In vitro anti-HIV and -HSV activity and safety of sodium rutin sulfate as a microbicide candidate

Sodium rutin sulfate (SRS) is a sulfated rutin modified from the natural flavonol glycoside rutin. Here, we investigated its in vitro anti-HIV and -HSV activities and its cytotoxic profile. Fifty percent inhibitory concentration (IC(50)) values of SRS against HIV-1 X4 virus IIIB, HIV-1 R5 isolates Ada-M and Ba-L were 2.3+/-0.2, 4.5+/-2.0 and 8.5+/-3.8 microM with a selectivity index (SI) of 563, 575 and 329, respectively. Its IC(50) against primary R5 HIV-1 isolate from Yunnan province in China was 13.1+/-5.5 microM, with a SI of 197. In contrast, unsulfated rutin had no activity against any of the HIV-1 isolates tested. Further study indicated that SRS blocked viral entry and virus-cell fusion likely through interacting with the HIV-1 envelope glycoprotein. SRS also demonstrated some activity against human herpes simplex virus (HSV) with an IC(50) of 88.3+/-0.1 microM and a SI of 30. The 50% cytotoxicity concentration (CC(50)) of SRS was >3.0 mM, as determined in human genital ME180, HeLa and primary human foreskin fibroblast cells. Minimum inhibitory concentration of SRS for vaginal lactobacilli was >3.0 mM. These results collectively indicate that SRS represents a novel candidate for anti-HIV-1/HSV microbicide development.     

In vitro anti-HIV activity of oleanolic acid on infected human mononuclear cells

Oleanolic acid is a triterpenoid which is quite common in nature in the form either of free acid or in triterpenoid saponin glycosides. This study describes the effect of oleanolic acid on the growth of human immunodeficiency virus-1 (HIV-1) in cultures of human peripheral mononuclear cells (PBMC) and of monocyte/macrophages (M/M). Its inhibitory activity was also evaluated on PBMC obtained from HIV-1 infected patients. Results obtained show that oleanolic acid inhibits the HIV-1 replication in all the cellular systems used (EC50 values: 22.7 microM, 24.6 microM and 57.4 microM for in vitro infected PBMC, naturally infected PBMC and M/M, respectively). As regards the mechanism of action, oleanolic acid inhibits in vitro the HIV-1 protease activity.     

Effect of limonin and nomilin on HIV-1 replication on infected human mononuclear cells

In the last years several plant-derived natural compounds have been screened for their anti-HIV activity in order to find lead compounds with novel structures or mechanisms of action. Among these, several triterpenoids have been found to exhibit an antiretroviral activity with different mechanisms of action. In this study the effect of two limonoids, limonin and nomilin, on the growth of human immunodeficiency virus-1 (HIV-1) in culture of human peripheral blood mononuclear cells (PBMC) and on monocytes/macrophages (M/M) is described. Limonin and nomilin were found to inhibit the HIV-1 replication in all cellular systems used. A dose-dependent inhibition of viral replication was observed in PBMC isolated from healthy donors and infected with HIV-1 strain after incubation with limonin and nomilin (EC (50) values: 60.0 microM and 52.2 microM, respectively). The two terpenoids inhibited at all concentrations studied the production of HIV-p24 antigen even when the PBMC employed were chronically infected (EC (50) values of 61.0 microM for limonin and 76.2 microM for nomilin). Moreover, these compounds inhibited the HIV-1 replication even in infected M/M. In this cellular system the inhibitory effect was significant at the concentrations of 20 microM, 40 microM and 80 microM starting from day 14 and reached the maximum effect after 18 days of incubation. As regards the mechanism of action, limonin and nomilin inhibit in vitro HIV-1 protease activity. In general, the results obtained point out a similar anti-HIV activity of limonin and nomilin indicating that this activity is not drastically influenced by the structural difference between the two compounds.     

基于假病毒技术探讨黄芩苷抗HIV-1活性研究

目的:基于假病毒技术探究黄芩苷抗HIV-1活性作用.方法:利用HIV-1骨架质粒NL4-3 mCherry Luciferase、HIV-1包膜蛋白质粒pCMV-VSV-G和人胚肾细胞HEK-293T构建HIV-1假病毒药物筛选体系,并对该体系进行优化和安全性验证.测定黄芩苷对293T细胞活性、HIV-1假病毒活性、H...     

Structure-specific inhibition of cholesteryl ester transfer protein by azaphilones.

