Lipidomic profiling of exosomes from colorectal cancer cells and patients reveals potential biomarkers.

Strong evidence suggests that differences in the molecular composition of lipids in exosomes depend on the cell type and has an influence on cancer initiation and progression. Here, we analyzed by liquid chromatography-mass spectrometry (LC-MS) the lipidomic signature of exosomes derived from the human cell lines normal colon mucosa (NCM460D), and colorectal cancer (CRC) nonmetastatic (HCT116) and metastatic (SW620), and exosomes isolated from the plasma of nonmetastatic and metastatic CRC patients and healthy donors. Analysis of this exhaustive lipid study highlighted changes in some molecular species that were found in the cell lines and confirmed in the patients. For example, exosomes from primary cancer patients and nonmetastatic cells compared with healthy donors and control cells displayed a common marked increase in phosphatidylcholine (PC) 34 : 1, phosphatidylethanolamine (PE) 36 : 2, sphingomyelin (SM) d18 : 1/16 : 0, hexosylceramide (HexCer) d18 : 1/24 : 0 and HexCer d18 : 1/24 : 1. Interestingly, these same lipids species were decreased in the metastatic cell line and patients. Further, levels of PE 34 : 2, PE 36 : 2, and phosphorylated PE p16 : 0/20 : 4 were also significantly decreased in metastatic conditions when compared to the nonmetastatic counterparts. The only molecule species found markedly increased in metastatic conditions (in both patients and cells) when compared to controls was ceramide (Cer) d18 : 1/24 : 1. These decreases in lipid species in the extracellular vesicles might reflect function-associated changes in the metastatic cell membrane. Although these potential biomarkers need to be validated in a larger cohort, they provide new insight toward the use of clusters of lipid biomarkers rather than a single molecule for the diagnosis of different stages of CRC.

Anticolitic activity of prodigiosin loaded with selenium nanoparticles on acetic acid-induced colitis in rats.

Ulcerative colitis (UC) is a chronic autoimmune inflammatory disease associated with extensive mucosal damage. Prodigiosins (PGs) are natural bacterial pigments with well-known antioxidant and immunosuppressive properties. In the current study, we examined the possible protective effect of PGs loaded with selenium nanoparticles (PGs-SeNPs) against acetic acid (AcOH)-induced UC in rats. Thirty-five rats were separated into five equal groups with seven animals/group: control, UC, PGs (300 mg/kg), sodium selenite (Na2SeO3, 2 mg/kg), PGs-SeNPs (0.5 mg/kg), and 5-aminosalicylates (5-ASA, 200 mg/kg). Interestingly, PGs-SeNPs administration lessened colon inflammation and mucosal damage as indicated by inhibiting inflammatory markers upon AcOH injection. Furthermore, PGs-SeNPs improved the colonic antioxidant capacity and prevented oxidative insults as evidenced by the upregulation of Nrf2- and its downstream antioxidants along with the decreased pro-oxidants [reactive oxygen species (ROS), carbonyl protein, malondialdehyde (MDA), inducible nitric oxide synthase (iNOS), and nitric oxide (NO] in the colon tissue. Furthermore, PGs-SeNPs protected intestinal cell loss through blockade apoptotic cascade by decreasing pro-apoptotic proteins [Bcl-2-associated X protein (Bax) and caspase-3] and increasing anti-apoptotic protein, B cell lymphoma 2 (Bcl2). Collectively, PGs-SeNPs could be used as an alternative anti-colitic option due to their strong anti-inflammatory, antioxidant, and anti-apoptotic activities.

Marine Actinomycetes-Derived Secondary Metabolites Overcome TRAIL-Resistance via the Intrinsic Pathway through Downregulation of Survivin and XIAP.

