Publications by authors named "Pei-Wen Hsieh"

79 Publications

Randialic acid B and tomentosolic acid block formyl peptide receptor 1 in human neutrophils and attenuate psoriasis-like inflammation in vivo.

Biochem Pharmacol 2021 May 6;190:114596. Epub 2021 May 6.

Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan 33302, Taiwan; Research Center for Chinese Herbal Medicine, Research Center for Food and Cosmetic Safety, and Graduate Institute of Health Industry Technology, College of Human Ecology, Chang Gung University of Science and Technology, Taoyuan 33302, Taiwan; Department of Anesthesiology, Chang Gung Memorial Hospital, Taoyuan 33305, Taiwan; Department of Chemical Engineering, Ming Chi University of Technology, New Taipei City 24301, Taiwan. Electronic address:

Psoriasis is a long-lasting inflammatory skin disease lacking proper cure. Dysregulated activation of neutrophils is a major pathogenic factor in psoriasis. Formyl peptide receptor 1 (FPR1) triggers neutrophil activation in response to bacteria- or mitochondria-derived N-formyl peptides, but its significance in neutrophilic psoriasis remains unknown. In this study, we discovered two derivatives of ursolic acid, 3β-hydroxyurs-12,18-dien-28-oic acid (randialic acid B, RAB) and 3β-hydroxyurs-12,19-dien-28-oic acid (tomentosolic acid, TA), as FPR1 inhibitors in human neutrophils with ability to suppress psoriatic symptoms in mice. Both RAB and TA, triterpenoids of traditional medicinal plant Ilex kaushue, selectively inhibited reactive oxygen species production, elastase release, and CD11b expression in human neutrophils activated by FPR1, but not non-FPR1 agonists. Importantly, RAB and TA inhibited the binding of N-formyl peptide to FPR1 in human neutrophils, neutrophil-like THP-1 cells, and hFPR1-transfected HEK293 cells, indicating FPR1 antagonism. Moreover, in assays induced by various concentrations of FPR1 agonist, both RAB and TA acted competitively for its binding to the FPR1 receptor. The FPR1-downstream signaling such as Ca mobilisation and activation of Akt and MAPKs was also competitively inhibited. In addition, imiquimod-induced psoriasis-like symptoms, including epidermal hyperplasia, desquamation with scaling, neutrophil skin infiltration, and transepidermal water loss were significantly reduced by both RAB and TA. The results illustrate a possible role of human neutrophils FPR1 receptor in psoriasis-like inflammation. Accordingly, triterpenoids RAB and TA represent novel FPR1 antagonists and exhibit therapeutic potential for treating neutrophilic inflammatory skin diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bcp.2021.114596DOI Listing
May 2021

Neutrophil elastase inhibitor (MPH-966) improves intestinal mucosal damage and gut microbiota in a mouse model of 5-fluorouracil-induced intestinal mucositis.

Biomed Pharmacother 2021 Feb 26;134:111152. Epub 2020 Dec 26.

Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan; Graduate Institute of Natural Products, School of Traditional Chinese Medicine, College of Medicine, Chang Gung University, Taoyuan, Taiwan; Department of Anesthesiology, Chang Gung Memorial Hospital, Linkou, Taiwan; Research Center for Chinese Herbal Medicine, Chang Gung University of Science and Technology, Taoyuan, Taiwan. Electronic address:

Background: 5-Fluorouracil (5-FU)-based chemotherapy is first-line chemotherapy for colorectal cancer. However, 5-FU-induced intestinal mucositis (FUIIM) is a common adverse effect that severely impairs drug tolerance and results in poor patient health.

Methods: Male C57BL/6 mice were given 5-FU (50 mg/kg/day, i.p.) and treated with MPH-966 (5 and 7.5 mg/kg/day, p.o.) for five days. The body weight loss and the amount of food intake, and histopathological findings were recorded and analyzed. In addition, the neutrophil infiltration, levels of neutrophil serine proteases and pro-inflammatory cytokines, and tight junction proteins expression in intestinal tissues were determined. The ecology of gut microbiota was performed through next-generation sequencing technologies.

Results: Neutrophil elastase (NE) overexpression is a key feature in FUIIM. This study showed that treatment with the specific NE inhibitor MPH-966 (7.5 mg/kg/day, p.o.) significantly reversed 5-FU-induced loss in body weight and food intake; reversed villous atrophy; significantly suppressed myeloperoxidase, NE, and proteinase 3 activity; and reduced pro-inflammatory cytokine expression in an FUIIM mouse model. In addition, MPH-966 prevented 5-FU-induced intestinal barrier dysfunction, as indicated by the modulated expression of the tight junction proteins zonula occludin-1 and occludin. MPH-966 also reversed 5-FU-induced changes in gut microbiota diversity and abundances, specifically the Firmicutes-to-Bacteroidetes ratio; Muribaculaceae, Ruminococcaceae, and Eggerthellaceae abundances at the family level; and Candidatus Arthromitus abundance at the genus level.

Conclusion: These data indicate that NE inhibitor is a key treatment candidate to alleviate FUIIM by regulating abnormal inflammatory responses, intestinal barrier dysfunction, and gut microbiota imbalance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.biopha.2020.111152DOI Listing
February 2021

Anti-inflammatory, hepatoprotective and antioxidant activity of ellagitannin isolated from Melaleuca styphelioides.

Phytochemistry 2020 Sep 16;177:112429. Epub 2020 Jun 16.

Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan, 33302, Taiwan; Research Center for Chinese Herbal Medicine, Research Center for Food and Cosmetic Safety, and Graduate Institute of Health Industry Technology, College of Human Ecology, Chang Gung University of Science and Technology, Taoyuan, 33302, Taiwan; Department of Anesthesiology, Chang Gung Memorial Hospital, Taoyuan, 33305, Taiwan; Chinese Herbal Medicine Research Team, Healthy Aging Research Center, Chang Gung University, Taoyuan, 33302, Taiwan; Department of Chemical Engineering, Ming Chi University of Technology, New Taipei City, 24301, Taiwan. Electronic address:

Ellagitannins have a marked antioxidant effect and can prevent liver injury induced by free radicals. An undescribed ellagitannin named styphelioidin was isolated from Melaleuca styphelioides Sm. The structure of styphelioidin was elucidated by using various spectroscopic methods. The hepatoprotective activity of styphelioidin (25, 50, and 100 μM) was tested using the CCl-challenged HepG2 cell model by measuring alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in HepG2 cells treated with styphelioidin for 1 h followed by 40 mM CCl. Glutathione (GSH), superoxide dismutase activity (SOD) and lipid peroxidation (MDA) were evaluated to determine the mechanisms of the hepatoprotective activity. Styphelioidin significantly reduced the levels of ALT, AST, and MDA at all tested concentrations. Moreover, it conferred a marked increase in the GSH levels and the SOD activity compared to the CCl-treated groups. Styphelioidin also exerted DPPH· radical-scavenging effects with an IC value of 3.67 μM. Results indicated the hepatoprotective therapeutic potential of styphelioidin comparable to silymarin. Moreover, anti-inflammatory activity was assessed and styphelioidin inhibited fMLF/CB-induced elastase release in human neutrophils with IC 2.51 μM. Cell-free experiments with human neutrophil elastase indicated a direct enzymatic inhibitory effect of styphelioidin on the enzyme activity (IC 2.58 μM). The potential of styphelioidin to interact with human neutrophil elastase binding sites was further confirmed by molecular docking of styphelioidin into human neutrophil elastase crystal structure using AutoDock 4.2. Styphelioidin represents a potent hepatoprotective and antioxidant agent with effects on ALT, AST, MDA, GSH, and SOD comparable to silymarin. The beneficial anti-elastase properties hold the potential for drug development against elastase-related inflammatory diseases. This study highlights a promising natural hepatoprotective and anti-inflammatory candidate derived from M. styphelioides.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.phytochem.2020.112429DOI Listing
September 2020

Rosmarinic acid exhibits broad anti-enterovirus A71 activity by inhibiting the interaction between the five-fold axis of capsid VP1 and cognate sulfated receptors.

