COBRE Pilot Project for Binhua P. Zhou: Center of Research in Obesity and Cardiovascular Disease

Breast cancer remains a major challenge, as it remains the second leading cause of cancer-related death in women, with a yearly toll of more than 40,000 deaths in the United States alone. Among five different subtypes of breast cancer, triple-negative breast cancer (TNBC) is the worst subtype that lacks effective targeted therapies and has a tendency toward early metastatic spread to brain and lung, sites known to be associated with a poor prognosis and short survival1-5. Ample epidemiological data indicating that obesity is a significant risk factor for the development and progression of TNBC6-12, particularly for women in Appalachia area12. A better understanding of the molecular events that contribute to pathogenesis of TNBC for women with obesity is crucial to the development of novel strategies to prevent and treat this deadly disease. TNBC intrinsically activates the epithelial-mesenchymal transition (EMT) program that imparts cells with increasing plasticity and fitness during embryonic development, wound healing, tissue fibrosis/remodeling, and metastasis13, 14. Twist, a highly conserved transcription factor that contains basic helix-loop-helix (bHLH) domain, is a key EMT inducer during embryonic development and tumor metastasis15. Overexpression of Twist positively correlates with metastatic potential and poor clinical outcome in patients. We recently found that Twist is di-acetylated in response to inflammatory cytokines and that the di-acetylated Twist binds to and recruits BRD4, a key regulator in transcription regulation, to control gene transcription during EMT and metastasis16. Pharmacologic inhibition of the Twist-BRD4 association using BET-specific inhibitors suppresses tumor cell invasion, cancer stem cell (CSC)-like properties, and tumorigenicity in breast cancer. Our study indicates that the interaction with BRD4 is critical for the oncogenic function of Twist in EMT and cancer. Intriguingly, Twist is abundantly expressed in white and brown adipose tissues from mice and human, as well as in mature adipocytes in comparison with preadipocytes in vitro. Mice with Twist overexpression in adipose tissues (AT) enhance weight gain compared to wild-type mice on a high-fat diet. Conversely, twist heterozygous mice are protected against dietary-induced obesity17-19. In addition, Twist expression is strongly correlated with BMI and insulin resistance in humans. Mechanistically, Twist inhibits PGC-1.. function in adipocytes and thus results in the suppression of mitochondrial biogenesis and energy dissipation, leading to obesity in mice and human. These findings reveal an unexpected physiological role for Twist in maintenance of energy homeostasis in AT and have important implications for understanding the pathogenesis of obesity and diabetes. AT is an endocrine organ characterized with significant plasticity and cellular heterogeneity, the expansion of AT in obesity shares several unique properties that are remarkably similar to the growth of solid tumors20. Both involve chronic inflammation, wound/fibrosis, and hypoxia. Given the key role of Twist in regulating inflammation, wound/fibrosis, fatty acid metabolism, and TNBC, we propose that Twist plays an essential role in the development and progression of both TNBC and obesity. The objective of this proposal is to delineate the critical function and regulation of Twist in TNBC and obesity. Understanding the common mechanism underlying the contribution of Twist in the pathogenesis of TNBC and AT expansion in obesity presents a unique opportunity and challenge for therapeutic intervention to treat these diseases. To test our hypothesis and achieve the objective of this proposal, we have planned experiments with the following Specific Aims: Aim 1: To determine Twist activation in TNBC with obesity from Appalachia population We propose that Twist activation (di-acetylation) is responsible for development and progression of TNBC in obese women from Appalachia area. We will: (1) examine Twist di-acetylation in 200 cases of TNBC specimens and correlates this data with BMI and other clinical information in patients from Appalachia; and (2) determine the expression of cytokines, fibrosis-response genes, and enzymes of fatty acid oxidation in these tumor samples. Aim 2: To determine whether JQ1/MS417 restore lipid metabolism in obese tissue We will address whether BET-specific bromodomain inhibitors, JQ1/MS417, inhibit cytokine secretion, fatty acid bio-synthesis, and oxygen consumption in adipocytes by disrupting the interaction of Twist with BRD4. Aim 3: To examine whether obesity facilitates the development of TNBC in vivo We will challenge the BRCA1-/- mice, which develop TNBC specifically, with high-fat diet and examine whether obesity facilitates the development, progression and metastasis of TNBC. We will also analyze whether JQ1/MS417 treatment prevents the progression and metastasis of tumors as well as the pathogenesis of obesity in this mouse model in vivo. The research proposed in this application is novel and significant, as we envision that the disruption of the function of Twist using BET-specific inhibitors will provide novel therapeutic modality for both TNBC and obesity. We will perform the studies in a highly supportive research environment that is conducive to the success of our proposed research.