AI Promoted Virtual Screening, Structure-Based Hit Optimization, and Synthesis of Novel COVID-19 S‑RBD Domain Inhibitors

Coronavirus disease
2019 (COVID-19) is caused by a new, highly
pathogenic severe-acute-respiratory syndrome coronavirus 2 (SARS-CoV-2)
that infects human cells through its transmembrane spike (S) glycoprotein.
The receptor-binding domain (RBD) of the S protein interacts with
the angiotensin-converting enzyme II (ACE2) receptor of the host cells.
Therefore, pharmacological targeting of this interaction might prevent
infection or spread of the virus. Here, we performed a virtual screening
to identify small molecules that block S-ACE2 interaction. Large compound
libraries were filtered for drug-like properties, promiscuity and
protein–protein interaction-targeting ability based on their
ADME-Tox descriptors and also to exclude pan-assay interfering compounds.
A properly designed AI-based virtual screening pipeline was applied
to the remaining compounds, comprising approximately 10% of the starting
data sets, searching for molecules that could bind to the RBD of the
S protein. All molecules were sorted according to their screening
score, grouped based on their structure and postfiltered for possible
interaction patterns with the ACE2 receptor, yielding 31 hits. These
hit molecules were further tested for their inhibitory effect on Spike
RBD/ACE2 (19–615) interaction. Six compounds inhibited the
S-ACE2 interaction in a dose-dependent manner while two of them also
prevented infection of human cells from a pseudotyped virus whose
entry is mediated by the S protein of SARS-CoV-2. Of the two compounds,
the benzimidazole derivative CKP-22 protected Vero E6
cells from infection with SARS-CoV-2, as well. Subsequent, hit-to-lead
optimization of CKP-22 was effected through the synthesis
of 29 new derivatives of which compound CKP-25 suppressed
the Spike RBD/ACE2 (19–615) interaction, reduced the cytopathic
effect of SARS-CoV-2 in Vero E6 cells (IC50 = 3.5 μM)
and reduced the viral load in cell culture supernatants. Early in
vitro ADME-Tox studies showed that CKP-25 does not possess
biodegradation or liver metabolism issues, while isozyme-specific
CYP450 experiments revealed that CKP-25 was a weak inhibitor
of the CYP450 system. Moreover, CKP-25 does not elicit
mutagenic effect on Escherichia coli WP2 uvrA strain. Thus, CKP-25 is considered a lead
compound against COVID-19 infection.