Abstract 5564: Dual inhibition of TTK and HSPA9 functions to synergistically decrease viability in triple-negative breast cancer cell lines
Authors: Lavender Nicole A. , Chow Donald , Yin Holly , Carpten John 
Published: May 2015

Abstract
Background: Triple-negative breast cancer (TNBC) is an aggressive subtype that is more likely to develop in African-American women and younger females. TNBCs have higher rates of metastasis, recurrence, and mortality and also lack effective targeted treatments. We have found that compared to normal tissue, expression levels of the mitotic kinase TTK are elevated in metastatic TNBC tumors, particularly in African-American patients. We have demonstrated in TNBC cell lines that inhibiting TTK reduces cell viability, proliferation, and survival. Unfortunately, none of TTK inhibitors currently available are suitable for clinical use. Therefore, in an effort to generate an accessible treatment, we targeted an interaction between TTK and the chaperone protein, HSPA9. This events leads to super-activation of TTK and we hypothesized that disrupting this event may serve as an alternative and more effective method for targeting TTK as a TNBC chemotherapeutic.
Methods: We analyzed the effects of individual and combined inhibition of TTK and HSPA9 on cell viability in eight TNBC cell lines (i.e., BT-20, BT-549, CAL-51, HCC38, MDA-MB-231, MDA-MB-436, MDA-MB-453, and MDA-MB-468). AZ3146 and MKT-077 were used to inhibit TTK and HSPA9, respectively. The cells were first treated with each agent individually to determine the dose response profile and IC50 values. Then, to achieve therapeutically relevant dose combinations, cells were treated with IC20 doses of AZ3146 and sub-nanomolar (&lt;IC10) doses of MKT-077.
Results: For the single agent treatments, cells were more sensitive to MKT-077 alone than AZ3146. MKT-077 IC50 values ranged between 0.96-4.24μM, while AZ3146 values were 1.55-52.4μM. AZ3146+MKT-077 resulted in significantly lower viability compared to treatment with either agent alone (p=0.04-2.0x10-10). The combination was particularly effective in BT-549, MDA-MB-231, and MDA-MB-436 cells since they exhibited little sensitivity to AZ3146 as a single agent (IC50≥22.8μM). However, when each of these cell lines was treated with an IC20 dose of AZ3146 and 0.05-100nM of MKT-077, we observed a statistically significant decease in cell viability (p = 0.03-8.5x10-10).
Conclusions: Our findings show that simultaneously treating TNBC cells with low doses of TTK and HSPA9 inhibitors results in a substantially greater reduction in viability. Although previous data suggest that TTK inhibitors cause selective cancer cell death, none of these agents are clinically available. Additionally, MKT-077 was shown to exhibit significant antitumor activity in a variety of in vitro and in vivo model systems. However, we have shown that TNBC cells can be effectively treated with a TTK inhibitor combined with very low doses of MKT-077. More importantly, since these proteins are both overexpressed in tumor cells, dual inhibition of TTK and HSPA9 may be a selective, safe, and highly effective chemotherapeutic strategy for TNBC.
Citation Format: Nicole A. Lavender, Donald Chow, Holly Yin, John Carpten. Dual inhibition of TTK and HSPA9 functions to synergistically decrease viability in triple-negative breast cancer cell lines. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5564. doi:10.1158/1538-7445.AM2014-5564