ABSTRACT
The design of an immunogenic scaffold that serves a role in treating a pathogen, and can be rapidly and predictively modeled, has remained an elusive feat. Here, we demonstrate that SARS-BLOCK™ synthetic peptide scaffolds act as antidotes to SARS-CoV-2 spike protein-mediated infection of human ACE2-expressing cells. Critically, SARS-BLOCK™ peptides are able to potently and competitively inhibit SARS-CoV-2 S1 spike protein receptor binding domain (RBD) binding to ACE2, the main cellular entry pathway for SARS-CoV-2, while also binding to neutralizing antibodies against SARS-CoV-2. In order to create this potential therapeutic antidote-vaccine, we designed, simulated, synthesized, modeled epitopes, predicted peptide folding, and characterized behavior of a novel set of synthetic peptides. The biomimetic technology is modeled off the receptor binding motif of the SARS-CoV-2 coronavirus, and modified to provide enhanced stability and folding versus the truncated wildtype sequence. These novel peptides attain single-micromolar binding affinities for ACE2 and a neutralizing antibody against the SARS-CoV-2 receptor binding domain (RBD), and demonstrate significant reduction of infection in nanomolar doses. We also demonstrate that soluble ACE2 abrogates binding of RBD to neutralizing antibodies, which we posit is an essential immune-evasive mechanism of the virus. SARS-BLOCK™ is designed to “uncloak” the viral ACE2 coating mechanism, while also binding to neutralizing antibodies with the intention of stimulating a specific neutralizing antibody response. Our peptide scaffolds demonstrate promise for future studies evaluating specificity and sensitivity of immune responses to our antidote-vaccine. In summary, SARS-BLOCK™ peptides are a promising COVID-19 antidote designed to combine the benefits of a therapeutic and vaccine, effectively creating a new generation of prophylactic and reactive antiviral therapeutics whereby immune responses can be enhanced rather than blunted.
Figure 2
Peptides simulated via RaptorX were aligned with the SARS-CoV-2 binding interface of ACE2 (ACE2 in red, with PDBePISA-predicted binding interfaces in green). Shown from left to right (top) are SARS-BLOCK™ Peptides 1 (a), 4 (b), 5 (c), and 6 (d) bound to ACE2. Of note, all peptides exhibited two mutations introducing a disulfide bond to recreate the beta sheet structure of the SARS-CoV-2 receptor binding motif (RBM). Otherwise, Peptides 1 and 4 utilized the wildtype sequence, while Peptide 5 utilized MHC-I and MHC-II epitopes, and Peptide 6 utilized a GSGSG linker (white) in one of its non-ACE2-interfacing loop regions. Peptides 4, 5 and 6 exhibited additional, proprietary modifications to their sequences to facilitate appropriate folding, while Peptide 1 lacked this modification. Taking into account the 9 possible folded states generated for each peptide, we utilized PyMOL align commands which take into account multiple potential conformations of each peptide and may serve as a basis for future studies exploring more advanced molecular dynamics approaches for relaxing and simulating intramolecular interactions at the binding interface (e). In essence, the overlay of many possible folded states represents an electron distribution cloud of possible states that can be simulated for the minimal interfacial free energy of binding, and this approach requires vastly fewer computational resources than typically required for modeling binding pockets of de novo peptides or protein-protein interfaces without existing interfacial structures....
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