title: Structural and functional characterization of an anti-West Nile virus monoclonal antibody and its single-chain variant produced in glycoengineered plants.
abstract: Previously, our group engineered a plant-derived monoclonal antibody (MAb pE16) that efficiently treated West Nile virus (WNV) infection in mice. In this study, we developed a pE16 variant consisting of a single-chain variable fragment (scFv) fused to the heavy chain constant domains (CH) of human IgG (pE16scFv-CH). pE16 and pE16scFv-CH were expressed and assembled efficiently in Nicotiana benthamiana ∆XF plants, a glycosylation mutant lacking plant-specific N-glycan residues. Glycan analysis revealed that ∆XF plant-derived pE16scFv-CH (∆XFpE16scFv-CH) and pE16 (∆XFpE16) both displayed a mammalian glycosylation profile. ∆XFpE16 and ∆XFpE16scFv-CH demonstrated equivalent antigen-binding affinity and kinetics, and slightly enhanced neutralization of WNV in vitro compared with the parent mammalian cell-produced E16 (mE16). A single dose of ∆XFpE16 or ∆XFpE16scFv-CH protected mice against WNV-induced mortality even 4 days after infection at equivalent rates as mE16. This study provides a detailed tandem comparison of the expression, structure and function of a therapeutic MAb and its single-chain variant produced in glycoengineered plants. Moreover, it demonstrates the development of anti-WNV MAb therapeutic variants that are equivalent in efficacy to pE16, simpler to produce, and likely safer to use as therapeutics due to their mammalian N-glycosylation. This platform may lead to a more robust and cost-effective production of antibody-based therapeutics against WNV infection and other infectious, inflammatory or neoplastic diseases.
authors: The Biodesign Institute, Arizona State University, Tempe, AZ, USA.
authors: Huafang
authors: H
authors: Lai
authors: Junyun
authors: J
authors: He
authors: Jonathan
authors: J
authors: Hurtado
authors: Jake
authors: J
authors: Stahnke
authors: Anja
authors: A
authors: Fuchs
authors: Erin
authors: E
authors: Mehlhop
authors: Sergey
authors: S
authors: Gorlatov
authors: Andreas
authors: A
authors: Loos
authors: Michael S
authors: MS
authors: Diamond
authors: Qiang
authors: Q
authors: Chen
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000914:Antibodies, Viral / Q000276:immunology*; D000956:Antigens, Viral / Q000276:immunology*; D015870:Gene Expression; D006031:Glycosylation; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D009500:Neutralization Tests; D046549:Plantibodies / Q000276:immunology; D057127:Single-Chain Antibodies / Q000276:immunology*; D020349:Surface Plasmon Resonance; D014026:Nicotiana / Q000378:metabolism*; D014759:Viral Envelope Proteins / Q000276:immunology; D014901:West Nile Fever / Q000517:prevention & control*; D014902:West Nile virus / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D046549:Plantibodies; D057127:Single-Chain Antibodies; D014759:Viral Envelope Proteins
keywords: West Nile virus; glycoengineered plants; glycosylation; plant-made biologics; scFv-Fc; single-chain antibody