Induced Cell Cytotoxicity Research Articles

Pterospermum acerifolium (L.) wild bark extract induces anticarcinogenic effect in human cancer cells through mitochondrial-mediated ROS generation.

Plants have many medicinal properties including anticancer activity due to the presence of several secondary metabolites. Current cancer treatment policies are not much effective because of side effects and resistance development. Therefore, the discovery of new phytotherapeutics with no or fewer side effects is highly needed. Pterospermum acerifolium (L.) wild, an angiosperm has a broad application in traditional Indian medicinal system including cancer treatment. Despite, there is no study available on the cytotoxic and apoptotic effect of P. acerifolium in human cancer cells. Exploring the medicinal properties of P. acerifolium plant by its traditional use will be helpful towards developing novel cancer therapeutics. Hence, we decided to demonstrate the anti-carcinogenic property of P. acerifolium ethanolic bark extract against lung (A549) and pancreatic (PANC-1) cancer cells. The cytotoxicity was demonstrated by MTT assay, morphological changes, and scratch invasion assay. Flow cytometry, fluorescence staining techniques, and cell cycle analysis were confirmed the apoptotic property of P. acerifolium plant. The cell viability assay revealed that P. acerifolium ethanolic bark extract significantly reduced the viability of both A549 and PANC-1 cells. Moreover, PANC-1 cells showed more sensitivity towards P. acerifolium ethanolic bark extract than A549 at higher concentrations. Clear visualization of changes such as cytoplasmic condensation, cellular morphology, cell shrinkage, and augmented number of dead cells in both the cancer cells was observed after treatment. Scratch and invasion assay showed that cell migration and invasion rate of both the cancer cells were significantly reduced. Fluorescence microscopic studies using acridine orange/ethidium bromide and DAPI (4', 6-diamidino-2-phenylindole) staining showed early and late apoptotic symptoms after treatment with bark extract. Rhodamine-123 and DCFH-DA staining analysis by fluorescence and flow cytometry showed that bark extract depolarized the mitochondria membrane potential and induced reactive oxygen species (ROS) generation. Cell cycle analysis through flow cytometry using propidium iodide stain showed that P. acerifolium bark extract arrested A549 and PANC-1 cells in sub-G1 phase stated early apoptosis. These findings collectively point to the fact that P. acerifolium bark extract induced cell cytotoxicity in lung and pancreatic cancer cells by modulating mitochondrial-mediated ROS generation, and cell cycle checkpoints.

Evaluation of phyto-medicinal efficacy of thymoquinone against Arsenic induced mitochondrial dysfunction and cytotoxicity in SH-SY5Y cells

BackgroundIt is evaluated that a few million individuals worldwide are experiencing Arsenic (As) harmfulness coming about because of anthropogenic discharges. There is likewise proof to propose that As can affect the peripheral, as well as, the central nervous system (CNS). On the contrary, thymoquinone (TQ), a biologically active ingredient of Nigella sativa has exhibited numerous neuro-pharmacological traits since ancient times. Hypothesis/PurposeIn the present study, the neuroprotective efficacy of TQ was explored by primarily studying its antioxidant and anti-apoptotic potential against Arsenic trioxide (As2O3) induced toxicity in SH-SY5Y human neuroblastoma cell lines. Study designFor experimentation, cells were seeded in 96 well tissue culture plates and kept undisturbed for 24 h to attain proper adhesion. After 75–80% confluence, cells were pretreated with 10 µM and 20 µM thymoquinone (TQ) for 1 h After adding 2 µM As, cells were set aside for incubation for 24 h without changing the medium. MethodsThe mitigatory effects of TQ with particular reference to cell viability and cytotoxicity, the generation of reactive oxygen species, DNA damage, and mitochondrial dynamics were studied. ResultsPretreatment of SH-SY5Y cells with TQ (10 and 20 μM) for an hour and subsequent exposure to 2 μM As2O3 protected the SH-SY5Y cells against the neuro-damaging effects of the latter. Also, the SH-SY5Y cells were better preserved with increased viability, repaired DNA, less free radical generation and balanced transmembrane potential than those exposed to As2O3 alone. TQ pretreatment also inhibited As2O3-induced exacerbation in protein levels of BAX and PARP-1 and restored the loss of Bcl2 levels. ConclusionThe findings of this study suggest that TQ may prevent neurotoxicity and As2O3-induced apoptosis and cytotoxicity. It is, therefore, worth studying further for its potential to reduce the risks of arsenic-related neurological implications.

