Abstract B08: ESR1 coregulator binding site inhibitors (ECBIs) as novel therapeutics to target hormone therapy-resistant breast cancer
Authors: Vadlamudi Ratna K. , Sareddy Gangadhara Reddy , Viswanadhapalli Suryavathi , Lee Tae-Kyung , Ma Shi-Hong , Lee Wan Ru , Mann Monica , Krishnan Samaya Rajeshwari , Gonugunta Vijay , Strand Douglas W. , Tekmal Rajeshwar Rao , Ahn JungMo , Raj Ganesh V. 
Published: May 2016

Abstract
Estrogens contribute to the progression of breast cancer via estrogen receptor 1 (ESR1) and current therapies involve either antiestrogens (AE) or aromatase inhibitors (AI). However, most patients develop resistance to these drugs. Critically, therapy-resistant tumors retain ESR1-signaling. Mechanisms of therapy resistance involve the activation of ESR1 in the absence of ligand or mutations in ESR1 that allow interaction between the ESR1 and coregulators leading to sustained ESR1 signaling and proliferation. For patients with therapy-resistant breast cancers, there is a critical unmet need for novel agents to disrupt ESR1 signaling by blocking ESR1 interactions with its coregulators.
Methods: Using rational design, we synthesized and evaluated a small organic molecule (ESR1 coregulator binding inhibitor, ECBI) that mimics the ESR1 coregulator nuclear receptor box motif. Using in vitro cell proliferation and apoptosis assays, we tested the effect of ECBI on several breast cancer cells and therapy-resistant model cells. Mechanistic studies were conducted using established biochemical assays, reporter gene assays, RTqPCR and RNASeq analysis. Gene differential expression lists were analyzed using Ingenuity Pathway Analysis (IPA). ESR1+ve (MCF7 and ZR75) xenografts were used for preclinical evaluation and toxicity. The efficacy of ECBI was tested using an ex vivo cultures of freshly extirpated prrimary human breast tissues.
Results: In estrogen induced proliferation assays using several ESR1+ve model cells, we found that ECBI inhibit growth (IC50=300-500 nM). Importantly, ECBI showed little or no activity on ESR1 negative cells. Further, ECBI also reduced the proliferation of several ESR1 positive hormonal therapy resistant cells, directly interacted with MT-ESR1 with high affinity and significantly inhibited MT-ESR1 driven oncogenic activity. Mechanistic studies showed that ECBI interacts with ESR1, efficiently blocks ESR1 interactions with coregulators and reduces the ESR1 reporter gene activity. RNA sequencing analysis revealed that ECBI blocks multiple ESR1 driven pathways, likely representing the ability of a single ECBI compound to block multiple ESR1-coregulator interactions. Treatment of ESR1-positive xenograft tumors with ECBI (10 mg/Kg/oral) reduced tumor volume by 67% compared to control. Further, ECBI also significantly reduced the proliferation of coregulator-overexpressed breast cancer cells in xenograft model. Using human primary breast tissue ex vivo cultures, we have provided evidence that ECBI has potential to dramatically reduce proliferation of human breast tumor cells.
Conclusions: The ECBI is a novel agent that targets ESR1 with a unique mechanism of action. ECBI has distinct pharmacologic advantages of oral bioavailability, in vivo stability, and is associated with minimal systemic side effects. Remarkably, ECBIs block both native and mutant forms of ESR1 and have activity against therapy resistant breast cancer cell proliferation both in vitro and in vivo and against primary human tissues ex vivo. Thus development of ECBI represents a quantum leap in therapies to target ESR1
Citation Format: Ratna K. Vadlamudi, Gangadhara Reddy Sareddy, Suryavathi Viswanadhapalli, Tae-Kyung Lee, Shi-Hong Ma, Wan Ru Lee, Monica Mann, Samaya Rajeshwari Krishnan, Vijay Gonugunta, Douglas W. Strand, Rajeshwar Rao Tekmal, JungMo Ahn, Ganesh V. Raj. ESR1 coregulator binding site inhibitors (ECBIs) as novel therapeutics to target hormone therapy-resistant breast cancer. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B08.