Resistance of cancer cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis represents the major hurdle to the clinical use of TRAIL or its derivatives. The discovery and development of lead compounds able to sensitize tumor cells to TRAIL-induced cell death is thus likely to overcome this limitation. We recently reported that marine actinomycetes' crude extracts could restore TRAIL sensitivity of the MDA-MB-231 resistant triple negative breast cancer cell line. We demonstrate in this study, that purified secondary metabolites originating from distinct marine actinomycetes (sharkquinone (1), resistomycin (2), undecylprodigiosin (3), butylcyclopentylprodigiosin (4), elloxizanone A (5) and B (6), carboxyexfoliazone (7), and exfoliazone (8)), alone, and in a concentration-dependent manner, induce killing in both MDA-MB-231 and HCT116 cell lines. Combined with TRAIL, these compounds displayed additive to synergistic apoptotic activity in the Jurkat, HCT116 and MDA-MB-231 cell lines. Mechanistically, these secondary metabolites induced and enhanced procaspase-10, -8, -9 and -3 activation leading to an increase in PARP and lamin A/C cleavage. Apoptosis induced by these compounds was blocked by the pan-caspase inhibitor QvD, but not by a deficiency in caspase-8, FADD or TRAIL agonist receptors. Activation of the intrinsic pathway, on the other hand, is likely to explain both their ability to trigger cell death and to restore sensitivity to TRAIL, as it was evidenced that these compounds could induce the downregulation of XIAP and survivin. Our data further highlight that compounds derived from marine sources may lead to novel anti-cancer drug discovery.

Membrane-anchored heat-shock protein 70 (Hsp70) in cancer.

Hsp70 is a highly conserved and inducible heat shock protein that belongs to the HSP70 family of molecular chaperones and plays a central role in protein homeostasis. The main function of Hsp70 is to protect cells from physiological, pathological and environmental insults, as it assists an ATP-dependent manner the process of protein folding. Since Hsp70 provides critical cell survival functions, cancer cells are assumed to rely on this chaperone. Strong evidence suggests that Hsp70 is upregulated in different type of cancers and is involved in tumor growth, invasion, migration and resistance to anti-cancer therapy. Interestingly, this Hsp70 upregulation induces Hsp70 re-location into plasma membrane. In this review, the role of Hsp70 in cancer will be discussed focusing particularly on the extracellular membrane-bound Hsp70. The mechanism by which Hsp70 is translocated to plasma membrane of tumor cells and the recent discoveries of drugs targeting this Hsp70 in cancer therapy will be also highlighted.

Prodigiosins from a marine sponge-associated actinomycete attenuate HCl/ethanol-induced gastric lesion via antioxidant and anti-inflammatory mechanisms.

Gastric ulcer is sores that form in the stomach mucosal layer because of erosion caused by high acid secretion and excessive use of non-steroidal anti-inflammatory drugs. Prodigiosins (PdGs) are red-pigmented secondary metabolites produced by bacteria, including actinomycetes. Butylcycloheptylprodigiosin (1) and undecylprodigiosin (2) were identified and isolated from a crude extract of the actinomycete RA2 isolated from the Red Sea Sponge Spheciospongia mastoidea. Chemical structure of 1 and 2 was determined by NMR and mass spectroscopy. Although their antioxidant and anti-inflammatory properties are known, their effect on gastric lesion is unknown. Therefore, this study aimed to investigate gastroprotective effects of PdGs against HCl/ethanol-induced gastric lesion in rats. Oral pretreatment with PdGs (100, 200, and 300 mg/kg) attenuated severity of HCl/ethanol-induced gastric mucosal injury, as evidenced by decreases in gastric lesion index scores, ulceration area, histopathologic abnormality, and neutrophil infiltration. These effects were comparable to those of omeprazole, a standard anti-gastric ulcer agent. HCl/ethanol-induced gastric erosions was associated with tremendous increases in lipid peroxidation, nitric oxide, and pro-inflammatory cytokines and mediators (myeloperoxidase, interleukin-1ß, tumor necrosis factor-α, and cyclooxygenase-2), and with significant decreases in enzymatic and non-enzymatic antioxidant activities. However, PdGs ameliorated gastric inflammation and oxidative stress by downregulating nuclear factor kappa B and inducible nitric oxide synthase expression and upregulating heme oxygenase-1 expression. PdGs prevented gastric mucosal apoptosis by downregulating Bax and caspase-3 expression and upregulating Bcl-2 expression, thereby increasing prostaglandin E2 production. Our results suggested that PdGs exerted gastroprotective effects by decreasing the levels of inflammatory mediators, apoptotic markers, and antioxidants.

Epigenetic Regulation of TRAIL Signaling: Implication for Cancer Therapy.