Photo-modulation of zinc phthalocyanine-treated breast cancer cell line ZR-75-1 inhibited the normal tumor activity in vitro.

Regarding post-complication of convenient therapies against breast cancer, the emergence of effective approaches is essential. Photodynamic therapy is touted as a novel invasive therapeutic approach by the application of a photosensitizer promoted by laser irradiation. This study aimed to investigate the combined regime of low-level laser irradiation with zinc phthalocyanine in human breast cancer ZR-75-1 cell line. Cells were treated with 0.01 and 5μg/ml of ZnPc for 24h and exposed to radiation (70mW) for 60s. Cell viability was evaluated by MTT and flow cytometry. Cell migration capacity was monitored by scratch test, Transwell migration insert, and gelatin zymography. The function of MDR in treated cells was examined by Rhodamine 123 exclusion test. The level of GALNT11 was measured by ELISA. The expression of Bax and Bcl-2 genes was evaluated by real-time PCR. Laser irradiation and zinc phthalocyanine induced cell cytotoxicity in a dose-dependent manner. Flow cytometry analysis showed the induction of apoptotic and necrotic changes in treated cells. We found a reduction in migration rate and MMP-9 activity in cells undergoing the experimental procedure (p < 0.05). Immunofluorescence imaging revealed the intracellular accumulation of Rhodamine 123 coincided with a reduction in the level of GALNT11 in treated cells, showing the reduction of MDR activity and tumor cell resistance. Similar to flow cytometry assay, the reduction of Bcl-2 (approximately twofold) and upregulation of Bax genes were found in treated cells. Photodynamic therapy could be as an effective and alternative method for the treatment of breast cancer in a human.

Degradation of fibrin-β amyloid co-aggregate: A novel function attributed to ubiquitin

Human placental extract contains numerous bioactive components that are effective wound healing, antimicrobial and anti-inflammatory agents. During our investigation on the therapeutic potency of human placental extract, we have purified ubiquitin-like molecules that showed strong fibrino(geno)lytic activity. Further investigation confirmed similar potency of ubiquitin purified from adult human erythrocyte. Additionally, ubiquitin efficiently degraded disordered amyloid β 42 peptide (Aβ42) aggregate and fibrin-Aβ42 co-aggregate in vitro and reduced co-aggregate induced cytotoxicity in SH-SY5Y human neuroblastoma cells as compared to plasmin. Ubiquitin also degraded abnormal co-aggregates of fibrin with other plasma proteins such as fibronectin, albumin, lysozyme, tranthyretin and α-synuclein. To elucidate the mechanism of degradation, synthetic peptides (ADG, GKT, DQQ, QRL, LIF, AGK, HLVL) derived from ubiquitin template as well as synthetic ubiquitin (8565.32 Da) were employed. Synthetic ubiquitin completely degraded preformed Aβ 42 aggregate and fibrin-Aβ42 co-aggregate, whereas, the smaller synthetic peptides showed varying degrees of degradation. These observations attribute a novel function of ubiquitin that may be used for degrading abnormal fibrin clots in human body. Thorough investigation might unfold a novel molecular mechanism of ubiquitin in protein homeostasis.

Cytotoxic effect of iron nanoparticles in vitro on some cell lines

Aim: Cancer nanotechnology offers great potential for cancer diagnosis, targeted treatment, and monitoring, Among the rapidly evolving types of NPs, magnetic NPs (MgNPs) – biocompatible and superparamagnetic nanomaterials with chemical stability and low toxicity – are especially promising. the biosafety of this material needs to be estimated. the aim of this study evaluate the cytotoxic effect with MgNPs-Fe3O4 on some cancer cell growth in vitro. Material and method: Six concentrations of iron oxide nanoparticles(IONP) [(100,200,400,600,800and1000 ) μg/ml] were prepared and tested on Hela, RD and ANM3 cancer cell line in compare with REF cell line as a normal control for( 24 and 48) hr. nine replicates for each concentration ,The optical density of cell growth read by Elisa reader at 500nm, MTT colorimetric assay was employed to estimate the percentage of viable cells after each treatment. Results: All tumor cell lines (RD, HeLa, and AMN3) demonstrated significant reduction in the values of OD and GI% after 24 and 48 hr exposure to all concentrations of iron oxide NPs, while those of Ref cell line revealed non-significant elevation, when correlated with control group .Conclusion: IONP(20-30nm) induced cell cytotoxicity at all concentration when exposed to (Hela ,RD ,and AMN3) cancer cell lines when compared with REF normal cell line.

