Abstract 64: Identification of novel kinases involved in Estrogen receptor-α (ERα) signalling and the development of endocrine resistance in breast cancer
Authors: Georgios Giamas, Hua Zhang, Wu Zhang, Aleksandra Filipovic, Heinz-Josef Lenz, Justin Stebbing
Journal: Cancer Research (2012)

Abstract Introduction: Therapies targeting estrogen receptor-α (ERα) have transformed the treatment of breast cancer (BC). ER+ BC is usually treated with hormone reduction or anti-estrogens. Still, only one third of the ER+ BC respond to tamoxifen. Mutations in ER are rarely found; instead other mechanisms, namely phosphorylation, have been associated with resistance. Since phosphorylation of ERα is important for the stimulation of its transcriptional activity, focusing on the role of kinases in modulating activity may reveal new druggable targets for exploitation in the clinic. We have performed a human kinome siRNA screening and identified novel kinases involved in ERα transcriptional activity (Giamas et al., Nat Med., 2011). Methodology: A systematic high-throughput RNAi screening using a Human Kinase siRNA Library was carried out to identify novel kinases able to modulate ERα activity, using qRT-PCR as readout. We assessed clinical significance of our observations by analyzing the expression of novel kinases in BC tissue microarrays (TMAs, n=1000) by immunohistochemistry. Pearson chi2 and Fisher's Exact Test were used to correlate kinase expression with various tumor biomarkers; Kaplan Meier analyses were used to investigate impact on disease-free (DFS), overall survival (OS) and response to endocrine therapy. We also determined two single nucleotide polymorphisms (SNPs), taking into consideration that the frequency of the polymorphism is sufficient enough that its impact in clinical outcome would be meaningful on a population level. SNPs have been tested using PCR-RFLP and correlated with the above parameters, notably OS, DFS, and response to endocrine therapy. Results: We identified a novel kinase ‘x’ widely expressed in a collection of breast cancer cell lines. siRNA knockdown inhibited proliferation in both ERα+ and ERα - BC cell lines, while over-expression increased ERα transcriptional activity. Moreover, the kinase of interest was expressed in low (60%) and high levels (40%) after analysis of the breast cancer TMAs. High expression correlated significantly with longer OS (p=0.012) and required more time to develop distant metastasis (p=0.014) in more than 20 yrs follow-up. Interestingly, kinase x expression was positively associated with BRCA1 (p=0.002) and reversely with p53 (p=0.038).Our data indicated that this novel kinase may be an independent prognostic biomarker by Cox Regression analysis (p=0.001). Furthermore, polymorphisms studies indentified 2 individual SNPs of kinase x that were independently associated with DFS (p1=0.029, p2=0.008) and OS (p1=0.006, p2=0.0024). Conclusions: Given that protein kinases have emerged as key regulators of all aspects of neoplasia, the development of drugs for molecular cancer treatment is considered imperative. We propose that our newly identified kinase x could be a successful target. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 64. doi:1538-7445.AM2012-64