Quinolizidines as Novel SARS-CoV-2 Entry Inhibitors

Abstract

COVID-19, caused by the highly transmissible severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), has rapidly spread and become a pandemic since its outbreak in 2019. We have previously discovered that aloperine is a new privileged scaffold that can be modified to become a specific antiviral compound with markedly improved potency against different viruses, such as the influenza virus. In this study, we have identified a collection of aloperine derivatives that can inhibit the entry of SARS-CoV-2 into host cells. Compound 5 is the most potent tested aloperine derivative that inhibited the entry of SARS-CoV-2 (D614G variant) spike protein-pseudotyped virus with an IC₅₀ of 0.5 µM. The compound was also active against several other SARS-CoV-2 variants including Delta and Omicron. Results of a confocal microscopy study suggest that compound 5 inhibited the viral entry before fusion to the cell or endosomal membrane. The results are consistent with the notion that aloperine is a privileged scaffold that can be used to develop potent anti-SARS-CoV-2 entry inhibitors.

Keywords: SARS-CoV-2 inhibitor; aloperine; aloperine derivatives.

Figure 1

Compound 5 arrests D614G spike-pseudotyped virus entry. D614G spike-pseudotyped viruses were used to infect 293T-ACE2 cells, which were stained with DAPI (nuclear stain, blue) 2 h post infection. Size bar = 10 µm. Confocal microscopy images were acquired using a Nikon A1R confocal microscope with a 60×/1.4 NA oil-immersion Plan-Apochromat lens. (A) 293T-ACE2 cells were infected with D614G spike-pseudotyped virus; (B) experiment was performed in the presence of compound 5 at 5 μM; (C) experiment was performed in the presence of chloroquine diphosphate at 5 μM.

Figure 2

Aloperine derivatives were inactive against cathepsin B and L. Compound 5 and aloperine were tested for their inhibitory activity against cathepsin B or L using a BPS bioscience assay kit and the protocol provided by the manufacturer (catlog#79590). The enzyme activity in the absence of compounds (control) was defined as 100%. Aloperine (Alop) and compound 5 (Cpd 5) were tested at 20 µM. The known cathepsin B inhibitor E64 was tested at 0.1 µM (E64-H) and 0.01 µM (E64-L), respectively. The data represent the average of a duplicated experiment.

Figure 3

Model for inhibition of SARS-Cov-2 entry by Compound 5.