IDENTIFICATION OF NOVEL NATURAL COMPOUNDS WITH ANTI-BREAST CANCER ACTIVITIES

Breast cancer continues to be the leading cause of cancer-related death in women worldwide. In this dissertation, natural compounds with anti-breast cancer activities were identified. We investigated the effect of Salinomycin, a potassium ionophore isolated from Streptomyces albus, on the survival of three human breast cancer cell lines MCF-7, T47D and MDA-MB-231. High concentrations of Salinomycin induced a G2 arrest, down regulation of survivin and triggered apoptosis. Interestingly, treatments with low concentrations of Salinomycin induced a transient G1 arrest at an earlier time and G2 arrest and senescence, associated with enlarged cell morphology, at a later time. There was also an upregulation of p21 protein, an increase in Histone H3 and H4 hyperacetylation and expression of SA-β-Gal activity. Furthermore, it was found that Salinomycin was able to potentiate the killing of the MCF-7 and MDA-MB-231 cells, by the chemotherapeutic agents, 4-Hydroxytamoxifen and frondoside A, We also investigated the effect of Origanum majorana, a culinary herb, ethanolic extract (OME) on the survival, migration, invasion and tumor growth of the highly proliferative and invasive triple-negative p53 mutant breast cancer cell line MDA-MB-231. We found that OME inhibited the viability of MDA-MB-231 breast cancer cells both in vitro and in vivo. OME was found to elicit anti-breast cancer activity by inducing mitotic arrest, DNA damage and triggering the extrinsic apoptotic pathway. Moreover, OME was found to induce down regulation of survivin, an important therapeutic target against breast cancer. v OME was also found to inhibit cell migration and invasion, two major events required for tumor metastasis. Our data are the first to link senescence and histone modifications to Salinomycin which provides a new insight to better understand the mechanism of action of Salinomycin, at least in breast cancer cells. Moreover, our findings provide strong evidence that O. majorana may be a promising chemopreventive and therapeutic candidate against cancer especially for highly invasive triple negative p53 mutant breast cancer; thus validating its use in alternative medicine

Effects of Medicinal Plants on Lead Induced Toxicity of Biotransformation and Redox Cycling Enzymes

All organisms including humans are constantly and unavoidably exposed to foreign chemicals (xenobiotics), which include both manufactured and natural molecules such as pollutants and plant metabolites. One such alarming pollutant is lead as it is constantly produced from automobiles exhaust products and food containers. It has been documented that lead, as the major heavy metal pollutant in the air, has notable effects on nervous systems and efficiency of several enzymes. There are some natural products such as medicinal plants that might have antagonistic effects against such pollutants. Several medicinal plants are used as traditional remedies for many ailments without knowing their detailed biochemical actions at the molecular level. This work focused on lead toxicity that had been tested in-vitro against some enzymes that are involved in biomolecules synthesis and on enzymes involved in liver detoxication and biotransformation. Furthermore, the antagonistic (or synergestic) effects of some medicinal plants extracts was tested against lead toxicity, as an example of a commonly present heavy metal, with regards to certain metabolic and biotransformation enzymes activities. In addition, enzyme kinetic parameters (Km and Vmax) in relation to substrate binding stoichiometry and inhibition (or activation) were investigated. In this study, three major enzymes that play important roles in the body were selected. Cytochrome P4502E1 (CYP2E1), Glutathione -S- Transferses (GSTs) enzymes were chosen as representative examples of phase I and phase II of biotransfomation enzymes systems respectively. Nicotinamide-adenine dinucleotide synthetase (NADS) was the third enzyme in our investigation which is involved in Nicotinamide-adenine dinucleotide (NAD+) synthesis. We have selected 4 indigenous species of plants which are commonly used as medicinal plants in the U.A.E for various ailments. These plants are Salvia officinalis, Hibiscus sabdariffa, Chamomilla recutita, and Nigella sativa. Results of this study showed that lead solution inhibited the three sets of enzymes selected for this study in vitro but the extent and sensitivity of such inhibition was different among the studied enzymes systems. The study revealed that increasing the concentrations of the four selected medicinal plants have the ability to decrease the inhibitory effect of lead solution on CYP2E1 activity. GSTs activity was also affected by the medicinal plants extracts within the concentration range used. It has been shown that all the plants extracts revealed inhibitory effects on GSTs activity in vitro. The extent of the inhibitory effect depends on plant species and concentrations of each plant extract. NADS activity in rat liver microsome was determined using High-performance chromatography (HPLC) and the formed NAD+ is separated from the substrates and the other microsome components in Retnetion time (Rt) of 13 minutes. It has been shown that the four plants extracts ameliorated the inhibitory effect of lead on NADS activity when low and high concentrations of each plant extract were used with lead solution. However, these plants extracts alone did not show any significant effect on NADS activity except for Hibiscus sabdariffa extract which caused decreased in NADS activity (11 % inhibition) and such decrease in activity may be due to some active compounds in the plant that cause this decrease in activity in vitro. Total phenols, flavonoids and proteins were determined and the results revealed that all the plant extracts contain phenolic compounds range from 76.4-28.6 mg GAE/g plant extract) and flavonoids content range from 7.6-28.1 mg QE/g plant extract). The four plants extracts also showed scavenging activity against DPPH free radical which may be related to the phenolic and flavonoids contents of these plants and this may-in part- contribute to the results we obtained in this study regarding lead toxicity and enzymes activities