Emerg Microbes Infect 2020 Dec;9(1):1194-1205

Department of Biochemistry and Molecular Biology, College of Medicine, Chang Gung University, Taoyuan, Taiwan.

Enterovirus A71 (EV-A71), a positive-stranded RNA virus of the Picornaviridae family, may cause neurological complications or fatality in children. We examined specific factors responsible for this virulence using a chemical genetics approach. Known compounds from an anti-EV-A71 herbal medicine, (Danshen), were screened for anti-EV-A71. We identified a natural product, rosmarinic acid (RA), as a potential inhibitor of EV-A71 by cell-based antiviral assay and mouse model. Results also show that RA may affect the early stage of viral infection and may target viral particles directly, thereby interfering with virus-P-selectin glycoprotein ligand-1 (PSGL1) and virus-heparan sulfate interactions without abolishing the interaction between the virus and scavenger receptor B2 (SCARB2). Sequencing of the plaque-purified RA-resistant viruses revealed a N104K mutation in the five-fold axis of the structural protein VP1, which contains positively charged amino acids reportedly associated with virus-PSGL1 and virus-heparan sulfate interactions via electrostatic attraction. The plasmid-derived recombinant virus harbouring this mutation was confirmed to be refractory to RA inhibition. Receptor pull-down showed that this non-positively charged VP1-N104 is critical for virus binding to heparan sulfate. As the VP1-N104 residue is conserved among different EV-A71 strains, RA may be useful for inhibiting EV-A71 infection, even for emergent virus variants. Our study provides insight into the molecular mechanism of virus-host interactions and identifies a promising new class of inhibitors based on its antiviral activity and broad spectrum effects against a range of EV-A71.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/22221751.2020.1767512DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7448925PMC
December 2020

Polysaccharide-containing fraction from Artemisia argyi inhibits tumor cell-induced platelet aggregation by blocking interaction of podoplanin with C-type lectin-like receptor 2.

J Food Drug Anal 2020 01 26;28(1):115-123. Epub 2019 Sep 26.

Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan; Graduate Institute of Natural Products, School of Traditional Chinese Medicine, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan; Department of Anesthesiology, Chang Gung Memorial Hospital, Linkou, Taiwan; Research Center for Chinese Herbal Medicine, Chang Gung University of Science and Technology, Taoyuan, Taiwan. Electronic address:

Tumor cell-induced platelet aggregation (TCIPA) is a mechanism that involves the protection of tumor cells in the circulation and the promotion of tumor cell invasion and metastases. The C-type lectin-like receptor 2 (CLEC-2) that binds podoplanin (PDPN) is on the platelet surface and facilitates the TCIPA. Selective blockage of the PDPN-mediated platelet-tumor cell interaction is thereby a plausible strategy for inhibiting metastases. In a search for antagonists of PDPN- and tumor cell-induced platelet aggregation, traditional Chinese medicines were screened and it was found that the water extract of Artemisia argyi leaves selectively inhibited the PDPN-induced platelet aggregation. Bioactivity-guided fractionation analysis was performed for defining a polysaccharide-containing fraction (AAWAP) characterized by inhibition of PDPN activity and tumor cell-induced platelet aggregation. The pharmacological effects of AAWAP on PDPN-activated CLEC-2 signaling were determined by using Western blot and alpha screening analyses. AAWAP was non-toxic to the cells and platelets and it suppressed PDPN- and tumor cell-induced platelet aggregation by irreversibly blocking the interaction between PDPN and CLEC-2 in a dose-dependent manner. These findings indicate that AAWAP is an antagonist of the PDPN-CLEC-2 interaction. This action by AAWAP may result in the prevention of tumor cell metastases, and if so, could become an effective pharmacological agent in treating cancer patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jfda.2019.08.002DOI Listing
January 2020

A pull-down and slot blot-based screening system for inhibitor compounds of the podoplanin-CLEC-2 interaction.

PLoS One 2019 25;14(9):e0222331. Epub 2019 Sep 25.

Department of Emergency and Critical Care Medicine, Tokai University School of Medicine, Isehara, Kanagawa, Japan.

Podoplanin, a transmembrane glycoprotein, is overexpressed in certain types of tumors and induces platelet aggregation by binding to C-type lectin-like receptor 2 (CLEC-2) on the platelet membrane. Activated platelets release granule components, which in turn, trigger epithelial-mesenchymal transition and confer invasive capacity to the tumor cells. Therefore, blocking the podoplanin-CLEC-2 interaction by a small-molecule compound is a potential therapeutic strategy to prevent cancer metastasis and invasion. To effectively identify such inhibitory compounds, we have developed a pull-down-based inhibitory compound screening system. An immunoglobulin Fc domain-CLEC-2 fusion protein was used as a bait to capture podoplanin derived from podoplanin-overexpressing HeLa cells in the presence and absence of the test compound. The protein complex was then pulled down using protein A beads. To shorten the turnaround time, increase throughput, and decrease the workload for the operators, centrifugal filter units were employed to separate free and bound podoplanin, instead of using customary aspiration-centrifugation washing cycles. Slot blotting was also utilized in lieu of gel electrophoresis and electrical transfer. Thus, the use of our pull down screening system could facilitate the effective selection of potential inhibitor compounds of the podoplanin-CLEC-2 interaction for cancer therapy. Importantly, our methodology is also applicable to targeting other protein-protein interactions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0222331PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6760769PMC
March 2020

Design, Synthesis, and Biological Evaluation of Itaconic Acid Derivatives as Potential Anti-Influenza Agents.

J Med Chem 2019 03 22;62(5):2390-2403. Epub 2019 Feb 22.

Research Center for Chinese Herbal Medicine , Chang Gung University of Science and Technology , Taoyuan 33303 , Taiwan.

Influenza A viruses (IAVs) have caused worldwide epidemics and pandemics by reassortment and generation of drug-resistant mutants, which render antivirals and current vaccinations no longer usable. In this study, an itaconic acid derivative 1 was identified from a chemical library of 20 000 compounds, by performing a cell-based screening assay, as a lead agent exhibiting anti-influenza A activity. Accordingly, a series of itaconic acid derivatives were designed and synthesized by adopting a rational design strategy to obtain more potent anti-influenza agents. The results of an in vitro pharmacological study showed that compounds 4 and 8 exhibited the most potent anti-IAV effect with half-maximal effective concentration values of 0.14 and 0.11 μM, respectively, in Madin-Darby canine kidney cells. The mechanism of action studies showed that lead agents 1 and 4 reduced virus replication by directly targeting IAV nucleoproteins and disrupting virus ribonucleoprotein export from the nucleus to the cytosol. On the basis of its high potential as an anti-IAV agent and its selectivity index >785, compound 4 was found to be a promising candidate for further development against IAVs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.jmedchem.8b01683DOI Listing
March 2019

Topical application of anthranilate derivatives ameliorates psoriatic inflammation in a mouse model by inhibiting keratinocyte-derived chemokine expression and neutrophil infiltration.

