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PDF - 68K, Supplementary Materials and Methods.

Selected functional pathways affected by changes in EMP2 expression.

Supplemental Methods

Modulation of EMP2 produces transcriptomic changes in breast cancer.

PDF - 855K, Supplementary Figure 1. Anti- EMP2 IgG1 can detect EMP2 in human tissue. Supplementary Figure 2. The serum levels of the human EMP2 IgG1 was compared to those of Trastuzamab.

PDF - 59K, Known Profile of EMP2 Upregulation in Breast Cancer.

PDF - 51K, Supplementary Table 3. Expression levels of EMP2 in triple negative breast cancer.

PDF - 54K, Supplementary Table 2. Clinical Data for Invasive Breast Cancer Patients used in the TMA.

Despite significant advances in biology and medicine, the incidence and mortality due to breast cancer worldwide is still unacceptably high. Thus, there is an urgent need to discover new molecular targets. In this article, we show evidence for a novel target in human breast cancer, the tetraspan protein epithelial membrane protein-2 (EMP2). Using tissue tumor arrays, protein expression of EMP2 was measured and found to be minimal in normal mammary tissue, but it was upregulated in 63% of invasive breast cancer tumors and in 73% of triple-negative tumors tested. To test the hypothesis that EMP2 may be a suitable target for therapy, we constructed a fully human immunoglobulin G1 (IgG1) antibody specific for a conserved domain of human and murine EMP2. Treatment of breast cancer cells with the anti-EMP2 IgG1 significantly inhibited EMP2-mediated signaling, blocked FAK/Src signaling, inhibited invasion, and promoted apoptosis in vitro. In both human xenograft and syngeneic metastatic tumor monotherapy models, anti-EMP2 IgG1 retarded tumor growth without detectable systemic toxicity. This antitumor effect was, in part, attributable to a potent antibody-dependent cell-mediated cytotoxicity response as well as direct cytotoxicity induced by the monoclonal antibody. Together, these results identify EMP2 as a novel therapeutic target for invasive breast cancer. Mol Cancer Ther; 13(4); 902–15. ©2014 AACR.

PDF - 64K, Supplementary Table 4. Evaluation of EMP2 IgG1 toxicity compared to control IgG following 7 weeks of treatment delivered systemically IP. Supplementary Table 5. Evaluation of EMP2 IgG1 toxicity compared to control IgG following systemic IP treatment of up to 40mg/kg.

Little is known about the role of epithelial membrane protein-2 (EMP2) in breast cancer development or progression. In this study, we tested the hypothesis that EMP2 may regulate the formation or self-renewal of breast cancer stem cells (BCSC) in the tumor microenvironment. In silico analysis of gene expression data demonstrated a correlation of EMP2 expression with known metastasis-related genes and markers of cancer stem cells (CSC) including aldehyde dehydrogenase (ALDH). In breast cancer cell lines, EMP2 overexpression increased and EMP2 knockdown decreased the proportion of stem-like cells as assessed by the expression of the CSC markers CD44+/CD24−, ALDH activity, or by tumor sphere formation. In vivo, upregulation of EMP2 promoted tumor growth, whereas knockdown reduced the ALDHhigh CSC population as well as retarded tumor growth. Mechanistically, EMP2 functionally regulated the response to hypoxia through the upregulation of HIF-1α, a transcription factor previously shown to regulate the self-renewal of ALDHhigh CSCs. Furthermore, in syngeneic mouse models and primary human tumor xenografts, mAbs directed against EMP2 effectively targeted CSCs, reducing the ALDH+ population and blocking their tumor-initiating capacity when implanted into secondary untreated mice. Collectively, our results show that EMP2 increases the proportion of tumor-initiating cells, providing a rationale for the continued development of EMP2-targeting agents.

PDF - 55K, Legends for Supplementary Tables 1 through 5.

Figure S4 Quantification of Western blot analysis (related to Figure 4) (B-G). Data are presented as mean fold increases ({plus minus}SD) in treated groups over basal values from three independent experiments. B, *p

Figure S6 The comparison of expression levels of phospho-ERK and phospho-Smad4 between lean and overweight/obesity patients, Related to Figure 7. Staining was analyzed by using a system based on the percentage of positively stained cells and the staining intensity. Integrated optical density of all the positive staining in each image was determined, and its ratio to total area of each photograph was calculated as density.

