Hammada scoparia flavonoids and rutin kill adherent and chemoresistant leukemic cells

doi:10.1016/j.leukres.2010.12.011

Leukemia Research

Volume 35, Issue 8, August 2011, Pages 1093-1101

https://doi.org/10.1016/j.leukres.2010.12.011 Get rights and content

Abstract

In search for compounds able to reduce cell adhesion-mediated drug resistance (CAM-DR), we studied effects of Hammada scoparia extracts on leukemic cells adherent or in suspension. We show that H. scoparia flavonoidic fraction and its compound rutin induce apoptosis specifically in adherent leukemic cells and abolish CAM-DR. Importantly, rutin inhibited survival of adherent leukemic progenitors (CD34⁺38⁻123⁺) but spared normal progenitors (CD34⁺38⁻). The pro-apoptotic effects of rutin were correlated with a decrease of active GSK3β and inhibitors of GSK3β reproduced rutin-induced cytotoxicity. This study uncovers the potential of H. scoparia flavonoids and rutin to overcome CAM-DR in acute myeloid leukemia.

Introduction

Acute myeloid leukemia (AML) is characterized by the aberrant accumulation of immature myeloid hematopoietic cells. The leukemia-initiating cells are hematopoietic stem cells which have either dysregulated self-renewal and differentiation functions or have committed progenitors transformed to leukemic stem cells (LSCs). Current chemotherapy for AML uses drugs such as nucleoside analogs and anthracyclines that interfere with DNA replication and induce apoptosis primarily in replicating cells. However, since LSCs are mostly quiescent like hematopoietic stem cells (HSCs) [1], relapse is common after initial remission in response to chemotherapy.

The hematopoietic niche plays a key role in maintaining HSCs quiescence. Adhesive interactions between HSCs and their microenvironment are required to maintain stem cell capacities. We and others have demonstrated that adhesive interactions, notably through integrin engagement on fibronectin, are responsible for cell adhesion-mediated drug resistance (CAM-DR) of leukemic cells [2]. Current research is focussing on determining specific cell survival mechanisms that distinguish LSCs from HSCs in order to develop LSC-specific targeted therapy.

Nuclear factor κB (NF-κB) and mammalian target of rapamycin (mTOR) were found overactivated in LSCs and have been proposed as targets for LSCs therapy [3], [4]. The phosphoinositide 3-kinase/Akt pathway regulates NF-κB and mTOR activities, and has been shown to mediate CAM-DR of AML [5]. Another key signaling molecule controlling the NF-κB and mTOR pathways is the glycogen synthase kinase 3β (GSK3β), a serine threonine kinase important for survival responses to various extracellular stress from the microenvironment [6]. Interestingly, GSK3β activation is linked to a cell quiescence status and regulates specifically NF-κB allowing the transcription of genes such as IL-6, a cytokine involved in the survival of malignant hematopoietic cells [7].

We have previously demonstrated that GSK3β activation supports CAM-DR [2] and resistance to TNF-α [8] of leukemic blasts. Having obtained preliminary results showing that aqueous extracts from Hammada scoparia had anti-inflammatory properties correlated with GSK3β downregulation in a rat model of hepatotoxicity (Bourogaa et al., in preparation), we raised the question of putative modulation of leukemic cell survival by H. scoparia compounds. H. scoparia is a medicinal plant whose anti-cancer, anti-inflammatory and anti-oxidant properties, as well as phytochemistry, are little known [9]. In this work, we tested the activity of H. scoparia extracts over leukemic or normal hematopoietic cells and identified one of the bioactive components as rutin.