FASEB J 2018 Jun 19:fj201800354. Epub 2018 Jun 19.

Graduate Institute of Natural Products, Chang Gung University, Kweishan, Taoyuan, Taiwan.

Psoriasis is an inflammatory autoimmune skin disorder possessing a complex etiology related to genetic and environmental triggers. Keratinocytes show a potential role for the origin of psoriasis. In this study, we estimated the efficiency of 2 anthranilate derivatives-(E)-4-( N-{2-[1-(hydroxyimino)ethyl]phenyl}sulfamoyl)phenyl pivalate (HFP031) and butyl 2-[2-(2-fluorophenyl)acetamido]benzoate (HFP034)-on psoriasis amelioration in a mouse model. The results showed that topical treatment with both compounds could attenuate epidermal thickness and scaling in an imiquimod (IMQ)-induced psoriasis mouse model via decreased expression of cytokines and chemokines [C-X-C chemokine ligand (CXCL)1 and CXCL2], leading to the reduction of neutrophilic abscess in the skin. The in vivo cutaneous absorption of HFP034 was 7.6-fold greater than that of HFP031. Both compounds caused negligible irritation on healthy mouse skin. In addition, we examined the effect of the anthranilate derivatives on chemokine expression in IMQ-treated HaCaT keratinocytes. Our results elucidated a mechanism for anti-inflammatory activity of HFP034 that involved the elevation of intracellular cAMP concentration, suppression of NF-κB activity, and attenuation of neutrophil chemoattractant expression. These results suggest that HFP034 could increase the cutaneous concentration of cAMP to suppress neutrophil infiltration into the skin. Topically applied HFP034 may demonstrate a potential for future clinical application as a novel therapy for psoriasis treatment.-Lin, Z.-C., Hsieh, P.-W., Hwang, T.-L., Chen, C.-Y., Sung, C. T., Fang, J.-Y. Topical application of anthranilate derivatives ameliorates psoriatic inflammation in a mouse model by inhibiting keratinocyte-derived chemokine expression and neutrophil infiltration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.201800354DOI Listing
June 2018

Design, synthesis & structure-activity relationships of a new class of antihuman enterovirus D68 & A71 agents.

Future Med Chem 2018 06 10;10(11):1333-1347. Epub 2018 May 10.

Graduate Institute of Biomedical Science, College of Medicine, Chang Gung University, Taoyuan, 33302, Taiwan.

Aim: No antiviral medications are currently approved to treat enterovirus (EV)-associated disease or prevent EV infection.

Methods: In this study, a series of probenecid derivatives were designed via a rational strategy and synthesized to obtain more potent anti-EV agents.

Results: Compounds 8 and 24 exhibited the most potent activity against EV D68 and A71, with half maximal effective concentration (EC) values of 2.49/2.09 and 2.59/2.41 μM, respectively, and revealed a broad inhibition spectrum toward other EV strains, with high selectivity indices. Additionally, compounds 8 and 24 showed good stability in rat serum, with half-lives of 48.39 and 60.26 min, respectively.

Conclusion: Compounds 8 and 24 are the promising candidates for the development of new agents against EV D68 and A71 viruses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4155/fmc-2017-0268DOI Listing
June 2018

β-Nitrostyrene derivatives attenuate LPS-mediated acute lung injury via the inhibition of neutrophil-platelet interactions and NET release.

Am J Physiol Lung Cell Mol Physiol 2018 04 11;314(4):L654-L669. Epub 2018 Jan 11.

Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University , Taoyuan , Taiwan.

Acute lung injury (ALI) and the acute respiratory distress syndrome (ARDS) are high-mortality and life-threatening diseases that are associated with neutrophil activation and accumulation within lung tissue. Emerging evidence indicates that neutrophil-platelet aggregates (NPAs) at sites of injury increase acute inflammation and contribute to the development of ALI. Although numerous studies have increased our understanding of the pathophysiology of ALI, there is still a lack of innovative and useful treatments that reduce mortality, emphasizing that there is an urgent need for novel treatment strategies. In this study, a new series of small compounds of β-nitrostyrene derivatives (BNSDs) were synthesized, and their anti-inflammatory bioactivities on neutrophils and platelets were evaluated. The new small compound C7 modulates neutrophil function by inhibiting superoxide generation and elastase release. Compound C7 elicits protective effects on LPS-induced paw edema and acute lung injury via the inhibition of neutrophil accumulation, proinflammatory mediator release, platelet aggregation, myeloperoxidase activity, and neutrophil extracellular trap (NET) release. NET formation was identified as the bridge for the critical interactions between neutrophils and platelets by confocal microscopy and flow cytometry. This research provides new insights for elucidating the complicated regulation of neutrophils and platelets in ALI and sheds further light on future drug development strategies for ALI/ARDS and acute inflammatory diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1152/ajplung.00501.2016DOI Listing
April 2018

CYT-Rx20 Inhibits Cervical Cancer Cell Growth and Migration Through Oxidative Stress-Induced DNA Damage, Cell Apoptosis, and Epithelial-to-Mesenchymal Transition Inhibition.

Int J Gynecol Cancer 2017 09;27(7):1306-1317

Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung.

Objective: The β-nitrostyrene family has been reported to possess anticancer properties. However, the anticancer activity of β-nitrostyrenes on cervical cancer cells and the underlying mechanisms involved remain unexplored. In this study, a β-nitrostyrene derivative CYT-Rx20 (3'-hydroxy-4'-methoxy-β-methyl-β-nitrostyrene) was synthesized, and its anticancer activity on cervical cancer cells and the mechanisms involved were investigated.

Methods: The effect of CYT-Rx20 on human cervical cancer cell growth was evaluated using cell viability assay. Reactive oxygen species (ROS) generation and annexin V staining were detected by flow cytometry. The protein expression levels of cleaved caspase-3, cleaved caspase-9, cleaved poly (ADPribose) polymerase, γH2AX, β-catenin, Vimentin, and Twist were measured by Western blotting. DNA double-strand breaks were determined by γ-H2AX foci formation and neutral comet assay. Migration assay was used to determine cancer cell migration. Nude mice xenograft was used to investigate the antitumor effects of CYT-Rx20 in vivo.

Results: CYT-Rx20 induced cytotoxicity in cervical cancer cells by promoting cell apoptosis via ROS generation and DNA damage. CYT-Rx20-induced cell apoptosis, ROS generation, and DNA damage were reversed by thiol antioxidants. In addition, CYT-Rx20 inhibited cervical cancer cell migration by regulating the expression of epithelial-to-mesenchymal transition markers. In nude mice, CYT-Rx20 inhibited cervical tumor growth accompanied by increased expression of DNA damage marker γH2AX and decreased expression of mesenchymal markers β-catenin and Twist.

Conclusions: CYT-Rx20 inhibits cervical cancer cells in vitro and in vivo and has the potential to be further developed into an anti-cervical cancer drug clinically.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/IGC.0000000000001033DOI Listing
September 2017

HPW-RX40 prevents human platelet activation by attenuating cell surface protein disulfide isomerases.