Obesity increases the risk of distant metastatic recurrence and reduces breast cancer survival. However, the mechanisms behind this pathology and identification of relevant therapeutic targets are poorly defined. Plasma free fatty acids (FFA) levels are elevated in obese individuals. Here we report that TGFβ transiently activates ERK and subsequently phosphorylates SMAD4 at Thr277, which facilitates a SMAD4–USP9x interaction, SMAD4 nuclear retention, and stimulates TGFβ/SMAD3–mediated transcription of Twist and Snail. USP9x inhibited the E3 ubiquitin-protein ligase TIF1γ from binding and monoubiquitinating SMAD4, hence maintaining the SMAD4 nuclear retention. FFA further facilitated TGFβ-induced ERK activation, SMAD4 phosphorylation, and nuclear retention, promoting TGFβ-dependent cancer progression. Inhibition of ERK and USP9x suppressed obesity-induced metastasis. In addition, clinical data indicated that phospho-ERK and -SMAD4 levels correlate with activated TGFβ signaling and metastasis in overweight/obese patient breast cancer specimens. Altogether, we demonstrate the vital interaction of USP9x and SMAD4 for governing TGFβ signaling and dyslipidemia-induced aberrant TGFβ activation during breast cancer metastasis. Cancer Res; 77(6); 1383–94. ©2017 AACR.

Figure S1 Quantification of Western blot analysis (related to Figure 1) (A-H). Data are presented as mean fold increases ({plus minus}SD) in treated groups over basal values from three independent experiments. *p < 0.01 versus controls; #p < 0.01 versus their corresponding DMSO controls. FFA facilitates TGF-beta-induced ERK activation and SAMD4 nuclear retention, Related to Figure 1. I, Nuclear extracts were made from MDA-MB-231 cells that were treated with 3 ng/ml of TGF-beta for the time points shown. They were analyzed by Western blotting with antibodies against Smad4 or phosphor-ERK and ERK. H3 was used as a loading control for nuclear fractions. We evaluated the cytoplasmic contamination of the nuclear fractions by blotting for α-vinculin. J, Nuclear extracts were made from MDA-MB-231 cells treated with 3 ng/ml of TGF-beta in the presence of palmitate (PA, 400 microM) for the time points shown. Phosphor-ERK and ERK and nuclear SMAD4 were analyzed by Western blotting.

Figure S3 FFA promotes TGF-beta-induced migration by activating ERK and USP9x, Related to Figure 3. A, BT549 cells were plated in Ibidi culture insert dishes (Madison, WI). Cells were cultured in complete medium to 80% confluence, inserts were removed to create a gap and cells were then incubated with BSA, PA, PA plus AZD6244 or WP1130 in the absence or presence of TGF-beta1. Migrating cells were photographed under a phase contrast microscope. B, The percentage of the wound closed was quantified from three independent replicates and is expressed as mean {plus minus} SD. *p

Estrogen signaling is critical in the progression of tumors that bear estrogen receptors. In most patients with breast cancer, inhibitors that block interactions of estrogen with its receptors or suppress the production of endogenous estrogens are important interventions in the clinic. Recent evidence now suggests that estrogen also contributes to the pathogenesis of non–small cell lung cancer (NSCLC). We used a human lung cancer xenograph model system to analyze the effect of aromatase or estradiol on tumor growth. We further examined the level of protein expression of aromatase in 422 patients with NSCLC using a high-density tissue microarray. Results were confirmed and validated on an independent patient cohort (n = 337). Lower levels of aromatase predicted a greater chance of survival in women 65 years and older. Within this population, the prognostic value of aromatase was greatest in earlier stage lung cancer (stage I/II). In addition, for women with no history of smoking, lower aromatase levels were a strong predictor of survival. Our findings implicate aromatase as an early-stage predictor of survival in some women with NSCLC. We predict that women whose lung cancers have higher levels of aromatase might be good candidates for targeted treatment with aromatase inhibitors. [Cancer Res 2007;67(21):10484–90]

Supplementary Tables 1-3, Figures 1-2 from Aromatase Expression Predicts Survival in Women with Early-Stage Non–Small Cell Lung Cancer

Skene’s glands are a pair of female paraurethral glands with similar embryologic, physiologic, and pathologic features to the male prostate. Cysts and abscesses represent the most common pathological changes affecting Skene’s gland, whereas adenofibromas represent an extremely rare entity. Herein, we present a rare case report of adenofibroma in the Skene’s gland of a postmenopausal female.

In type 1 endometrial cancer, unopposed estrogen stimulation is thought to lead to endometrial hyperplasia which precedes malignant progression. Recent data from our group and others suggest that ALDH activity mediates stemness in endometrial cancer, but while aldehyde dehydrogenase 1 (ALDH1) has been suggested as a putative cancer stem cell marker in several cancer types, its clinical and prognostic value in endometrial cancer remains debated. The aim of this study was to investigate the clinical value of ALDH1 expression in endometrial hyperplasia and to determine its ability to predict progression to endometrial cancer. Interrogation of the TCGA database revealed upregulation of several isoforms in endometrial cancer, of which the ALDH1 isoforms collectively constituted the largest group. To translate its expression, a tissue microarray was previously constructed which contained a wide sampling of benign and malignant endometrial samples. The array contained a metachronous cohort of samples from individuals who either developed or did not develop endometrial cancer. Immunohistochemical staining was used to determine the intensity and frequency of ALDH1 expression. While benign proliferative and secretory endometrium showed very low levels of ALDH1, slightly higher expression was observed within the stratum basalis. In disease progression, cytoplasmic ALDH1 expression showed a step-wise increase between endometrial hyperplasia, atypical hyperplasia, and endometrial cancer. ALDH1 was also shown to be an early predictor of EC development, suggesting that it can serve as an independent prognostic indicator of patients with endometrial hyperplasia with or without atypia who would progress to cancer (p = 0.012).