Section snippets

Cells

Leukemic cell lines U937, HL-60, KG1 and KG1a were purchased from the German Collection of Microorganisms and Cell Cultures (Braunschweig, Germany) and grown as previously described [8]. AML cells were obtained from bone marrow of six patients with informed consent at diagnostic, using Ficoll-Hypaque density-gradient centrifugation (Collection No. 1 HIMIP AC-2008-129/DC-2008-307 approved by CCPRB, Toulouse University Hospital Dr. C. Demur). Bone marrow samples contained more than 80% leukemic

Aqueous extracts and flavonoid-enriched fractions from H. scoparia are cytotoxic for leukemic cells

Aqueous extracts were prepared from dried H. scoparia leaves and serial dilutions were assayed for cytotoxic effects on leukemic cells either in suspension or in adhesion. As shown in Fig. 1A, pretreatment of U937 leukemic cells with aqueous extract from H. scoparia (EtOH/H₂O 1:9, v/v) reduced survival of adherent U937 (30% decrease of survival). The survival of leukemic cells in suspension was slightly affected. The cytotoxic effects of DNR were strongly increased by the H. scoparia aqueous

Discussion

Natural plant extracts and their flavonoids have been shown to display anti-cancer properties both in vitro and in vivo. Clinical trials with flavonoids have been performed for their development as anti-leukemic agents [17], [18], [19], [20]. Here, we have demonstrated that H. scoparia flavonoids and rutin have the capacity to specifically induce the cytotoxicity of adherent and chemoresistant leukemic progenitors.

Cytotoxic assays with flavonoids glycosides present in H. scoparia, or

Conflict of interest

All authors have no conflict of interest to declare.

Acknowledgements

The authors would thank Drs M. Allouche, A. Gargouri and C. Cambon for helpful discussion along the realization of this work. The orthopedic service of CHU Purpan (Toulouse, France) and Pr. P. Bonnevialle are greatly acknowledged for providing human bone fragments. The authors would thank also the «plateforme de cytométrie» in IFR150.

Funding source: Association de Recherche contre le Cancer (contracts no. 3638 and 8407), Institut National du Cancer (contract no. 07/3D1616/IABC-23-8/NC-NG),