Corrigendum: Furanodiene Induces Extrinsic and Intrinsic Apoptosis in Doxorubicin-Resistant MCF-7 Breast Cancer Cells via NF-κB-Independent Mechanism.

[This corrects the article on p. 648 in vol. 8, PMID: 28959205.].

Furanodiene Induces Extrinsic and Intrinsic Apoptosis in Doxorubicin-Resistant MCF-7 Breast Cancer Cells via NF-κB-Independent Mechanism.

Chemotherapy is used as a primary approach in cancer treatment after routine surgery. However, chemo-resistance tends to occur when chemotherapy is used clinically, resulting in poor prognosis and recurrence. Currently, Chinese medicine may provide insight into the design of new therapies to overcome chemo-resistance. Furanodiene, as a heat-sensitive sesquiterpene, is isolated from the essential oil of Rhizoma Curcumae. Even though mounting evidence claiming that furanodiene possesses anti-cancer activities in various types of cancers, the underlying mechanisms against chemo-resistant cancer are not fully clear. Our study found that furanodiene could display anti-cancer effects by inhibiting cell viability, inducing cell cytotoxicity, and suppressing cell proliferation in doxorubicin-resistant MCF-7 breast cancer cells. Furthermore, furanodiene preferentially causes apoptosis by interfering with intrinsic/extrinsic-dependent and NF-κB-independent pathways in doxorubicin-resistant MCF-7 cells. These observations also prompt that furanodiene may be developed as a promising natural product for multidrug-resistant cancer therapy in the future.

NEUROPROTECTIVE EFFECT OF TERMINALIA CHEBULA EXTRACTS AND ELLAGIC ACID IN PC12 CELLS.

Background:Alzheimer’s disease (AD) is one of the common neurodegenerative disorders among elderly. The purpose of this study was to determine the neuroprotective effect and mechanisms of action underlying the Terminalia chebula extracts and ellagic acid by using beta-amyloid25-35 (Aβ25-35)-induced cell toxicity in an undifferentiated pheochromocytoma (PC12) cell line.Materials and Methods:The T. chebula extracts were prepared using the methanol, water, and 95% ethanol. Specifically, the ellagic acid was obtained in our laboratory. Assays including cell toxicity and changes in intracellular reactive oxygen species (ROS) and calcium level were evaluated to examine the neuroprotective effects and mechanisms of the T. chebula extracts and ellagic acid.Results:The methanolic and water extracts of T. chebula and ellagic acid exhibited the strongest neuroprotective activity against Aβ25-35-induced PC12 cell damages at 0.5–5.0 μg/ml. The ellagic acid also exhibited partial neuroprotective activity against H2O2-induced PC12 cell damages at 0.5–5.0 μg/ml. The methanolic and water extracts of T. chebula and ellagic acid protected PC12 cells from Aβ25-35-mediated cell damages and enhanced cell viability thorough two key mechanisms by: (1) inhibiting ROS production and (2) reducing calcium ion influx.Conclusion:The T. chebula represents a promising plant-source as medicine in the application for the treatment of AD. Further investigation focusing on the active component of T. chebula extracts e.g., ellagic acid is crucial to verify the neuroprotective efficacy and mechanisms in vivo.