Non-Hodgkin lymphoma represents a heterogeneous group of tumors that constitute the seventh most common malignancy. Immunohistochemistry plays a major role in the detection of specific cell receptors. Transcription factors are a heterogeneous group of genes that play a critical role in the commitment, differentiation, and proliferation of specific cell types.Paraffin-embedded tissue sections of non-Hodgkin lymphoma cases were selected, classified, and evaluated before staining with immunohistochemical markers (PAX5, OCT2, BCL6, and P53). Expression of the aforementioned markers was compared with histological subtypes and grades of lymphoma cases. Means of expression were also compared among histological subtypes.A total of 55 cases of NHL including 26 cases of low-grade lymphomas and 29 cases of high-grade lymphomas were included in the study. DLBCL and FL were the most common subtypes of high-grade and low-grade lymphomas respectively. Both PAX5 and OCT2 were positive in 44 cases of NHL (80%) including all cases of B-cell lymphomas. BCL6 and P53 demonstrated positive expression in 29.1% and 67.3% respectively. Interestingly, we found a significant association between the histological subtypes and the aforementioned markers (P-value0.05).Expression of PAX5, OCT2, BCL, and P53 played a major role in the diagnosis and grading of non-Hodgkin lymphomas in our study. Both PAX5 and OCT2 provided more accuracy and specificity in the diagnosis of B-cell neoplasms compared to the classical B-cell markers. BCL6 expression reflected its role in germinal center formation in normal and malignant lymphoid tissues, and expression of P53 mirrored the accumulation of gene mutations in more aggressive lymphoma subtypes.In this manuscript, we aimed to present a unique study that highlights the immunohistochemical expression of all the aforementioned factors among various histological subtypes of non-Hodgkin lymphomas with disparities in histological aggressiveness, highlighting a promising diagnostic and prognostic panel for non-Hodgkin lymphomas.

The abnormal PI3K/AKT/mTOR pathway is one of the most common genomic abnormalities in breast cancers including triple-negative breast cancer (TNBC), and pharmacologic inhibition of these aberrations has shown activity in TNBC patients. Here, we designed and identified a small-molecule Comp34 that suppresses both AKT and mTOR protein expression and exhibits robust cytotoxicity towards TNBC cells but not nontumorigenic normal breast epithelial cells. Mechanically, long noncoding RNA (lncRNA) AL354740.1-204 (also named as NUDT3-AS4) acts as a microRNA sponge to compete with AKT1/mTOR mRNAs for binding to miR-99s, leading to decrease in degradation of AKT1/mTOR mRNAs and subsequent increase in AKT1/mTOR protein expression. Inhibition of lncRNA-NUDT3-AS4 and suppression of the NUDT3-AS4/miR-99s association contribute to Comp34-affected biologic pathways. In addition, Comp34 alone is effective in cells with secondary resistance to rapamycin, the best-known inhibitor of mTOR, and displays a greater in vivo antitumor efficacy and lower toxicity than rapamycin in TNBC xenografted models. In conclusion, NUDT3-AS4 may play a proproliferative role in TNBC and be considered a relevant therapeutic target, and Comp34 presents promising activity as a single agent to inhibit TNBC through regulation of NUDT3-AS4 and miR-99s.

Background and aims: Insulin-like Growth Factors (IGF-1) plays as mediator between metabolic syndrome (MetS), oxidative stress and breast cancer (BrCa) progression. The objective of this study was to examine the relationships between IGF-1 serum levels and metabolic profile biomarkers in a population group of BrCa patients. Material and methods: 126 Syrian women with breast lesions were assigned in 3 study groups: I. Malignant breast tumor group, II. Benign breast tumor group and a Normal (control) group. The following biochemical parameters were measured: IGF-1, HDL-cholesterol, LDL-cholesterol, triglycerides (TG) and glucose. Results: The mean levels of serum IGF- 1 in patients with breast cancer was significantly higher than those with benign tumors but we did not find any correlation between IGF-1 serum levels and tumor stage or lymph nodes metastases. Total cholesterol and LDL-cholesterol levels, along with TG were significantly higher in patients with BrCa versus benign and normal subjects. Conclusion: Results support the link of metabolic dysregulation and oxidative stress in BrCa progression as elevation of serum IGF-1 levels in BrCa patients are associated with metabolic syndrome markers which eventually adds more risk in cancer progression.

