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 Inhibitors of the NS3 Proteinase of West Nile Virus |
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Specific Aim: Identify drug-like small molecule inhibitors of the NS3 West Nile virus protease by high throughput screening of chemical libraries.
Future plans include optimization the structure of the novel, low-molecular weight, synthetic inhibitors of the West Nile and Dengue virus NS3 proteinase and to validate the selectivity and potency of the selected compound leads in additional in vitro tests and assays. In addition, we will determine, at the atomic resolution level, the structure of the NS3 protease bound to lead antagonists. Recent exciting discoveries directly related to this particular project [1] and vast amounts of structure-activity data that should flow from this project should facilitate creation of therapeutically important, anti-viral, drugs.
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PMLSC WNV NS2B-NS3 Probe Report
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| HTS Assays for Inhibitors of HIV RNase H |
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Specific Aim: The conversion of HIV genomic RNA into DNA comprises multiple steps, each catalyzed by the essential viral enzyme, reverse transcriptase (RT). To catalyze this conversion, RT is multifunctional, possessing both DNA polymerase activity (RNA-dependent and DNA-dependent) and ribonuclease H (RNH) activity. Eleven of the therapeutics currently used for the clinical treatment of HIV infection targets the viral DNA polymerase, reverse transcriptase (RT). Seemingly thousands of inhibitors of RT DNA polymerase activity have been identified, and all clinically used antivirals targeting RT are directed at this activity. Viral resistance to these drugs is an increasingly serious problem in the therapeutic management of HIV infection, and new drugs are needed, especially drugs that inhibit new targets of HIV. RT RNH is absolutely essential for HIV replication, and therefore represents a logical target for antiviral development. Unfortunately, very few inhibitors of HIV RT RNH have been described, and none are in preclinical development, let alone in clinical use. One of the reasons for the scarcity of RNH inhibitors has been the lack of an HTS assay to measure RNH activity.
Traditional gel based RNH assays are cumbersome and time consuming, unsuitable for high throughput screening (HTS) of large libraries of compounds. We have developed a rapid and simple fluorescence based assay for RNH and have validated this assay in HTS screens using 96- and 384-well microtiter plates. We therefore believe that this assay is suitable for use in the Molecular Libraries Screening Center Network (MLSCN). To this end, we propose the following Specific Aims to be carried out by the MLSCN:
1. To transfer and implement our HTS assay technology in the MLSCN.
2. To validate hits obtained in the primary screen.
3. To evaluate the cytotoxicity of validated hits.
4. To conduct preliminary structural optimization of selected validated hits.
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PMLSC HIV RNase H Probe Report
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