Structural basis for phospholipase A2-like toxin inhibition by the synthetic compound Varespladib (LY315920)

Abstract

The World Health Organization recently listed snakebite envenoming as a Neglected Tropical Disease, proposing strategies to significantly reduce the global burden of this complex pathology by 2030. In this context, effective adjuvant treatments to complement conventional antivenom therapy based on inhibitory molecules for specific venom toxins have gained renewed interest. Varespladib (LY315920) is a synthetic molecule clinically tested to block inflammatory cascades of several diseases associated with elevated levels of secreted phospholipase A2 (sPLA2). Most recently, Varespladib was tested against several whole snake venoms and isolated PLA2 toxins, demonstrating potent inhibitory activity. Herein, we describe the first structural and functional study of the complex between Varespladib and a PLA2-like snake venom toxin (MjTX-II). In vitro and in vivo experiments showed this compound’s capacity to inhibit the cytotoxic and myotoxic effects of MjTX-II from the medically important South American snake, Bothrops moojeni. Crystallographic and bioinformatics analyses revealed interactions of Varespladib with two specific regions of the toxin, suggesting inhibition occurs by physical blockage of its allosteric activation, preventing the alignment of its functional sites and, consequently, impairing its ability to disrupt membranes. Furthermore, based on the analysis of several crystallographic structures, a distinction between toxin activators and inhibitors is proposed.

Varespladib (LY315920) and Methyl Varespladib (LY333013) Abrogate or Delay Lethality Induced by Presynaptically Acting Neurotoxic Snake Venoms

“…Following the initial description of Varespladib's capacity to inhibit PLA 2 activity of many snake venoms in vitro [7], further studies demonstrated its ability to abrogate lethality in mouse and pig models of envenoming by the venoms of the elapid snakes Oxyuranus scutellatus and Micrurus fulvius, respectively, whose neurotoxic effect is based predominantly on the action of presynaptically acting neurotoxic PLA 2 s [18,19]. Furthermore, Varespladib was effective in the inhibition of myotoxicity induced by crude venoms and isolated PLA 2 s of viperid and elapid species [20,21]. This drug was reported to inhibit the in vitro coagulotoxic effects of venoms of African spitting cobras [22].…”

Sulfur Compounds as Inhibitors of Enzymatic Activity of a Snake Venom Phospholipase A2: Benzyl 4-nitrobenzenecarbodithioate as a Case of Study

“…Recently, Salvador et al reported the crystal structure of a complex formed by a catalytically inactive PLA 2 (MjTX-II from Bothrops moojeni) and a synthetic inhibitor called Varespladib. The complex showed the presence of the inhibitor in the hydrophobic channel of the toxin, interacting particularly with His48 and Lys49 residues [30]. We hypothesize that compounds I and II may have a similar inhibitory mechanism on the catalytically inactive PLA 2 since docking results suggest that these compounds interact with amino acids located at the hydrophobic channel of the toxin.…”

“…Recently, Salvador et al reported the crystal structure of a complex formed by a catalytically inactive PLA2 (MjTX-II from Bothrops moojeni) and a synthetic inhibitor called Varespladib. The complex showed the presence of the inhibitor in the hydrophobic channel of the toxin, interacting particularly with His48 and Lys49 residues [30]. We hypothesize that compounds Instead, docking results suggested several favorable interactions between compounds I and II and the studied PLA 2 ; the hydrogen bond found between the donor Nδ1 of His48 and one oxygen atom of the nitro group as the acceptor is a critical interaction to explain the biological activity of both compounds, since His48 is one of the catalytic residues (Figure 4).…”

“…We hypothesize that compounds I and II may have a similar inhibitory mechanism on the catalytically inactive PLA 2 since docking results suggest that these compounds interact with amino acids located at the hydrophobic channel of the toxin. Hence, compounds I and II could be classified as class I inhibitors of the homologous (Lys49) PLA 2 [30]. The activity of these compounds could be due to the blocking of the hydrophobic channel and prevention of the binding of the fatty acid necessary for the toxin allosteric activation.…”

BoaγPLI: Structural and functional characterization of the gamma phospholipase A2 plasma inhibitor from the non-venomous Brazilian snake Boa constrictor

“…Control groups received 20 μg of Asp-49 or Lys-49 PLA 2 or 20 μg of BoaγPLI or 40 μL of 0.85% saline. Mice blood samples were collected from the tip of the tail [36] into tubes containing citrate as anticoagulant, centrifuged at 1200 g for 15 minutes and the plasma was separated. The amount of Creatine Kinase (CK) present in the samples was estimated with a commercial CK kit (Sigma), according to the manufacture's instructions.…”