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Proprotein convertases and serine protease inhibitors : developing novel indirect-acting antiviral strategies against hepatitis c virus

University of British Columbia

Hepatitis C virus (HCV) utilizes host lipids for every stage of its lifecycle. HCV hijacks host lipid droplets (LDs) to coordinate assembly through the host lipoprotein assembly pathway; this facilitates uptake into hepatocytes through the low density lipoprotein receptor (LDLR). Induction of host lipid metabolism by HCV supports chronic infection and leads to steatosis, exacerbating liver dysfunction in infected patients. One pathway activated by HCV is the sterol regulatory element binding protein (SREBP) pathway which controls lipid metabolism gene expression. To activate genes in the nucleus, SREBPs must first be cleaved by host subtilisin kexin isozyme-1/site-1 protease (SKI-1/S1P). Proprotein convertase subtilisin/kexin type 9 (PCSK9) is one SREBP-regulated protein that post-translationally decreases LDLR expression in the liver. The overall aim of this thesis was to determine the potential application of these two important regulators of host lipid homeostasis, PCSK9 and SKI-1/S1P, as targets for inhibiting HCV infection. The first hypothesis tested was that inhibiting SKI-1/S1P would block HCV hijacking of the SREBP pathway and limit sequestration of host lipids by HCV, blocking virus propagation. To inhibit SKI-1/S1P function, an engineered serine protease inhibitor (serpin) and a small molecule inhibitor were employed. Both inhibitors were shown to block SKI-1/S1P cleavage of SREBPs, reduce LD accumulation in hepatoma cells and inhibit HCV infection. The next hypothesis explored was that amplifying PCSK9 expression or function in hepatoma cells would increase their resistance to HCV infection through downregulation of LDLR. It was confirmed, using overexpression of wild-type PCSK9 or treating cells with gain-of-function PCSK9, that PCSK9 can be used to prevent HCV entry into hepatoma cells. Finally, studies are presented detailing the discovery and characterization of a non-inhibitory serpin variant with potent antiviral activity against HCV infection. It is hypothesized that inhibition may be related to antiviral functions exhibited by other human serpins or serpin-derived peptides possessing diverse regulatory properties. Host lipid metabolism is a critical component of the lifecycle of HCV and many other viruses. These studies confirm that lipid metabolism pathways can be rationally targeted to inhibit viral infection and may lead to the development of novel, indirect-acting therapies against HCV and related viruses.

Text

2013-06-30

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/39818

Microbiology and Immunology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/