Abstract
BackgroundPorcine reproductive and respiratory syndrome (PRRS) is one of the most economically devastating diseases affecting the pork industry globally. PRRS is caused by PRRS virus (PRRSV). Currently there are no effective treatments against this swine disease.MethodsThrough artificial intelligence molecular screening, we obtained a set of small molecule compounds predicted to target the scavenger receptor cysteine-rich domain 5 (SRCR5) of CD163, which is a cell surface receptor specific for PRRSV infection. These compounds were screened using a cell-based bimolecular fluorescence complementation (BiFC) assay, and the function of positive hit was further evaluated and validated by PRRSV-infection assay using porcine alveolar macrophages (PAMs).ResultsUsing the BiFC assay, we identified one compound with previously unverified function, 4-Fluoro-2-methyl-N-[3-(3-morpholin-4-ylsulfonylanilino)quinoxalin-2-yl]benzenesulfonamide (designated here as B7), that significantly inhibits the interaction between the PRRSV glycoprotein (GP2a or GP4) and the CD163-SRCR5 domain. We further demonstrated that compound B7 inhibits PRRSV infection of PAMs, the primary target of PRRSV in a dose-dependent manner. B7 significantly inhibited the infection caused by both type I and type II PRRSV strains. Further comparison and functional evaluation of chemical compounds structurally related to B7 revealed that the 3-(morpholinosulfonyl)aniline moiety of B7 or the 3-(piperidinylsulfonyl)aniline moiety in a B7 analogue is important for the inhibitory function against PRRSV infection.ConclusionsOur study identified a novel strategy to potentially prevent PRRSV infection in pigs by blocking the PRRSV-CD163 interaction with small molecules.
Highlights
Porcine reproductive and respiratory syndrome (PRRS) is one of the most economically significant swine diseases, with over billion-dollar losses to the global pork industry annually
Development of bimolecular fluorescence complementation (BiFC) assays to identify small molecules that inhibit the Protein-protein interaction (PPI) between PRRS virus (PRRSV) and CD163 Using the previously described BiFC vector [33] based on the fragmented Venus protein (VN-155[I152L], hereafter named VN) and (VC-155, hereafter named VC), we established fusion protein constructs between the porcine CD163 protein scavenger receptor cysteine-rich domain 5 (SRCR5) or scavenger receptor cysteine-rich domain 2 (SRCR2) domain and VN, and between the PRRSV minor glycoproteins (GP2a or GP4) and VC (Figs. 1a, S1)
The fusion protein of CD163-SRCR2 domain (SRCR2-VN), which is dispensable for the PRRSV infection and PRRSV-CD163 interaction [18], only showed background fluorescence when co-expressed with GP2a- or GP4-VC (Fig. 1b, c, lower panel)
Summary
Porcine reproductive and respiratory syndrome (PRRS) is one of the most economically significant swine diseases, with over billion-dollar losses to the global pork industry annually. The causative virus of PRRS (PRRSV) is an enveloped, positive-sense, single-stranded RNA virus of the Arterivirus genus within the order Nidovirales [1, 2]. PRRSV infection results in severe reproductive failure in sows and respiratory disease in piglets [3]. This may be complicated by secondary infections with even greater clinical manifestations and mortality [4,5,6]. Porcine reproductive and respiratory syndrome (PRRS) is one of the most economically devastating diseases affecting the pork industry globally. There are no effective treatments against this swine disease
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