Structural-based virtual screening and in vitro assays for small molecules inhibiting the feline coronavirus 3CL protease as a surrogate platform for coronaviruses.

Feline infectious peritonitis (FIP) which is caused by feline infectious peritonitis virus (FIPV), a variant of feline coronavirus (FCoV), is a member of family Coronaviridae, together with severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-2. So far, neither effective vaccines nor approved antiviral therapeutics are currently available for the treatment of FIPV infection. Both human and animal CoVs shares similar functional proteins, particularly the 3CL protease (3CLpro), which plays the pivotal role on viral replication. We investigated the potential drug-liked compounds and their inhibitory interaction on the 3CLpro active sites of CoVs by the structural-bases virtual screening. Fluorescence resonance energy transfer (FRET) assay revealed that three out of twenty-eight compounds could hamper FIPV 3CLpro activities with IC50 of 3.57 ± 0.36 µM to 25.90 ± 1.40 µM, and Ki values of 2.04 ± 0.08 to 15.21 ± 1.76 µM, respectively. Evaluation of antiviral activity using cell-based assay showed that NSC629301 and NSC71097 could strongly inhibit the cytopathic effect and also reduced replication of FIPV in CRFK cells in all examined conditions with the low range of EC50 (6.11 ± 1.90 to 7.75 ± 0.48 µM and 1.99 ± 0.30 to 4.03 ± 0.60 µM, respectively), less than those of ribavirin and lopinavir. Analysis of FIPV 3CLpro-ligand interaction demonstrated that the selected compounds reacted to the crucial residues (His41 and Cys144) of catalytic dyad. Our investigations provide a fundamental knowledge for the further development of antiviral agents and increase the number of anti-CoV agent pools for feline coronavirus and other related CoVs.

In vitro antibacterial activity of mangosteen (Garcinia mangostana Linn.) crude extract against Staphylococcus pseudintermedius isolates from canine pyoderma.

BACKGROUND: Staphylococcus pseudintermedius, commonly involved in canine pyoderma, can be classified as meticillin-susceptible S. pseudintermedius (MSSP) or meticillin-resistant S. pseudintermedius (MRSP). MRSP infections may be difficult to treat due to broad ß-lactam resistance of MRSP and typically additional multidrug-resistance. Topical antibacterial treatment is the preferred treatment modality for surface and superficial skin infections. HYPOTHESIS/OBJECTIVES: Mangosteen crude extract containing the antibacterial compound α-mangostin will have in vitro activity against MSSP and MRSP isolated from canine pyoderma. BACTERIAL ISOLATES: Twenty-three samples, MSSP (n = 12) and MRSP (n = 11), isolated from canine pyoderma. METHODS AND MATERIALS: Minimum inhibitory concentrations (MICs) were determined for mangosteen crude extract by broth microdilution. High-performance liquid chromatography (HPLC) analysis was used to determine the amount of α-mangostin in mangosteen crude extract. A time-kill assay was performed at 30 min and 2 h after exposure to a high concentration of crude extract (100× MIC). Antibacterial activity for α-mangostin was calculated according to HPLC results. RESULTS: The concentration of α-mangostin was 17.72 ± 1.42% w/w. The mean MIC of α-mangostin towards MSSP was 0.53 ± 0.35 µg/mL, whereas the mean value for MRSP was 0.47 ± 0.27 µg/mL. There was no difference between the mean MIC of MRSP and MSSP (P = 0.84). After a 30 min exposure to 100× MIC of the crude extract, a 95% reduction in colony forming units was found. CONCLUSIONS AND CLINICAL IMPORTANCE: The results showed that α-mangostin in mangosteen crude extract was effective in inhibiting S. pseudintermedius (both MRSP and MSSP). Clinical studies are needed to investigate this effectiveness further in vivo.

Screening for lead compounds and herbal extracts with potential anti-influenza viral activity.

Nonstructural protein 1 (NS1) of the highly pathogenic avian influenza virus (H5N1) contains a conserved RNA binding domain (RBD) that inhibits antiviral functions of host-innate immune response. Dimerization of NS1 forms a central groove and binds to double stranded (ds) RNA. This region might serve as a potential drug target. In this study, three dimensional structure model of NS1 RBD protein was constructed and virtual screening was performed to identify lead compounds that bound within and around the central groove. The virtual screening showed that 5 compounds bound within the central groove with binding energy ranging between -16.05 and -17.36 Kcal/mol. Two commercially available compounds, estradiol and veratridine, were selected for using in an in vitro screening assay. The results showed that neither of the compounds could inhibit the association between dsRNA and NS1 RBD protein. In addition, 34 herbal extracts were examined for their inhibitory effects. Five of them were able to inhibit association between NS1 RBD and dsRNA in electrophoresis mobility shift assay. Four herbs, Terminalia belirica, Salacia chinensis, Zingiber montanum and Peltophorum pterocarpum, could reduce > 50% of infectivity of H5N1 in a cell-based assay, and it is worth further studying their potential use as source of antiviral drugs.