Eun-Hye Hong

Antiviral activity of ginsenosides against coxsackievirus B3, enterovirus 71, and human rhinovirus 3

Background Ginsenosides are the major components responsible for the biochemical and pharmacological actions of ginseng, and have been shown to have various biological activities. In this study, we investigated the antiviral activities of seven ginsenosides [protopanaxatriol (PT) type: Re, Rf, and Rg2; protopanaxadiol (PD) type: Rb1, Rb2, Rc, and Rd)] against coxsackievirus B3 (CVB3), enterovirus 71 (EV71), and human rhinovirus 3 (HRV3). Methods Assays of antiviral activity and cytotoxicity were evaluated by the sulforhodamine B method using the cytopathic effect (CPE) reduction assay. Results The antiviral assays demonstrated that, of the seven ginsenosides, the PT-type ginsenosides (Re, Rf, and Rg2) possess significant antiviral activities against CVB3 and HRV3 at a concentration of 100 μg/mL. Among the PT-type ginsenosides, only ginsenoside Rg2 showed significant anti-EV71 activity with no cytotoxicity to cells at 100 μg/mL. The PD-type ginsenosides (Rb1, Rb2, Rc, and Rd), by contrast, did not show any significant antiviral activity against CVB3, EV71, and HRV3, and exhibited cytotoxic effects to virus-infected cells. Notably, the antiviral efficacies of PT-type ginsenosides were comparable to those of ribavirin, a commonly used antiviral drug. Conclusion Collectively, our findings suggest that the ginsenosides Re, Rf, and Rg2 have the potential to be effective in the treatment of CVB3, EV71, and HRV3 infection.

Antiviral Activity of Hederasaponin B from Hedera helix against Enterovirus 71 Subgenotypes C3 and C4a.

Enterovirus 71 (EV71) is the predominant cause of hand, foot and mouth disease (HFMD). The antiviral activity of hederasaponin B from Hedera helix against EV71 subgenotypes C3 and C4a was evaluated in vero cells. In the current study, the antiviral activity of hederasaponin B against EV71 C3 and C4a was determined by cytopathic effect (CPE) reduction method and western blot assay. Our results demonstrated that hederasaponin B and 30% ethanol extract of Hedera helix containing hederasaponin B showed significant antiviral activity against EV71 subgenotypes C3 and C4a by reducing the formation of a visible CPE. Hederasaponin B also inhibited the viral VP2 protein expression, suggesting the inhibition of viral capsid protein synthesis.These results suggest that hederasaponin B and Hedera helix extract containing hederasaponin B can be novel drug candidates with broad-spectrum antiviral activity against various subgenotypes of EV71.

Blockade of Myd88 signaling induces antitumor effects by skewing the immunosuppressive function of myeloid‐derived suppressor cells

Myd88 is an important adaptor molecule for the activation of NADPH oxidase and arginase‐1, which are responsible for the suppressive function of myeloid‐derived suppressor cells (MDSCs). When wild‐type and Myd88−/− mice were subcutaneously injected with CT26 colon cancer cells expressing human Her‐2/neu, tumor growth was retarded in Myd88−/− mice than in wild‐type mice. Although the generation of CD11b+Gr‐1+ MDSCs was less in Myd88−/− mice than in wild‐type mice, Myd88−/− mice having tumor masses still had significant quantities of MDSCs, suggesting that MDSC generation might be independent of Myd88 signaling. However, MDSCs obtained from tumor‐bearing Myd88−/− mice failed to suppress antigen‐specific proliferation of CD8+ T cells and CD4+ T cells, whereas MDSCs from wild‐type mice significantly suppressed both types of T cells. Consistent with this, we found that the levels of costimulatory molecules and MHC class II were significantly increased in MDSCs obtained from Myd88−/− mice compared with wild‐type mice after tumor challenge. Furthermore, CD4+ T cells residing in tumor‐draining lymph nodes of Myd88−/− mice secreted more TNF‐α than those of wild‐type mice. Finally, the blockade of Myd88 signaling by treatment with Myd88 inhibitory peptide, during later tumor stages, significantly inhibited the growth of immunogenic tumors. Overall, these data suggest that signaling through the Myd88 adaptor molecule is critical for the direct suppressive function of MDSCs and approaches to block Myd88‐mediated signaling in MDSCs might be effective to inhibit the immunosuppressive function of MDSCs.

