Small molecule effects on Hepatitis C Virus

Abstract

In this thesis, chemical biology approaches were used to study Hepatitis C Virus (HCV) in non-infectious, subgenomic HCV replicon model systems. Specifically, small molecules were used as probes for studying host-virus interactions. Compared to genetic techniques, small molecules are interesting potential therapeutics for HCV because they can diffuse relatively quickly to their targets and also act on proteins after they have been posttranslationally modified. Small molecules are also much easier to deliver to cells. In our first study we looked on the effects of using a small molecule agonists and antagonist of the host receptor, peroxisome proliferator activated receptor alpha (PPARalpha) on the HCV viral life cycle. We found that impaired function of PPARalpha perturbs lipid homeostasis within the cells, causing inhibition of HCV replication. We confirmed this effect by silencing the PPARalpha gene using small interfering (si)RNA molecules. Our next project involved screening a library of abscisic acid (ABA) analogs for anti-HCV activity. ABA is a plant hormone, structurally similar to its counterpart in humans, retinoic acid. We found three ABA analogs that have anti-HCV activity comparable to known HCV inhibitors like lovastatin and 25-hydroxycholesterol. We identified the protein targets for one of the active ABA analogs by performing in-vivo experiments using an inhibitor-based profiling approach with a synthesized rhodamine fluorescent probe. Target identification allowed us to postulate a mechanism by which the ABA analog realizes its anti-HCV activity by interfering with the host protein responsible for both host and HCV protein folding. Finally, we demonstrated anti-HCV activity of the antitumor drug bleomycin (BLM). BLM causes HCV inhibition by directly targeting and degrading the HCV RNA. Although toxic to the host cell due to DNA degradation, we showed that BLM acts much faster on viral RNA located in the cytoplasm than on host DNA located in the nucleus.