The SOX11 transcription factor is a critical regulator of basal-like breast cancer growth, invasion, and basal-like gene expression.

Basal-like breast cancers (BLBCs) are aggressive breast cancers associated with poor survival. Defining the key drivers of BLBC growth will allow identification of molecules for targeted therapy. In this study, we performed a primary screen integrating multiple assays that compare transcription factor expression and activity in BLBC and non-BLBC at the RNA, DNA, and protein levels. This integrated screen identified 33 transcription factors that were elevated in BLBC in multiple assays comparing mRNA expression, DNA cis-element sequences, or protein DNA-binding activity. In a secondary screen to identify transcription factors critical for BLBC cell growth, 8 of the 33 candidate transcription factors (TFs) were found to be necessary for growth in at least two of three BLBC cell lines. Of these 8 transcription factors, SOX11 was the only transcription factor required for BLBC growth, but not for growth of non-BLBC cells. Our studies demonstrate that SOX11 is a critical regulator of multiple BLBC phenotypes, including growth, migration, invasion, and expression of signature BLBC genes. High SOX11 expression was also found to be an independent prognostic indicator of poor survival in women with breast cancer. These results identify SOX11 as a potential target for the treatment of BLBC, the most aggressive form of breast cancer

The SOX11 transcription factor is a critical regulator of basal-like breast cancer growth, invasion, and basal-like gene expression

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Analysis of phosphatases in ER-negative breast cancers identifies DUSP4 as a critical regulator of growth and invasion.

Estrogen receptor (ER)-negative cancers have a poor prognosis, and few targeted therapies are available for their treatment. Our previous analyses have identified potential kinase targets critical for the growth of ER-negative, progesterone receptor (PR)-negative and HER2-negative, or "triple-negative" breast cancer (TNBC). Because phosphatases regulate the function of kinase signaling pathways, in this study, we investigated whether phosphatases are also differentially expressed in ER-negative compared to those in ER-positive breast cancers. We compared RNA expression in 98 human breast cancers (56 ER-positive and 42 ER-negative) to identify phosphatases differentially expressed in ER-negative compared to those in ER-positive breast cancers. We then examined the effects of one selected phosphatase, dual specificity phosphatase 4 (DUSP4), on proliferation, cell growth, migration and invasion, and on signaling pathways using protein microarray analyses of 172 proteins, including phosphoproteins. We identified 48 phosphatase genes are significantly differentially expressed in ER-negative compared to those in ER-positive breast tumors. We discovered that 31 phosphatases were more highly expressed, while 11 were underexpressed specifically in ER-negative breast cancers. The DUSP4 gene is underexpressed in ER-negative breast cancer and is deleted in approximately 50 % of breast cancers. Induced DUSP4 expression suppresses both in vitro and in vivo growths of breast cancer cells. Our studies show that induced DUSP4 expression blocks the cell cycle at the G1/S checkpoint; inhibits ERK1/2, p38, JNK1, RB, and NFkB p65 phosphorylation; and inhibits invasiveness of TNBC cells. These results suggest that that DUSP4 is a critical regulator of the growth and invasion of triple-negative breast cancer cells

Elevated Expression of MAPK Phosphatase 3 in Breast Tumors—A Mechanism of Tamoxifen Resistance

Antiestrogen resistance is a major clinical problem in the treatment of breast cancer. Altered growth factor signaling with estrogen receptor (ER) α has been shown to be associated with the development of resistance. Gene expression profiling was utilized to identify MAPK phosphatase 3 (MKP3) whose expression was correlated with response to the antiestrogen tamoxifen in both patients and in vitro derived cell line models. Overexpression of MKP3 rendered ERα-positive breast cancer cells resistant to the growth inhibitory effects of tamoxifen, and enhanced tamoxifen agonist activity in endometrial cells. MKP3 overexpression was associated with lower levels of activated ERK1,2 phosphorylation in the presence of estrogen, but that estrogen deprivation and tamoxifen treatment decreased MKP3 phosphatase activity, leading to an up-regulation of pERK1,2 MAPK, phosphoserine 118 of ERα, and cyclin D1. The MEK inhibitor PD98059 blocked tamoxifen-resistant growth. Accumulation of reactive oxygen species was observed with tamoxifen treatment of MKP3 overexpressing cells, and antioxidant treatment increased MKP3 phosphatase activity, thereby blocking resistance. Furthermore, PD98059 increased the levels of phospho-JNK in tamoxifen-treated MKP3 overexpressing cells, suggesting an interaction between MKP3 levels, activation of ERK1,2 MAPK, and JNK signaling in human breast cancer cells. MKP3 represents a novel mechanism of resistance which may be a potential biomarker for the use of ERK1,2 and/or JNK inhibitors in combination with tamoxifen treatment

