38 We confirmed our data in cultures of primary human fetal hepatocytes, which are a more amenable in vitro culture system, given the more robust infection levels achieved compared with adult hepatocytes.39 Primary human fetal hepatocytes, transduced with the RFPnls-IPS HCV reporter system,43 were preincubated with different concentrations of mAb16-71. Parallel cultures were treated with an isotype-matched antibody (negative control) or JS81, an antibody that targets CD81 and click here prevents attachment and infection of HCV (positive control). As shown in Fig. 1C, infection of primary fetal hepatocytes by J6/JFH-1 HCVcc was also reduced in a dose-dependent manner. HCV can

spread directly from an infected Huh-7.5 cell to uninfected neighboring cells, with possibly all four HCV entry factors CD81, SR-BI, claudin, and occludin, being involved in this process.32-34 However, by using Huh-7.5 target cells in which CD81 was selectively knocked down (EGFP-IPS/CD81neg), we have recently shown that HCV cell-to-cell

spread can occur independently of CD81.43 We therefore used this cell line to investigate whether mAb16-71 is capable of inhibiting this alternative transmission route. To this end, HCVcc-infected (Jc1) Huh-7.5 cells were cocultured with uninfected EGFP-IPS/CD81neg cells. EGFP-IPS/CD81neg cells have been previously shown to be essentially nonpermissive to cell-free HCV infection.43 Nevertheless, mixing uninfected EGFP-IPS/CD81neg target cells with infected Huh7.5 cells resulted in an infection of 8%-10% of the target cells. However, in the presence Ibrutinib concentration of increasing concentrations of mAb16-71 a dose-dependent reduction in EGFP-IPS/CD81neg target cell infection was observed, whereas no significant changes

in HCV transmission were observed in the presence of an isotype-matched control antibody (Fig. 1D). This clearly proves that mAb16-71 not only prevents cell-free HCV infection, but also interferes 上海皓元医药股份有限公司 with the direct cell-to-cell transmission route. Given the encouraging results in cell culture, we investigated whether administration of mAb16-71 to “human liver urokinase-type plasminogen activator, severe combined immune deficiency (uPA-SCID)” mice (chimeric mice) could protect these animals from a subsequent challenge with serum-derived virus. These mice have a humanized liver (up to 90% chimerism) and are in addition to the chimpanzee the preferred animal model for reproducible infection with natural HCV isolates.40, 42, 45, 46 Two chimeric mice underwent a 2-week therapy consisting of five intraperitoneal injections, each containing 400 μg of mAb16-71. One day after the first injection, both chimeric mice were challenged with a 100% mouse infectious dose (MID100) of serum-derived genotype 1a HCV (mH77C). In contrast to nontreated control animals, which experienced a rapid increase of viral RNA in their plasma, HCV RNA remained undetectable (<375 IU/mL) in both treated mice in the 12-week observation period (Fig. 2A).