Anemoside B4 inhibits SARS-CoV-2 replication in vitro and in vivo / 中草药·英文版

OBJECTIVE@#Anemoside B4 (AB4), the most abundant triterpenoidal saponin isolated from Pulsatilla chinensis, inhibited influenza virus FM1 or Klebsiella pneumoniae-induced pneumonia. However, the anti-SARS-CoV-2 effect of AB4 has not been unraveled. Therefore, this study aimed to determine the antiviral activity and potential mechanism of AB4 in inhibiting human coronavirus SARS-CoV-2 in vivo and in vitro.@*METHODS@#The cytotoxicity of AB4 was evaluated using the Cell Counting Kit-8 (CCK8) assay. SARS-CoV-2 infected HEK293T, HPAEpiC, and Vero E6 cells were used for in vitro assays. The antiviral effect of AB4 in vivo was evaluated by SARS-CoV-2-infected hACE2-IRES-luc transgenic mouse model. Furthermore, label-free quantitative proteomics and bioinformatic analysis were performed to explore the potential antiviral mechanism of action of AB4. Type I IFN signaling-associated proteins were assessed using Western blotting or immumohistochemical staining.@*RESULTS@#The data showed that AB4 reduced the propagation of SARS-CoV-2 along with the decreased Nucleocapsid protein (N), Spike protein (S), and 3C-like protease (3CLpro) in HEK293T cells. In vivo antiviral activity data revealed that AB4 inhibited viral replication and relieved pneumonia in a SARS-CoV-2 infected mouse model. We further disclosed that the antiviral activity of AB4 was associated with the enhanced interferon (IFN)-β response via the activation of retinoic acid-inducible gene I (RIG-1) like receptor (RLP) pathways. Additionally, label-free quantitative proteomic analyses discovered that 17 proteins were significantly altered by AB4 in the SARS-CoV-2 coronavirus infections cells. These proteins mainly clustered in RNA metabolism.@*CONCLUSION@#Our results indicated that AB4 inhibited SARS-CoV-2 replication through the RLR pathways and moderated the RNA metabolism, suggesting that it would be a potential lead compound for the development of anti-SARS-CoV-2 drugs.

Synthesis and biological evaluation of a series of 2-(((5-akly/aryl-1-pyrazol-3-yl)methyl)thio)-5-alkyl-6-(cyclohexylmethyl)-pyrimidin-4(3)-ones as potential HIV-1 inhibitors

A series of 2-(((5-akly/aryl-1-pyrazol-3-yl)methyl)thio)-5-alkyl-6-(cyclohexylmethyl)-pyrimidin-4(3)-ones were synthesized and their anti-HIV-1 activities were evaluated. Most of these compounds were highly active against wild-type (WT) HIV-1 strain (IIIB) with EC values in the range of 0.0038-0.4759 μmol/L. Among those compounds, had an EC value of 3.8 nmol/L and SI (selectivity index) of up to 25,468 indicating excellent activity against WT HIV-1. anti-HIV-1 activity and resistance profile studies suggested that compounds and displayed potential anti-HIV-1 activity against laboratory adapted strains and primary isolated strains including different subtypes and tropism strains (ECs range from 4.3 to 63.6 nmol/L and 18.9-219.3 nmol/L, respectively). On the other hand, it was observed that those two compounds were less effective with EC values of 2.77 and 4.87 μmol/L for HIV-1A (K103N + Y181C). The activity against reverse transcriptase (RT) was also evaluated for those compounds. Both and obtained sub-micromolar IC values showing their potential in RT inhibition. The pharmacokinetics examination in rats indicated that compound has acceptable pharmacokinetic properties and bioavailability. Preliminary structure-activity relationships and molecular modeling studies were also discussed.

Optimization and application of VSVG/HIV-1 NL4-3 Luc system for screening of anti-HIV-1 compounds / 中国药理学通报

Aim To establish and optimize the VSVG/HIV-1NL4-3 Luc pseudovirus model for anti-HIV drugs screening. Methods The infectivity of VSVG/HIV-1 NL4-3 Luc in 4 different cell lines was investigated according to the method of the lucifer-ase activity analysis system of Promega company. 3 different ex-perimental settings were used to detect the activities of approved anti-HIV drugs to confirm the feasibility and effectiveness of the system. Finally, some potential compounds were screened for their anti-HIV activities, and their antiviral activities against the pseudovirus were compared with HIV-1ⅢB . Results The pseud-ovirus showed the strongest replication ability in CRFK cells, and a clear dose-effect relationship was found between the report gene expression level and the virus quantity. Comparing the EC50 of different positive inhibitors against VSVG/HIV-1 NL4-3 Luc on 3 kinds of experimental conditions, 3rd scheme is the best. Finally, the system was used to screen compounds, the EC50 s a-gainst pseudovirus were similar to those in HIV-1ⅢB . Conclusion An optimized VSVG/HIV-1 NL4-3 Luc anti-HIV screening sys-tem has been successfully developed.

