Identification of the phytobioactive Polygonum cuspidatum as an antiviral source for restricting dengue virus entry.

Dengue virus (DENV) is a mosquito-borne pathogen that is becoming a serious global threat, owing to its rising incidence in inter-tropical regions that yield over 50 million annual infections. There are currently no approved antiviral agents for the management of dengue, and recent shortcomings in its immunization called for immediate action to develop effective drugs with prophylactic ability to better manage its infection. In an attempt to discover novel antiviral sources, we identified the medicinal herb Polygonum cuspidatum (PC) as a bioactive botanical material against DENV infectivity. Specifically, the methanolic extract from PC rhizomes (PCME) potently inhibited DENV infection without causing significant cytotoxicity. Further examination on the viral life cycle demonstrated that PCME particularly targeted the initial stages of DENV infection, while pre- and post-infection treatments had no effect. More importantly, the PCME could efficiently inactivate DENV free virus particles and block the viral attachment and entry/fusion events without apparently influencing viral replication, egress, and cell-to-cell spread. The antiviral effect of PCME was also recapitulated in infection analysis using DENV pseudoparticles displaying viral structural proteins that mediate DENV particle entry. Besides, PCME treatment also inhibited direct DENV entry into several cell types relevant to its infection and reduced viral infectivity of other members of the Flaviviridae family, including the hepatitis C virus (HCV) and Zika virus (ZIKV). Due to its potency against DENV entry, we suggest that the phytobioactive extract from PC is an excellent starting point as an antiviral source material for further development of therapeutic strategies in the prophylactic management of DENV infection.

Targeting Autophagy Augments BBR-Mediated Cell Death in Human Hepatoma Cells Harboring Hepatitis C Virus RNA.

Hepatocellular carcinoma (HCC), including hepatitis C virus (HCV)-induced HCC, is a deadly disease highly refractory to chemotherapy, thus requiring the continuous identification of novel treatment strategies. Berberine (BBR) has been previously reported to inhibit hepatoma cell growth, but the main type of cell death elicited by BBR, and whether the alkaloid can inhibit hepatoma cells carrying HCV genomes, is unclear. Herein, we show that BBR treatment induced a biphasic cell death irrespective of the presence of HCV subgenomic replicon RNA, first triggering apoptosis that then progressed to necrosis between 24 and 48 h post-treatment. Furthermore, BBR treatment potentiated the HCV replicon-induced reactive oxygen species (ROS) production, inhibition of which with an antioxidant attenuated the cell death that was elicited by BBR in these cells. Moreover, BBR dampened the autophagic response in HCV RNA-positive or negative hepatoma cells, and pharmacological inhibition of autophagy conversely augmented the BBR-induced cell death. Finally, BBR inhibited the growth of Huh-7 cells that were persistently infected with the full-length genome HCV particles, and concomitant pharmacological inhibition of autophagy potentiated the killing of these cells by BBR. Our findings suggest that combining BBR with the inhibition of autophagy could be an attractive treatment strategy against HCC, irrespective of the presence of the HCV genome.

Berberine inhibits hepatitis C virus entry by targeting the viral E2 glycoprotein.

BACKGROUND: Despite the advent of direct-acting antivirals (DAAs), HCV remains an important public health problem globally. There is at present no effective vaccine against the virus, and the DAAs in current use cannot prevent de novo infection, including in liver transplant setting wherein donor livers inevitably become re-infected. Developing inhibitors to HCV entry using nature-derived small molecules may help to expand/complement the current treatment options. PURPOSE: In this study, we explored the effect of the plant alkaloid berberine (BBR) on HCV early viral entry. METHODS: Cell culture-derived HCV (HCVcc), viral pseudoparticles bearing HCV glycoproteins (HCVpp), and entry-related assays were employed to assess BBR's bioactivity. Molecular docking was used to predict BBR-HCV glycoproteins interaction, and the compound's antiviral activity was confirmed against HCVcc infection of primary human hepatocytes (PHHs). RESULTS: BBR specifically impeded HCVcc attachment and entry/fusion steps without inactivating the free virus particles or affecting the expression of host cell entry factors and post-entry viral replication. BBR also effectively inhibited infection by viral pseudoparticles expressing HCV E1/E2 glycoproteins and molecular docking analysis pointed at potential interaction with HCV E2. Finally, BBR could suppress HCVcc infection of PHHs. CONCLUSIONS: We identified BBR as a potent HCV entry inhibitor, which merits further evaluation particularly for use in transplant setting against graft re-infection by HCV.

Identification of baicalin from Bofutsushosan and Daisaikoto as a potent inducer of glucose uptake and modulator of insulin signaling-associated pathways.