The Effects of Frankincense on Oral Squamous Cell Carcinoma Cell Line

Background: Cytotoxic effects of Frankincense resin have been shown on some cancer cell lines. Due to its low side effects, this study was designed to evaluate the anticancer properties of water soluble elements of Frankincense oleo-gum-resin on human oral squamous cell carcinoma cell line. Methods: Oleo-gum-resin was macerated in ethanol. After filtration, the water soluble fraction of dried residue was extracted. KB cells were treated with 0, 62.5, 125, 250 and 500 μg/mL concentrations of obtained Frankincense aqueous fractions and with Doxorubicin as positive control. Frankincense induced cell cytotoxicity; apoptosis and proliferation were investigated using WST assay, Annexin V-FITC/PI, and Ki-67 staining, respectively. Data were analyzed using Kruskal-Wallis test by SPSS 17 software. Results: IC50 of 137.21 μg/mL was obtained from Frankincense aqueous fraction after 48 hours. The percentage of apoptotic cells was elevated in a time- and dose-dependent manner. There was no statistical difference in the Ki-67 expression of KB cells, using different concentration of Frankincense aqueous fraction after 24 and 48 hours (P = 0.083). Doxorubicin inhibited cells growth essentially through apoptosis. Conclusions: Frankincense aqueous fractions seem to suppress KB cell growth through the induction of apoptosis and necrosis rather than the inhibition of proliferation and hence might be a potential anticancer agent. Structural analysis and purification of potent components are suggested for determining more definitive results.

Antiplatelet Activity of a Newly Synthesized Novel Ruthenium (II): A Potential Role for Akt/JNK Signaling.

In oncotherapy, ruthenium complexes are considered as potential alternatives for platinum compounds, and have been proved as promising anticancer drugs with high efficacy and lesser side effects. Platelet activation plays a major role in cancer metastasis and progression. Hence, this study explored the effect of a newly synthesized ruthenium complex, [Ru(η6-cymene)(L)Cl]BF4(TQ5), where L = 4-phenyl-2-pyridin-2-yl-quinazoline), on human platelet activation. TQ5 (3–5 µM) inhibited concentration-dependent collagen-induced platelet aggregation in washed human platelets. However, this compound only inhibited platelet aggregation at a maximum concentration of 500 and 100 µM against thrombin and 9,11-dideoxy-11α, 9α-epoxymethanoprostaglandin (U46619)-induced stimulation, respectively. TQ5 inhibited collagen-induced ATP release and calcium mobilization ([Ca2+]i), without inducing cell cytotoxicity. In addition, neither SQ22536, an adenylate cyclase inhibitor, nor 1H-[1,2,4] oxadiazolo [4,3-a]quinoxalin-1-one (ODQ), a guanylate cyclase inhibitor, significantly reversed the TQ5-mediated inhibition of platelet aggregation. TQ5 inhibited the collagen-induced phosphorylation of protein kinase B (Akt) and c-Jun N-terminal kinase (JNK), but did not effectively inhibit extracellular signal-regulated kinase 1/2 (ERK1/2) and p38-mitogen-activated protein kinase (p38-MAPK) in human platelets. Additionally, TQ5 significantly prolonged the closure time in whole blood and increased the occlusion time of thrombotic platelet plug formation in mice. This study demonstrates, for the first time, that a newly synthesized ruthenium complex, TQ5, exhibits potent antiplatelet activity by hindering ATP release and [Ca2+]i, and by decreasing the activation of Akt/JNK signals. Together, these results suggest that TQ5 could be developed as a therapeutic agent that helps prevent or treat thromboembolic disorders, since it is found to be potently more effective than a well-established antithrombotic aspirin.

Novel endodontic sealers induce cell cytotoxicity and apoptosis in a dose-dependent behavior and favorable response in mice subcutaneous tissue.

The objective of the present study is to evaluate the in vitro cytotoxicity and in vivo biocompatibility of two novel endodontic sealers: RealSeal XT1 and Sealapex Xpress on the subcutaneous connective tissue of mice. The cytotoxicity was assessed by cell viability using the MTT assay (one-way ANOVA), trypan blue test (Mann-Whitney) and cell apoptosis by flow cytometer. For the subcutaneous study, polyethylene tubes filled with the sealers were implanted in 70 BALB/c mice: 6 experimental groups (n=10/group) and 2 control groups with empty tubes (n=5/group). At the end of experimental periods (7, 21, and 63days), the tissue was removed and histotechnically processed. Angioblastic proliferation and edema (Fisher's exact test) were evaluated, besides thickness measurement (μm) of the reactionary granulomatous tissue and neutrophil counts (Kruskal-Wallis and Dunn's post test; Mann-Whitney) (α=0.05). MTT assay, trypan blue, and analysis of apoptotic cells showed a dose-dependent direct effect: the more diluted the sealer, the less cytotoxic. Regarding the angioblastic proliferation and edema, difference between the sealers at 7 and 63days occurred (p<0.05). Both endodontic sealers initially promoted perimaterial tissue reaction as a foreign body granuloma and thus stimulated favorable tissue responses. Both sealers showed a dose-dependent effect and promoted satisfactory subcutaneous tissue response; the sealer Sealapex Xpress was less cytotoxic and more biocompatible than RealSeal XT. The step of root canal filling during endodontic treatment is highly important for the preservation of the periapical tissue integrity. Subcutaneous reaction to endodontic sealers enables scientific basis for clinical use.