Intratumoral injection of attenuated Salmonella vaccine can induce tumor microenvironmental shift from immune suppressive to immunogenic

Attenuated Salmonella vaccines show therapeutic anti-cancer effects, but the underlying mechanism has not been well investigated. In the current study, intratumoral (i.t.) injection of recombinant attenuated Salmonella enterica serovar Typhimurium vaccine (RASV) significantly inhibited Her-2/neu-expressing tumor growth. Although depletion of CD8(+) cells in RASV-treated mice significantly restored tumor growth, the induction of Her-2/neu-specific cytotoxic T lymphocytes (CTLs) was not well correlated with the generation of the anti-tumor effect. Therefore, we hypothesized that RASV might induce a tumor microenvironmental shift, from immune suppressive to immunogenic, to reduce the suppressive force and finally elicit a successful anti-tumor response. We found that i.t. injection of RASV significantly increased the level of CD11b(+)Gr-1(+) myeloid cells identified as myeloid-derived suppressor cell (MDSC), but a significant portion of these cells were TNF-α-secreting Ly6-G(high) subsets, which can function as antitumor effector cells. We further investigated whether RASV can modulate immunosuppressive Treg cells, and CD4(+)CD25(+) Foxp3(+) Tregs was significantly reduced in RASV-treated mice. Thus, i.t. injection of RASV may offer a novel anti-cancer approach by eliciting transformation of immunosuppressive MDSCs into TNF-α-secreting neutrophils and reducing the generation of Treg cells, especially in the presence of tumor-specific CTLs. Collectively, these data will provide us an insight for the development of new anti-tumor approaches to overcome the immunosuppressive environment generated by tumors.

α-Mangostin Reduced ER Stress-mediated Tumor Growth through Autophagy Activation

α-Mangostin is a xanthon derivative contained in the fruit hull of mangosteen (Garcinia mangostana L.), and the administration of α-Mangostin inhibited the growth of transplanted colon cancer, Her/CT26 cells which expressed Her-2/neu as tumor antigen. Although α-Mangostin was reported to have inhibitory activity against sarco/endoplasmic reticulum Ca2+ ATPase like thapsigargin, it showed different activity for autophagy regulation. In the current study, we found that α-Mangostin induced autophagy activation in mouse intestinal epithelial cells, as GFP-LC3 transgenic mice were orally administered with 20 mg/kg of α-Mangostin daily for three days. However, the activation of autophagy by α-Mangostin did not significantly increase OVA-specific T cell proliferation. As we assessed ER stress by using XBP-1 reporter system and phosphorylation of eIF2α, thapsigargin-induced ER stress was significantly reduced by α-Mangostin. However, coadministration of thapsigargin with α-Mangostin completely blocked the antitumor activity of α-Mangostin, suggesting ER stress with autophagy blockade accelerated tumor growth in mouse colon cancer model. Thus the antitumor activity of α-Mangostin can be ascribable to the autophagy activation rather than ER stress induction.

Boronic acid-tethered amphiphilic hyaluronic acid derivative-based nanoassemblies for tumor targeting and penetration.

