P4-01-18: AP-1 Blockade Potentiates the Anti-Tumor Effect of Endocrine Treatment and Reverts the Resistant Phenotype in Hormone Receptor-Positive Breast Cancer.
Authors: Malorni L , Giuliano M , Migliaccio I , Wang T , Creighton CJ , Lupien M , Hilsenbeck SG , Healy N , Mazumdar A , Trivedi MV , Jeselsohn R , He HH , Fu X , Gutierrez C , Brown M , Brown PH , Osborne CK , Schiff R 
Published: June 2012

Abstract
Background: Resistance to endocrine therapy is a major clinical issue. The transcription factor AP-1 is a key regulator of cell growth and survival as well as a downstream signaling component of several pathways deregulated in endocrine-resistant breast cancer. We have previously shown that acquired endocrine resistance is associated with increased AP-1 activity. AP-1 has also been shown to interact with and modulate the ER network and transcriptional program, especially under hyperactive growth factor signaling, which is commonly associated with endocrine resistance. We hypothesized that interfering with AP-1 function would circumvent endocrine resistance possibly due to its role in modulating ER transcriptional activity.
Methods and results: We inhibited AP-1 function by a genetic approach. We used two different MCF7 clones stably transfected with a Doxycycline (Dox)-inducible dominant-negative (DN) c-Jun (MCF7/Tet-Off Tam67 clones 62 and 67) and two vector-alone control MCF7 clones. Xenografts of these clones were established in ovariectomized nude mice supplemented with estrogen (E2). Mice were then randomized to continued E2 supplementation (control) or to endocrine therapy with either estrogen deprivation (ED) or tamoxifen (Tam), all in the presence or absence of Dox to induce the DN c-Jun expression. AP-1 blockade in both MCF7/Tet-Off Tam67 clones significantly enhanced sensitivity to Tam by reducing time to tumor size halving (p=.014 and p=.006 for clone 62 and 67, respectively) and time to complete tumor disappearance (p=.001 and p=.0034 for clone 62 and 67, respectively). Similar results were obtained with ED treatment. In addition, AP-1 blockade significantly delayed the onset of Tam resistance by increasing time to tumor size doubling (p=.0028). Furthermore, induction of DN c-Jun resulted in a dramatic shrinkage of growing tumors after long-term Tam treatment, suggesting reversal of endocrine resistance with AP-1 blockade. None of the above effects was observed in control clones upon Dox removal. Interestingly, no significant effect of AP-1 blockade was observed on E2-stimulated tumor growth. IHC analysis showed that AP-1 blockade induced tumor response by reducing proliferation (i.e., decreased % of Ki67- and phospho-Histone 3-positive cells) and by inducing apoptosis (i.e., increased % of cleaved caspase 3/7-positive cells). Bioinformatic analyses were conducted to intersect our MCF7 xenograft/Tam-resistant gene signature and the datasets of genes associated with ER DNA-binding sites obtained by whole-genome ER cistromic analysis under estrogen or epidermal growth factor (EGF) stimulation of MCF7 cells. A significant enrichment of the genes associated with the EGF-unique ER DNA-binding sites was observed within our Tam-resistant signature (p&lt;2E-16). Remarkably, 90% of these DNA binding sites harbored an AP-1 motif.
Conclusions: We show that AP-1 blockade increases tumor sensitivity and circumvents resistance to endocrine therapy, thus warranting the development of AP-1-targeted therapy to improve endocrine treatment outcomes. Overall, we suggest that AP-1 is critical in induction of a switch in the ER transcriptional program and may be a new hallmark of endocrine resistance.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-01-18.