The Screening Of Anti-hepatitis C Virus Reagents

Hepatitis C Virus (HCV) is an enveloped positive-strand RNA virusbelonging to the Flaviviridae family. There are6genotypes of the virus andmore subgenotypes and quasi species in every genotype. HCV can infecthuman beings and primate such as chimpanzee and Macaca rhesus. Theinfection is easy to develop to chronic infection because of the factors of virusand the host. HCV infection is one of the reasons which lead to chronichepatitis, liver cirrhosis and hepatic cellular carcinoma. So it is an importantpublic health problem.The standard therapy for anti-HCV is the combination of PEG-IFN andRBV. The SVR rate of the therapy is differed from the genotypes, which is lessthen50%in the genotype1infection and near80%in the genotype2and3infection. Recently the direct antiviral agents (DAA) was used for the treatmentof HCV infection. With or without the combination of PEG-IFN, the DAAimproved the SVR rate. But there are many side effect of the drugs. ThePEG-IFN can cause fever, influent-like response, arrest of bone marrow,dysfunction of thyroid gland (hyperthyroidism or hypothyroidism), ischemiamyocardial, hyperglycemia, and depression. RBV can cause hyemolysis,jaundice and anorexia. The DAA can also cause the dysfunction ofhematological system. And the PEG-IFN and DAA are expensive. The therapyof PEG-IFN is more than1000yuan every week, while the cost of DAA is2-3times more than PEG-IFN. These conditions inhibit the use of the therapy anddecrease the compliance of the patients. Some patients give up the therapybecause of the cost, and some patients decease the amount of the drugs or cannot complete the whole therapy because of the side effects. From the clinicalobservation, the course of the treatment, the dose and the total dose should be more then80%of the standard therapy which can get the better SVR rate. Sothe decreasing of the treatment make the SVR rate less. The development of theanti-viral reagent is still a hot pot of the research because of the short point ofthe anti-HCV therapy.Polyoxometalates (POMs) are negatively charged clusters of inorganicsubstances. The medicinal properties of POMs have been a subject of interestfor their low cytotoxicity and lower cost. Anti-virus, anti–becteria andanti-tumor activities of POMs have been reported by many groups. But there isstill no report for anti-HCV activity of POMs.Objective: To check the anti HCV activity of the POMs with differentstructures and screen the best POM for anti-HCV reagent, and discover themechanism of the anti-HCV function of POM.Methods: The genotype2a HCV infectious particle JFH1and thehepatoma cell line Huh7.5.1cells were used for the detection of HCV infection.The HCVpp and VSVpp were used for the checking of viral entry. The repliconcells were used for the checking of the HCV replication. The HCV RNA titerswere detected with RT-qPCR. The plaque assay was used for the infection ofFlu V and VSV. Different concentration of POMs were added to theJFH1/Huh7.5.1system. The HCV infectious focus was observed withimmunofluorescence analysis. POMs with different concentration were add toHCVpp and VSVpp to check the effect on the virus entry. HCV replicon cellswere treated with different concentration of POMs to check the effect on theHCV replication of the POMs. The POMs and JFH1viral particle wereincubated directly to check if the POMs can disrupt the HCV particle directely.The effect of POMs on the infection of VSV and Flu V was also checked by theinfection plaque assay of the infection in different concentration of POMs.Result: Polyoxometalates POM#4, POM#6and POM#12were checkedon JFH1/Huh7.5.1system and the result showed the best inhibitory activity of HCV infection of POM#12. The EC50was0.8μM. POM#12inhibited theinfection of HCVpp, in which the EC50was1.19μM, while the inhibitoryactivity on VSVpp was not so obvious. POM#12did not inhibit the replicationof HCV. POM#12disrupted the envelope of the HCV particle and the HCVRNA was not resistant to the RNase without the protection of the envelop. Theinhibitory activity of POM#12on the VSV and Flu V infection was obviouslylower than that on the HCV.Conclusion: POM#12inhibit the HCV infection by disrupting the virusenvelop directely. This activity was HCV specific.