cytoreductive surgery and either ‘standard’ first-line paclitaxel-carboplatin chemotherapy (n=1,158) or any first-line chemotherapy regimen (n=2,867). We also evaluated ABCB1 expression in primary tumours from 143 EOC patients. Result Fine-mapping revealed that rs1128503, rs2032582, and rs1045642 were the best candidates in optimally debulked patients. However, we observed no significant association between any SNP and either progression-free survival or overall survival in analysis of data from 14 studies. There was a marginal association between rs1128503 and overall survival in patients with nil residual disease (HR 0.88, 95% CI 0.77-1.01; p=0.07). In contrast, ABCB1 expression in the primary tumour may confer worse prognosis in patients with sub-optimally debulked tumours. Conclusion Our study represents the largest analysis of ABCB1 SNPs and EOC progression and survival to date, but has not identified additional signals, or validated reported associations with progression-free survival for rs1128503, rs2032582, and rs1045642. However, we cannot rule out the possibility of a subtle effect of rs1128503, or other SNPs linked to it, on overall survival.

Ovarian cancer is a devastating disease. More than half the women diagnosed with advanced disease will die within five years. Tumours displaying serous histology are the most common, and although initially responsive to current chemotherapy regimens, the disease is characterised by a frequent rate of relapse, often with disease that is resistant to further treatment. Tumours displaying serous histology are the most common and often have the worst prognosis. The ATP-binding cassette (ABC) transporter superfamily consists of 48 functional transmembrane proteins which are further classified into seven subfamilies according to sequence similarity, designated A through G. Various members of the B, C and G branches are well-known for their abilities to convey drug resistance to cancer cells in vitro, but their precise roles in determining clinical outcome are still unclear. In addition, ABC transporters have important and diverse physiological roles through active transport of a variety of endogenous substrates, which could also contribute to tumour phenotype and treatment response. This is the first study to explore the entire ABC transporter gene family in relation to clinical outcome of ovarian cancer. Using a Taqman low density array format for real-time PCR, we examined expression of all ABC transporter genes in a cohort of 150 serous ovarian cancers. Kaplan-Meier survival analysis revealed that expression of multiple ABCA family transporters was significantly associated with progression-free or overall survival and multivariate modelling identified ABCA1, ABCA5, ABCA6, ABCA8 and ABCA9 as new prognostic markers for serous ovarian cancer that are independent of established clinical indicators. The prognostic significance of ABCA family transporter expression was validated in an independent microarray dataset consisting of 350 serous tumours available through The Cancer Genome Atlas. These ABCA family transporters are not known to transport drugs but instead function in cellular lipid trafficking and cholesterol homeostasis, suggesting that treatment failure may involve mechanisms other than simple efflux of chemotherapeutic drugs. In light of accumulating evidence for the importance of cholesterol and bioactive lipids in several cancers, this study highlights an important area for investigation into the biology and treatment of serous ovarian cancer. 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 1660. doi:10.1158/1538-7445.AM2011-1660

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