Rhus coriaria induces senescence and autophagic cell death in breast cancer cells through a mechanism involving p38 and ERK1/2 activation

Here, we investigated the anticancer effect of Rhus coriaria on three breast cancer cell lines. We demonstrated that Rhus coriaria ethanolic extract (RCE) inhibits the proliferation of these cell lines in a time- and concentration-dependent manner. RCE induced senescence and cell cycle arrest at G1 phase. These changes were concomitant with upregulation of p21, downregulation of cyclin D1, p27, PCNA, c-myc, phospho-RB and expression of senescence-associated β-galactosidase activity. No proliferative recovery was detected after RCE removal. Annexin V staining and PARP cleavage analysis revealed a minimal induction of apoptosis in MDA-MB-231 cells. Electron microscopy revealed the presence of autophagic vacuoles in RCE-treated cells. Interestingly, blocking autophagy by 3-methyladenine (3-MA) or chloroquine (CQ) reduced RCE-induced cell death and senescence. RCE was also found to activate p38 and ERK1/2 signaling pathways which coincided with induction of autophagy. Furthermore, we found that while both autophagy inhibitors abolished p38 phosphorylation, only CQ led to significant decrease in pERK1/2. Finally, RCE induced DNA damage and reduced mutant p53, two events that preceded autophagy. Our findings provide strong evidence that R. coriaria possesses strong anti-breast cancer activity through induction of senescence and autophagic cell death, making it a promising alternative or adjunct therapeutic candidate against breast cancer.UAEU Program for Advanced Research (Grant 31S111-UPAR) and by the Zayed Center for Health Sciences (ZCHS) research grant (grant 31R021) and College of Science Individual Research Grant (grant 31S123) to Rabah Iratni

Perirenal Adipose Tissue Inflammation: Novel Insights Linking Metabolic Dysfunction to Renal Diseases

