Chrysovalantou Mihailidou

CHOP-dependent regulation of p21/waf1 during ER stress.

The transcription factor CHOP/GADD153 is induced during the unfolded protein response (UPR) and is associated to the induction of ER stress-related apoptosis. However, how the transition between the pro-survival and the pro-apoptotic role of ER stress is being orchestrated remains poorly understood. Here we show that tunicamycin, an antibiotic promoting ER stress, suppresses the expression of p21, a tumor suppressor that induces cell cycle arrest and inhibits apoptosis. This suppression of p21 levels was independent of p53 that is the major transcriptional regulator of p21, but could be reproduced by forced expression of CHOP. Consistently with these findings, siRNA-mediated inhibition of p21 levels restored the sensitivity of CHOP-deficient cells to tunicamycin. Our findings are consistent with a CHOP-dependent role for p21 in the shift from the pro-survival to the pro-apoptotic function of UPR.

Improvement of chemotherapeutic drug efficacy by endoplasmic reticulum stress

Tunicamycin (TUN), an inhibitor of protein glycosylation and therefore a potent stimulator of endoplasmic reticulum (ER) stress, has been used to improve anticancer drug efficacy, but the underlying mechanism remains obscure. In this study, we show that acute administration of TUN in mice induces the unfolded protein response and suppresses the levels of P21, a cell cycle regulator with anti-apoptotic activity. The inhibition of P21 after ER stress appears to be C/EBP homologous protein (CHOP)-dependent because in CHOP-deficient mice, TUN not only failed to suppress, but rather induced the expression of P21. Results of promoter-activity reporter assays using human cancer cells and mouse fibroblasts indicated that the regulation of P21 by CHOP operates at the level of transcription and involves direct binding of CHOP transcription factor to the P21 promoter. The results of cell viability and clonogenic assays indicate that ER-stress-related suppression of P21 expression potentiates caspase activation and sensitizes cells to doxorubicin treatment, while administration of TUN to mice increases the therapeutic efficacy of anticancer therapy for HepG2 liver and A549 lung cancers.

Regulation of P21 during diabetes-associated stress of the endoplasmic reticulum

Endoplasmic reticulum (ER) stress plays a major role in the pathogenesis of diabetes by inducing β-cell apoptosis in the islets of Langerhans. In this study, we show that the transcription factor CHOP, which is instrumental for the induction of ER-stress-associated apoptosis and the pancreatic dysfunction in diabetes, regulates the expression of P21 (WAF1), a cell cycle regulator with anti-apoptotic activity that promotes cell survival. Deficiency of P21 sensitizes pancreatic β-cells to glucotoxicity, while in mice genetic ablation of P21 accelerates experimental diet-induced diabetes, results indicative of a protective role for P21 in the development of the disease. Conversely, pharmacological stimulation of P21 expression by nutlin-3a, an inhibitor of P53-MDM2 interaction, restores pancreatic function and facilitates glucose homeostasis. These findings indicate that P21 acts as an inhibitor of ER-stress-associated tissue damage and that stimulation of P21 activity can be beneficial for the management of diabetes and probably of other conditions in which ER-stress-associated death is undesirable.

A crosstalk between p21 and UPR-induced transcription factor C/EBP homologous protein (chop) linked to type 2 diabetes

Type 2 diabetes (T2D) is a disease that is characterized by raised levels of glucose in the blood combined with insulin resistance and relative insulin deficiency. The pathogenesis of type 2 diabetes is associated with the induction of the unfolded protein response (UPR). While UPR aims to restore tissue homeostasis following stress of the endoplasmic reticulum (ER), prolonged ER stress triggers apoptosis at least in part through the unfolded protein response (UPR)-activated transcription factor C/EBP (CCAAT/enhancer binding protein) homologous protein (CHOP). CHOP has elevated as a critical mediator connecting accumulation and aggregation of unfolded proteins in the ER and oxidative stress and also contributes to the induction of apoptosis in β-cell (beta-cell) - cells under conditions of increased insulin demand. p21 is a cell cycle regulator that is implicated in the regulation of the UPR by various mechanisms involving inhibition of apoptosis and facilitation of the regeneration capacity of the β cells. In this review we summarize the role of ER stress in the pathogenesis of type 2 diabetes which is associated with the induction of the unfolded protein response (UPR). We also review recent evidence associating p21 activity with β cell health and regenerative capacity by mechanisms that may interfere with the effects of p21 in the UPR or operate independently of ER stress. Most likely understanding the molecular details of the pathogenesis of type 2 diabetes will be beneficial for the management of the disease.

