Phage Display-Derived Compounds Displace hACE2 from Its Complex with SARS-CoV-2 Spike Protein

Abstract

Severe respiratory syndrome coronavirus-2 (SARS-CoV-2) is a highly contagious beta-class coronavirus. Although vaccinations have shown high efficacy, the emergence of novel variants of concern (VOCs) has already exhibited traits of immune evasion. Thus, the development of tailored antiviral medications for patients with incomplete, inefficient, or non-existent immunization, is essential. The attachment of viral surface proteins to the cell surface is the first crucial step in the viral replication cycle, which for SARS-CoV-2 is mediated by the high affinity interaction of the viral trimeric spike with the host cell surface-located human angiotensin converting enzyme-2 (hACE2). Here, we used a novel and efficient next generation sequencing (NGS) supported phage display strategy for the selection of a set of SARS-CoV-2 receptor binding domain (RBD)-targeting peptide ligands that bind to the target protein with low µM to nM dissociation constants. Compound CVRBDL-3 inhibits the SARS-CoV-2 spike protein association to hACE2 in a concentration-dependent manner for pre- as well as post-complex formation conditions. Further rational optimization yielded a CVRBDL-3 based divalent compound, which demonstrated inhibitory efficacy with an IC50 value of 47 nM. The obtained compounds were not only efficient for the different spike constructs from the originally isolated “wt” SARS-CoV-2, but also for B.1.1.7 mutant trimeric spike protein. Our work demonstrates that phage display-derived peptide ligands are potential fusion inhibitors of viral cell entry. Moreover, we show that rational optimization of a combination of peptide sequences is a potential strategy in the further development of therapeutics for the treatment of acute COVID-19.

Receptor-binding domain-anchored peptides block binding of severe acute respiratory syndrome coronavirus 2 spike proteins with cell surface angiotensin-converting enzyme 2

“…Frontiers in Microbiology 06 frontiersin.org pathogen invasions in this lab (Zhao et al, 2012) or others (Hall et al, 2009;Wei et al, 2020). This study and several other efforts (Anand et al, 2021;Sokullu et al, 2021;Ballmann et al, 2022;Labriola et al, 2022;Petrenko et al, 2022;Sevenich et al, 2022;Yang et al, 2022) demonstrated that PhD was a plausible method for generating possible therapeutics to treat COVID-19 as well. For example, Petrenko used phage-displayed spike S1 protein mimotopes to search for "all" cellular receptors, including authentic and alternative ones.…”

“…Like us, Sevenich performed three rounds of screening on S-RBD proteins using a 16-aa phage library combined with high throughput sequencing. The five final sequences they obtained manifested variable affinity for S-RBD proteins in confirmation assays, with Kd from 1.3 to 89.4 μM ( Sevenich et al, 2022 ). Others also demonstrated the stringent dependence of efficacy of intended therapeutics on their molecular compositions and configuration ( Raghuvamsi et al, 2021 ).…”

“…Here, we report that two SEAL peptides were obtained via phage displaying against S-RBD and S-trimer proteins, and preliminary functional studies demonstrated weak blocking effects at high concentrations. Encouragingly, while this project was ongoing, three groups reported their results obtained by protocols mainly relying on PhD (Petrenko et al, 2022;Sevenich et al, 2022;Yang et al, 2022Yang et al, ). 10.3389/fmicb.2022 The promises and limitations of these studies were also discussed.…”

Discovery of All-d-Peptide Inhibitors of SARS-CoV-2 3C-like Protease

“…Even with the development of several vaccines against SARS-CoV-2, the development of suitable therapeutics against COVID-19 is important since no causative therapy is known. Instead of inhibiting the interaction of the spike protein with the cell receptors, , we intend to target one of the most important proteases in virus replication, the 3CL protease, by d -enantiomeric peptide ligands preventing the cleavage function of 3CL pro . To this end, a phage display selection was performed.…”

Discovery of all-D-peptide inhibitors of SARS CoV 2 3C-like protease

“…Even upon the development of several vaccines against SARS-CoV-2, the development of suitable therapeutics against COVID-19 is important, since no causative therapy is known. Instead of inhibiting the interaction of the spike protein with the cell receptors, 37,38 we intend to target one of the most important proteases in virus replication, the 3CL protease, by D-enantiomeric peptide ligands preventing the cleavage function of 3CL pro . To this end, a phage display selection was performed.…”