Triple Negative breast cancer (TNBC) is a subtype of breast cancer that lacks the expression of estrogen receptor (ER), progesterone receptor, and human epidermal growth factor receptor 2. TNBC accounts for 15-20% of all breast cancer cases but accounts for over 50% of mortality. We propose that Estrogen receptor-beta (ERβ) and IGF2 play a significant role in the pathogenesis of TNBCs, and could be important targets for future therapy. Tissue microarrays (TMAs) from over 250 TNBC patients' were analyzed for ERβ and IGF2 expression by immunohistochemistry. Expression was correlated with clinical outcomes. In addition, TNBC cell lines Caucasians (CA): MB-231/BT549 and African Americans (AAs): MB-468/HCC70/HCC1806 were used to investigate the effect of hormonal and growth factor regulation on cell proliferation. TMAs from AAs had higher expression of ERβ and IGF2 expression when compared to CA. ERβ and IGF2 were found to be upregulated in our TNBC cell lines when compared to other cell types. TNBC cells treated with ERβ agonist displayed significant increase in cell proliferation and migration when compared to controls. AA tissue samples from TNBC patients had higher expression of ERβ. African-American breast cancer TNBC tissue samples from TNBC patients have higher expression of ERβ. In addition, TNBC cell lines were also found to express high levels of ERβ. IGF2 increased transcription of ERβ in TNBC cells. Understanding the mechanisms of IGF2/ERβ axis in TNBC tumors could provide an opportunity to target this aggressive subtype of breast cancer.

Aims: Epidemiologic evidence indicates that diabetes may increase risk of breast cancer (BC) and mortality in patients with cancer. The pathophysiological relationships between diabetes and cancer are not fully understood, and personalized treatments for diabetes-associated BC are urgently needed. Results: We observed that high glucose (HG), via activation of nuclear phosphatase PP2Cδ, suppresses p53 function, and consequently promotes BC cell proliferation, migration, and invasion. PP2Cδ expression is higher in tumor tissues from BC patients with hyperglycemia than those with normoglycemia. The mechanisms underlying HG stimulation of PP2Cδ involve classical/novel protein kinase-C (PKC) activation and GSK3β phosphorylation. Reactive oxygen species (ROS)/NF-κB pathway also mediates HG induction of PP2Cδ. Furthermore, we identified a 1,5-diheteroarylpenta-1,4-dien-3-one (Compound 23, or C23) as a novel potent PP2Cδ inhibitor with a striking cytotoxicity on MCF-7 cells through cell-based screening assay for growth inhibition and activity of a group of curcumin mimics. Beside directly inhibiting PP2Cδ activity, C23 blocks HG induction of PP2Cδ expression via heat shock protein 27 (HSP27) induction and subsequent ablation of ROS/NF-κB activation. C23 can thus significantly block HG-triggered inhibition of p53 activity, leading to the inhibition of cancer cell proliferation, migration, and invasion. In addition, hyperglycemia promotes BC development in diabetic nude mice, and C23 inhibits the xenografted BC tumor growth. Conclusions and Innovation: Our findings elucidate mechanisms that may have contributed to diabetes-associated BC progression, and provide the first evidence to support the possible alternative therapeutic approach to BC patients with diabetes. Antioxid. Redox Signal. 30, 1983-1998.

This study investigated the inhibitory effect of arctigenin, a novel anti-inflammatory lignan, on prostate cancer in obese conditions both in vitro and in vivo. In vitro obese models were established by co-culture of mouse adipocytes 3T3-L1 with androgen-sensitive LNCaP human prostate cancer cells, or by culturing LNCaP cells in adipocytes-conditioned medium. Arctigenin significantly inhibited LNCaP proliferation, along with decreased androgen receptor (AR) and increased Nkx3.1 cellular expression. Male severe combined immunodeficiency mice were subcutaneously implanted with human prostate cancer LAPC-4 xenograft tumors for in vivo study. Mice were fed high-fat (HF) diet and orally given arctigenin at 50 mg/kg body weight daily or vehicle control for 6 weeks. Tumor bearing HF control mice showed a significant increase in serum free fatty acids (FFAs) and decrease in subcutaneous/peritoneal fat depots compared to non-tumor bearing control mice. Arctigenin intervention significantly reduced tumor growth by 45%, associated with decreased circulating FFAs and adipokines/cytokines including IGF-1, VEGF, and MCP-1, along with decreased AR, Ki67, and microvessel density and increased Nkx3.1 expression in tumors. These results indicate the strong ability of arctigenin to co-target obesity and tumor itself in inhibition of prostate tumor growth at a lower concentration compared to most phytochemicals.