One of the main characteristics of carcinogenesis relies on genetic alterations in DNA and epigenetic changes in histone and non-histone proteins. At the chromatin level, gene expression is tightly controlled by DNA methyl transferases, histone acetyltransferases (HATs), histone deacetylases (HDACs), and acetyl-binding proteins. In particular, the expression level and function of several tumor suppressor genes, or oncogenes such as c-Myc, p53 or TRAIL, have been found to be regulated by acetylation. For example, HATs are a group of enzymes, which are responsible for the acetylation of histone proteins, resulting in chromatin relaxation and transcriptional activation, whereas HDACs by deacetylating histones lead to chromatin compaction and the subsequent transcriptional repression of tumor suppressor genes. Direct acetylation of suppressor genes or oncogenes can affect their stability or function. Histone deacetylase inhibitors (HDACi) have thus been developed as a promising therapeutic target in oncology. While these inhibitors display anticancer properties in preclinical models, and despite the fact that some of them have been approved by the FDA, HDACi still have limited therapeutic efficacy in clinical terms. Nonetheless, combined with a wide range of structurally and functionally diverse chemical compounds or immune therapies, HDACi have been reported to work in synergy to induce tumor regression. In this review, the role of HDACs in cancer etiology and recent advances in the development of HDACi will be presented and put into perspective as potential drugs synergizing with TRAIL's pro-apoptotic potential.

Probiotics and yogurt modulate oxidative stress and fibrosis in livers of Schistosoma mansoni-infected mice.

BACKGROUND: Considerable morbidity, mortality, and economic loss result from schistosomiasis infection. Deposition of Schistosoma eggs in the hepatic portal vein is considered as the main causative agent for the development of liver fibrosis and subsequent liver cirrhosis. Probiotics are exogenous and beneficial microorganisms to living hosts against the harmful effect of many parasites. Strong evidence suggests the importance of probiotics in the control strategy of helminth. The ultimate goal of this study is to evaluate the protective effect of probiotics and yogurt on Schistosoma mansoni-induced oxidative stress and hepatic fibrosis in mice. METHODS: Mice were infected by tail immersion of schistosomal cercariae followed by an oral treatment with either probiotics or yogurt for one week before infection and immediately post-infection. Mice were scarified on day 56 following infection with S. mansoni and liver sample were obtained. RESULTS: We showed that oral administration of probiotics or yogurt revealed a significant reduction in worm number, egg load, and granuloma size in liver tissue, which is mainly assigned to the decreased expression level of matrix metalloproteinases 9 (MMP-9) in liver tissue. A significant reduction in the oxidative stress markers-induced by S. mansoni infection including lipid peroxidation and nitrite/nitrate was also detected. The level of some antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase) and reduced glutathione was greatly enhanced. Furthermore, treatment with probiotics or yogurt inhibited apoptosis in hepatic tissue, which is mainly assigned to the decreased expression level of caspases-3 in liver tissue. CONCLUSION: Our findings represent the promising anti-schistosomal activities of probiotics and yogurt.

Improving the catalytic, kinetic and thermodynamic properties of Bacillus subtilis KU710517 milk clotting enzyme via conjugation with polyethylene glycol.

Milk clotting enzyme (MCE) produced by Bacillus subtilis KU710517 was conjugated to several activated polysaccharides. Among all the conjugates, the enzyme conjugated with polyethylene glycol (PEG) exhibited the highest retained activity (551U/mg protein) with a recovered activity of 95.3%. The activation energy of PEG-conjugated enzyme was calculated as 24.56kJ·mol-1which was lower than that of the native one (29.27kJ·mol-1) however, the temperature quotient (Q10) was about 1.08 for the two forms of the enzyme. The calculated half-life times of PEG-conjugated enzyme at 55 and 60°C were 317.78 and 128.6min respectively, whereas at the same temperatures the native enzyme had lower half-life times (53 and 19.6min respectively). The data of thermodynamic analysis for substrate catalysis including the specificity constant (Vmax/Km), turnover number (kcat), catalytic efficiency (kcat/Km), enthalpy of activation (ΔH*), free energy of activation (ΔG*), free energy for transition state formation ΔG*E-T and free energy of substrate binding ΔG*E-S were determined for both native and PEG-conjugated enzyme. In addition, the thermodynamic parameters for irreversible inactivation (ΔH, ΔG, ΔS) were evaluated. The calculated results indicated that the catalytic properties after the PEG-conjugation were significantly improved.

Cytotoxic activity of alkyl benzoate and fatty acids from the red sea sponge Hyrtios erectus.

