Vitamin E metabolites as potent inhibitors of 5-lipoxygenase

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Screening of Vietnamese medicinal plants for NF-κB signaling inhibitors: Assessing the activity of flavonoids from the stem bark of Oroxylum indicum

AbstractEthnopharmacological relevanceSeventeen plants used in Vietnamese traditional medicine for the treatment of inflammatory disorders were screened for NF-κB inhibitory activity. Oroxylum indicum, which exhibited activity, was investigated in detail.Materials and methodsForty plant extracts from 17 species were prepared by maceration using dichloromethane and methanol and were tested (10µg/mL) to evaluate their ability to inhibit NF-κB activation using TNF-α-stimulated HEK-293 cells stably transfected with a NF-κB-driven luciferase reporter. The active extract of Oroxylum indicum was subsequently fractionated by different chromatographic techniques. After isolation, all single compounds were identified by spectroscopic methods and assessed for NF-κB inhibitory effects.ResultsThe dichloromethane extracts obtained from Chromolaena odorata leaves and the stem bark of Oroxylum indicum showed distinct inhibitory effects on NF-κB activation at a concentration of 10µg/mL. The active extract of Oroxylum indicum was subjected to further phytochemical studies resulting in identification of four flavonoid aglyca and six flavonoid glycosides. Pharmacological evaluation of the obtained compounds identified oroxylin A as the most active substance (IC50=3.9µM, 95% CI: 3.5–4.4µM), while chrysin and hispidulin showed lower activity with IC50=7.2µM (95% CI: 6.0–8.8µM) and 9.0µM (95% CI: 7.9–10.2µM), respectively. Interestingly, in this study the activity of baicalein (IC50=28.1µM, 95% CI: 24.6–32.0µM) was weak. The isolated glycosides showed no inhibitory activity when tested at a concentration of 30µM. Quantification of the four active flavonoids in extracts and plant materials suggested that oroxylin A contributes to the NF-κB inhibitory activity of the stem barks of Oroxylum indicum to a greater extent than baicalein which was thought to be responsible for the anti-inflammatory activity of this plant.ConclusionsThe screening presented in this study identified the dichloromethane extracts of Chromolaena odorata and Oroxylum indicum as promising sources for NF-κB inhibitors. Hispidulin, baicalein, chrysin and oroxylin A, isolated from Oroxylum indicum, were identified as inhibitors of NF- κB activation

A combination of trastuzumab and BAG-1 inhibition synergistically targets HER2 positive breast cancer cells

Treatment of HER2+ breast cancer with trastuzumab is effective and combination anti-HER2 therapies have demonstrated benefit over monotherapy in the neoadjuvant and metastatic settings. This study investigated the therapeutic potential of targeting the BAG-1 protein co-chaperone in trastuzumab-responsive or -resistant cells. In the METABRIC dataset, BAG-1 mRNA was significantly elevated in HER2+ breast tumors and predicted overall survival in a multivariate analysis (HR = 0.81; p = 0.022). In a breast cell line panel, BAG-1 protein was increased in HER2+ cells and was required for optimal growth as shown by siRNA knockdown. Overexpression of BAG-1S in HER2+ SKBR3 cells blocked growth inhibition by trastuzumab, whereas overexpression of a mutant BAG-1S protein (BAG-1S H3AB), defective in binding HSC70, potentiated the effect of trastuzumab. Injection of a Tet-On SKBR3 clone, induced to overexpress myc-BAG-1S into the mammary fat pads of immunocompromised mice, resulted in 2-fold larger tumors compared to uninduced controls. Induction of myc-BAG-1S expression in two Tet-On SKBR3 clones attenuated growth inhibition by trastuzumab in vitro. Targeting endogenous BAG-1 by siRNA enhanced growth inhibition of SKBR3 and BT474 cells by trastuzumab, while BAG-1 protein-protein interaction inhibitor (Thio-S or Thio-2) plus trastuzumab combination treatment synergistically attenuated growth. In BT474 cells this reduced protein synthesis, caused G1/S cell cycle arrest and targeted the ERK and AKT signaling pathways. In a SKBR3 subpopulation with acquired resistance to trastuzumab BAG-1 targeting remained effective and either Thio-2 or BAG-1 siRNA reduced growth more compared to trastuzumab-responsive parental cells. In summary, targeting BAG-1 function in combination with anti-HER2 therapy might prove beneficial

Exploration of Long-Chain Vitamin E Metabolites for the Discovery of a Highly Potent, Orally Effective, and Metabolically Stable 5-LOX Inhibitor that Limits Inflammation.

