Abstract P6-04-06: Targeting cancer stem cells in triple negative breast cancer by a combination treatment of LBH589 and salinomycin
Authors: Kai M , Kanaya N , Warden C , Luu T , Chen S 
Published: August 2014

Background and purpose:

Triple-negative breast cancer (TNBC) is aggressive with poorer prognosis compared to breast cancer that is positive for hormone receptors or HER2. This can be explained further due in part to the existence of breast cancer stem cells (BCSCs). Our lab has found that histone deacetylase (HDAC) inhibitors are effective in the growth suppression of TNBC in vitro and in vivo. To understand the mechanisms involved, we performed RNA-seq analysis for TNBC cells treated with two HDAC inhibitors (LBH589 and Entinostat). Our RNA-seq analysis and studies from other laboratories have found that HDAC inhibitors may modify multiple signaling pathways with undesired effects due to its broad reactivity. To improve the efficacy of HDAC inhibitors in the treatment of TNBCs, we have examined the effects of LBH589 in combination with several drugs. Among them, Salinomycin works effectively with LBH589. It has been used as an antibiotic for farm animals, but has been identified from a screen of a large library of chemicals to target BCSCs (Gupta et al., Cell, 2009). The purpose of this study is to examine the synergistic effect between LBH589 and Salinomycin in the anticipation of their clinical utility and to evaluate a new target therapy to treat TNBC more efficiently and completely.

Material and method:

Two TNBC cell lines (HCC1937, MDA-MB-231) were used as models to examine the combined effects of Salinomycin and LBH589. Cell proliferation studies were performed through 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. To evaluate the effects on BCSC population, mammosphere assay was used for estimating self-renewal capacity and ALDEFLUOR analysis was used for monitoring the distribution of BCSCs in treated samples. For tumor induction experiments, ALDH+ cells were injected into NSG mice. As a therapeutic study, single or combined use of LBH589 (10mg/kg) and Salinomycin (5mg/kg) were peritoneally injected into non-obese diabetic scid gamma (NSG) mice 3 days a week. Tumor volume, body weight and food intake were checked weekly. For mechanistic analysis, qPCR, Western blotting and IHC were performed for gene and protein expression using tumors from mice from different treatments.

Results:

Salinomycin and LBH589 worked synergistically in the suppression of the proliferation of TNBCs (IC50 was 68.8nM and 13.1nM, respectively). Similarly, both drugs inhibited mammosphere formation and ALDH positive population in a synergistic manner. The combination of LBH589 (16nM) with Salinomycin (60nM) reduced 60% of mammosphere formation and 94% of ALDH positive population compared with DMSO treatment. In a mouse model, the combination of LBH589 and Salinomycin had an inhibitory effect on tumor growth compared with the control group and the groups treated with single drug. In all groups, no side effect was seen. The results from mechanistic studies showed that the combination of LBH589 and Salinomycin regulated the Wnt/b-catenin pathway.

Discussion:

We have evidence that the combination of LBH589 and Salinomycin has a synergistic effect on TNBC through Wnt pathway and suggested this combination could potentially be a new therapeutic strategy for exploring targeting therapy in TNBC.

Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P6-04-06.