Antioxidant Activity and Cytotoxicity of Taraxacum Hispanicum Aqueous and Ethanolic Extracts on Hepg2 Cells

Introduction: Plants belonging to the genus Taraxacum have been used in traditional medicine. Nowadays, extracts of these plants have been reported for the treatment of diseases, including liver disorders. Increasing interest and research on these plants also revealed its potential for treating cancer. This study aims to evaluate the antioxidant activity and cytotoxic properties of crude extracts from aerial parts of Taraxacum hispanicum H.Lindb, against human hepatocarcinoma (HepG2). Material and methods: Evaluation of the antioxidant properties was performed using DPPH in vitro test, superoxide scavenging assay and Fe2+ chelating activity. MTT assay was used to determine metabolic activity, for 24 and 48 hours. Results: For antioxidant capacity of the ethanolic extract (overall the one with the best results), IC50 values were 62.4 ± 6.7 μg/ml (DPPH radical scavenging activity) 53.9 ± 10.3 μg/ml (Fe2+ chelating activity) and 2.0 ± 0.3 μg/ml (superoxide scavenging assay). The aqueous and ethanolic extracts had different effects on HepG2 cell viability. Aqueous extract induced cell cytotoxicity in a time and dose-dependent manner, leaving only 52.6% viable cells at a concentration of 200 μg/ml, after 48 h. An increase in the cell viability was seen in the ethanolic extract, from 24 h to 48 h at higher concentrations. Conclusions: Ethanolic extract of T. hispanicum was the most promising, presenting anti-oxidative capacity and only the aqueous extract of the plant presented more relevant cytotoxicity over HepG2 cell line. These activities may be related with the extract phenolic content. However, further studies are needed to elucidate the main mechanisms responsible for these potential effects.

Beauvericin, a cyclic peptide, inhibits inflammatory responses in macrophages by inhibiting the NF-κB pathway.

Beauvericin (BEA), a cyclic hexadepsipeptide produced by the fungus Beauveria bassiana, is known to have anti-cancer, anti-inflammatory, and anti-microbial actions. However, how BEA suppresses macrophage-induced inflammatory responses has not been fully elucidated. In this study, we explored the anti-inflammatory properties of BEA and the underlying molecular mechanisms using lipopolysaccharide (LPS)-treated macrophage-like RAW264.7 cells. Levels of nitric oxide (NO), mRNA levels of transcription factors and the inflammatory genes inducible NO synthase (iNOS) and interleukin (IL)-1, and protein levels of activated intracellular signaling molecules were determined by Griess assay, semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR), luciferase reporter gene assay, and immunoblotting analysis. BEA dose-dependently blocked the production of NO in LPS-treated RAW264.7 cells without inducing cell cytotoxicity. BEA also prevented LPS-triggered morphological changes. This compound significantly inhibited nuclear translocation of the NF-κB subunits p65 and p50. Luciferase reporter gene assays demonstrated that BEA suppresses MyD88-dependent NF-κB activation. By analyzing upstream signaling events for NF-κB activation and overexpressing Src and Syk, these two enzymes were revealed to be targets of BEA. Together, these results suggest that BEA suppresses NF-κB-dependent inflammatory responses by suppressing both Src and Syk.

Epigallocatechin-3-Gallate Upregulates miR-221 to Inhibit Osteopontin-Dependent Hepatic Fibrosis.