Endogenous long-chain metabolites of vitamin E (LCMs) mediate immune functions by targeting 5-lipoxygenase (5-LOX) and increasing the systemic concentrations of resolvin E3, a specialized proresolving lipid mediator. SAR studies on semisynthesized analogues highlight α-amplexichromanol (27a), which allosterically inhibits 5-LOX, being considerably more potent than endogenous LCMs in human primary immune cells and blood. Other enzymes within lipid mediator biosynthesis were not substantially inhibited, except for microsomal prostaglandin E2 synthase-1. Compound 27a is metabolized by sulfation and β-oxidation in human liver-on-chips and exhibits superior metabolic stability in mice over LCMs. Pharmacokinetic studies show distribution of 27a from plasma to the inflamed peritoneal cavity and lung. In parallel, 5-LOX-derived leukotriene levels decrease, and the inflammatory reaction is suppressed in reconstructed human epidermis, murine peritonitis, and experimental asthma in mice. Our study highlights 27a as an orally active, LCM-inspired drug candidate that limits inflammation with superior potency and metabolic stability to the endogenous lead

Lignan Derivatives from Krameria lappacea Roots Inhibit Acute Inflammation in Vivo and Pro-inflammatory Mediators in Vitro

The roots of Krameria lappacea are used traditionally against oropharyngeal inflammation. So far, the astringent and antimicrobial properties of its proanthocyanidin constituents are considered to account for the anti-inflammatory effect. The aim of the present study was to characterize pharmacologically a lipophilic extract of K. lappacea roots and several isolated lignan derivatives (111) in terms of their putative anti-inflammatory activity. The dichloromethane extract (ID50 77 \u3bcg/cm2) as well compounds 111 (ID50 0.310.60 \u3bcmol/cm2) exhibited topical antiedematous properties comparable to those of indomethacin (ID50 0.29 \u3bcmol/cm2) in a mouse ear in vivo model. Two of the most potent compounds, 2-(2-hydroxy-4-methoxyphenyl)-5-(3-hydroxypropyl)benzofuran (5) and (+)-conocarpan (7), were studied regarding their time-dependent edema development and leukocyte infiltration up to 48 h after croton oil-induced dermatitis induction, and they showed activity profiles similar to that of hydrocortisone. In vitro studies of the isolated lignan derivatives demonstrated the inhibition of NFkB, cyclooxygenase-1 and -2, 5-lipoxygenase, and microsomal prostaglandin E2 synthase-1 as well as antioxidant properties, as mechanisms possibly contributing to the observed in vivo effects. The present findings not only support the ethnopharmacological use of K. lappacea roots but also reveal that the isolated lignan derivatives contribute strongly to the anti-inflammatory activity of this herbal drug

Semisynthetic and Natural Garcinoic Acid Isoforms as New mPGES-1 Inhibitors

Over the last twenty years, tocotrienol analogues raised great interest because of their higher level and larger domain of biological activities when compared with tocopherols. Amongst the most promising therapeutic application, anti-inflammatory potency has been evaluated through the inhibition of various mediators of inflammation. Here, we worked on the isolation of two natural isoforms of garcinoic acid (i.e., δ and γ) from two different sources, respectively, Garcinia kola seeds and Garcinia amplexicaulis bark. We also developed semisynthetic strategies to access the other two non-natural α- and β-garcinoic acid isoforms. In the next stage of our work, microsomal prostaglandin E2 synthase was defined as a target to evaluate the anti-inflammatory potential of the four garcinoic acid isomers. Both dimethylated isoforms, β- and γ-garcinoic acid, exhibited the lowest IC50, 2.8 µM and 2.0 µM, respectively. These results showed that the affinity of tocotrienol analogues to microsomal prostaglandin E2 synthase-1 most probably contributes to the anti-inflammatory potential of this class of derivatives

Plant extracts in cell-based anti-inflammatory assays—Pitfalls and considerations related to removal of activity masking bulk components

Plants used in traditional medicine represent an important source of new lead compounds. However, cell-based in vitro screening assays with plant material are hampered by the complex nature of plant extracts as mixtures of active and inactive components. Bulk constituents, such as chlorophyll and polyphenols were previously shown to interfere with several biological in vitro assays. Their influence on anti-inflammatory cell-based testing systems has not been thoroughly investigated. Hence, the present study was aimed at comparing different procedures for the removal of bulk constituents from plant extracts and examining the influence of their elimination on selected cell-based anti-inflammatory assays. Malva sp. and Glechoma hederacea L., two plants used in traditional European medicine for the treatment of inflammatory disorders, were subjected to three different methods for the removal of chlorophyll and polyphenols, respectively. Removal of bulk constituents was confirmed by HPLC and mass spectrometry. Extracts were tested before and after the purification procedure, to determine their potential to inhibit the activation of the transcription factor NF-κB in reporter gene assay and to interfere with the secretion of the chemokine IL-8 after stimulation of endothelial cells with tumor necrosis factor (TNF-α) or lipopolysaccharide (LPS). Removal of chlorophyll from tested extracts led to a strong decrease in the anti-inflammatory activities, due to loss of bioactive constituents. In contrast, the effect of the polyphenol-free extracts was either not changed or significantly increased, depending on the purification method used. The study concluded that clearance of bulk compounds represents a valuable strategy for cell-based in vitro anti-inflammatory evaluation of plant extracts. Liquid–liquid partitioning was identified as the optimal method for the elimination of both chlorophyll and polyphenols. It is recommended that removal of chlorophyll from extracts always be accompanied by HPLC profiling to detect a possible loss of active constituents