A versatile structure-based therapeutic platform for development of VHH-based antitoxin and antiviral agents - ABSTRACT
In previous NIH sponsored research we successfully tested the hypothesis that integrating structural and
mechanistic information into heteromultimeric VHH-based neutralizing agent (VNA) design facilitated
development of antitoxins with even greater efficacy and versatility. In this renewal proposal, we will apply these
findings to test the hypothesis that our designer VNA platform, which is rapidly responsive to new threats, will
permit development of highly practical, next-generation antitoxin and antiviral products that possess excellent
potencies in treating intoxications or viral infections and are effective against a broad range of natural pathogen
variants. Our research will focus on two pathogens that are major current threats which could benefit from next-
generation therapeutics: botulinum neurotoxin (BoNT) and SARS-CoV-2. We propose two Specific Aims which
will be underway simultaneously throughout the five years of research. In Aim 1, we will develop a small pool of
antitoxin VNAs that protect against all subtypes of the three prevalent BoNT serotypes (A, B and E). BoNTs are
CDC Tier 1 select agents. However, the few available antitoxin treatments against BoNTs primarily derive from
large animal polyclonal antisera, such as the equine botulism antitoxin HBAT, which suffer from multiple
manufacturing and storage challenges. Our goal is to test the platform’s ability to produce highly practical VNAs
as a next-generation BoNT antitoxin product, likely delivered as RNA nanoparticles, which improves on the
potencies and natural variant specificities of the current HBAT antitoxin product and is rapidly responsive to
potential new BoNT threats. In Aim 2 we will develop a single VNA antiviral agent that protects against known
variants of SARS-CoV-1 and SARS-CoV-2. SARS-CoV-2 is the viral cause of the ongoing COVID-19 pandemic.
A promising strategy for rapid development of a therapy is development of SARS-CoV-2 neutralizing antibodies,
especially antibodies targeting the spike protein, for prophylactic or passive immunotherapies. However, novel
variants of SARS-CoV-2, which cause enhanced infection and transmission, have emerged, and more
dangerous variants are expected to evolve. Of immediate concern are variants that partially escape
neutralization by current Ab-based therapies and in vaccinated or previously-infected COVID-19 patients, leading
to reduced vaccine efficacy in certain areas with a high prevalence of these variants. We propose an mRNA-
based antiviral product that, once administered, elicits expression of a VNA with extremely high virus neutralizing
potency. The VNA will contain multiple covalently linked VHHs binding to conserved epitopes of the spike protein.
This approach will test the platform’s ability to develop a product that minimizes the risks of immune escape
through evolution and selection of clinical strains of SARS-CoV-2 and SARS-CoV-1. If successful, this
technology platform could have broad applications in creating practical therapeutics for a wide variety of
emerging and potential pandemic viral infections, bioterror threat agents, and other infectious diseases.
