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Page 276 Cancer Drug Resist 2018;1:266-302 I http://dx.doi.org/10.20517/cdr.2018.18

Despite good initial responses towards non-targeted drugs, ~50% of children with high-risk neuroblastoma
(NB) relapse. Drug-resistance is a major obstacle for successful chemotherapy in high-risk patients. For
high-risk subset, new chemotherapeutic agents are urgently required for better response and also to com-
bat resistance. We repurposed an Ewing’s sarcoma drug, YK-4-279 (targeted towards EWS-FLI1 oncogenic
fusion protein) to target ETS-transcription factors (effectors of ALK and Ras-pathways) which are highly
overexpressed in NB. YK-4-279 inhibited growth of wide range of NB genotypes with different oncogenic
drivers. Independent of ETS-transcription factors we discovered an unexpected mode of action. YK-4-279
rapidly induced a strong mitotic phenotype. Depending on the cell line, treatment resulted in multipolar,
monopolar and fragmented spindles, together leading to disrupted mitotic progression. Notably, YK-4-279
does not affect microtubule acetylation, unlike the conventional mitotic poisons paclitaxel and vincris-
tine. Mechanistically we show that YK-4-279 affected the distribution of Eg5, a key kinesin involved in the
fidelity of microtubule assembly. Interestingly YK-4-279 induced robust apoptosis independent of p53 in
a caspase-3 dependent manner as confirmed by QVD experiments. With respect to mechanism of toxic-
ity our flow cytometry analysis showed that the majority of cells died in subsequent interphase, not in
mitosis. Strikingly YK-4-279 overcame multi-drug resistance in vincristine resistant NB cell lines. Despite
extremely high levels of a MDR1/ABCB1 (drug transporter), YK-4-279 was unaffected, suggesting it is not a
substrate for MDR1 transporter. We also showed that YK-4-279 effectively synergized with clinical mitotic
inhibitors, highlighting the potential of YK-4-279 as an adjuvant chemotherapeutic agent. 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.

21. A potential role for IL6ST mediating endocrine resistance in breast cancer via interaction
with the ER signaling pathway

1,2
1
1,2
Duniya Mosly , Arran K. Turnbull , Simon P. Langdon , Andrew H. Sims 2
1 Cancer Research UK Edinburgh Centre and Division of Pathology Laboratory, MRC
Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh EH4 2XU, UK
2 Applied Bioinformatics of Cancer, University of Edinburgh Cancer Research Centre, MRC Institute of Genet-
ics and Molecular Medicine, Edinburgh EH4 2XR, UK

Aim: IL6ST (gp130) receptor has been identified as a predictive biomarker of endocrine treatment response
in breast cancer patients and is included in the “Endopredict” test. At least seven cytokines signal via
IL6ST. Interleukin-6 (IL-6) can mediate effects via two signaling pathways; classic signaling (through the
membrane-bound IL-6 receptor, IL-6R) and trans-signaling (via non-signaling membrane-bound soluble
IL-6R, sIL-6R). Both pathways may occur in parallel and activate cells.

Methods: Three ERa+ luminal breast cancer cell lines (MCF-7, T47D, ZR-75-1) were chosen to examine
IL6ST expression by western blot, gel electrophoresis and qRT-PCR. Proliferation assays were carried out
to investigate the effects of IL-6 family cytokines on cell growth. The action of both IL-6 and OSM on cell
migration and downstream signaling pathways was studied. The extent of trans-signaling occurrence was
investigated.

Results: Three cell lines were shown to express varying levels of full length IL6ST. IL6ST soluble forms
were identified in the three cell lines. Surprisingly, the growth response to the cytokines was variable
across the cell lines. IL6 caused a modest increase in growth in MCF-7 but produced inhibition in ZR-75-1.
OSM and LIF stimulated growth in MCF-7, whereas only OSM inhibited ZR-75-1. Interestingly, no signifi-
cant effect on growth was seen in T47D. Both STAT3 and MAPK/ERK pathways were activated to different