Compounds derived from Humulus lupulus inhibit SARS-CoV-2 papain-like protease and virus replication

Background: Selected natural compounds exhibit very good antiviral properties. Especially, the medicinal plant Humulus lupulus (hop) contains several secondary plant metabolites some of which have previously shown antiviral activities. Among them, the prenylated chalcone xanthohumol (XN) demonstrated to be a potent inhibitor of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease (M^pro).

Hypothesis/purpose: Following the finding that xanthohumol (XN) is a potent inhibitor of SARS-CoV-2 M^pro, the effect of XN and its major derivatives isoxanthohumol (IXN), 6-prenylnaringenin (6-PN), and 8-prenylnaringenin (8-PN) from hops on SARS-CoV-2 papain-like protease (PL^pro) were investigated.

Study design: The modulatory effect of the hop compounds on PL^pro were studied first in silico and then in vitro. In addition, the actual effect of hop compounds on the replication of SARS-CoV-2 in host cells was investigated.

Methods: In silico docking analysis was used to predict the binding affinity of hop compounds to the active site of PL^pro. A recombinant PL^pro was cloned, purified, characterized, and analyzed by small-angle X-ray scattering (SAXS), deISGylation assays, and kinetic analyses. Antiviral activity of hop compounds was assessed using the fluorescently labeled wildtype SARS-CoV-2 (icSARS-CoV-2-mNG) in Caco-2 host cells.

Results: Our in silico docking suggests that the purified hop compounds bind to the active site of SARS-CoV-2 PL^pro blocking the access of its natural substrates. The hop-derived compounds inhibit SARS-CoV-2 PL^pro with half maximal inhibitory concentration (IC₅₀) values in the range of 59-162 µM. Furthermore, we demonstrate that XN and 6-PN, in particular, impede viral replication with IC₅₀ values of 3.3 µM and 7.3 µM, respectively.

Conclusion: In addition to the already known inhibition of M^pro by XN, our results show, for the first time, that hop-derived compounds target also SARS-CoV-2 PL^pro which is a promising therapeutic target as it contributes to both viral replication and modulation of the immune system. These findings support the possibility to develop new hop-derived antiviral drugs targeting human coronaviruses.