The Keap1-Nrf2-ARE signaling pathway is one of the critical mechanisms for cellular defense against oxidative stress damage. The Nrf2 signaling pathway has emerged as a target for the prevention and treatment of oxidative stress-related diseases, such as neurodegenerative diseases, cancer, cardiovascular diseases, metabolic disorders, and inflammation.

Natural products play a significant role in drug discovery, with many medications directly or indirectly derived from them. Currently, the vast majority of Nrf2 activators from natural products are indirect inhibitors of the Keap1-Nrf2 interaction. These inhibitors react with the sulfhydryl functional groups on Keap1, modifying the cysteine residues, leading to structural changes in Keap1 and disruption of the Keap1-Nrf2 complex interaction, thereby activating Nrf2. Based on the structure of the activators and the type of reaction with cysteine, indirect inhibitors of the Keap1-Nrf2 interaction can be classified into several types, including Michael acceptors, oxidizable diphenols, isothiocyanates, dithiolethiones and diallyl sulfides, polyene compounds, and others.

1. Michael Reaction Acceptor Molecules

Michael addition acceptors containing α,β-unsaturated carbonyl groups are among the most common Nrf2 activators. They are considered as mild Lewis acids that can undergo Michael addition reactions with cysteine residues in Keap1, resulting in a conformational change in Keap1, which prevents the ubiquitination of Nrf2 and subsequently induces the expression of phase II detoxifying enzymes and antioxidant proteins. Michael reaction acceptor molecules typically exhibit beneficial cellular responses at low concentrations but fail to activate the Nrf2 signaling pathway at high concentrations, potentially leading to cellular toxicity. Common examples of Michael addition acceptors include curcumin, flavonoids, coumarins, cinnamic acid derivatives, terpenoids, and alkaloids.

Curcumin, containing α,β-unsaturated carbonyl groups, is a typical Michael acceptor that can react with cysteine residues, causing structural changes in Keap1, thereby activating the Keap1-Nrf2-ARE signaling pathway. This activation induces the expression of a series of phase II detoxifying enzymes with antioxidant functions, which play a cytoprotective role.

Caffeic acid phenethyl ester (CAPE), a derivative of cinnamic acid, is an active component found in bee propolis, exhibiting anti-mitotic, anti-inflammatory, and immunomodulatory functions. Ferulic acid and its derivatives, such as ferulic acid ethyl ester (EFE), are abundant in fruits and vegetables, showing strong protective effects against the oxidation of proteins and lipids. Both CAPE and EFE contain α,β-unsaturated ketone structures, and their abilities to induce ARE are comparable to that of curcumin.

Chalcones are a class of open-chain flavonoid compounds found in various plants, with a main structural chain consisting of two benzene rings connected by an α,β-unsaturated carbonyl group through three carbon atoms. Chalcones possess numerous biological properties, including anti-proliferative, anti-inflammatory, and anti-infective activities. Two typical chalcone compounds, sophoraflavone and isoliquiritigenin, can induce the expression of cytoprotective proteins by activating the Keap1-Nrf2-ARE pathway.

Sesquiterpenes such as isoalantolactone, terpenes like perillaldehyde and citral, all contain α,β-unsaturated aldehydes, which can induce the expression of Nrf2-mediated detoxifying enzymes. Andrographolide extracted from Andrographis paniculata activates the Keap1-Nrf2-ARE pathway. Additionally, some naturally occurring coumarin derivatives, such as imperatorin and auraptene, possess the ability to induce ARE-related enzyme activities through the Keap1-Nrf2-ARE signaling pathway. Many flavones also exhibit ARE-inducing properties, including 4-bromoflavone, β-naphthoflavone, and the natural isoflavone sappanone A.

Xanthohumol, a unique product found in hops, and 6-dehydrogingerdione, isolated from ginger, can activate the ARE system by promoting Nrf2 nuclear translocation. This mechanism is closely related to the α,β-unsaturated ketone structure present in their structures.