A healthy adipose tissue (AT) is indispensable to human wellbeing. Among other roles, it contributes to energy homeostasis and provides insulation for internal organs. Adipocytes were previously thought to be a passive store of excess calories, however this view evolved to include an endocrine role. Adipose tissue was shown to synthesize and secrete adipokines that are pertinent to glucose and lipid homeostasis, as well as inflammation. Importantly, the obesity-induced adipose tissue expansion stimulates a plethora of signals capable of triggering an inflammatory response. These inflammatory manifestations of obese AT have been linked to insulin resistance, metabolic syndrome, and type 2 diabetes, and proposed to evoke obesity-induced comorbidities including cardiovascular diseases (CVDs). A growing body of evidence suggests that metabolic disorders, characterized by AT inflammation and accumulation around organs may eventually induce organ dysfunction through a direct local mechanism. Interestingly, perirenal adipose tissue (PRAT), surrounding the kidney, influences renal function and metabolism. In this regard, PRAT emerged as an independent risk factor for chronic kidney disease (CKD) and is even correlated with CVD. Here, we review the available evidence on the impact of PRAT alteration in different metabolic states on the renal and cardiovascular function. We present a broad overview of novel insights linking cardiovascular derangements and CKD with a focus on metabolic disorders affecting PRAT. We also argue that the confluence among these pathways may open several perspectives for future pharmacological therapies against CKD and CVD possibly by modulating PRAT immunometabolism.This work was supported by AUB-Faculty of Medicine Medical Practice Plan Grant #320148 and an AUB President Collaborative Research Stimulus Grant to AE-Y

Rhus coriaria suppresses angiogenesis, metastasis and tumor growth of breast cancer through inhibition of STAT3, NFκB and nitric oxide pathways

Recently, we reported that Rhus coriaria exhibits anticancer activities by promoting cell cycle arrest and autophagic cell death of the metastatic triple negative MDA-MB-231 breast cancer cells. Here, we investigated the effect of Rhus coriaria on the migration, invasion, metastasis and tumor growth of TNBC cells. Our current study revealed that non-cytotoxic concentrations of Rhus coriaria significantly inhibited migration and invasion, blocked adhesion to fibronectin and downregulated MMP-9 and prostaglandin E2 (PgE2). Not only did Rhus coriaria decrease their adhesion to HUVECs and to lung microvascular endothelial (HMVEC-L) cells, but it also inhibited the transendothelial migration of MDA-MB-231 cells through TNF-α-activated HUVECs. Furthermore, we found that Rhus coriaria inhibited angiogenesis, reduced VEGF production in both MDA-MB-231 and HUVECs and downregulated the inflammatory cytokines TNF-α, IL-6 and IL-8. The underlying mechanism for Rhus coriaria effects appears to be through inhibiting NFκB, STAT3 and nitric oxide (NO) pathways. Most importantly, by using chick embryo tumor growth assay, we showed that Rhus coriaria suppressed tumor growth and metastasis in vivo. The results described in the present study identify Rhus coriaria as a promising chemopreventive and therapeutic candidate that modulate triple negative breast cancer growth and metastasis.UAEU Program for Advanced Research (Grant 31S111-UPAR), the Zayed Center for Health Sciences (ZCHS) research grant (grant 31R021) and the Terry Fox Foundation Grant (2013) to Rabah Iratni

Propolis: A Detailed Insight of Its Anticancer Molecular Mechanisms

Cancer is the second most life-threatening disease and has become a global health and economic problem worldwide. Due to the multifactorial nature of cancer, its pathophysiology is not completely understood so far, which makes it hard to treat. The current therapeutic strategies for cancer lack the efficacy due to the emergence of drug resistance and the toxic side effects associated with the treatment. Therefore, the search for more efficient and less toxic cancer treatment strategies is still at the forefront of current research. Propolis is a mixture of resinous compounds containing beeswax and partially digested exudates from plants leaves and buds. Its chemical composition varies widely depending on the bee species, geographic location, plant species, and weather conditions. Since ancient times, propolis has been used in many conditions and aliments for its healing properties. Propolis has well-known therapeutic actions including antioxidative, antimicrobial, anti-inflammatory, and anticancer properties. In recent years, extensive in vitro and in vivo studies have suggested that propolis possesses properties against several types of cancers. The present review highlights the recent progress made on the molecular targets and signaling pathways involved in the anticancer activities of propolis. Propolis exerts anticancer effects primarily by inhibiting cancer cell proliferation, inducing apoptosis through regulating various signaling pathways and arresting the tumor cell cycle, inducing autophagy, epigenetic modulations, and further inhibiting the invasion and metastasis of tumors. Propolis targets numerous signaling pathways associated with cancer therapy, including pathways mediated by p53, β-catenin, ERK1/2, MAPK, and NF-κB. Possible synergistic actions of a combination therapy of propolis with existing chemotherapies are also discussed in this review. Overall, propolis, by acting on diverse mechanisms simultaneously, can be considered to be a promising, multi-targeting, multi-pathways anticancer agent for the treatment of various types of cancers

