Identification of an ASC oligomerization inhibitor for the treatment of inflammatory diseases

The ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain (CARD)) protein is an scaffold component of different inflammasomes, intracellular multiprotein platforms of the innate immune system that are activated in response to pathogens or intracellular damage. The formation of ASC specks, initiated by different inflammasome receptors, promotes the recruitment and activation of procaspase-1, thereby triggering pyroptotic inflammatory cell death and pro-inflammatory cytokine release. Here we describe MM01 as the first-in-class small-molecule inhibitor of ASC that interferes with ASC speck formation. MM01 inhibition of ASC oligomerization prevents activation of procaspase-1 in vitro and inhibits the activation of different ASC-dependent inflammasomes in cell lines and primary cultures. Furthermore, MM01 inhibits inflammation in vivo in a mouse model of inflammasome-induced peritonitis. Overall, we highlight MM01 as a novel broad-spectrum inflammasome inhibitor for the potential treatment of multifactorial diseases involving the dysregulation of multiple inflammasomes

In vitro Anti-Proliferative Activity of the Rubia tinctorum and Alkanna tinctoria Root Extracts in Panel of Human Tumor Cell Lines hasson

Cancer is a devastating disease and is considered number one killer worldwide. Herbal formulations had played a key role over the past several decades in the development of anti-cancer drugs. Medicinal plants, which are endemic in Jordan, are known for several biological activities in particular their anti-cancer activity. However, the anti-cancer efficacy of the root extracts of Jordanian Rubia tinctorum and Alkanna tinctoria is not yet reported. To address this issue, this study assessed the anti-cancer activity of some root extracts obtained from Jordanian R. tinctorum and A. tinctoria in different tumor cell lines including the tongue, bladder, colon, gastric, lungs, breast, pancreas, and renal tissue origins by modified propidium iodide (PI) based monolayer assay. Among the tested root extracts obtained by different solvent systems, A. tinctoria in 100 % ethanol and methanol showed prominent anti-cancer activity against MDA-MB-231breast cancer cells (IC50: 2.98 µg/ml, IC70: 6.03 µg/ml), and CAL-27 tongue squamous carcinoma cells (IC50: 3.86 µg/ml, IC70: 5.97 µg/ml) respectively. Different solvent root extracts of R. tinctorum exhibited a similar trend of anti-tumor activity in both CAL-27 and MDA MB-231 cells. The anti-proliferative property of the extracts on CAL-27 and MDA-MB-231 cells is unclear. However, it can be concluded that the observed anti-cancer potential can be attributed to the phenolic compounds of the extracts as high polar solvents were used for extraction. The current study forms the rationale for isolating significant amount of anti-cancer active compounds from R. tinctorum and A. tinctori

Characterisation of metabolites of the putative cancer chemopreventive agent quercetin and their effect on cyclo-oxygenase activity

Quercetin (3,5,7,3′,4′-pentahydroxyflavone) is a flavone with putative ability to prevent cancer and cardiovascular diseases. Its metabolism was evaluated in rats and human. Rats received quercetin via the intravenous (i.v.) route and metabolites were isolated from the plasma, urine and bile. Analysis was by high-performance liquid chromatography and confirmation of species identity was achieved by mass spectrometry. Quercetin and isorhamnetin, the 3′-O-methyl analogue, were found in both the plasma and urine. In addition, several polar peaks were characterised as sulphated and glucuronidated conjugates of quercetin and isorhamnetin. Extension of the metabolism studies to a cancer patient who had received quercetin as an i.v. bolus showed that (Quercetin removed) isorhamnetin and quercetin 3′-O-sulphate were major plasma metabolites. As a catechol, quercetin can potentially be converted to a quinone and subsequently conjugated with glutathione (GSH). Oxidation of quercetin with mushroom tyrosinase in the presence of GSH furnished GSH conjugates of quercetin, two mono- and one bis-substituted conjugates. However, these species were not found in biomatrices in rats treated with quercetin. As cyclo-oxygenase-2 (COX-2) expression is mechanistically linked to carcinogenesis, we examined whether quercetin and its metabolites can inhibit COX-2 in a human colorectal cancer cell line (HCA-7). Isorhamnetin and its 4′-isomer tamarixetin were potent inhibitors, reflected in a 90% decrease in prostaglandin E-2 (PGE-2) levels, a marker of COX-2 activity. Quercetin was less effective, with a 50% decline. Quercetin 3- and 7-O-sulphate had no effect on PGE-2. The results indicate that quercetin may exert its pharmacological effects, at least in part, via its metabolites

