Molecular and structural characterization of broadly neutralizing anti-HCV antibodies

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Summary More than 70 million people worldwide are infected with hepatitis C virus (HCV), and development of a vaccine for HCV is essential for disease eradication. Although direct-acting antivirals are highly effective for treatment, the majority of countries surveyed are not on track to reach the WHO goal of eliminating HCV as a public health problem by 2030, with most countries seeing rising incidence of HCV. Fortunately, there is strong evidence that vaccine-induction of high titers of broadly neutralizing antibodies (bNAbs; antibodies capable of neutralizing diverse HCV variants) could provide protection against human HCV infection. However, we are unable to stimulate bNAbs against HCV with a vaccine because we have not defined the full spectrum of anti-HCV antibodies that are critical for neutralizing breadth or characterized the HCV envelope glycoprotein (E1 and E2) genetic and structural features that favor bNAb selection and maturation. The overarching goal of this proposal is to isolate a large and representative set of E1E2-specific bNAbs, bNAb unmutated ancestors, and bNAb intermediates from Elite Neutralizers (EN), individuals with broadly neutralizing plasma and spontaneous clearance of HCV infection. In Aim 1, we will isolate monoclonal antibodies (mAbs) from E1E2-specific B cells isolated from EN and from controls with chronic, persistent infection (CP). We will infer unmutated germline bNAb ancestors and use B cell receptor-sequencing of longitudinal E1E2-specific B cells to identify bNAb genetic intermediates. We will compare neutralizing breadth, potency, epitopes, and genetic features for EN vs. CP mAbs. In Aim 2, we will use X-ray crystallography or cryo-EM techniques to compare structures of EN bNAbs or CP mAbs in complex with soluble E2 or E1E2 heterodimers. By comparing mAb-E1E2 interactions among CP mAbs, bNAb unmutated ancestors, bNAb intermediates, and mature bNAbs, we will define structural and genetic features of E1E2 necessary for the development of neutralizing breadth. Together, these studies will identify a large, representative set of bNAbs associated with spontaneous clearance of HCV, defining key epitope residues and structural features in E1E2 that could be stabilized to optimize vaccine antigens. These studies will inform structure-based design efforts to improve E1E2-based vaccine candidates, which is an urgent challenge with global public health implications.