Ginkgolic Acid is a Multi-Target Inhibitor of Key Enzymes in Pro-Inflammatory Lipid Mediator Biosynthesis

Abstract

Introduction: Lipid mediators (LMs) comprise bioactive metabolites of polyunsaturated fatty acids, including pro-inflammatory prostaglandins (PGs), thromboxanes (TXs), and leukotrienes (LTs), as well as specialized pro-resolving mediators (SPMs). They are essentially biosynthesized via cyclooxygenase (COX) and lipoxygenase (LO) pathways in complex networks and regulate the progression as well as the resolution of inflammatory disorders including inflammation-triggered cancer. Ginkgolic acid (GA) is a phenolic acid contained in Ginkgo biloba L. with neuroprotective, antimicrobial, and antitumoral properties. Although LMs regulate microbial infections and tumor progression, whether GA affects LM biosynthesis is unknown and was investigated here in detail. Methods: Pharmacophore-based virtual screening was performed along with docking simulations. Activity assays were conducted for isolated human recombinant 5-LO, cytosolic phospholipase (PLA) 2 α, COX-2, and ovine COX-1. The activity of human mPGES-1 and thromboxane A 2 synthase (TXAS) was determined in crude cellular fractions. Cellular LM formation was studied using human monocytes, neutrophils, platelets, and M1- and M2-like macrophages. LMs were identified after (ultra)high-performance liquid chromatography by UV detection or ESI-tandem mass spectrometry. Results: GA was identified as virtual hit in an mPGES-1 pharmacophore-based virtual screening. Cell-free assays revealed potent suppression of mPGES-1 activity (IC 50 = 0.7 µM) that is fully reversible and essentially independent of the substrate concentration. Moreover, cell-free assays revealed COX-1 and TXAS as additional targets of GA with lower affinity (IC 50 = 8.1 and 5.2 µM). Notably, 5-LO, the key enzyme in LT biosynthesis, was potently inhibited by GA (IC 50 = 0.2 µM) in a reversible and substrate-independent manner. Docking simulations support the molecular interaction of GA with mPGES-1 and 5-LO and suggest concrete binding sites. Interestingly, interference of GA with mPGES-1, COX-1, TXAS, and 5-LO was evident also in intact cells with IC 50 values of 2.1–3.8 µM; no radical scavenging or cytotoxic properties were obvious. Analysis of LM profiles from bacteria-stimulated human M1- and M2-like macrophages confirmed the multi-target features of GA and revealed LM redirection towards the formation of 12-/15-LO products including SPM. Conclusions: We reveal GA as potent multi-target inhibitor of key enzymes in the biosynthesis of pro-inflammatory LMs that contribute to the complex pharmacological and toxicological properties of GA.

Prospects of multitarget drug designing strategies by linking molecular docking and molecular dynamics to explore the protein–ligand recognition process

“…should have multitarget activity (Figure 2a-f), and this was verified experimentally with IC 50 values shown of 385, 674, and 46 nM, respectively (Xu et al, 2019). Similarly, the well-known SUMOylation inhibitor, ginkgolic acid, has been identified as a potential hit in pharmacophore-based virtual screening for inhibitors of microsomal prostaglandin E synthase-1 (mPGES-1) and 5-lipoxygenase (5-LO) (Gerstmeier et al, 2019). The docking model supports the molecular interaction of ginkgolic acid with mPGES-1 and 5-LO and provides specific binding sites (Figure 2g,h), correlating with associated IC 50 values of 0.7 and 0.2 μM, respectively (Fukuda et al, 2009).…”

“…Ginkgolic acid fitted into the binding site of mPGES-1 (g) and 5-LO (h) using GOLD software having scores 74.53 and 68.9, respectively. Images in panels (a)-(e) were adapted from (Xu et al, 2019) with permission from Elsevier, while (g) and (h) were adapted from Gerstmeier et al (2019) with permission under a Creative Commons Attribution License (CC BY). EGFR, epidermal growth factor receptor; mPGES, microsomal prostaglandin E synthase; PDGFR, platelet-derived growth factor receptor; VEGFR, vascular endothelial growth factor receptor T A B L E 4 A list of popular force-fields used in molecular dynamics (MD) simulations…”

Ginkgo biloba Seeds—An Environmental Pollutant or a Functional Food

“…In addition, our experiment showed that after further treatment of the aqueous methanolic extract with hexane, the level of ginkgolic acid was reduced from 19 ppm to 4 ppm with minor changes in the concentrations of other bioactive constituents. However, the pharmacological properties of ginkgolic acids family are not limited to those mentioned above [29] and undoubtedly deserve further research.…”

Targeting SUMO Signaling to Wrestle Cancer

“…The limitations of these natural products are that they mostly function in the micromolar range and that they do not singly target SUMOylation. Ginkgolic acid is known to target proinflammatory molecules like prostaglandins and leukotrienes [69], and tannic acid can also induce cancer cell death via activation of apoptosis rather than via inhibition of cell cycle progression, as expected for a SUMOylation inhibitor [68]. Trends in Cancer…”