Anti-HBV Mechanism And Therapeutic Efficacy Of Reverse Transcriptase Inhibitor Foscarnet

The characteristic of reverse transcription in the replication procedure exists in different kinds of viruses including hepatitis B virus (HBV), human T lymphocyte virus (HILV), human immunodeficiency virus (HIV), Cauliflower mosaic virus (CaMV), Rous sarcoma virus (RSV) et al. It threatens human health and production. Specific antagonist targeting at reverse transcriptase (RT) is the important solution. The drug discovery process usually requires design of high-throughput screening assays against certain targets. Although the conventional isotopic RT assay is sensitive, it is a labor intensive procedure and becoming increasingly restrictive due to the use of radioactive material. It is difficult to be a candidate of high-throughput screening assay.We have established a simple and rapid ELISA method for screening and studying RT specific inhibitors. The template were immobilized onto Covalink microtiter plate, a synthesized biotinlated dUTP was incorporated by RT. The products were detected and quantified using a colorimetric streptavidin-horseradish peroxidase reporter system. Using the HIV-1 RT ELISA assay, we measured the inhibitory activity of foscarnet (PEA) and nevirapine (NVP) on HIV-1 RT (IC50: 0.23 ± 0.12 μ g/ml, 0.21±0.08 μ g/ml) and compared this method with the traditional isotope incorporation assay (r~2=0.997). At the same time, the stability and repeatability of the ELISA method were evaluated, the specificity and sensitivity were also done. The ELISA procedure was simple, quick and reliable to be used as an alternative method for high throughput screening of HIV RT inhibitors. Furthermore, the procedure is also used to detect the activity of Moloney murine leukemia virus reverse transcriptase (M-MLV-RT), Avian Myeloblastosis Virus reverse transcriptase (AMV-RT) and duck hepatitis B virus replication complex (DHBV RCs). The ELISA procedure may be applicable for detecting activity of different viral enzymes characterizing reverse transcription.The regimen of highly active anti-retroviral therapy (HAART) has been employed for HIV-infected patients. Whereas, the treatment of HBV infection falls into difficulties, because 1) there is not enough candidate drugs for combination therapy; 2) drug resistant viruses emerged along with long term monotherapy.