Co-targeting of EGFR and autophagy signaling is an emerging treatment strategy in metastatic colorectal cancer

The epidermal growth factor receptor (EGFR) and its associated pathway is a critical key regulator of CRC development and progression. The monoclonal antibodies (MoAbs) cetuximab and panitumumab, directed against EGFR, represent a major step forward in the treatment of metastatic colorectal cancer (mCRC), in terms of progression-free survival and overall survival in several clinical trials. However, the activity of anti-EGFR MoAbs appears to be limited to a subset of patients with mCRC. Studies have highlighted that acquired-resistance to anti-EGFR MoAbs biochemically converge into Ras/Raf/Mek/Erk and PI3K/Akt/mTOR pathways. Recent data also suggest that acquired-resistance to anti-EGFR MoAbs is accompanied by inhibition of EGFR internalization, ubiqutinization, degradation and prolonged downregulation. It is well established that autophagy, a self-cannibalization process, is considered to be associated with resistance to the anti-EGFR MoAbs therapy. Additionally, autophagy induced by anti-EGFR MoAbs acts as a protective response in cancer cells. Thus, inhibition of autophagy after treatment with EGFR MoAbs can result in autophagic cell death. A combination therapy comprising of anti-EGFR MoAbs and autophagy inhibitors would represent a multi-pronged approach that could be evolved into an active therapeutic strategy in mCRC patients.

Superior efficacy of the antifungal agent Ciclopirox Olamine over Gemcitabine in pancreatic cancer models

Ciclopirox olamine (CPX) is an antifungal agent that has recently demonstrated promising anti-neoplastic activity against hematologic and solid tumors. Here, we evaluated CPX compared with gemcitabine alone as well as their combination in human pancreatic cancer cell lines; BxPC-3, Panc-1, and MIA PaCa-2 and in humanized xenograft mouse models. We also examined the preclinical pharmacodynamic activity of CPX. CPX caused a pronounced decrease in cell proliferation and clonogenic growth potential. These inhibitory effects were accompanied by induction of reactive oxygen species (ROS), which were strongly associated with reduced Bcl-xL and survivin levels and activation of a panel of caspases, especially caspase-3, and finally resulted in apoptotic death. CPX-induced apoptosis was associated with reduced pEGFR (Y1068) and pAkt (Ser473) protein levels. Additionally, decreased proliferation was observed in CPX-treated xenografts tumors, demonstrating unique tumor regression and a profound survival benefit. Finally, we showed that CPX significantly abrogated gemcitabine-induced ROS levels in pancreatic tissues. These pre-clinical results have verified the superior antitumor efficacy of CPX over gemcitabine alone, while their combination is even more effective, providing the rationale for further clinical testing of CPX plus gemcitabine in pancreatic cancer patients.

Endoplasmic Reticulum stress is associated with the pathogenesis of Pemphigus Vulgaris

tion of Met during bleaching (s3). Trp, an amino acid that is susceptible to reactive oxygen species (s4) or thermal reaction (5), was depleted by the chemical treatments of hair. In addition to the individual alteration of these AAs, multivariate analysis using PCA revealed that combined assessment of Trp, Met, Cya and Cys/Cya may be useful in grading hair damage associated with chemical treatments. High levels of Trp, Met and Cys/ Cya suggested intact or healthy hairs, while their decreases indicated hair damage. Along with the recently reported proteomic approach (6), this methodology may overcome the limitation of morphological or mechanical evaluation by providing information on the compositional changes, which is crucial to the prevention or repairing of damaged hairs. Hair lipids are important components of hair cuticle. Hair lipids are mainly composed of FFAs, CH, cholesteryl sulphate, wax esters, 18-MEA, triglyceride and CERs (7). Of these hair lipids, FFAs like C22:1FA (erucic acid), C22FA (behenic acid), C24FA (lignoceric acid), C24:1FA (nervonic acid), C26FA (cerotic acid) and an integral hair lipid, 18-MEA, were depleted in proportion to the severity of hair damage. FFAs originated from sebum secretion coat the outside of hairs (8). Therefore, it is plausible that these FFAs are easily depleted during chemical treatment of hair. 18-MEA, which is biosynthesized in hair matrix cells, is linked by a cysteine thioester bond within the cuticle at the surface of human hairs (s5), which makes it susceptible to the breakage of cross-links during chemical treatment as well as in diseases (9). In conclusion, we demonstrate that hair AAs and lipids are markedly altered during chemical treatments of hair. The alteration of these AAs and lipids that are critical to impart mechanical strength and barrier function on hair may ultimately deteriorate the integrity or fibre quality of hair. Metabolomic analysis revealed AA or lipid biomarkers specific to hair damage, which may be useful for the quantitative grading of hair damage, and ultimately would be instrumental for the development of safer and undamaging hair care products. Author contributions KMJ, BMK, HKL, JHL and KML designed the study; KMJ, ARK and SNK involved in the research; KMJ, KML and SJB analysed the data; and KMJ and KML wrote and revised the manuscript. Conflict of interest None Supporting Information