Aberrant PP2Cδ inhibits p300-mediated p53 acetylation and activation via the ATM/BRCA1 pathway, leading to cancer progression., Although nuclear type 2C protein phosphatase (PP2Cδ) has been demonstrated to be pro-oncogenic with an important role in tumorigenesis, the underlying mechanisms that link aberrant PP2Cδ levels with cancer development remain elusive. Here, we found that aberrant PP2Cδ activity decreases p53 acetylation and its transcriptional activity and suppresses doxorubicin-induced cell apoptosis. Mechanistically, we show that BRCA1 facilitates p300-mediated p53 acetylation by complexing with these two proteins and that S1423/1524 phosphorylation is indispensable for this regulatory process. PP2Cδ, via dephosphorylation of ATM, suppresses DNA damage–induced BRCA1 phosphorylation, leading to inhibition of p300-mediated p53 acetylation. Furthermore, PP2Cδ levels correlate with histological grade and are inversely associated with BRCA1 phosphorylation and p53 acetylation in breast cancer specimens. C23, our newly developed PP2Cδ inhibitor, promotes the anticancer effect of doxorubicin in MCF-7 xenograft–bearing nude mice. Together, our data indicate that PP2Cδ impairs p53 acetylation and DNA damage response by compromising BRCA1 function.

Background Precision medicine and prediction of therapeutic response requires monitoring potential biomarkers before and after treatment. Liquid biopsies provide noninvasive prognostic markers such as circulating tumor DNA and RNA. Circulating tumor RNA (ctRNA) in blood is also used to identify mutations in genes of interest, but additionally, provides information about relative expression levels of important genes. In this study, we analyzed PD-L1 expression in ctRNA isolated from various cancer types. Tumors inhibit antitumor response by modulating the immune checkpoint proteins programmed death ligand 1 (PD-L1) and its cognate receptor PD1. The expression of these genes has been implicated in evasion of immune response and resistance to targeted therapies. Methods Blood samples were collected from gastric (GC), colorectal (CRC), lung (NSCLC), breast (BC), prostate cancer (PC) patients, and a healthy control group. ctRNA was purified from fractionated plasma, and following reverse transcription, levels of PD-L1 expression were analyzed using qPCR. Results PD-L1 expression was detected in the plasma ctRNA of all cancer types at varying frequencies but no PD-L1 mRNA was detected in cancer-free individuals. The frequencies of PD-L1 expression were significantly different among the various cancer types but the median relative PD-L1 expression values were not significantly different. In 12 cases where plasma and tumor tissue were available from the same patients, there was a high degree of concordance between expression of PD-L1 protein in tumor tissues and PD-L1 gene expression in plasma, and both methods were equally predictive of response to nivolumab. Conclusions PD-L1 mRNA can be detected and quantitated in ctRNA of cancer patients. These results pave the way for further studies aimed at determining whether monitoring the levels of PD-L1 mRNA in blood can identify patients who are most likely to benefit from the conventional treatment.

Triple-negative breast cancer (TNBC) affects only about 10-15% of women with breast cancer (BC) but accounts for almost 50% of all BC deaths. At this time, development of targeted treatments for TNBC are urgently needed. Insulin-like growth factor-2 (IGF2) is highly expressed in TNBCs and mediates its proliferative effects via two hormone receptors, insulin-like growth factor -1 receptor (IGF1R) and insulin receptor (IR). Using a TNBC tissue microarray (TMA) consisting of 159 archival clinical samples, we confirm that IGF2 is highly expressed in tumor tissue as compared to nearby normal tissue (P Citation Format: Nalo M. Hamilton, Diana Marquez-Garban, David Austin, Neda Moatamed, Lorena P. Burton, David Elashoff, Begonya Comin-Anduix, Tristan Grogan, Madhuri Wadehra, Helena Chang, Adrienne Martinez, Mary-Lynn Brecht, Isabel Jabara, Richard Pietras, Yahya Elshimali, Jaydutt Vadgama, Michael Jung. Insulin-like growth factor-2 a potential target for screening and treatment in patients with triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 943.

Cancer Drug resistance is a medical concern that requires extensive research and a thorough understanding in order to overcome. Remarkable achievements related to this field have been accomplished and further work is needed in order to optimize the cure for cancer and serve as the basis for precise medicine with few or no side effects.

