Search

Aim:To discover novel hepatitis C virus (HCV) inhibitors and elucidate the mechanism of action of the active compounds.Methods:HCV subgenomic replicon-based luciferase reporter cell line was used to screen 1200 synthetic compounds with novel structures. Huh7.5.1 cell line stably transfected with HCV NS3/4A protease reporter was established to investigate the anti-HCV mechanism of the active compounds. The active compounds were further examined in an in vitro HCV infection assay to confirm their anti-HCV activity.Results:After two-round screening in the anti-HCV replicon assay, some 2,4-diaminoquinazoline derivatives and carboxamide analogues were found to possess anti-HCV replicon activities (the IC50 values were less than 5 μmol/L). Among them, two representative compounds HZ-1157 and LZ-110618-6 inhibited HCV NS3/4A protease with IC50 values of 1.0 and 0.68 μmol/L, respectively. Furthermore, HZ-1157 and LZ-110618-6 inhibited HCV infection in vitro with IC50 values of 0.82 and 0.11 μmol/L, respectively.Conclusion:Some 2,4-diaminoquinazoline derivatives and carboxamide analogues have been identified as novel anti-HCV compounds. [ABSTRACT FROM AUTHOR]

Aim:To investigate the action of isothiafludine (NZ-4), a derivative of bis-heterocycle tandem pairs from the natural product leucamide A, on the replication cycle of hepatitis B virus (HBV) in vitro and in vivo.Methods:HBV replication cycle was monitored in HepG2.2.15 cells using qPCR, qRT-PCR, and Southern and Northern blotting. HBV protein expression and capsid assembly were detected using Western blotting and native agarose gel electrophoresis analysis. The interaction of pregenomic RNA (pgRNA) and the core protein was investigated by RNA immunoprecipitation. To evaluate the anti-HBV effect of NZ-4 in vivo, DHBV-infected ducks were orally administered NZ-4 (25, 50 or 100 mg·kg−1·d−1) for 15 d.Results:NZ-4 suppressed intracellular HBV replication in HepG2.2.15 cells with an IC50 value of 1.33 μmol/L, whereas the compound inhibited the cell viability with an IC50 value of 50.4 μmol/L. Furthermore, NZ-4 was active against the replication of various drug-resistant HBV mutants, including 3TC/ETV-dual-resistant and ADV-resistant HBV mutants. NZ-4 (5, 10, 20 μmol/L) concentration-dependently reduced the encapsidated HBV pgRNA, resulting in the assembly of replication-deficient capsids in HepG2.2.15 cells. Oral administration of NZ-4 dose-dependently inhibited DHBV DNA replication in the DHBV-infected ducks.Conclusion:NZ-4 inhibits HBV replication by interfering with the interaction between pgRNA and HBcAg in the capsid assembly process, thus increasing the replication-deficient HBV capsids. Such mechanism of action might provide a new therapeutic strategy to combat HBV infection. [ABSTRACT FROM AUTHOR]

Aim:To examine the therapeutic effects and underlying mechanisms of DZ2002, a reversible S-adenosyl-L-homocysteine hydrolase (SAHH) inhibitor, on lupus-prone female NZB×NZW F1 (NZB/W F1) mice.Methods:Female NZB/W F1 mice were treated orally with DZ2002 (0.5 mg·kg−1·d−1) for 11 weeks, and the proteinuria level and body weight were monitored. After the mice ware euthanized, serum biochemical parameters and renal damage were determined. Splenocytes of NZB/W F1 mice were isolated for ex vivo study. Toll-like receptor (TLR)-stimulated human peripheral blood mononuclear cells (PBMCs) or murine bone marrow-derived dendritic cells (BMDCs) were used for in vitro study.Results:Treatment of the mice with DZ2002 significantly attenuated the progression of glomerulonephritis and improved the overall health. The improvement was accompanied by decreased levels of nephritogenic anti-dsDNA IgG2a and IgG3 antibodies, serum IL-17, IL-23p19 and TGF-β. In ex vivo studies, treatment of the mice with DZ2002 suppressed the development of pathogenic Th17 cells, significantly decreased IL-17, TGF-β, IL-6, and IL-23p19 production and impeded activation of the STAT3 protein and JNK/NF-κB signaling in splenocytes. DZ2002 (500 μmol/L) significantly suppressed TLR agonists-stimulated up-regulation in IL-6, IL-12p40, TNF-α, and IgG and IgM secretion as well as in HLA-DR and CD40 expression of dendritic cells among human PBMCs in vitro. DZ2002 (100 μmol/L) also significantly suppressed TLR agonists-stimulated up-regulation in IL-6 and IL-23p19 production in murine BMDCs, and prevented Th17 differentiation and suppressed IL-17 secretion by the T cells in a BMDC-T cell co-culture system.Conclusion:DZ2002 effectively ameliorates lupus syndrome in NZB/W F1 mice by regulating TLR signaling-mediated antigen presenting cell (APC) responses. [ABSTRACT FROM AUTHOR]

