Abstract 2097: LIN9 regulation of NEK2 underlies taxol resistance in triple-negative breast cancer

Abstract

Abstract Currently, there are no targeted strategies to combat triple negative breast cancer, resulting in poor patient survival. This disease initially responds well to cytotoxic chemotherapies such as paclitaxel, yet resistance and metastatic recurrence are common. Taxanes cause defects in centrosome function and chromosome segregation leading to cell death. While greatly effective, they are also associated with high toxicity. Thus discovering new therapeutic targets that provide a selective vulnerability for TNBC should yield novel approaches for improving patient outcomes. We discovered that LIN9, a transcriptional regulator of mitosis, is overexpressed in 66% of TNBC and is associated with poor survival. We now report that both LIN9 mRNA and protein expression are upregulated in paclitaxel-resistant versus sensitive cells and directly correlates with paclitaxel IC50 values across eight breast cancer cell lines. In MDA-MB-231 and MDA-MB-468 cell lines, LIN9 silencing results in multi- and micronucleation, and supernumerary centrosomes. Moreover, LIN9 silencing increases sensitivity to paclitaxel in TNBC cells with intrinsic (BT549) or acquired (MDA-MB-231 and -468) resistance. We previously reported that Bromodomain and ExtraTerminal protein inhibitors (BETi) treatment reduces LIN9 expression, and thus determined if BETi could reverse paclitaxel resistance. Treatment with the BETi, JQ1, in conjunction with paclitaxel caused a greater induction of apoptosis, abnormal centrosomes, multi- and micronucleation compared to either drug alone. To identify the mechanism(s) by which suppression of LIN9 reverses paclitaxel resistance, we compared the transcriptomes of MDA-MB-231 and HCC70 TNBC cells transiently transfected with non-targeting or LIN9-targeted siRNAs. The resulting gene list was then filtered to include only genes whose expression correlates with LIN9 in breast cancer, are bound to LIN9 in a published ChIP-Seq dataset, and are associated with breast cancer survival. Using this approach, we identified NIMA-related Kinase 2 (NEK2), a serine/threonine kinase required for centrosome separation during mitosis, as a potential mediator of LIN9-associated paclitaxel resistance. NEK2 is overexpressed in 47% of basal breast cancers and is associated with poor patient outcomes. Additionally, NEK2 is upregulated in paclitaxel-resistant cells and LIN9 silencing decreases expression of NEK2. Importantly, silencing NEK2 expression also restores sensitivity to paclitaxel in resistant cells. Together, these data indicate that increased LIN9 expression in TNBC promotes paclitaxel resistance by upregulating NEK2 and that loss of LIN9 or NEK2 contributes to centrosome dysfunction, potentiating paclitaxel sensitivity. They also indicate that suppressing LIN9 expression using agents such as BET inhibitors may be a viable therapeutic approach for improving paclitaxel efficacy in TNBC patients. Citation Format: Melyssa S. Shively, Ruth A. Keri. LIN9 regulation of NEK2 underlies taxol resistance in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2097.