The chemical investigation of the methylene chloride fraction of marine sponge Hyrtios erectus led to the isolation of the known oxysterol (2) along with a new alkyl benzoate compound identified by spectroscopic methods (NMR and MS) as 4'-methylheptyl benzoate (1), whilst the n-butanol fraction afforded the known indole 3-carbaldehyde and ß-carboline derivatives. Moreover, the hexane fraction was analysed by GC-MS for their fatty acids (FAs). A total of 17 FAs with chain lengths between 14 and 25 carbons were identified. Methyl-branched FAs are predominated suggesting the presence of bacterial symbionts in the H. erectus sponge. Furthermore, compounds 1 and 2 displayed significant cytotoxicity against breast adenocarcinoma (MCF-7) with IC50 values of 2.4 and 3.8 µM, respectively, since compound 2 was also shown to have potent cytotoxic effect against hepatocellular carcinoma cells (HepG 2) with IC50 value of 1.3 µM.

Protective Effect of Fragaria ananassa Crude Extract on Cadmium-Induced Lipid Peroxidation, Antioxidant Enzymes Suppression, and Apoptosis in Rat Testes.

Cadmium is a deleterious environmental pollutant that threats both animals and human health. Oxidative stress and elevated levels of reactive oxygen species (ROS) have recently been reported to be the main cause of cellular damage as a result of cadmium exposure. We investigate, here, the protective effect of strawberry crude extracts on cadmium-induced oxidative damage of testes in rats. Four groups (n = 8) of 32 adult male Wistar rats weighing 160-180 g were used. The control group received 0.9% saline solution all over the experimental period (5 days). Group 2 was intraperitoneally injected with 6.5 mg/kg CdCl2. Group 3 was provided only with an oral administration of strawberry methanolic extract (SME) at a dose of 250 mg/kg. Group 4 was treated with SME before cadmium injection with the same mentioned doses. It was shown that cadmium exposure results in a significant decrease in both relative testicular weight and serum testosterone level. Analyzing the oxidative damaging effect of cadmium on the testicular tissue revealed the induction of oxidative stress markers represented in the elevated level of lipid peroxidation (LPO), nitric oxide (NO), and a decrease in the reduced glutathione (GSH) content. Considering cadmium toxicity, the level of the antioxidant enzyme activities including catalase (CAT), superoxide dismutase (SOD2), glutathione peroxidase (GPx1), and glutathione reductase (GR) were markedly decreased. Moreover, gene expression analysis indicated significant upregulation of the pro-apoptotic proteins, bcl-2-associated-X-protein (BAX), and tumor necrosis factor-α (TNFA) in response to cadmium intoxication, while significant downregulation of the anti-apoptotic, B-cell lymphoma 2 (BCL2) gene was detected. Immunohistochemistry of the testicular tissue possessed positive immunostaining for the increased level of TNF-α, but decreased number of proliferating cell nuclear antigen (PCNA) stained cells. Administration of SME debilitated the deleterious effect of cadmium via reduction of both LPO and NO levels followed by a significant enhancement in the gene expression level of CAT, SOD2, GPX1, GR, nuclear factor-erythroid 2-related factor 2 (NFE2L2), heme oxygenase-1 (HMOX1), Bcl-2, and PCNA. In addition, the SME treated group revealed a significant increase in the level of testosterone and GSH accompanied by a marked decrease in the gene expression level of Bax and TNF-α. In terms of the summarized results, the SME of Fragaria ananassa has a protective effect against cadmium-induced oxidative damage of testes.

Marine actinomycete crude extracts with potent TRAIL-resistance overcoming activity against breast cancer cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising anticancer agent, as it can kill tumor cells selectively. In our search of bioactive natural products to overcome TRAIL-resistance, we isolated 47 actinomycete strains from different sediments and seawater samples collected from the Red Sea coast in Egypt and found four crude extracts (EGY1, EGY3, EGY24 and EGY34) displaying TRAIL sensitizing activity in the resistant breast cancer cell line MDA-MB-231. None of these crude extracts exhibited cytotoxic effect on normal mouse embryonic fibroblasts (MEF), with the exception of EGY34. Analysis of the signaling pathways underlying the sensitization of MDA-MB-231 cells to TRAIL-induced apoptosis, by western blotting, revealed that all crude extracts facilitated initiator caspase­8/-10 activation upon TRAIL stimulation, but that in addition, EGY3 and EGY34, alone, induced strong ER-stress activation, with the appearance of BiP in the cytosolic extracts. Our results pave the way to the discovery and the development of marine-derived drugs for cancer therapy.

