Abstract
Escape variants of SARS-CoV-2 are threatening to prolong the COVID-19 pandemic. To address this challenge, we developed multivalent protein-based minibinders as potential prophylactic and therapeutic agents. Homotrimers of single minibinders and fusions of three distinct minibinders were designed to geometrically match the SARS-CoV-2 spike (S) trimer architecture and were optimized by cell-free expression and found to exhibit virtually no measurable dissociation upon binding. Cryo-electron microscopy (cryoEM) showed that these trivalent minibinders engage all three receptor binding domains on a single S trimer. The top candidates neutralize SARS-CoV-2 variants of concern with IC50 values in the low pM range, resist viral escape, and provide protection in highly vulnerable human ACE2-expressing transgenic mice, both prophylactically and therapeutically. Our integrated workflow promises to accelerate the design of mutationally resilient therapeutics for pandemic preparedness.
One-Sentence Summary We designed, developed, and characterized potent, trivalent miniprotein binders that provide prophylactic and therapeutic protection against emerging SARS-CoV-2 variants of concern.
Competing Interest Statement
Each contributor attests that they have no competing interests relating to the subject contribution, except as disclosed. A.C.H, L.C., I.G., J.B.C., L.M., L.K., L.C., K.W., J.B., N.E., A.C.W., B.V., R.R. Y-J.P, H-W.Y., L.S., D.V., M.D., M.C.J. and D.B. are co-inventors on provisional patent applications that incorporate discoveries described in this manuscript. D.B. is a cofounder of Neoleukin Therapeutics. M.S.D. is a consultant for Inbios, Vir Biotechnology, and NGM Biopharmaceuticals and is on the Scientific Advisory Board of Moderna. H.R-B. is a Scientific Advisor of CuriBio. D.V. is a consultant for Vir Biotechnology. The Veesler lab received an unrelated sponsored research agreement from Vir Biotechnology. M.C.J. is a cofounder of SwiftScale Biologics, Stemloop, Inc., Design Pharmaceuticals, and Pearl Bio. The interests of M.C.J. are reviewed and managed by Northwestern University in accordance with their conflict of interest policies. B.S.F. is an inventor on patent applications related to kidney organoid differentiation and application. J.D.B. consults for Moderna on viral evolution and epidemiology and Flagship Labs 77 on deep mutational scanning. J.D.B. may receive a share of IP revenue as an inventor on a Fred Hutchinson Cancer Research Center-optined technology/patent (application WO2020006494) related to deep mutational scanning of viral proteins. L.G., W.M. and C.T. are current employees of Amgen and own Amgen stock. Z.L. and S.P.J.W. received unrelated sponsored research agreements from VIR Biotechnology, AbbVie, and SAB therapeutics.
