Selective pressure of endocrine therapy activates the integrated stress response through NFκB signaling in a subpopulation of ER positive breast cancer cells.

BACKGROUND: While estrogen receptor (ER) positive breast tumors generally respond well to endocrine therapy (ET), up to 40% of patients will experience relapse, either while on endocrine therapy or after ET is completed. We previously demonstrated that the selective pressure of tamoxifen activates the NFκB pathway in ER + patient tumors, breast cancer cell lines, and breast cancer xenograft tumors, and that this activation allows for survival of a subpopulation of NFκB + cells that contribute to cell regrowth and tumor relapse after ET withdrawal. However, the mechanisms contributing to the expansion of an NFκB + cell population on ET are unknown. METHODS: Here, we utilized single-cell RNA sequencing and bioinformatics approaches to characterize the NFκB + cell population and its clinical relevance. Follow-up studies were conducted to validate our findings and assess the function of the integrated stress response pathway in breast cancer cell lines and patient-derived models. RESULTS: We found that the NFκB + population that arises in response to ET is a preexisting population is enriched under the selective pressure of ET. Based on the preexisting NFκB + cell population, we developed a gene signature and found that it is predictive of tumor relapse when expressed in primary ER + tumors and is retained in metastatic cell populations. Moreover, we identified that the integrated stress response (ISR), as indicated by increased phosphorylation of eIF2α, occurs in response to ET and contributes to clonogenic growth under the selective pressure of ET. CONCLUSIONS: Taken together, our findings suggest that a cell population with active NFκB and ISR signaling can survive and expand under the selective pressure of ET and that targeting this population may be a viable therapeutic strategy to improve patient outcome by eliminating cells that survive ET. Understanding the mechanisms by which breast cancer cells survive the selective pressure of ET may improve relapse rates and overall outcome for patients with ER + breast tumors.

Emerging Roles of Ceramides in Breast Cancer Biology and Therapy.

One of the classic hallmarks of cancer is the imbalance between elevated cell proliferation and reduced cell death. Ceramide, a bioactive sphingolipid that can regulate this balance, has long been implicated in cancer. While the effects of ceramide on cell death and therapeutic efficacy are well established, emerging evidence indicates that ceramide turnover to downstream sphingolipids, such as sphingomyelin, hexosylceramides, sphingosine-1-phosphate, and ceramide-1-phosphate, is equally important in driving pro-tumorigenic phenotypes, such as proliferation, survival, migration, stemness, and therapy resistance. The complex and dynamic sphingolipid network has been extensively studied in several cancers, including breast cancer, to find key sphingolipidomic alterations that can be exploited to develop new therapeutic strategies to improve patient outcomes. Here, we review how the current literature shapes our understanding of how ceramide synthesis and turnover are altered in breast cancer and how these changes offer potential strategies to improve breast cancer therapy.

Endocrine Therapy-Resistant Breast Cancer Cells Are More Sensitive to Ceramide Kinase Inhibition and Elevated Ceramide Levels Than Therapy-Sensitive Breast Cancer Cells.

ET resistance is a critical problem for estrogen receptor-positive (ER+) breast cancer. In this study, we have investigated how alterations in sphingolipids promote cell survival in ET-resistant breast cancer. We have performed LC-MS-based targeted sphingolipidomics of tamoxifen-sensitive and -resistant MCF-7 breast cancer cell lines. Follow-up studies included treatments of cell lines and patient-derived xenograft organoids (PDxO) with small molecule inhibitors; cytometric analyses to measure cell death, proliferation, and apoptosis; siRNA-mediated knockdown; RT-qPCR and Western blot for gene and protein expression; targeted lipid analysis; and lipid addback experiments. We found that tamoxifen-resistant cells have lower levels of ceramides and hexosylceramides compared to their tamoxifen-sensitive counterpart. Upon perturbing the sphingolipid pathway with small molecule inhibitors of key enzymes, we identified that CERK is essential for tamoxifen-resistant breast cancer cell survival, as well as a fulvestrant-resistant PDxO. CERK inhibition induces ceramide-mediated cell death in tamoxifen-resistant cells. Ceramide-1-phosphate (C1P) partially reverses CERK inhibition-induced cell death in tamoxifen-resistant cells, likely through lowering endogenous ceramide levels. Our findings suggest that ET-resistant breast cancer cells maintain lower ceramide levels as an essential pro-survival mechanism. Consequently, ET-resistant breast cancer models have a unique dependence on CERK as its activity can inhibit de novo ceramide production.

A Review of Principal Studies on the Development and Treatment of Epithelial Ovarian Cancer in the Laying Hen Gallus gallus.

Often referred to as the silent killer, ovarian cancer is the most lethal gynecologic malignancy. This disease rarely shows any physical symptoms until late stages and no known biomarkers are available for early detection. Because ovarian cancer is rarely detected early, the physiology behind the initiation, progression, treatment, and prevention of this disease remains largely unclear. Over the past 2 decades, the laying hen has emerged as a model that naturally develops epithelial ovarian cancer that is both pathologically and histologically similar to that of the human form of the disease. Different molecular signatures found in human ovarian cancer have also been identified in chicken ovarian cancer including increased CA125 and elevated E-cadherin expression, among others. Chemoprevention studies conducted in this model have shown that decreased ovulation and inflammation are associated with decreased incidence of ovarian cancer development. The purpose of this article is to review the major studies performed in laying hen model of ovarian cancer and discuss how these studies shape our current understanding of the pathophysiology, prevention, and treatment of epithelial ovarian cancer.