Osteopontin (OPN) promotes hepatic fibrosis, and developing therapies targeting OPN expression in settings of hepatic injury holds promise. The polyphenol epigallocatechin-3-gallate (EGCG), found in high concentrations in green tea, downregulates OPN expression through OPN mRNA degradation, but the mechanism is unknown. Previous work has shown that microRNAs can decrease OPN mRNA levels, and other studies have shown that EGCG modulates the expression of multiple microRNAs. In our study, we first demonstrated that OPN induces hepatic stellate cells to transform into an activated state. We then identified three microRNAs which target OPN mRNA: miR-181a, miR-10b, and miR-221. In vitro results show that EGCG upregulates all three microRNAs, and all three microRNAs are capable of down regulating OPN mRNA when administered alone. Interestingly, only miR-221 is necessary for EGCG-mediated OPN mRNA degradation and miR-221 inhibition reduces the effects of EGCG on cell function. In vivo experiments show that thioacetamide (TAA)-induced cell cytotoxicity upregulates OPN expression; treatment with EGCG blocks the effects of TAA. Furthermore, chronic treatment of EGCG in vivo upregulates all three microRNAs equally, suggesting that in more chronic treatment all three microRNAs are involved in modulating OPN expression. We conclude that in in vitro and in vivo models of TAA-induced hepatic fibrosis, EGCG inhibits OPN-dependent injury and fibrosis. EGCG works primarily by upregulating miR-221 to accelerate OPN degradation. EGCG may therefore have utility as a protective agent in settings of liver injury.

Combination Therapy with Bortezomib or Carfilzomib and Selinexor Induces Nuclear Localization of Ikbα and Overcomes Acquired Proteasome Inhibitor Resistance in Human Multiple Myeloma

Introduction: Acquired proteasome-inhibitor (PI) resistance is a major obstacle in the treatment of multiple myeloma (MM). We investigated whether the clinical XPO1-inhibitor selinexor, when combined with bortezomib or carfilzomib, could overcome acquired-resistance in MM.Materials and Methods: PI-resistant myeloma cell lines, RPMI8226-B25 and U226 PSR, and their respective parental cell lines RPMI8226 and U266, were treated both in vitro with selinexor/bortezomib or selinexor/carfilzomib and assayed for apoptosis. In vivo studies using U266 and U266PSR tumors were performed in NOD/SCID-gamma (NSG) mice. Mice were treated with selinexor/bortezomib and single agents. Bone marrow biopsies from refractory myeloma patients were treated ex vivo with selinexor/bortezomib or selinexor/carfilzomib and assayed for apoptosis. Mechanistic studies included NFkB pathway protein expression assays, immunofluorescence microscopy, ImageStream flow-cytometry and proximity-ligation assay. IkBα knockdown and NFkB transcriptional activity were measured in selinexor/bortezomib treated MM cells.Results: We found that selinexor restored sensitivity of PI-resistant RPMI8226-B25 and U266PSR MM cells to bortezomib (P = 0.00055) and carfilzomib (P = 0.0017). Bortezomib, when combined with selinexor reduced U266 MM tumor growth versus single-agent bortezomib (P = 0.022) in NSG mice. NSG mice challenged with PI-resistant U266PSR MM tumors also had reduced tumor growth with selinexor/bortezomib as compared to single agent bortezomib (P = 0.0006). Combining bortezomib and selinexor improved survival in mice with U266 MM tumors (P = 0.0072) and PI-resistant U266PSR when compared to single-agent bortezomib (P = 0.0072). Myeloma cells from PI-refractory MM patients (n=14) were sensitized by selinexor to bortezomib (P = 0.002) and carfilzomib (P = 0.001) without affecting non-myeloma cells. Immunofluorescence microscopy of PI-resistant human MM cell lines found a greater than 212% increase in IkBα when compared to untreated cells (confirmed by Western blot). A similar increase in IkBα immunofluorescence was found in newly diagnosed, relapsed and refractory patient MM cells. ImageStream analyses of MM cells showed an increase in total and nuclear IkBα from selinexor/bortezomib exposure. Proximity-ligation assays showed that IkBα-NFkB-complexes were increased 12-fold in bortezomib/selinexor treated MM cells. IkBα knockdown abrogated selinexor/bortezomib induced cytotoxicity in MM cells. Selinexor/bortezomib treatment decreased NFkB transcriptional activity in addition to a reduction of NFkB induced IAP-1, IAP-2, BCL-2, cyclin D2 and c-myc protein expression..Conclusions: Selinexor, when used with bortezomib or carfilzomib has the potential to overcome proteasome-inhibitor drug-resistance in MM. Sensitization may be due to inactivation of the NFkB pathway by IkBα. Selinexor, an orally active selective inhibitor of XPO1-mediated nuclear export (SINE), is currently undergoing phase I/II studies in a variety of indications, including a combination with carfilzomib, in both relapsed and refractory MM patients (NCT02199665). The results presented in this study support combinatorial clinical trials in relapsed and refractory MM that utilize PI therapies. DisclosuresKashyap:Karyopharm Therapeutics: Employment, Equity Ownership. Shain:Takeda/Millennium: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Signal Genetics: Research Funding; Novartis: Speakers Bureau; Amgen/Onyx: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Landesman:Karyopharm Therapeutics Inc: Employment, Other: stockholder.

