Abstract P1-07-30: FOXC1/FOXA1 transcriptional balance in breast cancer: From acquisition of mesenchymal and stem cell traits to occult lymph node independent breast cancer metastasis
Authors: Partha S Ray, Tor W Jensen, Tania Ray, Omid Gholamalamdari, Connie Y Tsai, Bomy Kim, Rohit Bhargava, K V Prasanth
Journal: Cancer Research (2015)

Abstract Background: Distant metastatic spread of cancer cells to other organs from the primary site of origin currently constitutes the most significant contributor to cancer-related morbidity and mortality. Epithelial-to-mesenchymal transition (EMT) is a biologic transformation of cancer cells from a non-migratory phenotype to a migratory one, and is thought to initiate the metastatic cascade in cancer. EMT has also been reported to trigger acquisition of stem cell traits in breast cancer. Transcription factor (TF) Forkhead box C1 (FOXC1), strongly associated with the basal-like and claudin-low breast cancer molecular subtypes, is a powerful EMT inducer and is also a marker of stem/progenitor cells. In contrast, TF forkhead box A1 (FOXA1), strongly associated with luminal subtypes, is an EMT repressor and a luminal differentiation marker, thus seemingly exerting reciprocally opposite transcriptional effects to that of FOXC1. We hypothesized that effective EMT program activation status in breast cancer might be better predicted by examining the expression ratio of an EMT inducer and EMT repressor, such as FOXC1/FOXA1, theoretically being more reflective of net transcriptional effect than either component alone. Methods: Herein we utilize RNA-Seq profiling of the HRAS-transformed MCF10A cell series, a well characterized and widely accepted in vitro model of breast cancer progression and metastasis, to correlate measured FOXC1/FOXA1 ratios to dynamic shifts in EMT marker expression in 3D matrigel cultures and to stem cell traits observed in primary and secondary mammosphere suspension cultures. We further test the ability of the FOXC1/FOXA1 expression ratio to predict lymph node independent breast cancer metastasis and death in independent human breast cancer gene expression datasets. Results: RNA-Seq and qRT-PCR profiling confirmed progressive increase in FOXC1/FOXA1 ratio to correlate with a progressive loss of E-cadherin expression and synchronous gain of EMT markers N-cadherin, Fibronectin, and Vimentin. FOXC1/FOXA1 ratio was found to be directly proportional to mammosphere formation efficiency, a surrogate indicator of stem cell enrichment. In patients without any evidence of nodal metastasis, elevated FOXC1/FOXA1 ratio was associated with significantly decreased 10 year Overall Survival (HR 2.58;95%CI 1.39 to 4.80, p = 0.003, 295 patient Van de Vijver dataset), 10 year Disease-specific Survival (HR 1.74;95%CI 1.16 to 2.61, p = 0.008,1992 patient Curtis dataset) and predicted the development of lung metastasis. Conclusion: Elevated FOXC1/FOXA1 expression ratio indicates EMT program activation in breast cancer, and predicts the associated occurrence of lymph-node-independent distant metastasis and death in human patients. These findings may allow for the early (pre-symptomatic) diagnosis of clinically occult (node negative) metastasis by using the FOXC1/FOXA1 ratio as a biomarker of metastasis and permit institution of appropriate therapy earlier than currently possible. The current study improves our understanding of EMT and highlights the importance of future studies geared towards unraveling mechanisms involved in regulating FOXC1 and FOXA1 expression in breast cancer. Citation Format: Partha S Ray, Tor W Jensen, Tania Ray, Omid Gholamalamdari, Connie Y Tsai, Bomy Kim, Rohit Bhargava, K V Prasanth. FOXC1/FOXA1 transcriptional balance in breast cancer: From acquisition of mesenchymal and stem cell traits to occult lymph node independent breast cancer metastasis [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P1-07-30.