Intratumoral Heterogeneity in a Trp53-Null Mouse Model of Human Breast Cancer

Intratumoral heterogeneity correlates with clinical outcome and reflects the cellular complexity and dynamics within a tumor. Such heterogeneity is thought to contribute to radio- and chemoresistance since many treatments may only target certain tumor cell subpopulations. A better understanding of the functional interactions between various subpopulations of cells, therefore, may help in the development of effective cancer treatments. We identified a unique subpopulation of tumor cells expressing mesenchymal-like markers in a p53 null mouse model of basal-like breast cancer using fluorescence-activated cell sorting and microarray analysis. Both in vitro and in vivo experiments revealed the existence of crosstalk between these “mesenchymal-like” cells and tumor-initiating cells. Knockdown of genes encoding ligands upregulated in the mesenchymal cells and their corresponding receptors in the tumor-initiating cells resulted in reduced tumorigenicity and increased tumor latency. These studies illustrate the non-cell autonomous properties and importance of cooperativity between tumor subpopulations

Prevention of Hypovolemic Circulatory Collapse by IL-6 Activated Stat3

Half of trauma deaths are attributable to hypovolemic circulatory collapse (HCC). We established a model of HCC in rats involving minor trauma plus severe hemorrhagic shock (HS). HCC in this model was accompanied by a 50% reduction in peak acceleration of aortic blood flow and cardiomyocyte apoptosis. HCC and apoptosis increased with increasing duration of hypotension. Apoptosis required resuscitation, which provided an opportunity to intervene therapeutically. Administration of IL-6 completely reversed HCC, prevented cardiac dysfunction and cardiomyocyte apoptosis, reduced mortality 5-fold and activated intracardiac signal transducer and activator of transcription (STAT) 3. Pre-treatment of rats with a selective inhibitor of Stat3, T40214, reduced the IL-6-mediated increase in cardiac Stat3 activity, blocked successful resuscitation by IL-6 and reversed IL-6-mediated protection from cardiac apoptosis. The hearts of mice deficient in the naturally occurring dominant negative isoform of Stat3, Stat3β, were completely resistant to HS-induced apoptosis. Microarray analysis of hearts focusing on apoptosis related genes revealed that expression of 29% of apoptosis related genes was altered in HS vs. sham rats. IL-6 treatment normalized the expression of these genes, while T40214 pretreatment prevented IL-6-mediated normalization. Thus, cardiac dysfunction, cardiomyocyte apoptosis and induction of apoptosis pathway genes are important components of HCC; IL-6 administration prevented HCC by blocking cardiomyocyte apoptosis and induction of apoptosis pathway genes via Stat3 and warrants further study as a resuscitation adjuvant for prevention of HCC and death in trauma patients

A Renewable Tissue Resource of Phenotypically Stable, Biologically and Ethnically Diverse, Patient-Derived Human Breast Cancer Xenograft Models

Breast cancer research is hampered by difficulties in obtaining and studying primary human breast tissue, and by the lack of in vivo preclinical models that reflect patient tumor biology accurately. To overcome these limitations, we propagated a cohort of human breast tumors grown in the epithelium-free mammary fat pad of SCID/Beige and NOD/SCID/IL2γ-receptor null (NSG) mice, under a series of transplant conditions. Both models yielded stably transplantable xenografts at comparably high rates (~21% and ~19%, respectively). Of the conditions tested, xenograft take rate was highest in the presence of a low-dose estradiol pellet. Overall, 32 stably transplantable xenograft lines were established, representing 25 unique patients. Most tumors yielding xenografts were “triple-negative” (ER-PR-HER2+) (n=19). However, we established lines from three ER-PR-HER2+ tumors, one ER+PR-HER2−, one ER+PR+HER2− and one “triple-positive” (ER+PR+HER2+) tumor. Serially passaged xenografts show biological consistency with the tumor of origin, are phenotypically stable across multiple transplant generations at the histologic, transcriptomic, proteomic, and genomic levels, and show comparable treatment responses as those observed clinically. Xenografts representing 12 patients, including two ER+ lines, showed metastasis to the mouse lung. These models thus serve as a renewable, quality-controlled tissue resource for preclinical studies investigating treatment response and metastasis

The SOX11 transcription factor is a critical regulator of basal-like breast cancer growth, invasion, and basal-like gene expression.

Basal-like breast cancers (BLBCs) are aggressive breast cancers associated with poor survival. Defining the key drivers of BLBC growth will allow identification of molecules for targeted therapy. In this study, we performed a primary screen integrating multiple assays that compare transcription factor expression and activity in BLBC and non-BLBC at the RNA, DNA, and protein levels. This integrated screen identified 33 transcription factors that were elevated in BLBC in multiple assays comparing mRNA expression, DNA cis-element sequences, or protein DNA-binding activity. In a secondary screen to identify transcription factors critical for BLBC cell growth, 8 of the 33 candidate transcription factors (TFs) were found to be necessary for growth in at least two of three BLBC cell lines. Of these 8 transcription factors, SOX11 was the only transcription factor required for BLBC growth, but not for growth of non-BLBC cells. Our studies demonstrate that SOX11 is a critical regulator of multiple BLBC phenotypes, including growth, migration, invasion, and expression of signature BLBC genes. High SOX11 expression was also found to be an independent prognostic indicator of poor survival in women with breast cancer. These results identify SOX11 as a potential target for the treatment of BLBC, the most aggressive form of breast cancer