(3-Aminomethylphenyl)boronic acid (AMPB)-installed hyaluronic acid-ceramide (HACE)-based nanoparticles (NPs), including manassantin B (MB), were fabricated for tumor-targeted delivery. The amine group of AMPB was conjugated to the carboxylic acid group of hyaluronic acid (HA) via amide bond formation, and synthesis was confirmed by spectroscopic methods. HACE-AMPB/MB NPs with a 239-nm mean diameter, narrow size distribution, negative zeta potential, and >90% drug encapsulation efficiency were fabricated. Exposed AMPB in the outer surface of HACE-AMPB NPs (in the aqueous environment) may react with sialic acid of cancer cells. The improved cellular accumulation efficiency, in vitro antitumor efficacy, and tumor penetration efficiency of HACE-AMPB/MB NPs, compared with HACE/MB NPs, in MDA-MB-231 cells (CD44 receptor-positive human breast adenocarcinoma cells) may be based on the CD44 receptor-mediated endocytosis and phenylboronic acid-sialic acid interaction. Enhanced in vivo tumor targetability, infiltration efficiency, and antitumor efficacies of HACE-AMPB NPs, compared with HACE NPs, were observed in a MDA-MB-231 tumor-xenografted mouse model. In addition to passive tumor targeting (based on an enhanced permeability and retention effect) and active tumor targeting (interaction between HA and CD44 receptor), the phenylboronic acid-sialic acid interaction can play important roles in augmented tumor targeting and penetration of HACE-AMPB NPs. STATEMENT OF SIGNIFICANCE: (3-Aminomethylphenyl)boronic acid (AMPB)-tethered hyaluronic acid-ceramide (HACE)-based nanoparticles (NPs), including manassantin B (MB), were fabricated and their tumor targeting and penetration efficiencies were assessed in MDA-MB-231 (CD44 receptor-positive human adenocarcinoma) tumor models. MB, which exhibited antitumor efficacies via the inhibition of angiogenesis and hypoxia inducible factor (HIF)-1, was entrapped in HACE-AMPB NPs in this study. Phenylboronic acid located in the outer surface of HACE-AMPB/MB NPs (in the aqueous milieu) may react with the sialic acid over-expressed in cancer cells and intramolecular B‒O bond can be formed. This phenylboronic acid-sialic acid interaction may provide additional tumor targeting and penetration potentials together with an enhanced permeability and retention (EPR) effect (passive tumor targeting) and HA-CD44 receptor interaction (active tumor targeting). Developed HACE-AMPB NP may be one of promising nanocarriers for the imaging and therapy of CD44 receptor-expressed cancers.

Interleukin-10 attenuates tumour growth by inhibiting interleukin-6/signal transducer and activator of transcription 3 signalling in myeloid-derived suppressor cells

Interleukin-10 (IL-10) is a well-characterized anti-inflammatory cytokine, but its role in anti-cancer immunity is controversial. After injection with TC-1 cancer cells, we observed more rapid tumour growth and significantly higher interleukin-6 (IL-6) production in IL-10 knockout (IL-10(-/-)) mice than wild-type (WT) mice. Blocking IL-6 with an anti-IL-6 receptor (IL-6R) monoclonal antibody (mAb) inhibited tumour growth and myeloid-derived suppressor cell (MDSC) generation, which were significantly increased in IL-10-deficient mice. MDSCs and tumour cells from IL-10(-/-) mice had increased phosphorylated signal transducer and activator of transcription 3 (p-STAT3) levels. Treatment with a STAT3 inhibitor, S3I, reduced tumour growth, inhibited MDSC expansion, reduced IL-6 in tumours, and relieved T cell suppression. The combination of anti-IL-6R mAb and S3I further inhibited tumour growth compared to S3I treatment alone. These results suggested that the inhibition of the IL-6/STAT3 signalling axis is a candidate anti-cancer strategy, especially under systemic inflammatory conditions with high IL-6.

Antiviral Activity of Oroxylin A against Coxsackievirus B3 Alleviates Virus-Induced Acute Pancreatic Damage in Mice

The flavonoids mosloflavone, oroxylin A, and norwogonin, which were purified from Scutellaria baicalensis Georgi, significantly protected Vero cells against Coxsackievirus B3 (CVB3)-induced cell death. To investigate the in vivo antiviral activity of oroxylin A, we intraperitoneally inoculated CVB3 into 4-week-old BALB/c mice. Body weights and blood glucose levels of the mice were decreased after CVB3 infection, and these changes were attenuated by the administration of oroxylin A. Importantly, treatment of mice with oroxylin A reduced viral titers in the pancreas and decreased the serum levels of the inflammatory cytokines including interleukin-6 (IL-6) and tumor necrosis factor (TNF)-α. Additionally, the administration of oroxylin A mitigated the histological pancreatic lesions and apoptotic cell death induced by CVB3 infection and increased the levels of phospho-eIF2α in infected pancreata. The results suggest that oroxylin A may represent a potent antiviral agent against CVB3 infection.

