NS4B Amphipathic Helix

All positive strand RNA viruses replicate their genome in intimate association with a subset of host intracellular membranes. In HCV, the membranes are derived in part from the endoplasmic reticulum and form a structure termed the membranous web due to its appearance on electron microscopy consisting of aggregations of membranous vesicles. Recently, a second target within NS4B has been genetically-validated and consists of a conserved amphipathic helix (AH) essential for viral genome replication. This AH, termed 4BAH2, has a dramatic biochemical activity of promoting the aggregation of lipid vesicles, with likely relevance to the establishment of the membranous web.

This biochemical activity was leveraged into a new high throughput screening assay for pharmacologic inhibitors of 4BAH2 function. Analysis of one of the inhibitors from a preliminary screen revealed that its mechanism of action involves inhibition of 4BAH2 induced vesicle aggregation, but not 4BAH2 association with membranes. A set of primary and secondary screening assays has been validated for identifying inhibitors of HCV replication. These screens and the compounds identified in them are the subject of a pending patent application, and SAR optimization of the existing leads as well as a larger small molecule screening effort are underway.