The clerodane diterpene casearin J induces apoptosis of T-ALL cells through SERCA inhibition, oxidative stress, and interference with Notch1 signaling

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy that preferentially affects children and adolescents. Over 50% of human T-ALLs possess activating mutations of Notch1. The clerodane diterpene casearin J (CJ) is a natural product that inhibits the sarcoendoplasmatic reticulum calcium ATPase (SERCA) pump and induces cell death in leukemia cells, but the molecular mechanism of cytotoxicity remains poorly understood. Here we show that owing to SERCA pump inhibition, CJ induces depletion of the endoplasmic reticulum calcium pools, oxidative stress, and apoptosis via the intrinsic signaling pathway. Moreover, Notch1 signaling is reduced in T-ALL cells with auto-activating mutations in the HD-domain of Notch1, but not in cells that do not depend on Notch1 signaling. CJ also provoked a slight activation of NF-κB, and consistent with this notion a combined treatment of CJ and the NF-κB inhibitor parthenolide (Pt) led to a remarkable synergistic cell death in T-ALL cells. Altogether, our data support the concept that inhibition of the SERCA pump may be a novel strategy for the treatment of T-ALL with HD-domain-mutant Notch1 receptors and that additional treatment with the NF-κB inhibitor parthenolide may have further therapeutic benefits.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigaciones en Productos Naturales (CIPRONA)UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Químic

Flavonoid glucosides and dicaffeoylquinic acids from flowerheads of Buphthalmum salicifolium

From flowerheads of Buphthalmum salicifolium the new acylated flavonol glycosides quercetagetin 7-O-(6''-O-isobutyryl)-beta -glucoside, quercetagetin 7-O-(6''-O-isovaleryl)-beta -glucoside, quercetagetin 7-O-(6''-O-2-methylbutyryl)-beta -glucoside, patuletin 7-O-(6''-O-isobutyryl)-beta -glucoside and the new acylated caffeoylquinic acid deriv., 1-isobutyryl-3,5-dicaffeoylquinic acid were isolated, as well as ten common flavonoid glycosides together with three sesquiterpenes, 10-acetoxy-8,9-epoxythymolisobutyrate and S-(+)-dehydrovomifoliol. All structures were mainly established on the basis of UV-, MS- and NMR (1H, 13C, 1H-1H COSY, HMQC and HMBC) spectroscopic data

Determination of wound healing effect of caledula extracts using the scratch assy with 3T3 fibroblasts

Pharmacological relevance: Presentation of the scratch assay as a convenient and inexpensive in vitro tool to gain first insights in the wound healing potential of plant extracts and natural compounds.Aim of the study: The present study deals with the optimization of the scratch assay which can be used as an in vitro model for quantification of fibroblast migration to and proliferation into the wounded area. It is suitable for the first evaluation of the wound re-epithelialization potential of crude herbal extracts, isolated compounds and pharmaceutical preparations. As a proof of concept three preparations from traditional medicinal plants were investigated.Materials and methods: Swiss 3T3 albino mouse fibroblasts were used in monolayers and platelet derived growth factor as positive control. Hexane and ethanolic extracts from Calendula officinalis and Matricaria recutita, Hypericum oil as well as the triterpenoids faradiol myristate and palmitate were studied. To differentiate between proliferation and migration antimitotic mitomycin C was added.Results: Both extracts of Calendula officinalis stimulated proliferation and migration of fibroblasts at low concentrations, e.g. 10 μg/ml enhanced cell numbers by 64.35% and 70.53%, respectively. Inhibition of proliferation showed that this effect is mainly due to stimulation of migration. Faradiol myristate and palmitate gave comparable stimulation rates at an almost 50 μg/ml concentration, indicating that they contribute partially, but not most significantly to the wound healing effects of Calendula preparations. Extracts from Matricaria recutita were only moderately active. Hypericum oil was cytotoxic at concentrations higher than 0.5 μg/ml.Conclusions: The scratch assay in the present form can be used as a promising scientific approach and platform to differentiate between plant extracts known for their wound healing and their anti-inflammatory properties