Steamed and Fermented Ethanolic Extract from Codonopsis lanceolata Attenuates Amyloid-β-Induced Memory Impairment in Mice.

Codonopsis lanceolata (C. lanceolata) is a traditional medicinal plant used for the treatment of certain inflammatory diseases such as asthma, tonsillitis, and pharyngitis. We evaluated whether steamed and fermented C. lanceolata (SFC) extract improves amyloid-β- (Aβ-) induced learning and memory impairment in mice. The Morris water maze and passive avoidance tests were used to evaluate the effect of SFC extract. Moreover, we investigated acetylcholinesterase (AChE) activity and brain-derived neurotrophic factor (BDNF), cyclic AMP response element-binding protein (CREB), and extracellular signal-regulated kinase (ERK) signaling in the hippocampus of mice to determine a possible mechanism for the cognitive-enhancing effect. Saponin compounds in SFC were identified by Ultra Performance Liquid Chromatography-Quadrupole-Time-of-Flight Mass Spectrometry (UPLC-Q-TOF-MS). SFC extract ameliorated amyloid-β-induced memory impairment in the Morris water maze and passive avoidance tests. SFC extract inhibited AChE activity and also significantly increased the level of CREB phosphorylation, BDNF expression, and ERK activation in hippocampal tissue of amyloid-β-treated mice. Lancemasides A, B, C, D, E, and G and foetidissimoside A compounds present in SFC were determined by UPLC-Q-TOF-MS. These results indicate that SFC extract improves Aβ-induced memory deficits and that AChE inhibition and CREB/BDNF/ERK expression is important for the effect of the SFC extract. In addition, lancemaside A specifically may be responsible for efficacious effect of SFC.

Coadministration of Hedera helix L. Extract Enabled Mice to Overcome Insufficient Protection against Influenza A/PR/8 Virus Infection under Suboptimal Treatment with Oseltamivir

Several anti-influenza drugs that reduce disease manifestation exist, and although these drugs provide clinical benefits in infected patients, their efficacy is limited by the emergence of drug-resistant influenza viruses. In the current study, we assessed the therapeutic strategy of enhancing the antiviral efficacy of an existing neuraminidase inhibitor, oseltamivir, by coadministering with the leaf extract from Hedera helix L, commonly known as ivy. Ivy extract has anti-inflammatory, antibacterial, antifungal, and antihelminthic properties. In the present study, we investigated its potential antiviral properties against influenza A/PR/8 (PR8) virus in a mouse model with suboptimal oseltamivir that mimics a poor clinical response to antiviral drug treatment. Suboptimal oseltamivir resulted in insufficient protection against PR8 infection. Oral administration of ivy extract with suboptimal oseltamivir increased the antiviral activity of oseltamivir. Ivy extract and its compounds, particularly hedrasaponin F, significantly reduced the cytopathic effect in PR8-infected A549 cells in the presence of oseltamivir. Compared with oseltamivir treatment alone, coadministration of the fraction of ivy extract that contained the highest proportion of hedrasaponin F with oseltamivir decreased pulmonary inflammation in PR8-infected mice. Inflammatory cytokines and chemokines, including tumor necrosis factor-alpha and chemokine (C-C motif) ligand 2, were reduced by treatment with oseltamivir and the fraction of ivy extract. Analysis of inflammatory cell infiltration in the bronchial alveolar of PR8-infected mice revealed that CD11b+Ly6G+ and CD11b+Ly6Cint cells were recruited after virus infection; coadministration of the ivy extract fraction with oseltamivir reduced infiltration of these inflammatory cells. In a model of suboptimal oseltamivir treatment, coadministration of ivy extract fraction that includes hedrasaponin F increased protection against PR8 infection that could be explained by its antiviral and anti-inflammatory activities.