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Evaluation of anti-inflammatory, diuretic and oral glucose tolerance activities of phenolic compound-rich fractions of Hammada articulata in Albino Wistar rats
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Hammada articulata is a plant widely used by the locals of the Algerian Sahara for multiple medicinal purposes. However, little was known about its chemical composition and the mechanisms of its bioactivity. For this purpose, the derived extracts [chloroform (CHCl₃), ethyl acetate (EtOAc) and n-butanol (n-BuOH)] of the 80% ethanol extract of its aerial parts, were evaluated for their anti-inflammatory, diuretic, and anti-hyperglycemic activities in vivo. A preliminary phytochemical screening of H. articulata extracts showed the presence of a variety of secondary metabolites. RP-HPLC/DAD was used to analyze some fractions obtained by fractionation of the three derived extracts, by column chromatography and chosen because of the abundance and simplicity of their chemical composition. The fractions obtained from EtOAc and n-BuOH extracts showed a particular richness in phenolic compounds mainly naringenin, quercetin, kaempferol, myricetin, and rutin, which were known for their many interesting biological activities. The three derived extracts from H. articulata were assessed for their anti-inflammatory activity in the carrageenan-induced edema model in rats and their diuretic activity using hydrochlorothiazide (HCTZ) as a diuretic reference. All extracts showed considerable anti-inflammatory activity; the highest was registered in the group treated with the n-BuOH extract. However, for the diuretic activity, only the chloroform extract was active, with a diuretic spectrum similar to that of the standard diuretic HCTZ. The anti-hyperglycemic effect was carried out on the three derived extracts administered orally at a dose of 200 mg/kg, using the glucose tolerance test after gavage with the extracts. The EtOAc and n-BuOH extracts showed significant anti-hyperglycemic activity, improving oral glucose tolerance in normal rats.
Leonurus japonicus Houttuyn induces reactive oxygen species-mediated apoptosis via regulation of miR-19a-3p/PTEN/PI3K/AKT in U937 and THP-1 cells
2022, Journal of Ethnopharmacology
Citation Excerpt :
Up to 1.165 μg/mL of rutin which is equally in the 200 μg/mL of Leonurus japonicus Houttuyn, there was no cytotoxicity in U937, THP-1 cancer cells nor in MDBK, IEC-18 normal cells. The cytotoxic concentration of rutin in U937, HL-60, and KG cell lines was 20–160 μg/mL (Bourogaa et al., 2011). The cytotoxicity of Leonurus japonicus Houttuyn is not due to rutin its constituent.
Leonurus japonicus Houttuyn is a medicinal ingredient in more than 300 prescriptions in traditional Korean medicine. It is especially important for women's health and blood-related diseases. Recent research revealed that Leonurus japonicus Houttuyn extracts have antioxidative, anticancer, analgesic, anti-inflammatory, and neuroprotective properties.
However, its underlying anti-cancerous mechanisms remain unclear. This study elucidated the anticancer mechanism of Leonurus japonicus Houttuyn in U937 and THP-1 cancer cells.
High-performance liquid chromatography (HPLC) was used for detecting main compound of Leonurus japonicus Houttuyn, rutin. EZ-Cytox cell viability assay, Western blot analysis, live and dead cell assay, 2′, 7’ dichlorofluorescin diacetate (DCFDA) assay, quantitative real-time PCR (qRT-PCR) analysis, and microRNA (miR) mimic transfection assay were applied to further investigate anti-cancer efficacies and underlying mechanism in U937 and THP-1 cells.
The main compound of Leonurus japonicus Houttuyn, rutin was detected using HPLC. The cytotoxic effect of Leonurus japonicus Houttuyn was exerted in U937 and THP-1 cancer cells but not in MDBK and IEC-6 normal cells. Leonurus japonicus Houttuyn decreased mitochondria membrane potential (ΔΨm). Consistently, Leonurus japonicus Houttuyn reduced the expression of survivin and cleaved caspase-9, caspase-3, and poly (ADP-ribose) polymerase (PARP). Cell death was increased in Leonurus japonicus Houttuyn treated groups. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and CCAAT-enhancer-binding protein homologous protein (CHOP) was increased and phosphatidylinositol-3-kinase (PI3K) and Protein kinase B (AKT) were decreased by Leonurus japonicus Houttuyn. Reactive oxygen speices generation was elevated by Leonurus japonicus Houttuyn and its cytotoxicity was reversed by N-acetyl-l-cysteine (NAC) pretreatment. Moreover, onco-microRNA (miR), miR-19a-3p was suppressed by Leonurus japonicus Houttuyn and transfection of miR-19a-3p mimic reversed the regulated PTEN, p-AKT, CHOP expression, attenuating Leonurus japonicus Houttuyn induced apoptosis.
These findings indicated that Leonurus japonicus Houttuyn has anti-cancer effects by regulation of PTEN/PI3K/AKT signal pathway and ROS-related ER stress-induced apoptosis via regulation of miR-19a-3p. Leonurus japonicus Houttuyn may be an effective candidate for triggering PTEN-dependent apoptosis of cancer cells related to acute myeloid leukemia.
Hydroalcoholic leaves extract of Vaccinium ashei Reade promotes cell cycle arrest and apoptosis on T-cell acute lymphoblastic leukemia
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Vaccinium ashei Reade, popularly known as Rabbiteye blueberry, has several therapeutic properties attributed to the phenolic compounds present in its leaves and fruits. Here, we sought to evaluate the effects of the hydroalcoholic extract from V. ashei leaves (Bluegem cultivar, VAB) in T-cell Acute lymphoblastic leukemia (T-ALL). The VAB extract was toxic to T-ALL cells at the ∼60 µg/ml concentration. T-ALL cell death occurred through apoptosis. VAB extract was found to induce micronuclei formation, p53 pathway activation, and cell cycle arrest. Those mutagenic effects were evidenced through microscopy analysis and molecular p53 pathway activation. A series of phenolic compounds were identified in VAB extract by mass spectrometry, such as vanillic acid, catechin, caffeic acid, chlorogenic acid, rutin, coumaric acid, taxifolin, quercetin and naringenin, some of which are presumed to induce DNA damage. In conclusion, the V. ashei leaves extract may have important secondary metabolites with antileukemic properties.
Thermodynamic and structural description of relative solubility of the flavonoid rutin by DFT calculations and molecular dynamics simulations
2021, Journal of Molecular Liquids
Citation Excerpt :
Similarly, [22] found that rutin significantly decreased the migration of human A549 cells (lung cancer) [22]. These and other properties have attracted the attention of researchers for the rutin and other flavonoids [23–26]. The chemical structure of the rutin is shown in Fig. 1, together with the thirty-second rutin geometry of our recent conformational analysis, based on Density Functional Theory (DFT) [27] calculations of thermodynamic and spectroscopic properties [28].