Green tea polyphenols affect invasiveness of human gastric MKN-28 cells by inhibition of LPS or TNF-α induced Matrix Metalloproteinase-9/2

Several studies demonstrated a correlation between green tea consumption and a reduced cancer risk. Among different components, green tea polyphenols have been identified as molecules responsible for the beneficial effects showed by the green tea against oxidative stress and cell invasiveness. In this study, we investigated the effects of green tea polyphenol extracts (GTPs) in human gastric MKN-28 cell line. To this aim, we have first evaluated the effect of GTPs on oxidative stress induced cell injury. The pre-treatment with 10−4 M catechin equivalents of GTPs exerts a protective effect on xanthine–xanthine oxidase induced cell cytotoxicity, thus confirming the anti-oxidant properties of GTPs. The effect of GTPs was also extended to the invasive ability of MKN-28 cells stimulated with TNF-α or LPS, as pro-inflammatory factors. Migration and matrigel invasion assays demonstrated that GTPs exposure (10−6 M) prevents the increase in cell invasiveness induced by TNF-α or LPS. Finally, we have analyzed the effect of GTPs on the levels of Matrix Metalloproteinases (MMP)-9/2, whose expression is up-regulated by TNF-α or LPS.Our results indicated that the pre-treatment with GTPs was able to reduce MMP-9/2 expression at both protein and enzyme activity levels in the conditioned media of TNF-α or LPS stimulated MKN-28 cells.In conclusion, our results demonstrated that green tea polyphenol extract reduces the invasiveness of gastric MKN-28 cancer cells through the reduction of TNF-α or LPS induced MMP-9/2 up-regulation. Therefore, these data support the hypothesis that GTPs could exert a protective role against the metastatic process in gastric cancer.

Polyphenol‑rich extract of Aronia melanocarpa inhibits TNF‑α induced apoptosis in H9c2 cells

Introduction. In certain pathological states within cardiovascular system, cardiomyocytes may exhibit overexpression of TNF‑α that results in induction of myocardial oxidative stress and cardiac cells apoptosis. Thus, they may participate in development and progression of post‑infarct congestive heart failure. The aim of this study was to evaluate the effect of polyphenol‑rich Aronia melanocarpa extract (AME) on TNF‑α induced apoptosis in cardiomyoblast H9c2 cells. Material and Methods. Apoptosis was measured in H9c2 cells preincubated with increasing concentrations of commercial extract of aronia – Aronox (10–50 μg/mL) for 24h and then treated with TNF‑α: 100 ng/mL for 24h as well. The MTT assay was used to determine cardiomyoblasts viability. Alteration of the mitochondrial membrane potential, specific for apoptotic cells, was evaluated with caspase-3 activity assay kit. Results. Our results showed that AME significantly inhibited TNF‑α induced apoptosis (IC50 = 55.84 µg/mL) and cytotoxicity in H9c2 cells. Significant inhibition of apoptosis was observed in all tested concentrations of AME. The highest anti‑apoptotic and cytoprotective effect was observed at the highest concentration (50 μg/mL), while in lower the concentrations cytoprotective effect was statistically insignificant. Conclusions. Polyphenol‑rich AME exhibits anti‑apoptotic and cytoprotective effect in H9c2 cardiomyoblasts treated with TNF‑α. Further studies are required in context of its possible application in prevention and/or therapy of cardiovascular diseases.

Effects of combination of melatonin and laser irradiation on ovarian cancer cells and endothelial lineage viability.

