Novel entry inhibitors against SARS-CoV-2 based on interface of spike RBD

ABSTRACT

Background: Since the outbreak of COVID-19, globally, more than 63 million people have been infected and 1.46 million people succumbed to death, and the number is still growing. It is well-established that attachment of spike glycoprotein of SARS-CoV-2 with ACE-2 is crucial for initiating infection. While a vaccine is awaited, alternate strategies can be adopted for blocking viral entry to host cells. Herein, we report two lead compounds that block the attachment of spike with ACE2 using lung epithelial cells. Methods: Methods: To identify the lead compounds, we conducted virtual screening of ∼3 million compounds that had potential to bind a site at ACE2/ Spike interface (PDB file 6M0J) using 'Glide' program of Schrödinger Suite. Then the combination of visual inspection and redocking with AutoDock Vina (to determine binding energy)was used to select 5 potential inhibitors of ACE2/ Spike interaction. These five compounds were then tested for their inhibitory activity in virological and biophysical assays. The inhibitory activity of these five compounds was measured using Vero-STAT1 knockout cells and a human bronchial epithelial cell line (UNCN1T). Results: Results: Of the five, two compounds, MU-UNMC-1 and MU-UNMC-2 with binding energy of-6.9kcal/mol and-7.8kcal/mol respectively, showed antiviral activity in two cell lines. In Vero-STAT1 cells, MU-UNMC-1 had IC50 of 5.35μM and 2.94μM, whereas MU-UNMC-2 had IC50 of 1.63μM and 0.54μM, after 24 and 48 hrs post infection (hpi), respectively. In UNCN1T cells, both compounds had significantly better efficacy. MU-UNMC-1 had an IC50 of 0.67μM and 1.16μM and at 24 and 48 hpi, respectively. MU-UNMC-2 had IC50 of 1.72μM and 0.89μM after 24 and 48 hpi, respectively. In Vero-STAT1 cells, the selectivity index (SI) (defined as CC50/IC50) of the compounds was 2.11(MU-UNMC-1) and 13.22(MU-UNMC-2), whereas in UNCN1T cells, the SI of the compounds was 9.27(MU-UNMC-1) and 4.15(MU-UNMC-2). Conclusion: Conclusion: We report the identification of two lead compounds (MU-UNMC-1 and MU-UNMC-2) that block the entry of SARS-CoV-2 in submicromolar concentration in biologically relevant human bronchial epithelial cells. Further, using structure-based similarity searches, we identified three additional chemotypes of these two compounds. These chemotypes of MUUNMC-3, MU-UNMC-4 and MU-UNMC-5) showed ∼2-fold better binding affinity with ACE2/Spike complex. These compounds are under investigation for their inhibitory effect in virological and biophysical assays.