Redox Biol 2017 10 29;13:266-277. Epub 2017 May 29.

Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Marine Biotechnology and Resources, National Sun Yat-sen University, Kaohsiung, Taiwan. Electronic address:

Protein disulfide isomerase (PDI) present at platelet surfaces has been considered to play an important role in the conformational change and activation of the integrin glycoprotein IIb/IIIa (GPIIb/IIIa) and thus enhances platelet aggregation. Growing evidences indicated that platelet surface PDI may serve as a potential target for developing of a new class of antithrombotic agents. In the present study, we investigated the effects of HPW-RX40, a chemical derivative of β-nitrostyrene, on platelet activation and PDI activity. HPW-RX40 inhibited platelet aggregation, GPIIb/IIIa activation, and P-selectin expression in human platelets. Moreover, HPW-RX40 reduced thrombus formation in human whole blood under flow conditions, and protects mice from FeCl-induced carotid artery occlusion. HPW-RX40 inhibited the activity of recombinant PDI family proteins (PDI, ERp57, and ERp5) as well as suppressed cell surface PDI activity of platelets in a reversible manner. Exogenous addition of PDI attenuated the inhibitory effect of HPW-RX40 on GPIIb/IIIa activation. Structure-based molecular docking simulations indicated that HPW-RX40 binds to the active site of PDI by forming hydrogen bonds. In addition, HPW-RX40 neither affected the cell viability nor induced endoplasmic reticulum stress in human cancer A549 and MDA-MB-231 cells. Taken together, our results suggest that HPW-RX40 is a reversible and non-cytotoxic PDI inhibitor with antiplatelet effects, and it may have a potential for development of novel antithrombotic agents.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.redox.2017.05.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5466588PMC
October 2017

CYT-Rx20 inhibits ovarian cancer cells in vitro and in vivo through oxidative stress-induced DNA damage and cell apoptosis.

Cancer Chemother Pharmacol 2017 Jun 12;79(6):1129-1140. Epub 2017 May 12.

Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.

Purpose: The β-nitrostyrene family has been previously reported to possess anticancer property. However, the biological effects of β-nitrostyrenes on ovarian cancer and the underlying mechanisms involved remain unclear. In the present study, we synthesized a β-nitrostyrene derivative, CYT-Rx20 3'-hydroxy-4'-methoxy-β-methyl-β-nitrostyrene), and investigated its anticancer effects and the putative pathways of action in ovarian cancer.

Methods: The effects of CYT-Rx20 were analyzed using cell viability assay, reactive oxygen species (ROS) generation assay, FACS analysis, annexin V staining, immunostaining, comet assay, immunoblotting, soft agar assay, migration assay, nude mice xenograft study and immunohistochemistry.

Results: CYT-Rx20 induced cytotoxicity in ovarian cancer cells by promoting cell apoptosis via ROS generation and DNA damage. CYT-Rx20-induced cell apoptosis, ROS generation and DNA damage were reversed by thiol antioxidants. In addition, CYT-Rx20 inhibited ovarian cancer cell migration by regulating the expression of epithelial to mesenchymal transition (EMT) markers. In nude mice, CYT-Rx20 inhibited ovarian tumor growth accompanied by increased expression of DNA damage marker γH2AX and decreased expression of EMT marker Vimentin.

Conclusions: CYT-Rx20 inhibits ovarian cancer cells in vitro and in vivo, and has the potential to be further developed into an anti-ovarian cancer drug clinically.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00280-017-3330-9DOI Listing
June 2017

Synthetic β-nitrostyrene derivative CYT-Rx20 as inhibitor of oral cancer cell proliferation and tumor growth through glutathione suppression and reactive oxygen species induction.

Head Neck 2017 06 27;39(6):1055-1064. Epub 2017 Mar 27.

Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.

Background: The β-nitrostyrene family possesses anticancer properties. In this study, β-nitrostyrene derivative CYT-Rx20 (3'-hydroxy-4'-methoxy-β-methyl-β-nitrostyrene) was synthesized and investigated its anticancer activity in oral cancer.

Methods: Anticancer activity of CYT-Rx20 and the underlying mechanisms were analyzed using cell viability assay, reactive oxygen species (ROS) generation assay, fluorescence-activated cell sorter analysis, annexin V staining, comet assay, glutathione (GSH)/glutathione disulfide (GSSG) ratio, immunoblotting, soft agar assay, nude mice xenograft study, and immunohistochemistry.

Results: CYT-Rx20-induced cell apoptosis via ROS generation and mitochondrial membrane potential reduction, associated with release of mitochondrial cytochrome C to cytosol and activation of downstream caspases and poly ADP-ribose polymerase (PARP). Furthermore, CYT-Rx20 induced mitochondrial ROS accumulation and mitochondrial dysfunction, followed by GSH downregulation. CYT-Rx20-induced cell apoptosis, ROS generation, and DNA damage were reversed by thiol antioxidants. In nude mice, CYT-Rx20 inhibited oral tumor growth accompanied by increased expression of γH2AX, GSH reductase, and cleaved-caspase-3.

Conclusion: CYT-Rx20 has the potential to be further developed into an antioral cancer drug clinically. © 2017 Wiley Periodicals, Inc. Head Neck 39: 1055-1064, 2017.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/hed.24664DOI Listing
June 2017

Dipeptide HCH6-1 inhibits neutrophil activation and protects against acute lung injury by blocking FPR1.

Free Radic Biol Med 2017 05 21;106:254-269. Epub 2017 Feb 21.

Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan; Division of Natural Products, Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan; Research Center for Industry of Human Ecology, Research Center for Chinese Herbal Medicine, and Graduate Institute of Health Industry Technology, Chang Gung University of Science and Technology, Taoyuan 333, Taiwan; Chinese Herbal Medicine Research Team, Healthy Aging Research Center, Chang Gung University, Taoyuan 333, Taiwan; Department of Anesthesiology, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan. Electronic address:

Formyl peptide receptor 1 (FPR1) is an emerging therapeutic target for the discovery of drugs to treat neutrophilic inflammatory diseases. However, development of FPR1 antagonists for clinical use is still inadequate. The purpose of this study was to identify a synthetic dipeptide N-(N-benzoyl-L-tryptophanyl)-D-phenylanlanine methyl ester (HCH6-1) as a FPR1 inhibitor and to investigate its protective effects against acute lung injury (ALI). HCH6-1 inhibited superoxide anion generation, elastase release, and chemotaxis in human neutrophils specifically activated by formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLF), an FPR1 agonist. HCH6-1 produced right shifts in the concentration-response curves of fMLF, suggesting that HCH6-1 was a competitive antagonist of FPR1. Indeed, HCH6-1 bound to FPR1 in human neutrophils and neutrophil-like THP-1 as well as hFPR1-transfected HEK293 cells. Also, the FPR1 downstream signaling pathways were competitively inhibited by HCH6-1. Furthermore, HCH6-1 prevented pulmonary neutrophil infiltration and edema along with alveolar damage in LPS-induced ALI in mice. Our findings suggest that HCH6-1, a FPR1 antagonist, may have potential as a new therapeutic agent for treating FPR1-involved inflammatory lung diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.freeradbiomed.2017.02.038DOI Listing
May 2017

3'-hydroxy-4'-methoxy-β-methyl-β-nitrostyrene inhibits tumorigenesis in colorectal cancer cells through ROS-mediated DNA damage and mitochondrial dysfunction.