Cardiac autonomic neuropathy: A progressive consequence of chronic low-grade inflammation in type 2 diabetes and related metabolic disorders

Cardiac autonomic neuropathy (CAN) is one of the earliest complications of type 2 diabetes (T2D), presenting a silent cause of cardiovascular morbidity and mortality. Recent research relates the pathogenesis of cardiovascular disease in T2D to an ensuing chronic, low-grade proinflammatory and pro-oxidative environment, being the hallmark of the metabolic syndrome. Metabolic inflammation emerges as adipose tissue inflammatory changes extending systemically, on the advent of hyperglycemia, to reach central regions of the brain. In light of changes in glucose and insulin homeostasis, dysbiosis or alteration of the gut microbiome (GM) emerges, further contributing to inflammatory processes through increased gut and blood–brain barrier permeability. Interestingly, studies reveal that the determinants of oxidative stress and inflammation progression exist at the crossroad of CAN manifestations, dictating their evolution along the natural course of T2D development. Indeed, sympathetic and parasympathetic deterioration was shown to correlate with markers of adipose, vascular, and systemic inflammation. Additionally, evidence points out that dysbiosis could promote a sympatho-excitatory state through differentially affecting the secretion of hormones and neuromodulators, such as norepinephrine, serotonin, and γ-aminobutyric acid, and acting along the renin–angiotensin–aldosterone axis. Emerging neuronal inflammation and concomitant autophagic defects in brainstem nuclei were described as possible underlying mechanisms of CAN in experimental models of metabolic syndrome and T2D. Drugs with anti-inflammatory characteristics provide potential avenues for targeting pathways involved in CAN initiation and progression. The aim of this review is to delineate the etiology of CAN in the context of a metabolic disorder characterized by elevated oxidative and inflammatory load.Funding: This work was funded by the Faculty of Medicine at the American University of Beirut, Medical Practice Plan, grant #320148 to A.F.E. and UAEU Program for Advanced Research, grant number 31S398-UPAR to Y.A.-D.Scopu

Carnosol Is a Novel Inhibitor of p300 Acetyltransferase in Breast Cancer

Carnosol, a natural polyphenol abundant in edible plants such as sage, rosemary, and oregano, has shown promising anticancer activity against various types of cancers. Nonetheless, very little is known about its molecular mechanism of action or its downstream target(s). We have previously shown that carnosol inhibits cellular proliferation, migration, invasion, and metastasis as well as triggers autophagy and apoptosis in the highly invasive MDA-MB-231 breast cancer cells. Here, we report that carnosol induces histone hypoacetylation in MDA-MB-231 and Hs578T breast cancer cells. We show that, while carnosol does not affect HDACs, it promotes a ROS-dependent proteasome degradation of p300 and PCAF histone acetyl transferases (HATs) without affecting other HATs such as GCN5 and hMOF. Carnosol-induced histone hypoacetylation remains persistent even when p300 and PCAF protein levels were rescued from degradation by (i) the inhibition of the proteasome activity by the proteasome inhibitors MG-132 and bortezomib, and (ii) the inhibition of ROS accumulation by the ROS scavenger, N-acetylcysteine. In addition, we report that, in a cell-free system, carnosol efficiently inhibits histone acetyltransferase activity of recombinant p300 but not that of PCAF or GCN5. Molecular docking studies reveal that carnosol inhibits p300 HAT activity by blocking the entry of the acetyl-CoA binding pocket of the catalytic domain. The superimposition of the docked conformation of the p300 HAT domain in complex with carnosol shows a similar orientation as the p300 structure with acetyl-CoA. Carnosol occupies the region where the pantetheine arm of the acetyl-CoA is bound. This study further confirms carnosol as a promising anti-breast cancer therapeutic compound and identifies it as a novel natural p300 inhibitor that could be added to the existing panel of inhibitors.This work was supported by the Zayed Center for Health Sciences (ZCHS) research grant (Grant # 31R086) and by Al Jalila Foundation Research Grant (Grant # 21S102-AJF2018007)