Furthermore, some naturally occurring coumarin derivatives, such as imperatorin and auraptene, possess the ability to induce ARE-related enzyme activities through the Keap1-Nrf2-ARE signaling pathway. Many flavones also exhibit ARE-inducing properties, including 4-bromoflavone, β-naphthoflavone, and the natural isoflavone sappanone A.

2.Phenols and Quinones with Oxidative Properties

Polyhydroxyphenol compounds are widely present in nature, and some of them can be oxidized in vivo into electrophilic quinone compounds. These quinones react with the sulfhydryl group of cysteine, causing a conformational change in Keap1, which leads to the translocation of Nrf2 into the nucleus. In the nucleus, Nrf2 binds to ARE and induces the expression of downstream phase II detoxifying enzymes, thereby protecting cells against oxidative stress. Common oxidizable diphenols that act as Nrf2 activators include catechins, paeonol, and resveratrol.

Quercetin, found in many plants, is a flavonoid antioxidant derived from plants. Quercetin can inhibit the degradation of Nrf2 and promote its translocation into the nucleus, thereby increasing the level of Nrf2 in the nucleus. Resveratrol, a stilbenoid and non-flavonoid polyphenol compound primarily found in the skins of red grapes and other fruits, is similar to quercetin in that it can induce Nrf2-mediated expression of ARE genes.Green tea is rich in catechin polyphenols, among which epigallocatechin gallate (EGCG) is the most abundant and is the primary active ingredient responsible for green tea's antioxidant effects. Studies have shown that EGCG can increase the level of Nrf2 in the nucleus and induce the transcription of ARE luciferase reporter genes.Carnosol and its acid derivative carnosic acid, both catechol-type diterpenes obtained from rosemary, are used as food preservatives and antioxidants. They can also elevate Nrf2 levels and reduce lipid peroxidation and ROS generation.Baicalein, the main component of the traditional Chinese medicine Scutellaria baicalensis, has been shown in studies to significantly enhance the transcriptional activation of Nrf2.

3.Isothiocyanates

Natural isothiocyanate compounds primarily exist in the form of glucosinolates in cruciferous plants such as broccoli and cabbage, exhibiting anti-tumor and antioxidant activities. Isothiocyanates contain a typical functional group structure (-N=C=S), where the carbon atom is subjected to the electron-withdrawing inductive effect of the N and S atoms on both sides of the double bond, making the C atom highly electrophilic. This electrophilicity enables isothiocyanates to react with the sulfhydryl group of cysteine residues, forming dithiocarbamates, which alter the conformation of Keap1 and thereby activate Nrf2.

Among the isothiocyanate compounds, natural isothiocyanates sulforaphane (SFN) and phenethyl isothiocyanate (PEITC) have been the most extensively studied. Sulfoxythiocarbamate (STCA), which retains the structural features that induce ARE activity, is another notable compound. The electrophilicity of STCA analogues is significantly higher than that of sulforaphane. While isothiocyanates form reversible complexes with sulfhydryl groups, the reaction between STCA and sulfhydryl groups is irreversible.

To be continued in the "next part"...

"Advances in Research on Nrf2 Activators Based on Natural Products and Their Derivatives (Part II)"

References:

[1] Yao Juan, Wu Ping'an, Li Yun, Li Yuefeng, Liu Dongling, Liu Xuefeng. Research Progress on Keap1-Nrf2-ARE Signaling Pathway and Its Activators [J]. Chinese Pharmacological Bulletin, 2019, 35(10): 1342-1346.

[2] Cheng Maojun, Guo Jie, Liu Jing, Xie Saisai. Research Progress on Nrf2 and Natural Product-Derived Nrf2 Activators [J]. Natural Product Research and Development, 2021, 33(01): 165-177.

About the Author

Xiaonisha, a food technology professional holding a Master's degree in Food Science, is currently employed at a prominent domestic pharmaceutical research and development company. Her primary focus lies in the development and research of nutritional foods, where she contributes her expertise and passion to create innovative products.