Aim:To investigate the immunomodulating activity of astragalosides, the active compounds from a traditional tonic herb Astragalus membranaceus Bge, and to explore the molecular mechanisms underlying the actions, focusing on CD45 protein tyrosine phosphatase (CD45 PTPase), which plays a critical role in T lymphocyte activation.Methods:Primary splenocytes and T cells were prepared from mice. CD45 PTPase activity was assessed using a colorimetric assay. Cell proliferation was measured using a [3H]-thymidine incorporation assay. Cytokine proteins and mRNAs were examined with ELISA and RT-PCR, respectively. Activation markers, including CD25 and CD69, were analyzed using flow cytometry. Activation of LCK (Tyr505) was detected using Western blot analysis. Mice were injected with the immunosuppressant cyclophosphamide (CTX, 80 mg/kg), and administered astragaloside II (50 mg/kg).Results:Astragaloside I, II, III, and IV concentration-dependently increased the CD45-mediated of pNPP/OMFP hydrolysis with the EC50 values ranged from 3.33 to 10.42 μg/mL. Astragaloside II (10 and 30 nmol/L) significantly enhanced the proliferation of primary splenocytes induced by ConA, alloantigen or anti-CD3. Astragaloside II (30 nmol/L) significantly increased IL-2 and IFN-γ secretion, upregulated the mRNA levels of IFN-γ and T-bet in primary splenocytes, and promoted CD25 and CD69 expression on primary CD4+ T cells upon TCR stimulation. Furthermore, astragaloside II (100 nmol/L) promoted CD45-mediated dephosphorylation of LCK (Tyr505) in primary T cells, which could be blocked by a specific CD45 PTPase inhibitor. In CTX-induced immunosuppressed mice, oral administration of astragaloside II restored the proliferation of splenic T cells and the production of IFN-γ and IL-2. However, astragaloside II had no apparent effects on B cell proliferation.Conclusion:Astragaloside II enhances T cell activation by regulating the activity of CD45 PTPase, which may explain why Astragalus membranaceus Bge is used as a tonic herb in treating immunosuppressive diseases. [ABSTRACT FROM AUTHOR]

Objective SM934, an artemisinin derivative, possesses potent antiproliferative and antiinflammatory properties. The aim of this study was to examine the effects and explore the mechanisms of SM934 to treat autoimmune disease in lupus-prone female MRL/ lpr mice. Methods In vitro, the effects of SM934 on the activation of polyclonal CD4+ T cells and the differentiation of naive CD4+ T cells were examined. In vivo, the preventative or therapeutic effects of SM934 in MRL/ lpr mice were investigated. Ex vivo, the mechanisms of treatment were explored according to the immunologic correlates of disease. Results In vitro, SM934 inhibited interferon-γ (IFNγ) and interleukin-17 (IL-17) production from polyclonal CD4+ T cells activated by T cell receptor engagement and the differentiation of naive CD4+ T cells into Th1 and Th17 cells, but not Treg cells. In vivo, 12-week-old MRL/ lpr mice treated with SM934 for 4 weeks showed significantly ameliorated proteinuria and renal lesion severity; decreased levels of blood urea nitrogen, serum IFNγ, and serum anti-double-stranded DNA antibodies; decreased spleen size; and a lower percentage of CD3+B220+CD4−CD8− T cells; 16-week-old MRL/ lpr mice treated with SM934 for 8 weeks avoided severe proteinuria and survived longer. Ex vivo, SM934 treatment elevated the percentage of Treg cells, inhibited the development of Th1 and Th17 cells, and impeded the comprehensive activation of STAT-1, STAT-3, and STAT-5 proteins in splenocytes. Conclusion Taken together, the results of this study demonstrated that the artemisinin analog SM934 had therapeutic effects in lupus-prone female MRL /lpr mice by inhibiting both Th1 cell and Th17 cell responses. Moreover, this study indicated that both IFNγ and IL-17 are required for the elicitation and development of murine lupus. [ABSTRACT FROM AUTHOR]

Abstract: In the present study, we investigated the immunosuppressive effects and underlying mechanisms of β-aminoarteether maleate (SM934), a derivative of artemisinin, against T cell activation in vitro and in vivo. In vitro, SM934 significantly inhibited the proliferation of splenocytes induced by concanavalin A (Con A), lipopolysaccharide (LPS), mixed lymphocyte reaction (MLR), and anti-CD3 plus anti-CD28 (anti-CD3/28). SM934 significantly inhibited interferon (IFN)-γ production and CD4+ T cell division stimulated by anti-CD3/28. SM934 also promoted apoptosis of CD69+ population in CD4+ T cells stimulated by anti-CD3/28. Furthermore, SM934 inhibited interleukin (IL)-2 mediated proliferation and survival through blocking Akt phosphorylation in activated T cells. In ovalbumin (OVA)-immunized mice, oral administration of SM934 suppressed OVA-specific T cell proliferation and IFN-γ production. SM934 treatment also significantly inhibited the sheep red blood cell (SRBC)-induced delayed type hypersensitivity (DTH) reactions in mice. Taken together, SM934 showed potent immunosuppressive activities in vitro and in vivo. Our results demonstrated that SM934 might be a potential therapeutic agent for immune-related diseases. [Copyright &y& Elsevier]

Abstract: Sixteen novel 4-triazole-modified zanamivir (1) analogues were synthesized using the click reactions, and their inhibitory activities against avian influenza virus (AIV, H5N1) were determined. Compound 3b exerts promising inhibitory activity with EC50 of 6.4μM, which is very close to that of zanamivir (EC50 =2.8μM). Molecular modeling provided the information about the binding model between inhibitors and neuraminidase, which are in good agreement with inhibitory activities. [Copyright &y& Elsevier]