Effect of steviol, steviol glycosides and stevia extract on glucocorticoid receptor signaling in normal and cancer blood cells

The use of steviol glycosides as non-caloric sweeteners has proven to be beneficial for patients with type 2 diabetes mellitus (T2D), obesity, and metabolic syndrome. However, recent data demonstrate that steviol and stevioside might act as glucocorticoid receptor (GR) agonists and thus correlate with adverse effects on metabolism. Herein, we evaluated the impact of steviol, steviol glycosides, and a Greek-derived stevia extract on a number of key steps of GR signaling cascade in peripheral blood mononuclear cells (PBMCs) and in Jurkat leukemia cells. Our results revealed that none of the tested compounds altered the expression of primary GR-target genes (GILZ, FKPB5), GR protein levels or GR subcellular localization in PBMCs; those compounds increased GILZ and FKPB5 mRNA levels as well as GRE-mediated luciferase activity, inducing in parallel GR nuclear translocation in Jurkat cells. The GR-modulatory activity demonstrated by stevia-compounds in Jurkat cells but not in PBMCs may be due to a cell-type specific effect.

Exosomes in lung cancer diagnosis and treatment. From the translating research into future clinical practice.

Lung cancer is one of the main causes of cancer-related death worldwide. Despite advances in lung cancer pathophysiology, diagnosis and prognosis, a better understanding of the disease is strongly needed in order to establish novel diagnostic and therapeutic approaches that should improve treatment outcomes. Exosomes are a type of cell-secreted extracellular vesicles, which transfer a wide variety of biomolecules, such as proteins, mRNAs, microRNAs, and lipids, are implicated in intercellular communication and modulate tumor-host interactions. The potential value of exosomes and their contents in lung cancer diagnosis, prognosis and prediction of treatment outcome is supported by ample literature. Growing attention has been drawn specifically to the critical role of exosomal miRNAs in lung cancer pathogenesis and their potential clinical utility, especially due to their ability to modulate gene expression post-transcriptionally. Owing to their universal presence in the blood and other bodily fluids, exosomes are considered candidate biomarkers. Furthermore, their ability to deliver biomolecules and drugs to recipient cells renders them possible drug delivery vehicles in lung cancer. Here we review the pathological functions of exosomes in cancer and discuss their possible clinical utility as biomarkers and therapeutic agents in the management of lung cancer.

Ciclopirox enhances pancreatic islet health by modulating the unfolded protein response in diabetes

Pancreatic dysfunction during diabetes is linked to the induction of endoplasmic reticulum (ER) stress on pancreatic beta (β) cells. Our laboratory recently discovered that p21 protects from diabetes by modifying the outcome of ER stress response. In the present study, we explored the antidiabetic activity of ciclopirox (CPX), an iron chelator and recently described activator of p21 expression. The effects of CPX in beta cell survival and function were assessed in cultured islets in vitro as well as in diabetic mice in vivo. The consequences of CPX in high glucose-induced insulin release and reactive oxygen species (ROS) production were also evaluated. Islet survival assays confirmed the significance of p21 in the regulation of glucotoxicity and suggested that CPX counteracts glucotoxicity in a manner that depends on p21. In vivo, administration of CPX in wild-type (WT) diabetic mice restored glucose homeostasis. In WT-cultured islets, CPX suppressed the expression of ER stress markers BiP, GRP94, and CHOP and reduced the levels of ROS during culture at high glucose. This reduction of ER stress may be associated with the ability of CPX to inhibit insulin release. Iron citrate stimulated insulin release, which was inhibited by CPX that functions as an iron chelator. It is conceivable that inhibition of insulin production constrains ER stress in islets promoting their survival and thus protecting from diabetes in vivo. This unfolded protein response (UPR)-antagonizing activity of CPX suggests application for the management not only of diabetes but also of other conditions related to ER stress.