ABSTRACT
Breast cancer is the most common cancer and the second leading cause of cancer mortality in women in the
U.S. Approximately 75% of breast cancers are estrogen receptor ¿ (ER¿) positive. The current standard of
care for postmenopausal women with ER¿+ breast cancers involves the choice of an upfront aromatase
inhibitor (AI), a tamoxifen (Tam)-AI, or AI-Tam switch strategy. Tam is the first line therapy for premenopausal
women, second line for postmenopausal women, and a chemopreventive agent for all age groups. Thus, Tam
continues to remain an important selective ER modulator for treatment of ER¿+ breast cancer patients.
However, ~ 30% of ER¿+ patients do not respond to Tam because of de novo resistance, and most who do
respond eventually acquire Tam resistance (TamR). Breast cancer patients who relapse on Tam retain
functional ER¿ signaling, suggesting that endocrine resistance involves agonistic function of Tam or estrogen
independent ER¿ activity. Crosstalk between ER¿ and growth factor receptor (GFR) signaling pathways is
believed to contribute to endocrine resistance. However, clinical trials with GFR inhibitors have yielded mixed
results, indicating complex ER biology and involvement of novel crosstalk or additional/alternate mechanisms
in endocrine resistance. In this application, based on our data from isogenic models of acquired and de novo
TamR, we propose a novel transcription reprogramming induced mechanism of TamR that ensues from ERa-
dependent/Tam-induced upregulation of ß-catenin/TCF-mediated transcriptional activity. This is further
supported by our data that show ectopic ß-catenin expression in Tam-sensitive ERa+ breast cancer cells
transforms Tam from an antagonist to an agonist with resultant ERa-dependent Tam-induced transcriptional
activation of ß-catenin/TCF-responsive genes. Rad6B (a.k.a UBE2B), an ubiquitin conjugating enzyme (UBC),
stabilizes and activates ¿-catenin by K63-linked polyubiquitination (polyUb) of its lysine 394 residue, a
ubiquitination event that protects ¿-catenin from 26S proteasomal degradation. Rad6B is itself a ¿-catenin
transcriptional target, thus creating a vicious positive feedback loop between Rad6B gene expression and ¿-
catenin oncogenic activation. Based on these data, we hypothesize that Rad6B-mediated ß-catenin
polyubiquitination and transcriptional activation drive TamR by facilitating Tam-induced ubiquitinated-ß-
catenin/ERa complex formation and transcriptional reprogramming of ß-catenin/TCF gene targets. Hence a
hyperactive Rad6B/ß-catenin axis would drive cells from a Tam-sensitive to a Tam-refractory state, whereas
inhibition of Rad6B catalytic activity will inhibit ß-catenin ubiquitination/activation and uncouple TCF/ß-
catenin/ERa coregulated transcriptional programs resulting in restoration of Tam sensitivity. We will test this
hypothesis with two specific aims. Specific Aim 1. Establish the causative role of Rad6B in tamoxifen-induced
ß-catenin/TCF transcriptional reprogramming and TamR breast cancer development and progression. Specific
Aim 2. Identify and characterize ß-catenin/ERa complex regulated transcriptional networks and underlying
alterations in epigenetic landscape associated with TamR and assess their vulnerability to Rad6B or ß-catenin
inhibition. This study will uncover the molecular underpinnings of a novel paradigm shifting hypothesis that has
the potential to impact clinical management of endocrine resistant breast cancers. It will lay the foundation for
testing a new nontoxic strategy for treating TamR breast cancers, as well as identify markers with
diagnostic/prognostic and/or therapeutic potential.
