Bioactive α-Pyrone Derivatives from the Endophytic Fungus Diaporthe sp. CB10100 as Inducible Nitric Oxide Synthase Inhibitors

Hong Pu,Xueshuang Huang,Jianxin Liu,Yong Huang,Chunmei Nie,Yanwen Duan,Wanying Zheng,Yeji Wang,Yuhui Peng,Yang Tang,Boping Hui

doi:10.3389/fchem.2021.679592

Abstract

Inducible nitric oxide synthase (iNOS) produces NO from l-arginine and plays critical roles in inflammation and immune activation. Selective and potent iNOS inhibitors may be potentially used in many indications, such as rheumatoid arthritis, pain, and neurodegeration. In the current study, five new compounds, including a dibenzo-α- pyrone derivative ellagic acid B (5) and four α-pyrones diaporpyrone A–D (9–12), together with three known compounds (6–8), were isolated from the endophytic fungus Diaporthe sp. CB10100. The structures of these new natural products were unambiguously elucidated using NMR, HRESIMS or electronic circular dichroism calculations. Ellagic acid B (5) features a tetracyclic 6/6/6/6 ring system with a fused 2H-chromene, which is different from ellagic acid (4) with a fused 2H-chromen-2-one. Both 2-hydroxy-alternariol (6) and alternariol (7) reduced the expression of iNOS at protein levels in a dose-dependent manner, using a lipopolysaccharide (LPS)-induced RAW264.7 cell models. Also, they decreased the protein expression levels of pro-inflammatory cytokines, such as tumor necrosis factor-α, interleukin-6 and monocyte chemotactic protein 1. Importantly, 6 and 7 significantly reduced the production of NO as low as 10 μM in LPS-induced RAW264.7 cells. Molecular docking of 6 and 7 to iNOS further suggests that both of them may interact with iNOS. Our study suggests that 6 and 7, as well as the alternariol scaffold may be further developed as potential iNOS inhibitors.

Highlights

Inflammation plays important roles in the occurring and development of many diseases, including rheumatoid arthritis (RA) (Choy and Panayi, 2001), osteoarthritis (OA) (Ahmad et al, 2020), diabetes (Purkayastha and Cai, 2015), and cancers (Zhong et al, 2016)
Since endophytes from medical plants are attractive sources of bioactive natural products, we initiated an endeavor to isolate natural products from endophytes colonized in the stem and root of S. acutum, in our continuous search for new bioactive metabolites of microorganisms from un- or underexplored niches (Jiang et al, 2018; Jiang et al, 2021a). We report that these accumulating efforts have resulted in the isolation of multiple endophytes from S. acutum, and bioactivity-guided natural product isolation have yielded eight compounds, including a new dibenzo-α-pyrone derivative, ellagic acid B (5) and four new α-pyrone diaporpyrone A–D (9–12), together with three known compounds (6–8)
The fungus CB10100 was identified as a Diaporthe species based on DNA sequencing of its internal transcribed spacer 4 (ITS4) and the phylogenetic analysis of its ITS4 with selected Diaporthe strains in GenBank (Supplementary Figure S2)

Summary

Introduction

Inflammation plays important roles in the occurring and development of many diseases, including rheumatoid arthritis (RA) (Choy and Panayi, 2001), osteoarthritis (OA) (Ahmad et al, 2020), diabetes (Purkayastha and Cai, 2015), and cancers (Zhong et al, 2016). Excessive inflammatory mediators are produced by inducible nitric oxide synthase (iNOS) or cyclooxygenase-2 (COX-2), such as NO and prostaglandin E2 (PGE2) (Yu et al, 2019). Bioactive α-Pyrone Derivatives against iNOS is a mammalian protein composed of a C-terminal reductase and an N-terminal oxygenase domain, which produces micromolar NO by oxidizing L-arginine to L-citrulline in the presence of bacterial lipopolysaccharide (LPS) and/or proinflammatory cytokines. A considerable number of iNOS inhibitors, such as arginine derivatives, pyrimidines and aminopyrimidines, as well as aliphatic, aromatic, and cyclic amidines, have been developed (Cinelli et al, 2020). Many of them showed promise in the treatment of arthritis or inflammatory and neuropathic pain in animal models, there are no iNOS inhibitors on the market. There is strong need to discover new iNOS inhibitors as anti-inflammation agents

Methods

Results

Conclusion