The main goal of anti-cancer therapeutic approaches is to induce apoptosis in tumor masses but not in the normal tissues. Nevertheless, the combination of photodynamic irradiation with complementary oncostatic agents contributes to better therapeutic performance. Here, we applied two different cell lines; SKOV3 ovarian carcinoma cells and HUVECs umbilical cord cells as in vitro models to pinpoint whether pharmacological concentration of melatonin in combination with photodynamic therapy induces cell cytotoxicity. The cells were separately treated with various concentrations of melatonin (0 to 10mM) and photodynamic irradiation alone or in combination. Cells were preliminary exposed to increasing concentrations of melatonin for 24h and subsequently underwent laser irradiation for 60s with an output power of 80mW in continuous mode at 675nm wavelength and a total light dose of 13.22J/cm2. Cell viability, apoptosis/necrosis rates, and reactive oxygen species levels as well as heat shock protein 70 expression were monitored after single and combined treatments. A statistical analysis was performed by applying one-way analysis of variance (ANOVA) and post hoc Tukey's test. Combination treatment of both cell lines caused a marked increase in apoptosis/necrosis rate, reactive oxygen species generation, and heat shock protein 70 expression compared to incubation of the cells with each agent alone (p < 0.05). SKOV3 cancer cells expressed higher level of heat shock protein 70 under experimental procedure as compared to HUVECs (p < 0.05). Our results introduce melatonin as a potent stimulus for enhancing the efficacy of laser on induction of apoptosis in tumor cells.

Evaluations of thyme extract effects in human normal bronchial and tracheal epithelial cell lines and in human lung cancer cell line

Thyme (Thymus vulgaris) is used traditionally to prepare herbal remedies possessing expectorant, mucolytic, antitussive and antispasmodic properties.The aim of the present study was to investigate the effects of a standardized hydroalcoholic extract of thyme on primary human airway (bronchial/tracheal) epithelial cell lines in a model of lung inflammation induced by LPS. In addition, the effects of thyme extract on human lung cancer cell line (H460) were analysed. Thyme extract showed significant anti-inflammatory properties by reducing the NF-κB p65 and NF-κB p52 transcription factors protein levels followed by the decrease of pro-inflammatory cytokines (IL-1 beta and IL-8), and Muc5ac secretion in human normal bronchial and tracheal epithelial cells. Moreover, the extract showed cytotoxic effects on H460 cancer cells, modulated the release of IL-1 beta, IL-8 and down-regulated NF-κB p65 and NF-κB p52 proteins.Taken together, these results substantiated the traditional uses of thyme in the treatment of respiratory diseases. Thyme extract might be an effective treatment of chronic diseases based on inflammatory processes when hypersecretion of mucus overwhelms the ciliary clearance and obstructs airways, causing morbidity and mortality. Moreover thyme extract, evaluated in H460 lung cancer cell line, demonstrated to induce cell cytotoxicity in addition to reduce inflammatory cell signals.

Follicular lymphoma: in vitro effects of combining lymphokine-activated killer (LAK) cell-induced cytotoxicity and rituximab- and obinutuzumab-dependent cellular cytotoxicity (ADCC) activity.

Follicular lymphoma (FL) is a disease of paradoxes-incurable but with a long natural history. We hypothesized that a combination of lymphokine-activated killer (LAK) cells and monoclonal antibodies might provide a robust synergistic treatment and tested this hypothesis in a phase II clinical trial (NCT01329354). In this trial, in addition to R-CHOP, we alternated the administration of only rituximab with rituximab and autologous LAK cells that were expanded ex vivo. Our objective was to determine the in vitro capability of LAK cells generated from FL patients to produce cytotoxicity against tumor cell lines and to determine rituximab- and obinutuzumab-induced cytotoxicity via antibody-dependent cellular cytotoxicity (ADCC) activity. We analyzed the LAK cell-induced cytotoxicity and rituximab (R)- and obinutuzumab (GA101)-induced ADCC activity. We show that LAK cells generated from FL patients induce cytotoxicity against tumor cell lines. R and GA101 enhance cytolysis through ADCC activity of LAK cells. Impaired LAK cell cytotoxicity and ADCC activity were detected in 50 % of patients. Percentage of NK cells in LAK infusions were correlated with the R- and GA101-induced ADCC. Our results indicate that the combination of R or GA101 and LAK cells should be an option as frontline maintenance therapy in patients with FL.