Oncotarget 2017 Mar;8(11):18106-18117

Translational Research Center, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.

The β-nitrostyrene family has been shown to suppress cell proliferation and induce apoptosis in types of various cancers. However, the mechanisms underlying the anticancer effects of β-nitrostyrenes in colorectal cancer remain poorly understood. In this study, we synthesized a β-nitrostyrene derivative, CYT-Rx20 (3'-hydroxy-4'-methoxy-β-methyl-β-nitrostyrene), and investigated its anticancer activities in human colorectal cancer cells both in vitro and in vivo. Our findings showed that treatment with CYT-Rx20 reduced cell viability and induced DNA damage in colorectal cancer cells. In addition, CYT-Rx20 induced cell cycle arrest of colorectal cancer cells at the G2/M phase and upregulated the protein expression of phospho-ERK, cyclin B1, phospho-cdc2 (Tyr15), aurora A, and aurora B, while it downregulated the expression of cdc25A and cdc25C. Furthermore, we found that CYT-Rx20 caused accumulation of intracellular reactive oxygen species (ROS) and reduction of mitochondrial membrane potential. The effects of CYT-Rx20 on cell viability, DNA damage, and mitochondrial membrane potential were reversed by pretreatment with the thiol antioxidant N-acetyl-L-cysteine (NAC), suggesting that ROS-mediated DNA damage and mitochondrial dysregulation play a critical role in these events. Finally, the nude mice xenograft study showed that CYT-Rx20 significantly reduced tumor growth of implanted colorectal cancer cells accompanied by elevated protein expression of aurora A, aurora B, γH2AX, phosphor-ERK, and MDA in the tumor tissues. Taken together, these results suggest that CYT-Rx20 may potentially be developed as a novel β-nitrostyrene-based anticancer agent for colorectal cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.14996DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5392311PMC
March 2017

Anti-melasma codrug of retinoic acid assists cutaneous absorption with attenuated skin irritation.

Eur J Pharm Biopharm 2017 May 1;114:154-163. Epub 2017 Feb 1.

Research Center for Food and Cosmetic Safety and Research Center for Chinese Herbal Medicine, Chang Gung University of Science and Technology, Kweishan, Taoyuan, Taiwan; Department of Anesthesiology, Chang Gung Memorial Hospital, Kweishan, Taoyuan, Taiwan; Pharmaceutics Laboratory, Graduate Institute of Natural Products, Chang Gung University, Kweishan, Taoyuan, Taiwan; Chinese Herbal Medicine Research Team, Healthy Aging Research Center, Chang Gung University, Kweishan, Taoyuan, Taiwan. Electronic address:

Melasma treatment with combined retinoic acid (RA) and hydroquinone (HQ) usually causes unsatisfactory outcomes and safety concerns. This study attempted to evaluate the cutaneous absorption and skin tolerance of the codrug conjugated with RA and HQ via ester linkage. The codrug's permeation of the pig skin was estimated using Franz diffusion cell. The codrug and parent drugs were comparatively examined for anti-inflammatory activity and tyrosinase inhibition. In vivo cutaneous irritation was assessed on nude mouse skin. Chemical conjugation of RA with HQ increased the lipophilicity and thus the skin absorption. The codrug absorption produced a 5.5- and a 60.8-fold increment compared to RA skin deposition at an equimolar (1.2mM) and saturated solubility dose, respectively. The cumulative amount of HQ derived from the codrug in the receptor was comparable to or less than that of topically applied HQ. The RA-HQ codrug was partly hydrolyzed on penetrating the skin. The hydrolysis rate in intact skin was significantly lower than that in esterase medium and skin homogenates. The codrug showed an interleukin (IL)-6 inhibition activity comparable to RA. A therapeutic index 6-fold greater than RA was obtained with the topical codrug. The tyrosinase inhibition percentage of the codrug and HQ was 13% and 21%, respectively. The skin tolerance test determined by transepidermal water loss (TEWL), redness, and histopathology had exhibited minor skin irritation caused by the codrug compared to the physical mixture of RA and HQ at an equivalent dose. Topical codrug delivery not only promoted RA absorption, but also diminished the adverse effects of the parent agents.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejpb.2017.01.016DOI Listing
May 2017

3'-Hydroxy-4'-methoxy-β-methyl-β-nitrostyrene inhibits tumor growth through ROS generation and GSH depletion in lung cancer cells.

Life Sci 2017 Mar 20;172:19-26. Epub 2016 Dec 20.

Translational Research Center, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan; Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Obstetrics and Gynecology, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan. Electronic address:

Aims: Members of the β-nitrostyrene family are known to suppress tumor growth, with the underlying mechanisms of β-nitrostyrene remain mostly unclear. Herein, we synthesized a β-nitrostyrene derivative, 3'-hydroxy-4'-methoxy-β-methyl-β-nitrostyrene (CYT-Rx20), and explored its anticancer activities in human lung cancer cells in vitro and in vivo.

Main Methods: Cell viability was measured by XTT assay. Apoptosis was detected by Annexin V/PI staining. Caspase activation was determined by western blotting. ROS (reactive oxygen species), MMP (mitochondrial membrane potential) and mitochondrial mass were determined by flow cytometry. GSH level was detected by ELISA assay.

Key Findings: In this study, we found that CYT-Rx20 significantly reduced cell viability, accompanied by G2/M arrest in lung cancer cells. Increased protein levels of cleaved-caspase families indicated apoptotic cell death upon CYT-Rx20 treatment. Furthermore, increased level of intracellular reactive oxygen species (ROS), loss of mitochondrial membrane potential (ΔΨm), glutathione (GSH) depletion and inhibition of GSH reductase were observed after CYT-Rx20 treatment. The effects of CYT-Rx20 on cell viability and the loss of ΔΨm were significantly reversed when cells were pretreated with thiol antioxidants NAC, GSH, or 2-ME. Finally, xenograft animal study demonstrated that CYT-Rx20 significantly suppressed lung tumor growth in vivo.

Significance: Our data demonstrated that CYT-Rx20 triggered apoptotic cell death in lung cancer cells and suppressed lung tumor growth through GSH depletion, suggesting that CYT-Rx20 may have the potential to be further developed as an anticancer compound for treating lung cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.lfs.2016.12.009DOI Listing
March 2017

The Synthetic β-Nitrostyrene Derivative CYT-Rx20 Inhibits Esophageal Tumor Growth and Metastasis via PI3K/AKT and STAT3 Pathways.

PLoS One 2016 22;11(11):e0166453. Epub 2016 Nov 22.

Translational Research Center, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.

