Highly Specific Sigma Receptor Ligands Exhibit Anti-Viral Properties in SARS-CoV-2 Infected Cells

Controls included total LDH release as measured by lysis of all cells, spontaneous release from uninfected cells, and media alone. The toxicity of sigma ligands alone were also determined in parallel to discriminate the amount of SARS-CoV-2-induced cytotoxicity occurring in the presence of a given treatment. After spontaneous and background subtraction, OD 450 values were transformed to a percent of SARS-CoV-2 infected cells (100%) in the absence of any drug treatment to obtain percent of SARS-CoV-2-induced cytotoxicity. These experiments were carried out twice. Plaque Reduction Assay Vero E6 cells were plated in 24-well plates with triplicate replicates on different plates. Virus master mix was used to dilute down to 20-200 PFU/mL and aliquoted in separate tubes with drugs at the final indicated drug concentrations. The drug virus mixtures were immediately used to infect Vero E6 monolayers for 1 h with rocking every 10 min. Monolayers were then overlaid with MEM in 1.5% low-melt agarose containing drugs at the final concentrations indicated. Plaques were counted at 72 hours post infection and used to calculate the apparent reduction in viral concentration compared to the starting volume. Data presented is representative of two independent experiments. Generation of ACE-2 Lentivirus Particles The lentivirus containing ACE2 were generated by co-transfecting psPAX2, pMD2.G, and an ACE expression vector that also contained a blasticidin selection gene EX-U1285-Lv197..