A combined in silico / in vitro approach unveils common molecular requirements for efficient BVDV RdRp binding of linear aromatic N-polycyclic systems

Abstract

In this work, we present and discuss a comprehensive set of both newly and previously synthesized compounds belonging to 5 distinct molecular classes of linear aromatic N-polycyclic systems that efficiently inhibits bovine viral diarrhea virus (BVDV) infection. A coupled in silico/in vitro investigation was employed to formulate a molecular rationale explaining the notable affinity of all molecules to BVDV RNA dependent RNA polymerase (RdRp) NS5B. We initially developed a three-dimensional common-feature pharmacophore model according to which two hydrogen bond acceptors and one hydrophobic aromatic feature are shared by all molecular series in binding the viral polymerase. The pharmacophoric information was used to retrieve a putative binding site on the surface of the BVDV RdRp and to guide compound docking within the protein binding site. The affinity of all compounds towards the enzyme was scored via molecular dynamics-based simulations, showing high correlation with in vitro EC50 data. The determination of the interaction spectra of the protein residues involved in inhibitor binding highlighted amino acids R295 and Y674 as the two fundamental H-bond donors, while two hydrophobic cavities HC1 (residues A221, I261, I287, and Y289) and HC2 (residues V216, Y303, V306, K307, P408, and A412) fulfill the third pharmacophoric requirement. Three RdRp (K263, R295 and Y674) residues critical for drug binding were selected and mutagenized, both in silico and in vitro, into alanine, and the affinity of a set of selected compounds towards the mutant RdRp isoforms was determined accordingly. The agreement between predicted and experimental data confirmed the proposed common molecular rationale shared by molecules characterized by different chemical scaffolds in binding to the BVDV RdRp, ultimately yielding compound 6b (EC50 = 0.3 μM; IC50 = 0.48 μM) as a new, potent inhibitor of this Pestivirus.

Anti-BVDV Activity Evaluation of Naphthoimidazole Derivatives Compared with Parental Imidazoquinoline Compounds

“…As described above, the driven idea for this project is the isosteric substitution of a nitrogen atom with a carbon atom and evaluate the antiviral activity of the resulted molecules compared with the parental compounds. Naphthoimidazole derivatives ( 3a -j ) were designed and synthesised in order to confirm the relevance of nitrogen atom in position 5 of the imidazoquinoline derivatives for the interaction with the target, these molecule's antiviral results confirmed the predicted affinity data previously published since they showed a consistent loss of antiviral activity with respect to parental compounds [32]. Derivative 3a still maintains a weak antiviral activity, EC 50 at 16 µM, but related at cytotoxicity CC 50 value of 37 µM.…”

“…In detail, arginine 295 interacts with N5 of the lead compound as H-bond donor, while N1 resulted as H-bond acceptor in the interaction with tyrosine 674 and the nitrogen of the benzonitrile moiety gives H-bond with serine 411. Has also been predicted the decrease of affinity of some molecules with the substrate when R295 and Y674 -critical residues for compound binding -are mutated into alanine and the prediction has been confirmed by in vitro assays on the mutant RdRp [32]. In order to have a further negative control for these simulations, we now designed some chemical modification on the lead compounds and parent compounds similarly active in BVDV RdRp inhibition, and we synthesised the designed compounds.…”

“…In the framework of a long-lasting antiviral research program, our group designed and synthesised a series of angular and linear N-polycyclic derivatives active against BVDV, YFV, HCV and other related viruses [23 -30]. The molecular target for BVDV inhibition was identified in the viral RNA dependent RNA polymerase (RdRp) [24,28,31,32]. Imidazo[4,5-g]quinoline derivatives were published as interesting BVDV inhibitors and the most active derivative of that series, 4-(4-chloro-3H-imidazo[4,5-g]quinolin-2yl)benzonitrile (A) (EC 50 0.3µM; CC 50 >100 µM) [32,33] was docked and scored for affinity towards the binding site of the BVDV RdRp, the resulting receptor/ligand complex was relaxed by energy minimisation, followed by MD simulation.…”

Design, Synthesis, Antiviral Evaluation, and SAR Studies of New 1-(Phenylsulfonyl)-1H-Pyrazol−4-yl-Methylaniline Derivatives

“…As a part of our researches on heterocyclic compounds with antiviral activity (Conti et al, 2014, 2017; Carta et al, 2016; Fioravanti et al, 2017), we recently identified ( N -(1,3-diphenyl-1 H -pyrazol-4-yl)methyl)anilines ( 1a-v ) as a new class of potent and selective inhibitors of human respiratory syncytial virus (RSV) replication (Fioravanti et al, 2015). Some derivatives were also endowed with a moderate activity against YFV and BVDV representative type members of the Flavivirus genus and the Pestivirus genus, respectively, within the Flaviviridae family (Fioravanti et al, 2015).…”

Selection and characterization of specific nanobody against bovine virus diarrhea virus (BVDV) E2 protein

“…Bovine viral diarrhea-mucosal disease (BVD-MD), also known as bovine viral diarrhea-mucosal disease virus or mucosal virus, is a contagious disease that often occurs in domestic animals[1, 2]. BVD-MD is caused by BVDV viral infection from viruses in the Pestivirus genus of the Flaviviridae family.…”