Ziziphus nummularia attenuates the malignant phenotype of human pancreatic cancer cells: Role of ros

Pancreatic cancer (PC) is the fourth leading cause of all cancer-related deaths. Despite major improvements in treating PC, low survival rate remains a major challenge, indicating the need for alternative approaches, including herbal medicine. Among medicinal plants is Ziziphus nummu-laria (family Rhamnaceae), which is a thorny shrub rich in bioactive molecules. Leaves of Ziziphus nummularia have been used to treat many pathological conditions, including cancer. However, their effects on human PC are still unknown. Here, we show that the treatment of human pancreatic ductal adenocarcinoma cells (Capan-2) with Ziziphus nummularia ethanolic extract (ZNE) (100–300 µg/mL) attenuated cell proliferation in a time-and concentration-dependent manner. Pretreatment with N-acetylcysteine, an ROS scavenger, attenuated the anti-proliferative effect of ZNE. In addition, ZNE significantly decreased the migratory and invasive capacity of Capan-2 with a concomitant downregulation of integrin α2 and increased cell–cell aggregation. In addition, ZNE inhibited in ovo angiogenesis as well as reduced VEGF and nitric oxide levels. Furthermore, ZNE downregulated the ERK1/2 and NF-κB signaling pathways, which are known to drive tumorigenic and metastatic events. Taken together, our results suggest that ZNE can attenuate the malignant phenotype of Capan-2 by inhibiting hallmarks of PC. Our data also provide evidence for the potential anticancer effect of Ziziphus nummularia, which may represent a new resource of novel anticancer compounds, especially ones that can be utilized for the management of PC

Inhibitory Effects of Salinomycin on Cell Survival, Colony Growth, Migration, and Invasion of Human Non-Small Cell Lung Cancer A549 and LNM35: Involvement of NAG-1.

A major challenge for oncologists and pharmacologists is to develop more potent and less toxic drugs that will decrease the tumor growth and improve the survival of lung cancer patients. Salinomycin is a polyether antibiotic used to kill gram-positive bacteria including mycobacteria, protozoans such as plasmodium falciparum, and the parasites responsible for the poultry disease coccidiosis. This old agent is now a serious anti-cancer drug candidate that selectively inhibits the growth of cancer stem cells. We investigated the impact of salinomycin on survival, colony growth, migration and invasion of the differentiated human non-small cell lung cancer lines LNM35 and A549. Salinomycin caused concentration- and time-dependent reduction in viability of LNM35 and A549 cells through a caspase 3/7-associated cell death pathway. Similarly, salinomycin (2.5-5 µM for 7 days) significantly decreased the growth of LNM35 and A549 colonies in soft agar. Metastasis is the main cause of death related to lung cancer. In this context, salinomycin induced a time- and concentration-dependent inhibition of cell migration and invasion. We also demonstrated for the first time that salinomycin induced a marked increase in the expression of the pro-apoptotic protein NAG-1 leading to the inhibition of lung cancer cell invasion but not cell survival. These findings identify salinomycin as a promising novel therapeutic agent for lung cancer