Abstract P2-09-01: Reactivation of epigenetically silenced retinoic acid receptor-beta for therapy of breast cancer- from molecular mechanism to potential clinical applications
Authors: Ordentlich P , Nguyen N , Jin K , Sadik H , Han L , Sukumar S 
Published: August 2014

Background: Triple negative breast cancer (TNBC) is a subgroup of breast cancer that is characterized by a lack of expression of targets for validated targeted therapies. Although responses to chemotherapy are observed, resistance develops rapidly. Genomic and molecular profiling of TNBC has identified multiple contributors to their uncontrolled growth including epigenetic dysregulation of genes important for normal cell growth and differentiation. Recent studies have shown that histone deacetylase (HDAC) inhibitors reverse the epigenetic profile of tumors, resulting in the re-expression of silenced genes encoding proteins such as ERα, EGFR, and RARβ. Entinostat is an oral, class 1 isoform selective HDACi recently shown in a phase 2 study to be active in ER+ breast cancer. Clinical trials with entinostat in HER2+ and TNBC are also in progress. We hypothesized that combining epigenetic therapy using entinostat, with differentiation therapy using a retinoic acid receptor agonist- All Trans Retinoic Acid (ATRA) will provide an effective strategy for impeding the growth of TNBC and potentially sensitize tumors to commonly used chemotherapies (doxorubicin, carboplatin, paclitaxel).

Methods: Combination studies of entinostat combined with ATRA with and without non-toxic doses (half clinical) of chemotherapy were carried out in vitro and in xenograft models of TNBC. Expression of RARb and downstream effectors were measured in cell lines and tumors from xenograft studies.

Results: Nontoxic (half clinical) doses of doxorubicin when combined with entinostat and ATRA in vitro and in vivo exerted the best response when compared with combinations using paclitaxel and carboplatin. Re-expression of the silenced RARβ and downstream effectors was observed in the cell lines treated in vitro and in tumor xenografts of MDA-MB-231 and SUM159 cells in vivo. To gain insight into mechanism of action for the entinostat – doxorubicin combinations we examined the expression of enzyme targets of doxorubicin and determined that entinostat treatment reduced the expression of topoisomerase (Topo) II enzymes. Besides its conventional role in cell replication and division, Topo II has an important function in regulating transcription, especially in the nuclear receptor pathways such as ER and RAR. Our results demonstrated that Topo IIβ has a dual role in regulating the expression of RARβ. Inhibition of Topo IIβ in a time- and dose dependent manner up-regulated the expression of RARβ and its downstream genes. However, completely eliminating Topo IIβ decreased expression of RARβ.

Conclusions: Our detailed study provides a new approach to treating TNBC using combinations of nontoxic doses of chemotherapeutic drugs with epigenetic therapy, and a deeper understanding of the molecular pathways involved in this response. These results suggest that combination of entinostat and a retinoic acid receptor agonist with low dose doxorubicin will provide an effective strategy for treatment of TNBC.

Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P2-09-01.