(D and E) Live-virus infection of Vero E6 cells Quantification of total GFP signal resulting from successful pseudovirus infection, (B) Infection of ACE2 293T cells with SARS-CoV-2 spike pseudotyped lentivirus Visualize replication sites formed during infection. Cells were stained with anti-ds-RNA (red) antibody to Of SARS-CoV-2 (WA1/2020)-infected Vero E6 cells treated with 25 μg/mL CBDA,ĬBGA, or vehicle (control). (A) Representative images of high-resolution microscopy Of spike protein pseudotyped lentivirus and multiple variants of live SARS-CoV-2 virusīy cannabinoids CBDA and CBGA. Negative controls using denatured S1 showed no significant bindingĬBD compounds block viral entry of SARS-CoV-2 through spike binding. Of cannabinoid acids (0.10 μM each in this confirmatory chromatogram) as ligandsįrom hemp extracts. (D) MagMASS was used for the affinity selection and identification
(equivalent to 0.17 μM) (positive control) but not to immobilized denatured S1 (C) During AS-MS, the SBP-1 peptide bound to immobilized S1 Ligands were released using organic solvent and thenĪnalyzed using UHPLC-MS. Probe retained the microbeads containing the S1 subunit and bound ligands, while unboundĬompounds were washed away. Used to isolate and identify natural ligands to the spike protein S1 subunit.
Was immobilized on magnetic microbeads for affinity selection of ligands. Subunit binds to human ACE2 to initiate cell entry. (A) The spike protein of SARS-CoV-2 consists of trimers ofĪ protein containing an S1 subunit, an S2 subunit, and a transmembrane domain. Orally bioavailable and with a long history of safe human use, these cannabinoids, isolated or in hemp extracts, have the potential to prevent as well as treat infection by SARS-CoV-2.Īffinity selection–mass spectrometric (AS-MS) discovery of natural ligands to Importantly, cannabigerolic acid and cannabidiolic acid were equally effective against the SARS-CoV-2 alpha variant B.1.1.7 and the beta variant B.1.351. In follow-up virus neutralization assays, cannabigerolic acid and cannabidiolic acid prevented infection of human epithelial cells by a pseudovirus expressing the SARS-CoV-2 spike protein and prevented entry of live SARS-CoV-2 into cells. Cannabinoid acids from hemp ( Cannabis sativa) were found to be allosteric as well as orthosteric ligands with micromolar affinity for the spike protein.
Affinity selection-mass spectrometry was used for the discovery of botanical ligands to the SARS-CoV-2 spike protein. As a complement to vaccines, small-molecule therapeutic agents are needed to treat or prevent infections by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and its variants, which cause COVID-19.
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