Understanding the solubility of flavonoids is of relevance in view of the broad application such as antioxidant and antitumor agents. The solubility of flavonoid rutin has been experimentally investigated in a homologous series of alcohols, acetone, n-heptane and water solvents. In this work theoretical evaluation of Gibbs free energy of solvation was performed at the Density Functional Theory (DFT) and Molecular Dynamics (MD) levels, with the $ω$ B97x-D functional and multistate Bennett acceptance ratio (MBAR) method, respectively. The degree of solubility of a given solute among various solvents is dictated by solute-solvent interactions, which may be represented by the solvation energy. Our calculated relative energy values due to solvent effects, using quantum chemical methods (PCM and SMD solvent models) do not follow strictly the observed solubility trend. However, the PCM model including solute-solvent dispersion interaction, solute-solvent repulsion interaction and solute cavitation energy contribution to the total energy (named DIS,REP,CAV) yielded an overall agreement with experimental solubility profile, with exception of ethanol solvent. A similar satisfactory accordance with experiment was found for MBresults, with a discrepancy also found for 1-butanol, which deserve further investigation. Analysis of spatial and radial distribution functions (SDF and RDF) for pure solvents and the analysis of the contributions of vdW and electrostatic terms for the total free energy reveal that a balance between dispersive and electrostatic interactions between the rutin and the solvent is necessary to solubilize the flavonoid, which corroborates with the values of these energy contributions to the solvation-free energy calculated by the MBAR method. The use of explicit solvent molecules in DFT calculations is precluded due to the size, flexibility, and many sites for solvent interactions of rutin molecule. Our results indicate that in general DFT-PCM (DIS,REP,CAV) and MBAR levels of calculations can satisfactorily predict the solvation free energy and solubility trend for flavonoid rutin along a series of solvents, and also provide a reasonable estimate of relative solubilities of other flavonoids, information that can be relevant for further molecular modeling studies.
Rutin prevents cisplatin-induced ovarian damage via antioxidant activity and regulation of PTEN and FOXO3a phosphorylation in mouse model
2020, Reproductive Toxicology
Citation Excerpt :
These results can be related to the antioxidant effects of the rutin like inhibition of ROS generation, increase in the activity of antioxidant enzymes, such as glutathione (GSH), and prevention of lipid peroxidation, attenuating DNA and mitochondrial damage [21–23]. Rutin actions can be regulated by phosphatidylinositol 3-kinase (PI3K) signaling pathway members, which are important for the maintenance of cell survival and growth, such as phosphatase and tension homolog (PTEN) and Forkhead box O3 (FOXO3a) [24–27]. PTEN phosphorylation can effectively inhibit cell proliferation and induce apoptosis [28], while down-regulation of phosphorylated PTEN (p-PTEN) induces PI3K activation and consequent FOXO3a phosphorylation (p-FOXO3a), increasing its nuclear export, which promotes follicle activation and/or decreases apoptosis [29–31].
The aims of the present study were to evaluate the protective effects of rutin during cisplatin-induced ovarian toxicity in mice and to verify the possible involvement of the phosphatase and tension homolog (PTEN)/Forkhead box O3a (FOXO3a) pathway in the rutin actions. Mice received saline solution (control, 0.15 M, i.p.) or cisplatin (5 mg/Kg body weight, i.p.) or they were pretreated with N-acetylcysteine (positive control; 150 mg/Kg of body weight [p.o.]) or with rutin (10, 30 or 50 mg/Kg body weight, p.o.) before cisplatin (5 mg/Kg body weight, i.p.) once daily for 3 days. Next, the ovaries were harvested and destined to histological (follicular morphology and activation), immunohistochemical (cell proliferation and apoptosis) and fluorescence (reactive oxygen species [ROS], glutathione [GSH] and mitochondrial activity) analyses. Moreover, the expression of phosphorylated PTEN (p-PTEN) and FOXO3a (p-FOXO3a) were evaluated to investigate a molecular mechanism by which rutin would prevent the cisplatin-induced ovarian damage. The results showed that pretreatment with N-acetylcysteine or 10 mg/Kg rutin before cisplatin preserved the percentage of normal follicles and cell proliferation, reduced apoptosis and ROS levels and increased active mitochondria and GSH levels compared to the cisplatin treatment (P < 0.05). Cisplatin treatment increased p-PTEN and decreased p-FOXO3a expression in follicles, which was prevented by 10 mg/kg rutin. In conclusion, treatment with 10 mg/Kg rutin has the potential to protect the ovarian follicles against cisplatin-induced toxicity through its antioxidant effects and PTEN/FOXO3a pathway.
Rutin- serum albumin interaction in different media and its effective dose selection in radiation-induced cytotoxicity on human blood cells
2020, Journal of Herbal Medicine
Citation Excerpt :
DNA protective action of rutin has also been demonstrated (Marcarini et al., 2011). Rutin also has an inhibitory effect on the survival of leukemic progenitor cells (Bourogaa et al., 2011) and has been shown to inhibit inflammatory responses in UVB-irradiated mouse skin by inhibiting the increase in phosphorylated levels of STAT3; p38 MAPK and JNK (Choi et al., 2014; Perk et al., 2014). All these findings support the beneficial biological actions of rutin regarding its anticancer and anti-inflammatory responses with significant information about the signaling pathway involved.
Increases in general circulatory time, half-life and bioavailability increase the efficacy of any drug molecule. Rutin (3, 3′, 4′, 5, 7 -pentahydroxyflavone-3-rhamnoglucoside) is a flavonoid known for its ability to regulate cell metabolism, gene expression, and protect against oxidative stress, although certain biophysical aspects of their functioning are not yet clear. Non protein and small protein molecules with electrophilic properties are transported through the blood to their target site by binding with albumin. The authors therefore hypothesized that modification of rutin-albumin binding by changing the microenvironment in which it is delivered can increase the efficacy of the molecule. Therefore, the interaction of rutin in different solvent systems with albumin have been studied to evaluate the bioavailability and efficacy of the molecule to introduce it as a drug for combating UVB radiation-induced damage in biological systems. We evaluated the effect of rutin–albumin interaction in blood plasma in various modes of delivery (in DMSO, alcohol and PEG). The biological activity and efficacy of different solvents for rutin delivery and its bioavailability were studied using UVB irradiated human peripheral blood mononuclear cells in vitro. Amelioration of UVB radiation-induced oxidative damage and cell death by rutin has been studied by MTT assay and Tryphan blue exclusion assay and LDH-cytotoxicity assay. In these three studied systems the most effective concentration of rutin was also determined.