Study on Vitro Anti-HIV-1 Activity ofTaiqi PeiyuanGranules / 中国中医药信息杂志

ObjectiveTo evaluate the vitro anti-HIV-1 activity ofTaiqi Peiyuan Granules.MethodsMTT was used to detect cytotoxicity ofTaiqi Peiyuan Granules; cytopathy method was used to detect the inhibitory activity of Taiqi Peiyuan Granules on acute infection of HIV-1; HIV-1 p24 antigen ELISA detection was used to detect inhibitory activity ofTaiqi Peiyuan Granules for virus replication of HIV-1 acute infection cells, and count the medical therapeutic indexes.ResultsCC50ofTaiqi Peiyuan Granules in cytotoxicity test was 3.761±0.370 mg/mL; the EC50 of inhibition syncytial test was 0.454 5±0.204 6 mg/mL; the therapeutic index was between 5.84 and 12.97; p24 ofTaiqi Peiyuan Granules in inhibition experiments was 0.56±0.27 mg/mL, and the therapeutic index was between 5.30 and 8.74.ConclusionAnti-HIV-1 activity ofTaiqi Peiyuan Granules is relatively weak.

Screening of Active Compounds Against HIV-1 Reverse Transcriptase Based on Molecular Docking / 广州中医药大学学报

Objective To screen the non-nucleoside compounds against HIV-1 reverse transcriptase by molecular modeling and bioactivity assay. Methods Surflex-Dock module of Tripos SYBYL software was used to simulate the binding pattern of 22 000 compounds in SPECS database with the active pocket of HIV-1 reverse transcriptase. Based on the simulation results, the interaction mode between the above compounds and the crystal structure of HIV-1 reverse transcriptase was analyzed. The compounds with higher docking scores and better binding pattern were determined by anti-HIV-1 ac tivities test in vitro. Results The virtual screening results showed that the docking conformation of 1- (4-fluorophenyl) -3- [2- (1H-indol-3-yl) ethyl] thiourea was similar to the embedded ligand in Rilpivirine crystal structure. 1- ( 4-fluorophenyl) -3- [ 2- ( 1H-indol-3-yl) ethyl] thiourea was held together with the key residue Lys101 in docking pocket of HIV-1 reverse transcriptase by hydrogen bonds, and hadπ-πstacking action together with the conservative residue Trp229 and the aromatic residue Tyr181 respectively. The bioassay in vitro results showed that when the proliferation rate of C8166 lymphocyte syncytium infected by HIV-1ⅢB arrived 50% ( EC50) , the concentration of 1- ( 4-fluorophenyl) -3- [ 2- ( 1H-indol-3-yl) ethyl] thiourea was 5.45μg/mL. Conclusion Molecule docking technology is an effective approach to reducing the screening of candidate compounds with micromolecular activity, and can be used to predict the interaction mode between the compound and the target receptor. In the study, active compound 1- (4-fluorophenyl) -3- [2- (1H-indol-3-yl) ethyl] thiourea has been screened out by molecule docking technology.

Antiviral drug screening based on pseudoviruses:research advances / 国际药学研究杂志

The pseudotype virus is a virus nucleic acids packaged by another virus envelope protein, to form the virus with the exogenous virus envelope and maintain virus genome characteristics of itself. Due to the loss of its ability to self-replicate and high safety, pseudoviruses are widely used as safe substitutes for infectious virus in virology studies, such as HIV, H5N1 and HCV Here we will introduce the advantages and limitations of pseudoviruses, and through summarizing a large number of research achievements, we prove pseudoviruses are feasible and effective as an antiviral drug screening platform. The platform is safe and reliable and it will make more contribution in antiviral drug research.

Establishment and optimization of a method for screening HIV-1 integrase 3′-processing inhibitor / 中国药理学通报

Aim To establish and optimize a method for screening HIV-1 integrase 3′-processing inhibitor. Methods Fluorescence resonance energy transfer ( FRET) was used to create an assay for screening in-tegrase 3′-processing inhibitors; wavelength was de-fined by DNaseⅠ; factors affecting IN activity were optimized, including buffer composition, substrate con-centration, enzyme concentration, metal ion concentra-tion. Results Integrase 3′-processing optimizing reac-tion conditions were buffer 1 , 500 nmol · L-1 sub-strate, 1 μmol·L-1 integrase, 20mmol·L-1 magne-sium ion. Positive drug raltegravir and myricetin could effectively inhibit integrase 3′-processing activity using this assay. Two integrase 3′-processing inhibitors were screened by this method. Conclusion The method for screening HIV-1 integrase 3′-processing inhibitor is successfully established and optimized.