Sharkquinone, a new ana-quinonoid tetracene derivative from marine-derived Streptomyces sp. EGY1 with TRAIL resistance-overcoming activity.

A new ana-quinonoid tetracene metabolite, named sharkquinone (1), and the known SS-228R (2) have been isolated from the ethyl acetate extract of the culture of marine Streptomyces sp. EGY1. The strain was isolated from sediment sample collected from the Red Sea coast of Egypt. The structure of sharkquinone (1) was elucidated using detailed spectral (HRESI-MS, 1D and 2D NMR) analyses and quantum chemical calculations. This is the first report of the isolation of ana-quinonoid tetracene derivative from a natural source. Compound 1 showed the ability to overcome tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) resistance at a concentration of 10 µM in human gastric adenocarcinoma (AGS) cells.

Marine Drugs Regulating Apoptosis Induced by Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL).

Marine biomass diversity is a tremendous source of potential anticancer compounds. Several natural marine products have been described to restore tumor cell sensitivity to TNF-related apoptosis inducing ligand (TRAIL)-induced cell death. TRAIL is involved during tumor immune surveillance. Its selectivity for cancer cells has attracted much attention in oncology. This review aims at discussing the main mechanisms by which TRAIL signaling is regulated and presenting how marine bioactive compounds have been found, so far, to overcome TRAIL resistance in tumor cells.

Impact of methionine oxidation as an initial event on the pathway of human prion protein conversion.

Prion diseases comprise a group of fatal neurodegenerative disorders characterized by the autocatalytic conversion of the cellular prion protein PrP(C) into the infectious misfolded isoform PrP(Sc). Increasing evidence supports a specific role of oxidative stress in the onset of pathogenesis. Although the associated molecular mechanisms remain to be elucidated in detail, several studies currently suggest that methionine oxidation already detected in misfolded PrP(Sc) destabilizes the native PrP fold as an early event in the conversion pathway. To obtain more insights about the specific impact of surface-exposed methionine residues on the oxidative-induced conversion of human PrP we designed, produced, and comparatively investigated two new pseudosulfoxidation mutants of human PrP 121-231 that comprises the well-folded C-terminal domain. Applying circular dichroism spectroscopy and dynamic light scattering techniques we showed that pseudosulfoxidation of all surface exposed Met residues formed a monomeric molten globule-like species with striking similarities to misfolding intermediates recently reported by other groups. However, individual pseudosulfoxidation at the polymorphic M129 site did not significantly contribute to the structural destabilization. Further metal-induced oxidation of the partly unfolded pseudosulfoxidation mutant resulted in the formation of an oligomeric state that shares a comparable size and stability with PrP oligomers detected after the application of different other triggers for structural conversion, indicating a generic misfolding pathway of PrP. The obtained results highlight the specific importance of methionine oxidation at surface exposed residues for PrP misfolding, strongly supporting the hypothesis that increased oxidative stress could be one causative event for sporadic prion diseases and other neurodegenerative disorders.

UV-light-induced conversion and aggregation of prion proteins.

Increasing evidence suggests a central role for oxidative stress in the pathology of prion diseases, a group of fatal neurodegenerative disorders associated with structural conversion of the prion protein (PrP). Because UV-light-induced protein damage is mediated by direct photo-oxidation and radical reactions, we investigated the structural consequences of UVB radiation on recombinant murine and human prion proteins at pH 7.4 and pH 5.0. As revealed by circular dichroism and dynamic light scattering measurements, the observed PrP aggregation follows two independent pathways: (i) complete unfolding of the protein structure associated with rapid precipitation or (ii) specific structural conversion into distinct soluble beta-oligomers. The choice of pathway was directly attributed to the chromophoric properties of the PrP species and the susceptibility to oxidation. Regarding size, the oligomers characterized in this study share a high degree of identity with oligomeric species formed after structural destabilization induced by other triggers, which significantly strengthens the theory that partly unfolded intermediates represent initial precursor molecules directing the pathway of PrP aggregation. Moreover, we identified the first suitable photo-trigger capable of inducing refolding of PrP, which has an important biotechnological impact in terms of analyzing the conversion process on small time scales.