Insulin and leptin have an overlapping anorexigenic action as well as opposite effects on glucose and lipid metabolism. The study focuses on the biochemical and clinical relevance of new indices of insulin-leptin axis utilized in the study of the relationships between leptinemia, insulin sensitivity and oxidative stress, in elderly subjects with metabolic syndrome. We conducted clinical studies on elderly people with metabolic syndrome versus control subjects by creating new insulin-adipogenic indices, namely Insulin-to-Leptin Ratio (ILR) and Insulin-Adipogenic Resistance index (IAR-index). Inflammation and oxidative stress biomarkers evaluated were the high-sensitivity C-reactive protein (hsCRP), the advanced oxidation protein products (AOPP), and the serum antioxidant capacity measured as ferric reducing antioxidant potential (FRAP). The metabolic syndrome group showed significantly (p

Hormone therapy targeting estrogen receptor α (ERα) is the most effective treatment for breast cancer. However, this treatment eventually fails as the tumor develops resistance. Although reduced expression of ER-α is a known contributing factor to endocrine resistance, the mechanism of ER-α downregulation in endocrine resistance is still not fully understood. The present study shows that Slug has an inverse relationship with ERα in breast and prostate cancer patient samples. Also the inhibition of Slug blocks mammary stem cell activity in primary mammary epithelial cells. We hypothesize that Slug may be a key transcription factor in the regulation of ERα expression. To understand the Slug-ERα signaling pathway, we employed resistant cell line MCF-TAMR (ERα relatively negative) derived from its parental MCF-7 (ERα positive) cell line and assessed changes in cell phenotype, activity and response to therapy. Conversely, we performed knockdown of Slug in the high-Slug expressing cell line MDA-MB-231 and assessed reversal of the mesenchymal phenotype. Microarray analysis showed that Slug is overexpressed in high grade breast and prostate cancer tissues. Additionally, Slug overexpression leads to drug resistance. Furthermore, we demonstrated that Slug binds directly to ERα promoter E-boxes and represses ERα expression. This resulted in decrease in epithelial-to-mesenchymal transition in cancer cells. These findings demonstrate that Slug, by regulation of ERα expression, contributes to tumor progression and could serve as an important target for cancer therapy.

Triple-negative breast cancers (TNBCs) lack estrogen receptor-α (ERα), progesterone receptor (PR), and human epidermal growth factor receptor-2 (HER2) amplification and account for almost half of all breast cancer deaths. This breast cancer subtype largely affects women who are premenopausal, African-American, or have BRCA1/2 mutations. Women with TNBC are plagued with higher rates of distant metastasis that significantly diminish their overall survival and quality of life. Due to their poor response to chemotherapy, patients with TNBC would significantly benefit from development of new targeted therapeutics. Research suggests that the insulin-like growth factor (IGF) family and estrogen receptor beta-1 (ERβ1), due to their roles in metabolism and cellular regulation, might be attractive targets to pursue for TNBC management. Here, we review the current state of the science addressing the roles of ERβ1 and the IGF family in TNBC. Further, the potential benefit of metformin treatment in patients with TNBC as well as areas of therapeutic potential in the IGF-ERβ1 pathway are highlighted.

Obesity increases the risk of distant metastatic recurrence and reduces breast cancer survival. However, the mechanisms behind this pathology and identification of relevant therapeutic targets are poorly defined. Plasma free fatty acids (FFA) levels are elevated in obese individuals. Here we report that TGFβ transiently activates ERK and subsequently phosphorylates SMAD4 at Thr277, which facilitates a SMAD4–USP9x interaction, SMAD4 nuclear retention, and stimulates TGFβ/SMAD3–mediated transcription of Twist and Snail. USP9x inhibited the E3 ubiquitin-protein ligase TIF1γ from binding and monoubiquitinating SMAD4, hence maintaining the SMAD4 nuclear retention. FFA further facilitated TGFβ-induced ERK activation, SMAD4 phosphorylation, and nuclear retention, promoting TGFβ-dependent cancer progression. Inhibition of ERK and USP9x suppressed obesity-induced metastasis. In addition, clinical data indicated that phospho-ERK and -SMAD4 levels correlate with activated TGFβ signaling and metastasis in overweight/obese patient breast cancer specimens. Altogether, we demonstrate the vital interaction of USP9x and SMAD4 for governing TGFβ signaling and dyslipidemia-induced aberrant TGFβ activation during breast cancer metastasis. Cancer Res; 77(6); 1383–94. ©2017 AACR.

