SARS-CoV-2 is a novel virus that has rapidly spread, causing a global pandemic. In the majority of infected patients, SARS-CoV-2 leads to mild disease; however, in a significant proportion of infections, individuals develop severe symptoms that can lead to long-lasting lung damage or death. These severe cases are often associated with high levels of pro-inflammatory cytokines and low antiviral responses, which can cause systemic complications. Here, we have evaluated transcriptional and cytokine secretion profiles and detected a distinct upregulation of inflammatory cytokines in infected cell cultures and samples taken from infected patients. Building on these observations, we found a specific activation of NF-κB and a block of IRF3 nuclear translocation in SARS-CoV-2 infected cells. This NF-κB response was mediated by cGAS-STING activation and could be attenuated through several STING-targeting drugs. Our results show that SARS-CoV-2 directs a cGAS-STING mediated, NF-κB-driven inflammatory immune response in human epithelial cells that likely contributes to inflammatory responses seen in patients and could be therapeutically targeted to suppress severe disease symptoms.

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The phosphoramidite 9 has been particularly applied to the S-phosphorylation of nucleosides.9 For example, 3'-azido-3'deoxythymidine (AZT) was converted into its S-di-(ferr-butyl)phosphate triester upon reaction with 9 and lWtetrazole followed by oxidation with m-chloroperbenzoic acid (MCPBA). The tertiary alkyl phosphate protecting groups were cleaved with trifluoroacetic acid to produce AZT S-phosphate in quantitative yield. 9 Interestingly, the fert-butyl group could also be quantitatively removed, within 2 h at 75 OC, with trimethylsilyl chloride and triethylamine in acetonWe.10 These conditions permitted the synthesis of IV-protected-SO -(4,4'dimetho Yrity1)-2deoxyribonucleoside-3'-phosphates with no loss of the 4,4'-dimethoxytrityl and IV-acyl groups.1 One disadvantage in using the phosphoramidites l-8 for the 5'-phosphorylation of oligodeovibonucleotides was the lack of a convenient way to monitor the coupling reaction. Horn and Urdea circumvented this problem by developing the phosphitylating reagent 10 from sulfonyldiethanol.ll The phosphoramidite derivative was activated with VI-tetrazole and coupled to the 5'-hydroxyl function of thymidine bound to a controlled-pore glass (CPG) support. Following oxidation, treatment of the solid support with dichloroacetic acid released the bright orange 4,4'dimethoxytrityl cation which accounted for a coupling efficiency of 96%. Deprotection of the terminal phosphate triester with concentrated ammonium hydroxide at 60 OC afforded thymidine 5'monophosphate as the sole product. The S-phosphorylation of oligodeoxyribonucleotides can alternatively be achieved by the phosphotriester methodology via a solid-phase approachl5917 or in sohrtion.t4-reJ*-~ The s-triphenylmethyl-2-mercaptoethyl phosphoramidite 11 has also been used for the 5'phosphorylation of oligonucleotides.21 Typically, oligomers anchored to a solid support were phosphitylated with 11 in the presence of VI-tetrazole and oxidized under standard conditions. After deprotection and purification, the trityl-sulfur link was cleaved with aqueous silver nitrate affording the corresponding oligonucleotide-5'-mercaptoethyl~Ihosphodiester which decomposed quantitatively at pH 8.5 to the desired oligonucleotide-S-phosphate. The facile 5'-phosphorylation of oligonucleotides covalently linked to a solid support provided the impetus for the development of specific phosphoramidites that would enable the direct or indirect incorporation of a number of ligands at the S-terminus of oligonucleotides.

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