The effect of plasma protein binding on the anti-HBV activity and pharmacokinetic properties of NVR 3-778. [Antiviral Agents]

High plasma protein binding (PPB) not only affects drug-target engagement but can also impact exposure of hepatocytes to antivirals and thereby affect antiviral activity. In this study, we assessed the effect of PPB on the antiviral activity of NVR 3-778, a sulfamoylbenzamide capsid assembly modulator (CAM). To this end, primary human hepatocyte (PHH) medium was spiked with plasma proteins. At first, the effect of plasma proteins on the HBV-infection assay was evaluated. The addition of plasma proteins neither decreased cell viability nor affected HBV DNA secretion or intracellular HBV RNA accumulation. In contrast, the secretion and intracellular amount of HBV proteins were induced with increasing amounts of plasma proteins. Next, the antiviral activity of NVR 3-778 was demonstrated by multiple assays while PPB and the time-dependent disappearance of the parent drug were quantified by LC-MS/MS. Plasma proteins strongly decreased the free fraction of NVR 3-778 resulting in a physiologically relevant in vitro hepatocyte exposure. NVR 3-778 displayed high PPB, while the antiviral activity was only approximately 4-fold reduced. The disconnect between the high PPB and the only moderate shift of the antiviral activity was explained by the rapid hepatic clearance of NVR 3-778 in the absence of plasma proteins. This study highlights the use of PHHs as a model to accurately determine the antiviral activity by capturing PPB, clearance and liver distribution. It is advantageous to consider both pharmacokinetics and pharmacodynamics for selection of HBV antiviral drug candidates and successful extrapolation of in vitro data to clinical studies.

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