Potential Role of Pargyline in Overcoming Adaptive Resistance in Breast Cancer Cells.

Background: Estrogen-induced breast carcinogenesis is shown to be characterized by global changes in histone modifications. LSD1 (KDM1), a histone demethylase enzyme, play a key role in establishing specific histone methyl marks at target gene promoters. Recent evidence suggest that LSD1 is recruited to a significant fraction of estrogen receptor (ER) target genes and is required to demethylate proximal histones to enable productive ER transcription. These emerging findings also suggest that deregulation of LSD1 epigenetic pathway could contribute to hormonal independence and adaptive resistance in breast cancer cells. In this study, we examined the therapeutic efficacy of treating breast tumor cells with Pargyline, an FDA approved drug for blocking LSD1 functions, and evaluated the therapeutic benefit.Methods: To test this hypothesis, we used model cells that acquired resistance to hormonal therapy including (a) MCF7-HER2 that overexpress oncogene neu/HER2, (b) MCF7-Tam that have acquired resistance to Tamoxifen, (c) MCF7-Ca-LTLT cells that have acquired resistance to Letrozole, (d) MCF7 cells that overexpress proto-oncogene PELP1 (MCF7-PELP1). Parental MCF7 cells were used as a control. Cells were treated with LSD1 inhibitor Pargyline either alone or in combination with Letrozole and Dasatinib. Activation of ER genomic functions was studied using luciferase reporter gene assays. Epigenetic modifications at target promoters were analyzed by Chromatin immune precipitation (CHIP) assays using H3 methyl (di and Tri -H3K4, -K9) specific antibodies. Biological significance and hormonal therapy sensitivity was measured by in vitro cell proliferation assays. Xenograft studies were used to validate the drug effect in vivo. Pilot studies were performed for delivery of drug combinations using nanoparticle formulation.Results: Reporter gene assays showed that LSD1 has potential to enhance ER mediated transcription. LSD1 functionally interacts with ER coregulator PELP1 and is recruited to ER target genes. Pargyline substantially inhibited ER transactivation functions. ChIP analysis revealed that aggressively growing breast cancer cells and therapy resistant cells have distinct activating histone methyl modifications at growth regulatory ER target genes. Treatment of breast cancer models with LSD1 inhibitor Pargyline facilitated reversal of these specific modifications and thereby inhibited the growth of breast cancer cells in vitro and in vivo models. Combinatorial therapy using three agents; (a) that block Estrogen receptor genomic actions (Tamoxifen or Letrozole), (b) ER nongenomic actions (Dasatinib) and (c) ER epigenetic modifications (Pargyline) showed the most promising therapeutic effect compared to single agent therapy on the growth of therapy resistant cells.Conclusions: Our results suggest that histone methyl modification plays a role in therapy resistance and validates the therapeutic potential of Pargyline in combinational therapies. Collectively, these results suggest that targeting LSD1 axis with Pargyline in combination with current endocrine therapies will have better therapeutic effect and may inhibit or delay development of hormonal resistance, thus providing major benefits to patients care. This study is funded by Komen grant KG090447. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 409.