The pro-apoptotic actions of 2-methoxyestradiol against ovarian cancer involve catalytic activation of PKCδ signaling.

BACKGROUND: 2-methoxyestradiol (2MeOE2) is a natural metabolite of estradiol, which is generated by the action of CYP1A1 enzyme in the liver. We have previously shown that a flaxseed-supplemented diet decreases both the incidence and severity of ovarian cancer in laying hens, also induces CYP1A1 expression in liver. Recently, we have shown that as a biologically derived active component of flax diet, 2MeOE2 induces apoptosis in ovarian cancer cells which is partially dependent on p38 MAPK. The objective of this study was to elucidate the molecular mechanism of actions of 2MeOE2, a known microtubule disrupting agent, in inducing apoptosis in ovarian tumors. RESULTS: 2MeOE2 induces γH2Ax expression and apoptotic histone modifications in ovarian cancer cells, which are predicted downstream targets of protein kinase Cδ (PKCδ) during apoptosis. Overexpressing full length PKCδ alone does not induce apoptosis but potentiates 2MeOE2-mediated apoptosis. C3-domain mutated dominant-negative PKCδ (PKCδDN) significantly reduces 2MeOE2-induced caspase-3 cleavage and apoptotic histone modification. Silencing PKCδ diminishes 2MeOE2-mediated apoptosis. The catalytic fragment of PKCδ (PKCδCAT) evokes pro-apoptotic effects which are principally dependent on p38 MAPK phosphorylation. CONCLUSIONS: The pro-apoptotic actions of 2MeOE2 are in part dependent on catalytic activation of PKCδ. Catalytic activation of PKCδ accelerates the 2MeOE2-induced apoptotic cascade. This study describes a novel molecular action of flaxseed diet in ovarian cancer.

Pro-apoptotic and anti-angiogenic actions of 2-methoxyestradiol and docosahexaenoic acid, the biologically derived active compounds from flaxseed diet, in preventing ovarian cancer.

BACKGROUND: We have previously shown that a whole flaxseed supplemented diet decreased the onset and severity of ovarian cancer in the laying hen, the only known animal model of spontaneous ovarian cancer. Flaxseed is rich in omega-3 fatty acids (OM3FA), mostly α-Linoleic acid (ALA), which gets converted to Docosahexaenoic acid (DHA) by the action of delta-6 desaturase enzyme. Ingestion of flaxseed also causes an increase in production of 2-methoxyestradiol (2MeOE2) via the induction of the CYP1A1 pathway of estrogen metabolism. We have previously reported that the flaxseed diet induces apoptosis via p38-MAPK pathway in chicken tumors. The objective of this study was to investigate the effect of the flaxseed diet on ovarian cancer in chickens, focusing on two hallmarks of cancer, apoptosis and angiogenesis. RESULTS: The anti-cancer effects of two active biologically derived compounds of flax diet, 2MeOE2 and DHA, were individually tested on human ovarian cancer cells and in vivo by the Chick Chorioallantoic Membrane (CAM) assay. Our results indicate that a flaxseed-supplemented diet promotes apoptosis and inhibits angiogenesis in chicken tumors but not in normal ovaries. 2MeOE2 promotes apoptosis in human ovarian cancer cells, inhibits angiogenesis on CAM and its actions are dependent on the p38-MAPK pathway. DHA does not have any pro-apoptotic effect on human ovarian cancer cells but has strong anti-angiogenic effects as seen on CAM, but not dependent on the p38-MAPK pathway. CONCLUSIONS: Dietary flaxseed supplementation promotes a pro-apoptotic and anti-angiogenic effect in ovarian tumors, not in normal ovaries. The biologically derived active compounds from flaxseed diet act through different pathways to elicit their respective anti-cancer effects. A flaxseed-supplemented diet is a promising approach for prevention of ovarian cancer as well as having a significant potential as an adjuvant treatment to supplement chemotherapeutic agents for treatment of advanced stages of ovarian cancer.

Structural analysis of sigma E interactions with core RNA polymerase and its cognate P-hsp20 promoter of Mycobacterium tuberculosis.

Alternate sigma factor plays an important role for the survival of Mycobacterium tuberculosis in adverse environmental condition. Stress-induced sigma factors are major cause for expression of genes involved in pathogenesis, dormancy and various unusual environmental conditions. In the present work, an attempt has been made to characterize one of such M. tuberculosis (Mtb) sigma factor, SigE. The structures of Mtb-SigE and Mtb-ß have been predicted using comparative modelling techniques and validated. Effort has also been implied to understand the nature of interaction of SigE with the core RNA polymerase subunits which have well identified the amino acid residues in the binding interface and prompted the fact that Mtb-ß' and Mtb-ß interact with domain 2 and domain 4 of Mtb-SigE, respectively. Furthermore, intermolecular docking study predicted the interface between the Mtb-SigE and its putative promoter P-hsp20. The report confers the probable amino acid residues and the nitrogenous bases involved in the recognition of P-hsp20 by the sigma factor to initiate the transcription process.