Background Chemotherapy with docetaxel (Doc) remains the standard treatment for metastatic and castration-resistance prostate cancer (CRPC). However, the clinical success of Doc is limited by its chemoresistance and side effects. This study investigated whether natural products green tea (GT) and quercetin (Q) enhance the therapeutic efficacy of Doc in CRPC in mouse models. Methods Male severe combined immunodeficiency (SCID) mice (n = 10 per group) were inoculated with androgen-independent prostate cancer PC-3 cells subcutaneously. When tumors were established the intervention started. Mice were administered with GT + Q, Doc 5 mg/kg (LD), GT + Q + LD Doc, Doc 10 mg/kg (HD) or control. The concentration of GT polyphenols in brewed tea administered as drinking water was 0.07 % and Q was supplemented in diet at 0.4 %. Doc was intravenously injected weekly for 4 weeks, GT and Q given throughout the study. Results GT + Q or LD Doc slightly inhibited tumor growth compared to control. However, the combination of GT and Q with LD Doc significantly enhanced the potency of Doc 2-fold and reduced tumor growth by 62 % compared to LD Doc in 7-weeks intervention. A decrease of Ki67 and increase of cleaved caspase 7 were observed in tumors by the mixture, along with lowered blood concentrations of growth factors like VEGF and EGF. The mixture significantly elevated the levels of tumor suppressor mir15a and mir330 in tumor tissues. An increased risk of liver toxicity was only observed with HD Doc treatment. Conclusions These results provide a promising regimen to enhance the therapeutic effect of Doc in a less toxic manner.

Increasing body of evidence suggests that there exists a connection between diabetes and cancer. Nevertheless, to date, the potential reasons for this association are still poorly understood and currently there is no clinical evidence available to direct the proper management of patients presenting with these two diseases concomitantly. Both cancer and diabetes have been associated with abnormal lactate metabolism and high level of lactate production is the key biological property of these diseases. Conversely, high lactate contribute to a higher insulin resistant status and a more malignant phenotype of cancer cells, promoting diabetes and cancer development and progression. In view of associations between diabetes and cancers, the role of high lactate production in diabetes and cancer interaction should not be neglected. Here, we review the available evidence of lactate’s role in different biological characteristics of diabetes and cancer and interactive relationship between them. Understanding the molecular mechanisms behind metabolic remodeling of diabetes- and cancer-related signaling would endow novel preventive and therapeutic approaches for diabetes and cancer treatment.

Epidemiologic evidence suggests that women with diabetes have increased risk of breast cancer. Moreover, diabetes may dramatically increase mortality in patients with cancer, and there exists some important distinction between the breast cancer patients with and without diabetes in the regimen selection and outcomes of cancer therapy. There is an urgent need for more effective and specific clinical treatment for diabetes-associated breast cancer. Our previous studies have suggested that hyperglycemia, via PKC/GSK3β and ROS/NF-κB pathways that are involved in PP2Cδ activation, suppresses the tumor-suppressor p53 function and inhibits DNA damage-induced apoptosis, inducing proliferation in the epithelium and the development of breast cancer. HG inhibition of p53 activity and induction of cancer cell proliferation, migration, and invasion were significantly blocked by CCT007093, a known PP2Cδ inhibitor. The structure similarity between a group of curcumin mimics (compounds 1-33) and CCT007093 spurred us to investigate the inhibitory ability of compounds 1-33 on PP2Cδ. The growth inhibition assay screening identified that compound 23 (C19H26N4O) has a striking cytotoxicity on MCF-7 cells instead of MCF-12A cells versus CCT007093. More interestingly, the impressive cytotoxicity of 23 is fully mirrored by their much stronger inhibitory ability on PP2Cδ in cells (as assessed by phospho-P38 measurement) than CCT007093. Docking simulation for binding modes of these inhibitors to PP2Cδ phosphatase domain suggests that the carbonyl oxygen and the nitrogen on the heteroaromatic ring of compound 23 form two hydrogen bonds with side chains of Arg258 and Asp105, respectively. This leads to a stable binding model of compound 23, which is inconsistent with its stronger inhibition on PP2Cδ. Furthermore, compound 23 dramatically induces heat shock protein 27 (Hsp27), ablating HG-induced ROS production and NF-κB activation, which partially blocked HG induction of PP2Cδ protein expression. Thus, compound 23 significantly blocked HG inhibition of p53 activity and induction of cancer cell proliferation, migration, and invasion. Finally, using streptozotocin (STZ)-induced diabetic nude mice bearing MCF-7 cells, we demonstrate that hyperglycemia promoted the development of breast cancer in vivo and that compound 23 significantly inhibited xenografted breast cancer growth in mice. Taken together, these results suggest that the identified novel PP2Cδ inhibitor compound 23 inhibits HG-induced breast cancer progression in vivo through regulating expression and activity of PP2Cδ. These findings provide the first evidence proposing an alternative approach for the therapy of breast cancer patients with diabetes. Citation Format: Ke Wu, Xiaoting Yu, Xianghua Yi, Zhimin Huang, Ying-Li Wu, Yahya Elshimali, Yanjun Liu, Donghui Zhu, Shilong Zheng, Qiao-Hong Chen, Guangdi Wang, Yong Wu, Jaydutt V. Vadgama. Compound 23 inhibits high glucose-induced breast cancer progression in vivo through regulating activity and expression of PP2Cδ [abstract]. In: Proceedings of the AACR Special Conference: Advances in Breast Cancer Research; 2017 Oct 7-10; Hollywood, CA. Philadelphia (PA): AACR; Mol Cancer Res 2018;16(8_Suppl):Abstract nr B19.

