Abstract 3204: The identification of AURKB, BUB1B and CDK18 as putative molecular targets for breast cancer using kinome focused RNAi screens.

Abstract

Abstract The signal transduction events regulated by the kinase family of proteins control many cellular processes including normal cell proliferation and survival. The disruption of signaling pathways regulated either directly or indirectly by protein kinases is frequently observed in cancer cells, including breast cancer, and thus the development of inhibitors of specific kinases has become a major focus of drug discovery in oncology. Functional genomic RNAi screens of the kinome conducted within the context of a specific tumor type and/or a specific cancer-associated genetic background have the potential to identify kinases required for cancer cell survival. A comparison of RNAi screening data that we have obtained targeting the human kinome in three breast cancer cell lines with other published RNAi screens led us to perform a detailed analysis of the phenotypic effects of silencing AURKB, BUB1B, and CDK18 in multiple breast cancer cell lines with the aim of further characterizing kinase genes required for the growth of breast tumor cells. The non-tumorigenic mammary epithelial cell line MCF-10A exhibited minimal BUB1B protein expression and no detectable expression of AURKB protein, while all of the breast cancer cell lines studied (a total of eight), exhibited expression of both of these proteins. This differential expression was associated with functional differences in silencing of AURKB or BUB1B resulting in suppression of colony formation in almost all of the breast cancer cell lines tested, but not in MCF-10A cells. Silencing of AURKB also induced cell cycle arrest, caspase 3/7-activation and PARP cleavage in MDA-MB-468 and HCC-38 cells, but not in MCF-10A cells. These results were phenocopied by an ATP-competitive inhibitor of aurora kinase with selectivity for aurora B kinase ZM447439. BUB1B silencing resulted in cell cycle arrest in both MDA-MB-468 and HCC-38 cells, but caspase 3/7-activation and PARP cleavage were only observed in MDA-MB-468 cells. None of these effects were observed in MCF-10A cells. Little is known as to the functional role of CDK18, but its silencing induced cell cycle arrest, caspase 3/7-activation and PARP cleavage in MDA-MB-468. In contrast we saw minimal phenotypic effects in HCC-38 cells and MCF-10A cells, even though both of these cell lines express CDK18. To further elucidate the functional role of CDK18, we performed genome-wide expression profiling of MDA-MB-468 cells in which CDK18 had been silenced. Interestingly, one of the transcripts showing the greatest decrease in expression following silencing of CDK18 was CCNE1 that encodes cyclin E1. Another cyclin gene, CCND1, also exhibited a decrease in expression following silencing of CDK18, and we are currently examining this potential link between CDK18 function and cyclin E1 and cyclin D1 in more detail. Citation Format: Lihui Ou, Kristen Gehlhaus, Tamara Jones, Konrad Huppi, Natasha Caplen. The identification of AURKB, BUB1B and CDK18 as putative molecular targets for breast cancer using kinome focused RNAi screens. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3204. doi:10.1158/1538-7445.AM2013-3204