Together, these results indicate that blocking the LDLR with mAb MK-8669 mw C7 affects the ratio of several host cell lipids. To further investigate the role of the LDLR in HCV entry, we used sLDLR to determine its effect on HCV infectivity. sLDLR inhibited HCVcc infectivity in a dose-dependent manner (Fig. 4). There was no effect of sLDLR on SINV infectivity, a virus that is not dependent on LDLR for its entry, and sLDLR had minimal effect on HCVpp infectivity (Fig. 4).

Together, these results suggest that the LDLR can potentially interact with HCVcc. In the blood, lipoproteins are known to be processed by LPL, an enzyme that modifies triglyceride-rich lipoproteins and targets them to the liver. LPL hydrolyzes triglycerides in VLDL and mediates interactions with HSPGs, enabling lipoprotein clearance from the circulation.28 Because of the potential role of lipoproteins in HCV entry, LPL might potentially affect HCV entry. Indeed, LPL has been shown to increase serum-derived HCV binding to target cells and to inhibit virus infection,29 suggesting that LPL might promote HCV internalization

by the same mechanism as lipoprotein uptake, leading to nonproductive virus uptake. Furthermore, the modification of HCVcc-associated lipoproteins by LPL seems to account for the loss of infectivity.30 To further investigate the effect of LPL on HCV entry, HCVcc were preincubated with LPL at 4°C or 37°C in the presence PAK6 or absence of the LPL inhibitor, GSI-IX order THL, followed by viral infection at 4°C or 37°C, respectively. When HCVcc were incubated with LPL at 4°C, infectivity did not change, unless the THL inhibitor was present. In the latter condition, a 1 log10 increase in infectivity was observed (Fig. 5A). In contrast, when HCVcc were incubated with LPL at 37°C, infectivity was reduced by nearly 1 log10 in the absence of THL,

whereas the level of infectivity was similar as the treatment at 4°C when the THL was added (Fig. 5A). Similar experiments were performed in the HCVpp system. In contrast to the effects observed on HCVcc, treatment with LPL in the absence of THL at 37°C did not reduce HCVpp infectivity (Fig. 5B). Rather, a slight increase in virus entry was observed, suggesting that the enzymatic activity of LPL does not affect HCVpp infectivity. However, a stronger increase in HCVpp infectivity was observed in the presence of THL. This effect seems to be specific for the HCVpp, because a similar treatment of pseudoparticles containing vesicular stomatitis virus (VSV)-G envelope protein had no effect (Fig. 5C). One reason for this effect could be that THL stabilizes LPL in a conformation that is better in facilitating HCVpp entry. It is worth noting that LPL treatment strongly increased IDL binding to Huh-7 cells (Supporting Fig. 1). Furthermore, the majority of LPL-treated IDL were internalized after incubation for 1 hour at 37°C (data not shown).