The β-nitrostyrene family have been implicated for anti-cancer property. However, the pharmacological role of β-nitrostyrene in esophageal cancer remain unclear. Here, a β-nitrostyrene derivative, CYT-Rx20, was synthesized and assessed for its anti-cancer activities and underlying mechanism in esophageal cancer. CYT-Rx20 induced cytotoxicity in esophageal cancer cells by promoting apoptosis through activation of caspase cascade and poly(ADP-ribose) polymerase (PARP) cleavage. Besides, CYT-Rx20 inhibited esophageal cancer cell migration and invasion by regulating the expression of epithelial to mesenchymal transition (EMT) markers. CYT-Rx20 decreased cell viability and migration through suppression of the PI3K/AKT and STAT3 pathways. Of note, the cytotoxicity and anti-migratory effect of CYT-Rx20 were enhanced by co-treatment with SC79 (AKT activator) or colivelin (STAT3 activator), suggesting the dependency of esophageal cancer cells on AKT and STAT3 for survival and migration, an oncogene addiction phenomenon. In xenograft tumor-bearing mice, CYT-Rx20 significantly reduced tumor growth of the implanted esophageal cancer cells accompanied by decreased Ki-67, phospho-AKT, and phospho-STAT3 expression. In orthotopic esophageal cancer mouse model, decreased tumor growth and lung metastasis with reduced Ki-67 and phospho-STAT3 expression were observed in mice treated with CYT-Rx20. Together, our results suggest that CYT-Rx20 is a potential β-nitrostyrene-based anticancer compound against the tumor growth and metastasis of esophageal cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0166453PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5119777PMC
June 2017

An extract from Taxodium distichum targets hemagglutinin- and neuraminidase-related activities of influenza virus in vitro.

Sci Rep 2016 10 31;6:36015. Epub 2016 Oct 31.

Department of Biochemistry and Molecular Biology, College of Medicine, Chang Gung University, Taoyuan, Taiwan.

Influenza virus remains an emerging virus and causes pandemics with high levels of fatality. After screening different plant extracts with potential anti-influenza activity, a water extract of Taxodium distichum stems (TDSWex) showed excellent activity against influenza viruses. The EC of TDSWex was 0.051 ± 0.024 mg/mL against influenza virus A/WSN/33. TDSWex had excellent antiviral efficacy against various strains of human influenza A and B viruses, particularly oseltamivir-resistant clinical isolates and a swine-origin influenza strain. We observed that the synthesis of viral RNA and protein were inhibited in the presence of TDSWex. The results of the time-of-addition assay suggested that TDSWex inhibited viral entry and budding. In the hemagglutination inhibition assay, TDSWex inhibited the hemagglutination of red blood cells, implying that the extract targeted hemagglutin-related functions such as viral entry. In the attachment and penetration assay, TDSWex showed antiviral activity with ECs of 0.045 ± 0.026 and 0.012 ± 0.003 mg/mL, respectively. In addition, TDSWex blocked neuraminidase activity. We conclude that TDSWex has bimodal activities against both hemagglutinin and neuraminidase during viral replication.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/srep36015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5086851PMC
October 2016

Ilex kaushue and Its Bioactive Component 3,5-Dicaffeoylquinic Acid Protected Mice from Lipopolysaccharide-Induced Acute Lung Injury.

Sci Rep 2016 Sep 29;6:34243. Epub 2016 Sep 29.

Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan.

Acute lung injury (ALI) is a severe respiratory disease with high mortality rates worldwide. Recent reports suggest that human neutrophil elastase (HNE) plays a key role in the inflammatory response that is characteristic of ALI, which indicates that the development of HNE inhibitors could be an efficient treatment strategy. In the current study, an enzyme-based screening assay was used to identify effective HNE inhibitors from a number of traditional Chinese medicines (TCMs). Among them, a water extract of Ilex kaushue (IKWE) effectively inhibited HNE activity (IC, 11.37 ± 1.59 μg/mL). Using bioactivity-guided fractionation, one new compound and 23 known compounds were identified. Compound 6 (identified as 3,5-dicaffeoylquinic acid; 3,5-DCQA) exerted the most potent and selective inhibitory effect on HNE activity (IC, 1.86 ± 0.06 μM). In a cell-based assay, 3,5-DCQA not only directly reduced superoxide generation and elastase activity but also attenuated the Src family kinase (SRKs)/Vav signaling pathway in N-formyl-L-Met-L-Leu-L-Phe (fMLF)-stimulated human neutrophils. In an animal disease model, both 3,5-DCQA and standardized IKWE protected against lipopolysaccharide-induced ALI in mice, which provides support for their potential as candidates in the development of new therapeutic agents for neutrophilic inflammatory diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5041076PMC
http://dx.doi.org/10.1038/srep34243DOI Listing
September 2016

Handheld analyzer with on-chip molecularly-imprinted biosensors for electrical detection of propofol in plasma samples.

Biosens Bioelectron 2016 Dec 11;86:623-629. Epub 2016 Jul 11.

Department of Anesthesiology, Chang Gung Memorial Hospital, Taoyuan, Taiwan; Graduate Institute of Natural Products, School of Traditional Chinese Medicine, and Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan.

This paper proposes a novel handheld analyzer with disposable lab-on-a-chip technology for the electrical detection of the anesthetic propofol in human plasma samples for clinical diagnoses. The developed on-chip biosensors are based on the conduction of molecularly imprinted polymers (MIPs) that employ label-free electrical detection techniques. Propofol in total intravenous anesthesia is widely used with a target-controlled infusion system. At present, the methods employed for detecting blood propofol concentrations in hospitals comprise high-performance liquid chromatography and ion mobility spectrometry. These conventional instruments are bulky, expensive, and difficult to access. In this study, we developed a novel plastic microfluidic biochip with an on-chip anesthetic biosensor that was characterized for the rapid detection of propofol concentrations. The experimental results revealed that the response time of the developed propofol biosensors was 25s. The specific binding of an MIP to a nonimprinted polymer (NIP) reached up to 560%. Moreover, the detection limit of the biosensors was 0.1μg/mL, with a linear detection range of 0.1-30μg/mL. The proposed disposable microfluidic biochip with an on-chip anesthetic biosensor using MIPs exhibited excellent performance in the separation and sensing of propofol molecules in the human plasma samples. Compared with large-scale conventional instruments, the developed microfluidic biochips with on-chip MIP biosensors present the advantages of a compact size, high selectivity, low cost, rapid response, and single-step detection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bios.2016.07.032DOI Listing
December 2016

In vitro and in vivo studies of a potent capsid-binding inhibitor of enterovirus 71.

J Antimicrob Chemother 2016 07 19;71(7):1922-32. Epub 2016 Apr 19.

Department of Biochemistry and Molecular Biology, College of Medicine, Chang Gung University, Kweishan, Taoyuan, Taiwan Research Center for Emerging Viral Infections, College of Medicine, Chang Gung University, Kweishan, Taoyuan, Taiwan Research Center for Industry of Human Ecology and Graduate Institute of Health Industry Technology, Chang Gung University of Science and Technology, Taoyuan, Taiwan Molecular Infectious Disease Research Center, Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Taoyuan, Taiwan

Objectives: Enterovirus 71 (EV-A71) is an important pathogen that can cause severe neurological symptoms and even death. Our aim was to identify potent anti-EV-A71 compounds and study their underlying mechanisms and in vivo activity.

Methods: We identified a potent imidazolidinone derivative (abbreviated to PR66) as an inhibitor of EV-A71 infection from the screening of compounds and subsequent structure-based modification. Time-course treatments and resistant virus selection of PR66 were employed to study the mode of mechanism of PR66. In vivo activity of PR66 was tested in the ICR strain of new-born mice challenged with EV-A71/4643/MP4.