Type 2 diabetes mellitus (T2DM) is a chronic disease characterized by hyperglycemia that can lead to long-term complications including heart diseases, stroke, retinopathy, and renal failure. Treatment strategies include stimulating glucose uptake and controlling blood glucose level. Bofutsushosan (BOF) and Daisaikoto (DAI) are two herb-based kampo medicines that have been demonstrated to improve metabolism-associated disorders including obesity, hyperlipidemia, and nonalcoholic fatty liver. Given their bioactivities against metabolic syndromes, we explored in this study the effect of BOF and DAI extracts on glucose absorption and used them as source to identify phytochemical stimulator of glucose absorption. Glucose uptake and mechanistic studies were evaluated in differentiated C2C12 skeletal muscle cells, and HPLC analysis was used to determine the molecular bioactive constituents. Our results indicated that the ethanolic extracts of BOF and DAI (BOFEE and DAIEE, respectively) enhanced the glucose uptake ratio in the differentiated C2C12 cells, and further analysis identified the flavone baicalin as a major constituent capable of efficiently stimulating glucose absorption. Mechanistic studies revealed that the effect from baicalin involved the activation of IRS-1 and GLUT-4, and implicated the AMPK, PI3K/Akt, and MAPK/ERK signaling cascades. Due to its potency, we suggest that baicalin merit further evaluation as a potential candidate anti-hyperglycemic agent for the treatment and management of T2DM.

Methanolic Extract of Rhizoma Coptidis Inhibits the Early Viral Entry Steps of Hepatitis C Virus Infection.

Hepatitis C Virus (HCV) remains an important public health threat with approximately 170 million carriers worldwide who are at risk of developing hepatitis C-associated end-stage liver diseases. Despite improvement of HCV treatment using the novel direct-acting antivirals (DAAs) targeting viral replication, there is a lack of prophylactic measures for protection against HCV infection. Identifying novel antivirals such as those that target viral entry could help broaden the therapeutic arsenal against HCV. Herein, we investigated the anti-HCV activity of the methanolic extract from Rhizoma coptidis (RC), a widely used traditional Chinese medicine documented by the WHO and experimentally reported to possess several pharmacological functions including antiviral effects. Using the cell culture-derived HCV system, we demonstrated that RC dose-dependently inhibited HCV infection of Huh-7.5 cells at non-cytotoxic concentrations. In particular, RC blocked HCV attachment and entry/fusion into the host cells without exerting any significant effect on the cell-free viral particles or modulating key host cell entry factors to HCV. Moreover, RC robustly suppressed HCV pseudoparticles infection of Huh-7.5 cells and impeded infection by several HCV genotypes. Collectively, our results identified RC as a potent antagonist to HCV entry with potential pan-genotypic properties, which deserves further evaluation for use as an anti-HCV agent.

Activity-based and fraction-guided analysis of Phyllanthus urinaria identifies loliolide as a potent inhibitor of hepatitis C virus entry.

Without a vaccine, hepatitis C virus (HCV) remains a global medical and socio-economic burden, predisposing about 170 million carriers worldwide to end-stage liver diseases including cirrhosis and hepatocellular carcinoma. Although the recently developed direct-acting antivirals (DAAs) have revolutionized hepatitis C treatment, most of them are unsuitable for monotherapy due to risks of resistance, thus necessitating combination with interferon (IFN)-alpha, ribavirin, or additional DAAs. More importantly, the high cost associated with the DAAs restricts their accessibility to most parts of the world. Developing novel cost-effective anti-HCV therapeutics may help expand the scope of antivirals and treatment strategies against hepatitis C. Herein, we applied an activity-based and fraction-guided analysis of extracts from the medicinal plant Phyllanthus urinaria (P. urinaria), which yielded fraction 13 (F13) as possessing the most potent inhibitory activity against early viral entry of cell-culture HCV infection. Chemical analysis (silica gel chromatography followed by ESI LC-MS plus (1)H and (13)C NMR) of F13 identified loliolide (LOD), a monoterpenoid lactone, as a novel inhibitor of HCV entry. Specifically, LOD could efficiently inactivate HCV free virus particles, abrogate viral attachment, and impede viral entry/fusion, with minimal effect on viral replication/translation, particle production, and induction of type I IFN host antiviral immune response. ELISA-based binding analysis confirmed the monoterpenoid's ability in efficiently blocking HCV particle attachment to the host cell surface. Furthermore, LOD could inhibit infection by several genotypic strains of HCV. This is the first report characterizing P. urinaria and its bioactive compound LOD as potent HCV entry inhibitors, which merit further evaluation for development as candidate antiviral agents against hepatitis C.