The small molecule inhibitor YK-4-279 disrupts mitotic progression of neuroblastoma cells, overcomes drug resistance and synergizes with inhibitors of mitosis

Madhu Kollareddy,Alice Sherrard,Ji Hyun Park,Marianna Szemes,Kelli Gallacher,Zsombor Melegh,Sebastian Oltean,Martin Michaelis,Jindrich Cinatl,Abderrahmane Kaidi,Karim Malik

doi:10.1016/j.canlet.2017.05.027

Madhu Kollareddy, Alice Sherrard + Show 9 more

Open Access

https://doi.org/10.1016/j.canlet.2017.05.027

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Journal: Cancer Letters	Publication Date: Jun 7, 2017
Citations: 31	License type: cc-by

Affiliation: University of Bristol, University of Kent

Abstract

Neuroblastoma is a biologically and clinically heterogeneous pediatric malignancy that includes a high-risk subset for which new therapeutic agents are urgently required. As well as MYCN amplification, activating point mutations of ALK and NRAS are associated with high-risk and relapsing neuroblastoma. As both ALK and RAS signal through the MEK/ERK pathway, we sought to evaluate two previously reported inhibitors of ETS-related transcription factors, which are transcriptional mediators of the Ras-MEK/ERK pathway in other cancers. Here we show that YK-4-279 suppressed growth and triggered apoptosis in nine neuroblastoma cell lines, while BRD32048, another ETV1 inhibitor, was ineffective. These results suggest that YK-4-279 acts independently of ETS-related transcription factors. Further analysis reveals that YK-4-279 induces mitotic arrest in prometaphase, resulting in subsequent cell death. Mechanistically, we show that YK-4-279 inhibits the formation of kinetochore microtubules, with treated cells showing a broad range of abnormalities including multipolar, fragmented and unseparated spindles, together leading to disrupted progression through mitosis. Notably, YK-4-279 does not affect microtubule acetylation, unlike the conventional mitotic poisons paclitaxel and vincristine. Consistent with this, we demonstrate that YK-4-279 overcomes vincristine-induced resistance in two neuroblastoma cell-line models. Furthermore, combinations of YK-4-279 with vincristine, paclitaxel or the Aurora kinase A inhibitor MLN8237/Alisertib show strong synergy, particularly at low doses. Thus, YK-4-279 could potentially be used as a single-agent or in combination therapies for the treatment of high-risk and relapsing neuroblastoma, as well as other cancers.

Highlights

BRD32048 was reported to act as a perturbagen of ETV1 [16] while YK-4-279 was initially characterized as a disruptor of the EWS-FLI/DHX9 interaction [17], and later shown to affect ERG and ETV1 [18,21]
Our initial screening of these compounds showed that BRD32048 treatment of SK-N-AS, SK-N-BE(2)-C and Kelly neuroblastoma cell lines had no effect on cell proliferation (Fig. 1A), even at high micromolar concentrations (Supplementary Fig. S1)
We have shown that the small molecule inhibitor YK-4-279 causes apoptosis of neuroblastoma cell lines, irrespective of their diverse oncogenic genotypes

Summary

Introduction

M. Kollareddy et al / Cancer Letters 403 (2017) 74e85 in non-MNA high-risk neuroblastoma [3,4]. There still remain non-MNA high-risk neuroblastomas for which oncogenic drivers remain unclear, even taking into account activating point mutations of the Anaplastic Lymphoma Kinase (ALK) gene, which are apparent in less than 10% of neuroblastomas, encompassing the non-MNA and MNA high risk subtypes [5]. As a consequence of the lack of common druggable targets, chemotherapy for neuroblastoma still relies largely on DNA damaging agents and inhibitors of mitosis [6]. We sought to rationalize a broader, but relatively specific target class for new neuroblastoma therapeutics

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