The effect of thirteen different fungal azaphilones, which have a common 6-iso-chromane-like ring, was tested on cholesteryl ester transfer protein (CETP) activity in vitro. Chaetoviridin B showed the most potent inhibitory activity with an IC50 value of < 6.2 microM, followed by sclerotiorin with an IC50 value of 19.4 microM. Rotiorin, chaetoviridin A and rubrorotiorin had moderate inhibitory activity (IC50 ; 30 approximately 40 microM), but others showed very weak or no inhibitory activity. The relationship between the structures and their inhibitory activity indicated that the presence of an electrophilic ketone(s) and/or enone(s) at both C-6 and C-8 positions in the isochromane-like ring is essential for eliciting CETP inhibitory activity. The transfer activity of both CE and TG was inhibited by sclerotiorin to approximately the same extent (IC50: 14.4 and 10.3 microM, respectively). A model of the reaction suggested that sclerotiorin reacts with a primary amine of amino acids such as lysine in the protein to form a covalent bond.     

Styrylquinazoline derivatives as HIV-1 integrase inhibitors

Styrylquinazoline derivatives were prepared by Perkin condensation and evaluated for inhibitory activity against HIV-1 integrase. Among them, compound 5c containing a free catechol ring was the most potent (IC(50)=0.8 +/- 1.9 microM)and showed 6-fold more potency than the corresponding styrylquinoline compound (IC(50) = 130.7 +/- 8.6 microM).     

Synthesis of styrylbenzofuran derivatives as styrylquinoline analogues for HIV-1 integrase inhibitor

A series of styrylbenzofuran derivatives (8a-i) as styrylquinoline isosters were efficiently prepared by Wittig reaction and evaluated for inhibitory activity against HIV-1 integrase. In this series, compounds 8g, 8h and 8i containing a free catechol ring showed moderate inhibitory activities (IC50= approximately 40 microM) against HIV-1 integrase, while less than the corresponding styrylquinoline compound (I).     

Indolocarbazoles exhibit strong antiviral activity against human cytomegalovirus and are potent inhibitors of the pUL97 protein kinase

We have analyzed a panel of protein kinase inhibitors (PKIs) and found that some indolocarbazoles (G?6976, K252a, K252c) proved to be highly effective inhibitors of GCV-sensitive and -resistant human cytomegalovirus (HCMV) strains, but did not show any effect against herpes simplex virus. Antiviral activity was determined by focus reduction assays (IC(50) ranging from 0.009 to 0.4 microM). Other inhibitors of serine/threonine kinases (G?6850, H-7, roscovitine) were found to be ineffective. Virus yield at 5 days after infection was reduced by three orders of magnitude with nanomolar concentrations of the indolocarbazoles. These compounds were fully effective when added up to 24 h post infection and showed reduced activity up to 72 h post infection. Cytotoxicity assays in proliferating and non-proliferating cells demonstrated that the effective antiviral concentration of these compounds was significantly lower than either antiproliferative (IC(50)/CC(50) ranging from 6.5 to 390) or cytotoxic (IC(50)/CC(50) ranging from 72. 5 to 1000) doses. The effects of PKIs on the virus-encoded protein kinase pUL97 were studied using recombinant vaccinia viruses. Indolocarbazoles strongly inhibited both pUL97 autophosphorylation (IC(50) ranging from 0.0012 to 0.013 microM) and pUL97-dependent ganciclovir phosphorylation (IC(50) ranging from 0.05 to 0.26 microM). Other inhibitors of serine/threonine kinases showed only weak (G?6850) or no (H-7, roscovitine) effect on these pUL97 functions, while oxoflavone tyrosine kinase inhibitors had no effect at all.     

Inhibition of VHR dual-specificity protein tyrosine phosphatase activity by flavonoids isolated from Scutellaria baicalensis: structure-activity relationships

Three flavonoids: norwogonin, dihydronorwogonin and baicalein, were isolated from the roots of Scutellaria baicalensis, as potential inhibitors of VHR dual-specificity protein tyrosine phosphatase (DS-PTPase). Norwogonin (IC 50 = 1.1 microM), dihydronorwogonin (IC 50 = 2.9 microM) and baicalein (IC 50 = 2.4 microM) showed potent inhibitory activity toward VHR, but had no inhibitory activity against T-cell protein tyrosine phosphatase or serine/threonine protein phosphatase 1. From comparisons to the inhibitory activities of other similar flavonoids, it could be suggested that the presence of a hydroxy group in the B ring of flavonoids interferes with the inhibitory activity toward VHR DS-PTPase.     