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Hammada scoparia flavonoids and rutin kill adherent and chemoresistant leukemic cells

Abstract

Introduction

Section snippets

Cells

Aqueous extracts and flavonoid-enriched fractions from H. scoparia are cytotoxic for leukemic cells

Discussion

Conflict of interest

Acknowledgements

Blood

Blood

Journal of the American Society for Mass Spectrometry

Blood

Food and Chemical Toxicology

Biochemical Pharmacology

Leukemia Research

Leukemia Research

Chemotherapy-resistant human AML stem cells home to and engraft within the bone-marrow endosteal region

Nature Biotechnology

A crosstalk between the Wnt and the adhesion-dependent signaling pathways governs the chemosensitivity of acute myeloid leukemia

Oncogene

Interaction between leukemic-cell VLA-4 and stromal fibronectin is a decisive factor for minimal residual disease of acute myelogenous leukemia

Nature Medicine

Requirement for glycogen synthase kinase-3beta in cell survival and NF-kappaB activation

Nature

Glycogen synthase kinase 3beta functions to specify gene-specific, NF-kappaB-dependent transcription

Molecular and Cellular Biology

A new α5β1 integrin-dependent survival pathway through GSK3β activation in leukemic cells

PLoS One

Flavonol triglycosides from the leaves of Hammada scoparia (POMEL) ILJIN

Chemical & Pharmaceutical Bulletin (Tokyo)

A new α₅β₁ integrin-dependent survival pathway through GSK3β activation in leukemic cells