Breast cancer has a high incidence worldwide. African-American and Hispanic/Latina women have higher mortality from breast cancer than other ethnic groups. Epidemiologic evidence suggests that women with diabetes have increased risk of breast cancer. Diabetes and cancer share many risk factors, but the pathophysiologic relationship between the two diseases is incompletely understood in detail. We observed that exposure of cultured transformed (MCF-7) and normal (MCF-12A) breast epithelial cells to clinically relevant levels of glucose (HG, 22 mM) dramatically suppresses the tumor suppressor p53 acetylation, and, consequently, additively promotes tumor cell proliferation, migration, and invasion. Importantly, we found that activation of nuclear phosphatase PP2Cδ (Ppm1d, WIP1) plays a role in the enhancing effects of HG on aggressive phenotypes of these cells. The mechanisms underlying high-glucose stimulation of PP2Cδ involve classical/novel PKCs activation and its downstream target protein GSK3β phosphorylation and inactivation. In addition, HG-induced reactive oxygen species (ROS) generation and subsequent NF-κB activation play a partial role in HG induction of PP2Cδ. HG inhibition of p53 activity and DNA damage-induced apoptosis, as well as induction of cancer cell proliferation, migration, and invasion, were significantly blocked by CCT007093, a known PP2Cδ inhibitor. We conclude that hyperglycemia, via PKC/GSK3β and ROS/NF-κB pathways that are involved in PP2Cδ activation, suppresses the tumor suppressor p53 function and inhibits DNA damage-induced apoptosis, inducing proliferation in the epithelium and the development of breast cancer. Citation Format: Ke Wu, Xiaoting Yu, Xianghua Yi, Lynn Ma, Yahya Elshimali, Yanjun Liu, Donghui Zhu, Yong Wu, Jaydutt V. Vadgama. High glucose induces breast cancer progression through upregulating PP2Cδ [abstract]. In: Proceedings of the Tenth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2017 Sep 25-28; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2018;27(7 Suppl):Abstract nr B76.

The protein 14-3-3σ is involved in the regulation of cellular processes such as apoptosis, cell cycle progression and proliferation. Disruption of protein expression has been implicated in a number of malignancies. Here we examine the expression pattern of 14-3-3σ in breast cancer and specifically consider whether expression in ductal carcinoma in situ (DCIS) lesions is predictive of disease outcome. We examined 14-3-3σ protein expression and localization using immunohistochemical staining on a high-density tissue microarray consisting of 157 invasive breast cancer patients. Statistical analyses were used to assess the correlation of 14-3-3σ expression with clinico-pathological parameters and patient outcome. We observed a statistically significant increase in 14-3-3σ protein expression in ductal hyperplasia, DCIS, and invasive ductal carcinoma (IDC) as compared normal glandular epithelium. In IDC, lower expression of 14-3-3σ tended to predicted poorer survival time while in DCIS lesions, there was a stronger correlation between relatively higher levels of 14-3-3σ predicting shorter survival time. Further, of patients who had concurrent DCIS and IDC lesions, those that exhibited a decrease of 14-3-3σ expression from DCIS to IDC had significantly shorter survival time. Our findings indicate that 14-3-3σ expression may be a useful prognostic indicator for survival in patients with breast cancer with an elevated 14-3-3σ in earlier disease predicting a less favorable disease outcome. To our knowledge this is the first published study associating 14-3-3σ protein expression with breast cancer survival.

Estrogen signaling is critical in the progression of tumors that bear estrogen receptors. In most patients with breast cancer, inhibitors that block interactions of estrogen with its receptors or suppress the production of endogenous estrogens are important interventions in the clinic. Recent evidence now suggests that estrogen also contributes to the pathogenesis of non–small cell lung cancer (NSCLC). We used a human lung cancer xenograph model system to analyze the effect of aromatase or estradiol on tumor growth. We further examined the level of protein expression of aromatase in 422 patients with NSCLC using a high-density tissue microarray. Results were confirmed and validated on an independent patient cohort (n = 337). Lower levels of aromatase predicted a greater chance of survival in women 65 years and older. Within this population, the prognostic value of aromatase was greatest in earlier stage lung cancer (stage I/II). In addition, for women with no history of smoking, lower aromatase levels were a strong predictor of survival. Our findings implicate aromatase as an early-stage predictor of survival in some women with NSCLC. We predict that women whose lung cancers have higher levels of aromatase might be good candidates for targeted treatment with aromatase inhibitors. [Cancer Res 2007;67(21):10484–90]