SKI-1/S1P inhibition: A promising surrogate to statins to block Hepatitis C virus replication Matthieu Blanchet a , Nabil G. Seidah b , Patrick Labonté a,⇑ a INRS-Institut Armand-Frappier, Institut National de la Recherche Scientifique, Laval, Canada b Laboratory of Biochemical Neuroendocrinology, Institut de Recherches Cliniques de Montréal, Affiliated to the Université de Montréal, Montréal, Canada article info Article history: Received 2 February 2012 Revised 10 May 2012 Accepted 13 May 2012 Available online 22 May 2012 Keywords: HCV SKI-1/S1P HMG-CoAR FASn Statins Lipids metabolism abstract Hepatitis C virus (HCV) is often associated with steatosis, cirrhosis and hepatocellular carcinoma (HCC). Statins (HMG-CoAR inhibitors) have been shown to exert an antiviral effect in vitro, principally on repli- con harboring cells, but the effect of their use alone in vivo remains controversial. In clinical trials, when used in combination with the standards of care (SOC), they led to an increased proportion of sustained virological responder (SVR). Here we investigated the implication of SKI-1/S1P, a master lipogenic path- ways regulator upstream of HMG-CoAR, on different steps of HCV life cycle. We compared the HCV anti- viral effect of the most potent SKI-1/S1P small molecule inhibitor (PF-429242) with a set of two statins on different steps of the viral life cycle, and showed that SKI-1/S1P inhibitor blocked HCVcc (strain JFH-1) RNA replication (EC 50 = 5.8 lM) more efficiently than statins. Moreover, we showed that PF-429242 could reduce lipid droplets accumulation in Huh7 cells. Interestingly, PF-429242 dramatically reduced infec- tious particles production (EC 90 = 4.8 lM). Such inhibition could not be achieved with statins. SKI-1/ S1P activity is thus essential for viral production and its inhibition should be considered for antiviral drug development. Ó 2012 Published by Elsevier B.V. 1. Introduction The hepatitis C virus (HCV) is known to be a prominent factor for the onset of end-stage liver diseases such as liver cirrhosis and hepatocellular carcinoma (HCC) (Di Bisceglie, 1997). World- wide, approximately 170 million patients are chronically infected, and it has been estimated that over 300,000 individuals die annu- ally from HCV infection. The recent addition of HCV protease inhib- itors to the standard treatment consisting of pegylated interferon- a (PEG-IFN) and ribavirin has significantly increased viral clear- ance in patients (Hezode et al., 2009; Manns et al., 2012). However, 15–30% of patients still do not respond to treatments and severe side effects are common in treated individuals (Fried et al., 2002). Many studies have highlighted the close relationship between HCV life cycle and lipids metabolism (Herker and Ott, 2011). Statins, HMG-CoA reductase inhibitors that are widely used in human to lower cholesterol levels (Carroll et al., 2005), have been tested for HCV-antiviral properties. In vitro, several statins have shown potent antiviral activity in cells harboring HCV replicon, but lower effect were obtained in cell infected with HCVcc (Delang et al., 2009; Ikeda et al., 2006). Because of their low toxicity profile in vivo, statins have been studied in humans. While their antiviral properties alone re- mains controversial (Bader et al., 2008; Harrison et al., 2010; Mila- zzo et al., 2009; O’Leary et al., 2007), their use in combination with PEG-IFN and ribavirin seems to have positive effect (Harrison et al., 2010). The discrepancy observed in human could be partially explained by the wide antiviral effectiveness of different statins. However the divergence between various statins in their ability to inhibit HCV in vitro is still not understood, emphasizing the complex interrelation of HCV lifecycle and lipids metabolism. Because of the variable and limited antiviral effects obtained with statins so far, we sought to investigate the antiviral potency of an upstream regulator of cholesterol and fatty acid metabolic pathways. Thus, we concentrated on the eighth member of the pro- protein convertase family, namely subtilisin/kexin-isoenzyme-1 (SKI-1) or site-1 protease (S1P), now designated as SKI-1/S1P (Sei- dah and Prat, 2007). This pyrolysin-like serine protease is a mem- brane-bound enzyme, which after autocatalytic activation cleaves a defined set of Golgi-associated membrane-bound transcription factors (Seidah and Prat, 2007). SKI-1/S1P plays a key role in regu- lating cholesterol and fatty acids pathways by cleaving SREBP-1a, -1c, and -2 (Fig. 1) (reviewed in Horton et al., 2002). Additionally, 0166-3542/$ - see front matter Ó 2012 Published by Elsevier B.V. http://dx.doi.org/10.1016/j.antiviral.2012.05.006 Abbreviations: HCV, hepatitis C virus; HCC, hepatocellular carcinoma; SOC, standards of care; SKI-1/S1P, subtilisin/kexin-isoenzyme-1/site-1 protease; SREBP, sterol regulatory-element binding protein; LD, lipid droplet; VLDL, very low density lipoprotein; PEG-IFN-a, pegylated interferon-a. ⇑ Corresponding author. Address : INRS-Institut Armand-Frappier, Institut National de la Recherche Scientifique, 531 Boulevard des Prairies, Laval, Canada H7 V 1B7. Tel.: +1 450 687 5010; fax: +1 450 686 5314. E-mail address: patrick.labonte@iaf.inrs.ca (P. Labonté). Antiviral Research 95 (2012) 159–166 Contents lists available at SciVerse ScienceDirect Antiviral Research journal homepage: www.elsevier.com/locate/antiviral