Inhibition of prostaglandin E(2) production by taiwanin C isolated from the root of Acanthopanax chiisanensis and the mechanism of action

Five lignans, l-sesamin, savinin, helioxanthin, taiwanin C, and cis-dibenzylbutyrolactone, were isolated from the root of Acanthopanax chiisanensis (Araliaceae), a Korean medicinal plant, and their inhibitory effects on the production of prostaglandin (PG) E(2) stimulated by 12-O-tetradecanoylphorbol 13-acetate (TPA) in rat peritoneal macrophages were examined. Among the five lignans, taiwanin C was the most potent (IC(50)=0.12 microM), followed by helioxanthin, cis-dibenzylbutyrolactone, and savinin. l-Sesamin had no effect. Taiwanin C showed no inhibitory effect on the TPA-induced release of radioactivity from [3H]arachidonic acid-labeled macrophages, nor did it inhibit the expression of cyclooxygenase (COX)-2 protein induced by TPA. However, the activities of isolated COX-1 and COX-2 were inhibited by taiwanin C (IC(50)=1.06 and 9.31 microM, respectively), reflecting the inhibition of both COX-1- and COX-2-dependent PGE(2) production in the cell culture system. These findings suggest that the mechanism of action of taiwanin C in the inhibition of PGE(2) production is the direct inhibition of COX enzymatic activity.     

Inhibitory activity of flavonoids and tannins against HIV-1 protease

Twenty-nine flavonoids and six hydrolyzable tannins were studied for their inhibitory activity against human immunodeficiency virus (HIV)-1 protease using fluorescence and HPLC assays. Among the flavonoids, flavones, flavanones, flavonols, catechols and chalcones, the flavonols were the most active category while flavanones and catechols displayed low activity. Quercetin was the most potent inhibitor of the target enzyme with an IC50 value of 58.8 microM, while butein and luteolin showed moderate activity. Of the hydrolyzable tannins tested, three ellagitannins which contain a hexahydroxvdiphenoyl (HHDP) unit linked to the O-3 and 0-6 positions of the sugar, were found to strongly inhibit HIV-1 protease. The IC50 values of corilagin and repandusinic acid on HIV-1 protease were 20.7 and 12.5 microM, respectively.     

Inhibition of lipopolysaccharide-induced nitric oxide production by flavonoids in RAW264.7 macrophages involves heme oxygenase-1

The role of heme oxygenase-1 (HO-1) played in the inhibitory mechanism of flavonoids in lipopolysaccharide (LPS)-induced responses remained unresolved. In the present study, flavonoids, including 3-OH flavone, baicalein, kaempferol, and quercetin, induced HO-1 gene expression at the protein and mRNA levels in the presence or absence of LPS in RAW264.7 macrophages. This effect was associated with suppression of LPS-induced nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) protein expression. Hemin induced HO-1 protein expression and this was associated with the suppression of LPS-induced NO production and iNOS protein expression in a dose-dependent manner. In addition, an increase in bilirubin production was found in flavonoid- and hemin-treated cells. Hemin, at the doses of 10, 20, and 50 microM, dose-dependently stimulated the flavonoid (50 microM)-induced HO-1 protein expression, and enhanced their inhibitory effects on LPS-induced NO production and iNOS protein expression. Pretreatment of the HO-1 inhibitor, tin protoporphyrin (10 microM), attenuated the inhibitory activities of the indicated flavonoids on LPS-induced NO production. Morphologic analysis showed that 3-OH flavone, baicalein, kaempferol, quercetin, hemin, and tin protoporphyrin did not cause any change in cell viability in the presence or absence of LPS. In contrast, only 3-OH flavone showed a significant inhibition of cell growth using the MTT assay. Transfection of an HO-1 vector in macrophages (HO-1/RAW264.7) resulted in a 3-fold increase in HO-1 protein compared with that the parental RAW264.7 cells. NO production mediated by LPS in HO-1 over-expressed RAW264.7 cells (HO-1/RAW264.7) was significant less than that in parental RAW264.7 cells. 3-OH Flavone, baicalein, kaempferol, and quercetin showed a more significant inhibition on LPS-induced NO production in HO-1/RAW264.7 cells than in parental RAW264.7 cells. These results provide evidence on the role of HO-1 in the inhibition of LPS-induced NO production by flavonoids. A combination of HO-1 inducers (i.e. hemin) and flavonoids might be an effective strategy for the suppression of LPS-induced NO production.     

Inhibitory effects of some traditional medicines on proliferation of HIV-1 and its protease

In attempts to discover anti-HIV agents from natural sources, various traditional medicine extracts were tested for their inhibitory effects on HIV-1 proliferation and its protease. An extract of the seeds of Croton tiglium showed potent inhibitory effects on the proliferation of HIV-1. The active principle was determined to be phorbol esters. Several derivatives of phorbol ester were evaluated for inhibition of proliferation as well as activation of protein kinase C (PKC). Of these compounds, 12-O-acetylphorbol 13-decanoate (6) showed the most potent inhibition of proliferation without activating PKC. Some triterpenes from the stems of Cynomorium songaricum and the woody part of Xanthoceras sorbifolia showed inhibitory activity against HIV-1 protease. Various derivatives of oleanolic acid were synthesized and evaluated for their inhibitory activity against HIV-1 protease. Their inhibitory mechanism was also investigated.