The anti-HIV activity of three 2-alkylsulfanyl-6-benzyl-3, 4-dihydropyrimidin-4 (3H)-one derivatives acting as non-nucleoside reverse transcriptase inhibitor in vitro / 药学学报

It was recently shown that several new synthetic 2-alkylsulfanyl-6-benzyl-3, 4-dihydropyrimidin-4(3H)-one (S-DABO) derivatives demonstrated anti-HIV-1 activity. Three of the derivatives namely RZK-4, RZK-5 and RZK-6 were used in this study to explore their inhibitory effects on a variety of HIV strains. These compounds at a concentration of 200 microg mL(-1) almost completely inhibited the activity of recombinant HIV-1 reverse transcriptase. All of the three compounds reduced replication of HIV-1 laboratory-derived strains, low-passage clinical isolated strain, and the drug resistant strain. In particular RZK-6 showed potent activity against the HIV-1 drug resistant strain. In general, the antiviral activities are similar in magnitude to nevirapine (NVP), which is a non-nucleoside reverse transcriptase inhibitor approved by FDA. The therapeutic indexes of these compounds were remarkable, ranging from 3704 to 38462 indicating extremely low cytotoxicity. These results suggest that the three S-DABO derivatives in this study have good potential for further development in anti-HIV-1 therapy. It may be particularly useful to target at the non-nucleoside reverse transcriptase inhibitors resistant HIV-1 strain.

Advances in the study of anti-HIV natural compounds derived from traditional Chinese medicines / 药学学报

Anti-HIV drugs still remain as the dominant role in the treatment of acquired immunodeficiency syndrome (AIDS), because no vaccine was found till today. Owing to structural diversity, few side effects, and abundant resources, natural compounds from traditional Chinese medicines and medicinal plants have unique advantages and good potential in prevention and treatment of AIDS. Many researchers have made great efforts in the field of anti-HIV natural compounds, and have found some natural compounds from traditional Chinese medicines with potent anti-HIV activities. These compounds can be classified into the following categories: alkaloids, coumarins, lignans, flavonoids, terpenoids, tannins, polysaccharides, proteins and peptides, and polyphenols. However, most of these researches are performed in vitro, and most natural compounds show weak anti-HIV activities and indefinite acting targets. In the paper, we reviewed some natural compounds derived from traditional Chinese medicines with potent anti-HIV activities in recent years.

Establishment of AIDS Animal Model with SIVmac239 Infected Chinese Rhesus Monkey / 中国病毒学·英文版

In the present research, two Chinese rhesus monkeys were inoculated intravenously with 5000 TCID50 of SIVmac239. The changes in the numbers of CD4+T lymphocyte in peripheral blood,plasma viral loads, proviral DNA and humoral antibodies against virus were periodically monitored during 121 days. At the early stage of infection, proviral DNA had been detected in PBMCs, and infectious SIVmac239 virus had been isolated from PBMCs. At the same period, the numbers of CD4+T lymphocytes were significantly decreased, and maintained at low level during the 121-day period of infection. Plasma viral loads reached the peak at week 2 post-inoculation and kept at a steady state subsequently. Moreover, antibodies against viral proteins were detected from plasma. All the results showed that the two Chinese rhesus monkeys had been infected with SIVmac239 successfully. This animal model can be applied for further AIDS researches.

HIV protease inhibitors and apoptosis / 中国药理学通报

CD4 +T cells from patients with human immunodeficiency virus (HIV) infection undergo apoptosis at an enhanced rate, which leads to immune system damage and invalidation to withstand the virus intrusion. The current antiviral drugs aim at the virus can not clear the chronically infected cells which are the virus reservoirs. But treatment of HIV/AIDS patients with the HIV protease inhibitors (PIs) can reduce the cell apoptosis induced by HIV infection, which can improve immune function, and that is separate from the antiviral effects of PIs. Such suggests that immune reconstruction is a new strategy to cure AIDS. This text reviews HIV PIs research and development, their function characteristics and the effects on cell apoptosis. To define the relationship between HIV PIs and cell apoptosis may inspire new ways of curing the AIDS.-