Results: PR66 could impede the uncoating process during viral infection via interaction with capsid protein VP1, as shown by a resistant virus selection assay. Using site-directed mutagenesis, we confirmed that a change from valine to phenylalanine in the 179th amino acid residue of the cDNA-derived resistant virus resulted in resistance to PR66. PR66 increased the virion stability of WT viruses, but not the PR66-resistant mutant, in a particle stability thermal release assay. We further showed that PR66 had excellent anti-EV-A71 activity in an in vivo mouse model of disease, with a dose-dependent increase in survival rate and in protection against virus-induced hind-limb paralysis following oral or intraperitoneal administration. This was associated with reductions of viral titres in brain and muscle tissues.

Conclusions: We demonstrated here for the first time that an imidazolidinone derivative (PR66) could protect against EV-A71-induced neurological symptoms in vivo by suppressing EV-A71 replication. This involved binding to and restricting viral uncoating.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/jac/dkw101DOI Listing
July 2016

The synthetic β-nitrostyrene derivative CYT-Rx20 induces breast cancer cell death and autophagy via ROS-mediated MEK/ERK pathway.

Cancer Lett 2016 Feb 9;371(2):251-61. Epub 2015 Dec 9.

Translational Research Center, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan; Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Obstetrics and Gynecology, Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan; Lipid Science and Aging Research Center, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan. Electronic address:

The β-nitrostyrene family has been shown to suppress cancer cell proliferation and induce programmed cell death. However, mechanisms underlying β-nitrostyrenes remain less evaluated. Here, we synthesized a β-nitrostyrene derivative, CYT-Rx20, and characterized its anticancer effect and involving mechanisms in breast cancer. We found that CYT-Rx20 arrested breast cancer cells at G2/M phase and decreased cell viability by activating the caspase cascade, accompanying with increases of poly (ADP-ribose) polymerase (PARP) cleavage and γ-H2AX expression. On the other hand, up-regulation of Beclin-1, ATG5, and LC-3 was observed in CYT-Rx20-induced autophagy, which was evidently shown by transmission electron microscopy. In addition to these, CYT-Rx20-induced breast cancer cell death, intracellular reactive oxygen species (ROS) formation and expression of phospho-ERK1/2, Beclin-1, and LC-3 were significantly reversed in the presence of N-acetyl-l-cysteine (NAC), a thiol antioxidant. Furthermore, the cytotoxicity of CYT-Rx20 was enhanced by co-treatment with the autophagy inhibitor chloroquine or bafilomycin A1, suggesting that an incomplete autophagy process could deteriorate CYT-Rx20-induced cytotoxicity. In nude mice xenograft study, CYT-Rx20 significantly reduced orthotopic tumor growth. Immunohistochemical analysis revealed elevated expression of phospho-ERK1/2 and LC-3 in tumor tissues of the mice treated with CYT-Rx20. Together, we propose that CYT-Rx20 may have potential to be further developed into a β-nitrostyrene-based anticancer compound for the treatment of breast cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.canlet.2015.11.035DOI Listing
February 2016

Inhibition of the interactions between metastatic human breast cancer cells and platelets by β-nitrostyrene derivatives.

Life Sci 2015 Dec 10;143:147-55. Epub 2015 Nov 10.

Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung, Taiwan; Research Center for Natural Products and Drug Development, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Marine Biotechnology and Resources, National Sun Yat-sen University, Kaohsiung 80708, Taiwan. Electronic address:

Aims: The interactions between cancer cells and platelets have been recognized to play an important role in cancer progress as well as metastasis, and interference with cancer-platelet interactions is an attractive strategy for cancer therapy. In the present study, two β-nitrostyrene derivatives: 3, 4-methylene-dioxy-β-nitrostyrene (MNS) and 4-O-benzoyl-3-methoxyl-β-nitrostyrene (BMNS) have been tested for their inhibitory effect on platelet activation caused by metastatic human breast cancer MDA-MB-231 and Hs578T cells.

Main Methods: Washed human platelets were co-incubated with breast cancer cells, and platelet aggregation was determined turbidimetrically. Platelet adhesion to cancer cells and P-selectin expression were measured by flow cytometry. Platelet-derived growth factor (PDGF) released from cancer cell-stimulated platelets was determined by enzyme-linked immunosorbent assay (ELISA).

Key Findings: MNS and BMNS prevented cancer cell-induced platelet aggregation, P-selectin expression, and PDGF secretion. Moreover, the β-nitrostyrenes reduced platelet adhesion to cancer cells, suggesting the initial cancer-platelet interactions are inhibited. In contrast to current antiplatelet strategies, the glycoprotein IIb/IIIa (GPIIb/IIIa) antagonist RGDS peptide only prevented cancer cells-induced platelet aggregation, but not platelet adhesion and secretion; whereas the cyclooxygenase inhibitor aspirin and the adenosine diphosphate (ADP) scavenger apyrase affected neither platelet aggregation nor platelet secretion.

Significance: The inhibitory effects of the β-nitrostyrene derivatives on cancer-platelet interactions may offer a potential approach for repressing cancer metastasis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.lfs.2015.11.003DOI Listing
December 2015

Identification of a novel platelet antagonist that binds to CLEC-2 and suppresses podoplanin-induced platelet aggregation and cancer metastasis.

Oncotarget 2015 Dec;6(40):42733-48

Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan, Republic of China (ROC).

Podoplanin (PDPN) enhances tumor metastases by eliciting tumor cell-induced platelet aggregation (TCIPA) through activation of platelet C-type lectin-like receptor 2 (CLEC-2). A novel and non-cytotoxic 5-nitrobenzoate compound 2CP was synthesized that specifically inhibited the PDPN/CLEC-2 interaction and TCIPA with no effect on platelet aggregation stimulated by other platelet agonists. 2CP possessed anti-cancer metastatic activity in vivo and augmented the therapeutic efficacy of cisplatin in the experimental animal model without causing a bleeding risk. Analysis of the molecular action of 2CP further revealed that Akt1/PDK1 and PKCμ were two alternative CLEC-2 signaling pathways mediating PDPN-induced platelet activation. 2CP directly bound to CLEC-2 and, by competing with the same binding pocket of PDPN in CLEC-2, inhibited PDPN-mediated platelet activation. This study provides evidence that 2CP is the first defined platelet antagonist with CLEC-2 binding activity. The augmentation in the therapeutic efficacy of cisplatin by 2CP suggests that a combination of a chemotherapeutic agent and a drug with anti-TCIPA activity such as 2CP may prove clinically effective.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.5811DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4767466PMC
December 2015

HPW-RX40 restores anoikis sensitivity of human breast cancer cells by inhibiting integrin/FAK signaling.

Toxicol Appl Pharmacol 2015 Dec 18;289(2):330-40. Epub 2015 Sep 18.

Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Marine Biotechnology and Resources, National Sun Yat-sen University, Kaohsiung 80708, Taiwan; Research Center for Natural Products and Drug Development, Kaohsiung Medical University, Kaohsiung, Taiwan. Electronic address:

Anoikis is defined as apoptosis, which is induced by inappropriate cell-matrix interactions. Cancer cells with anoikis resistance tend to undergo metastasis, and this phenomenon has been reported to be associated with integrin and FAK activity. HPW-RX40 is a derivative of 3,4-methylenedioxy-β-nitrostyrene, which is known to prevent platelet aggregation by inhibition of integrin. In the present study, we investigated the effect of HPW-RX40 on an anoikis-resistant human breast cancer cell line MDA-MB-231. HPW-RX40 inhibited cell aggregation and induced cell death in suspending MDA-MB-231 cells, but had only little effect on the monolayer growth of adherent cells. Analysis of caspase activation and poly (ADP-ribose) polymerase (PARP) cleavage confirmed anoikis in HPW-RX40-treated suspending cancer cells. HPW-RX40 also affected the Bcl-2 family proteins in detached cancer cells. Furthermore, HPW-RX40 inhibited detachment-induced activation of FAK and the downstream phosphorylation of Src and paxillin, but did not affect this pathway in adherent cancer cells. We also found that the expression and activation of β1 integrin in MDA-MB-231 cells were reduced by HPW-RX40. The combination of HPW-RX40 with an EGFR inhibitor led to enhanced anoikis and inhibition of the FAK pathway in breast cancer cells. Taken together, our results suggest that HPW-RX40 restores the anoikis sensitivity in the metastatic breast cancer cells by inhibiting integrin and subsequent FAK activation, and reveal a potential strategy for prevention of tumor metastasis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.taap.2015.09.011DOI Listing
December 2015

Synthesis and pharmacological characterization of 2-aminobenzaldehyde oxime analogs as dual inhibitors of neutrophil elastase and proteinase 3.

Bioorg Med Chem 2015 Mar 16;23(5):1123-34. Epub 2015 Jan 16.

Graduate Institute of Natural Products, School of Traditional Chinese Medicine, and Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan; Chinese Herbal Medicine Research Team, Healthy Aging Research Center, Chang Gung University, Taoyuan, Taiwan; Research Center for Industry of Human Ecology, Chang Gung University of Science and Technology, Kweishan, Taoyuan, Taiwan. Electronic address:

Proteinase 3 (Pr3), and human neutrophil elastase (HNE) are two major neutrophilic serine proteases (NsPs) expressed in neutrophil azurophil granules. Emerging data suggest that excessive release of proteases mediates tissue damage, and therefore prolonged neutrophil accumulation has an important role in the pathogenesis of many diseases. Thus, HNE and Pr3 inhibitors may prove to be targets for the generation of agents in the treatment of neutrophilic inflammatory disease. Sivelestat is the only commercially available selective HNE inhibitor. Therefore, sivelestat was chosen as the model structure in an attempt to obtain more potent anti-NsPs agents. In the present study, a series 2-aminobenzaldehyde oxime and 2-aminobenzoate analogs were synthesized and their inhibitory effects on NsPs (CatG, Pr3, and HNE) were determined, respectively. The results of structure-activity relationships studies concluded that a hydroxyl oxime moiety plays an important role in ligand-enzyme affinity through hydrogen bonding. As compound 6 had more potency and showed dual inhibitory effects on NE and Pr3, both in vitro and in vivo experiments were carried out to evaluate its selectivity, effects in cell-based assays, and efficacy in models of inflammation and damage. Compound 6 had the potential to reduce paw edema induced by LPS and HNE, as well as acute lung injury, and may be approved as a candidate for the development of new agents in the treatment of neutrophilic inflammatory diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bmc.2014.12.056DOI Listing
March 2015

Hydroquinone-salicylic acid conjugates as novel anti-melasma actives show superior skin targeting compared to the parent drugs.

J Dermatol Sci 2014 Nov 4;76(2):120-31. Epub 2014 Sep 4.

Pharmaceutics Laboratory, Graduate Institute of Natural Products, Chang Gung University, Kweishan, Taoyuan, Taiwan; Chinese Herbal Medicine Research Team, Healthy Aging Research Center, Chang Gung University, Kweishan, Taoyuan, Taiwan; Research Center for Industry of Human Ecology, Chang Gung University of Science and Technology, Kweishan, Taoyuan, Taiwan. Electronic address:

Background: Hydroquinone (HQ) and salicylic acid (SA) are drugs for treating melasma through the mechanisms of tyrosinase inhibition and chemical peeling, respectively. Their high frequency of causing skin irritation has led to limited use of both drugs.

Objectives: We designed the new conjugates obtained by joining HQ and SA by the co-drug concept for evaluating cutaneous absorption capability.

Methods: Monoester (4-hydroxyphenyl 2-hydroxybenzoate, HPH) and diester (1,4-phenylene bis(2-hydroxybenzoate), PBH) forms of the conjugates were synthesized and physicochemically characterized. The enzymatic hydrolysis to the parent drugs was examined. Both an equimolar dose and a saturated solubility were utilized as the applied dose for testing cutaneous absorption via pig and nude mouse skins.

Results: The conjugates had higher lipophilicity, less aqueous solubility, and a lower melting point/crystallinity than the parent drugs. Both conjugates showed a quick conversion into the parent drugs in esterases and skin homogenates, with PBH showing the greater hydrolysis. The hydrolysis level in skin after topical application was less as compared to that in esterases and homogenates. The tyrosinase inhibition (%) and molecular docking demonstrated that the conjugates possessed skin-lightening capability (3% for HPH and 7% for PBH) although this activity was lower than that of HQ (23%). The conjugates showed an increased skin deposition compared to the respective parent drugs. Total absorption of HPH and PBH led to a 13- and 19-fold enhancement in cutaneous retention compared to HQ alone. A similar increment of skin deposition was shown for the conjugates when compared to SA. Contrary to skin reservoir retention, transdermal transport across the skin was decreased by the conjugates, especially for PBH. This indicates the maximization of cutaneous targeting by the conjugates.

Conclusions: Topically applied HPH and PBH can be the new candidates for treating melasma due to efficient skin absorption and acceptable skin tolerance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jdermsci.2014.08.013DOI Listing
November 2014

Nanostructured lipid carriers containing a high percentage of a Pluronic copolymer increase the biodistribution of novel PDE4 inhibitors for the treatment of traumatic hemorrhage.

J Biomed Nanotechnol 2014 Aug;10(8):1520-35

Although the application of nanotechnology to drug therapy has been widely investigated, very few nanomedicine-based treatments for traumatic hemorrhage have been reported so far. The aim of this work was to develop nanostructured lipid carriers (NLCs) loaded with phosphodiesterase 4 (PDE4) inhibitors to treat acute inflammation in peripheral organs. The pharmacokinetics and biodistribution of DSM-RX78 and EFB-1, two novel PDE4 inhibitors, were examined using rats as an animal model. Entrapment by NLCs resulted in sustained drug release. The plasma concentrations of DSM-RX78 and EFB-1 in NLCs were lower, and their half-lives were much shorter in the NLC condition than in the control condition. PDE4 inhibitors delivered in NLCs accumulated with high abundance in many organs, especially the brain and lungs. Polyethylene glycol (PEG) coating on the particulate surface (P-NLCs) significantly reduced brain delivery of the drugs. P-NLCs enhanced drug distribution to the lungs by 5-fold compared to free control. In vivo real-time imaging confirmed rapid escape of nanoparticles from the blood circulation. Histological examination and aminotransferase measurement revealed that P-NLCs containing EFB-1 improved hemorrhagic shock-induced injuries in the lungs, intestines, and liver. P-NLCs even reversed acute lung inflammation to the level observed in an uninjured condition. Our results indicate that NLC-based delivery of PDE4 inhibitors is a candidate treatment for traumatic hemorrhage.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1166/jbn.2014.1858DOI Listing
August 2014