Targeting epigenetic and posttranslational modifications regulating ferroptosis for the treatment of diseases

Abstract

Ferroptosis, a unique modality of cell death with mechanistic and morphological differences from other cell death modes, plays a pivotal role in regulating tumorigenesis and offers a new opportunity for modulating anticancer drug resistance. Aberrant epigenetic modifications and posttranslational modifications (PTMs) promote anticancer drug resistance, cancer progression, and metastasis. Accumulating studies indicate that epigenetic modifications can transcriptionally and translationally determine cancer cell vulnerability to ferroptosis and that ferroptosis functions as a driver in nervous system diseases (NSDs), cardiovascular diseases (CVDs), liver diseases, lung diseases, and kidney diseases. In this review, we first summarize the core molecular mechanisms of ferroptosis. Then, the roles of epigenetic processes, including histone PTMs, DNA methylation, and noncoding RNA regulation and PTMs, such as phosphorylation, ubiquitination, SUMOylation, acetylation, methylation, and ADP-ribosylation, are concisely discussed. The roles of epigenetic modifications and PTMs in ferroptosis regulation in the genesis of diseases, including cancers, NSD, CVDs, liver diseases, lung diseases, and kidney diseases, as well as the application of epigenetic and PTM modulators in the therapy of these diseases, are then discussed in detail. Elucidating the mechanisms of ferroptosis regulation mediated by epigenetic modifications and PTMs in cancer and other diseases will facilitate the development of promising combination therapeutic regimens containing epigenetic or PTM-targeting agents and ferroptosis inducers that can be used to overcome chemotherapeutic resistance in cancer and could be used to prevent other diseases. In addition, these mechanisms highlight potential therapeutic approaches to overcome chemoresistance in cancer or halt the genesis of other diseases.

Introduction

Ferroptosis, a new form of regulated cell death (RCD), is driven by iron-dependent lipid peroxidation (LPO) of polyunsaturated fatty acid-containing phospholipids (PUFA-PLs) in cellular membranes.^1,2,3,4 Ferroptosis was first reported in 2012 and was found to be induced by erastin, an oncogenic RAS-selective lethal chemical.⁴ Ferroptosis was officially identified as a non-apoptotic RCD triggered by intracellular iron perturbations and oxidative stress.⁵ A imbalance between ferroptosis defense and systems dictates the execution and induction of ferroptosis.⁶ Many metabolic pathways and degradation pathways orchestrate the complex response to ferroptosis by indirectly or directly regulating LPO or iron accumulation.⁷ The metabolic pathways include pathways related to lipid, iron, and amino acid metabolism, and the degradation pathways include pathways such as the ubiquitin–proteasome system (UPS) and macroautophagy/autophagy.

Accumulating evidence suggests that ferroptosis is precisely regulated at multiple levels that include protein posttranslational modifications (PTMs) and epigenetic modifications.⁸ Epigenetic modification that includes DNA methylation, histone modification, and noncoding RNA (ncRNA) regulation is a dynamic and reversible process, which regulates gene expression without changing the DNA sequence.^9,10,11 PTMs covalently or enzymaticly modify or introduce functional groups to dynamically modulate protein localization, activity, and molecular interactions.^12,13 PTMs include phosphorylation, ubiquitination, SUMOylation, acetylation, among others.¹³ Aberrant epigenetic modifications and PTMs dynamically drive abnormal transcription or translation processes to promote anticancer drug resistance, cancer progression, metastasis, etc. The epigenetic modifications regulate the expression levels of ferroptosis-related genes, consequently determining the vulnerability of cancer cells to ferroptosis at both the transcriptional and translational levels.^8,14,15,16 Moreover, emerging evidence has revealed the roles of epigenetic modifications and PTMs in the regulation of ferroptosis in NSDs, CVDs, liver diseases, lung diseases, and kidney diseases.¹⁷

Dysregulated ferroptosis is increasingly recognized as a significant contributor to the pathogenesis of diseases, including cancers,¹⁸ nervous system diseases (NSDs),^{19,20,21,22,23} cardiovascular diseases (CVDs),^{24,25,26,27,28} liver diseases,^29,30 lung diseases,^31,32,33,34, and kidney diseases.³⁵ Recently, ferroptosis has been recognized to play an important role in halting tumor growth.³⁶ In the last decade, accumulating evidence has revealed the role of ferroptosis in tumor growth suppression and shown that ferroptosis induction partially mediates the tumor-suppressive effects of chemotherapy.^3,37 Ferroptosis determines the efficacy of chemotherapy, immunotherapy, and radiotherapy, and thus, combination treatments with ferroptosis inducers could boost the efficacy of those therapies.^38,39 Accumulating studies have shown that pharmacologically modulating ferroptosis may be a therapeutic approach for NSDs, CVDs, liver diseases, lung diseases, and kidney diseases.^{19,25,40,41,42,43,44,45}

In this review, we first summarize the core molecular mechanisms of ferroptosis. Then, the role of epigenetic processes, including histone PTMs, DNA methylation, and ncRNA regulation, are concisely discussed. This discussion is followed by a detailed description of the roles of epigenetic regulation of ferroptosis in the genesis of diseases, including NSD, CVD, liver diseases, lung diseases, and kidney diseases, as well as the application of epigenetic modulators in the treatment of these diseases. Elucidating the epigenetic regulatory mechanisms of ferroptosis in cancer and other diseases will accelerate the development of promising combination therapeutic regimens containing epigenetic agents and ferroptosis inducers that can be used to overcome chemotherapeutic resistance in cancer and could be used to prevent other diseases. In addition, these mechanisms and highlight promising therapeutic approaches that may be used to overcome chemotherapy drug resistance in cancer or halt the genesis of other diseases.

Molecular mechanisms of ferroptosis

The term ferroptosis was coined by the Stockwell laboratory in 2012 based on the three major research areas that provided a foundational understanding of ferroptosis.: the control of ROS,^46,47 the mechanisms of amino acid and lipid metabolism,^48,49,50 and the regulation of iron² (Fig. 1). Iron accumulation and LPO trigger ferroptosis, resulting in plasma membrane rupture.⁵¹ The initiation of ferroptosis requires two key signals, namely, the accumulation of free iron and the inhibition of defense systems, mainly the solute carrier family seven members 11–glutathione–glutathione peroxidase 4 (SLC7A11–GSH–GPX4) system.⁵² The activation of ferroptosis indicates a delicate imbalance between ferroptosis-promoting factors and defense systems. When the former factors significantly override the latter antioxidant defense systems, lethal accumulation of lipid peroxides on cellular membranes leads to membrane rupture and results in ferroptosis-related cell death^3,6 (Fig. 2). Currently, the main ferroptosis defense systems constitute the SLC7A11–GSH–GPX4 system,^6,53 the GTP cyclohydrolase 1–tetrahydrobiopterin (GCH1–BH₄) system,^54,55 the ferroptosis suppressor protein 1–ubiquinol (FSP1–CoQH₂) system,^56,57 the dihydroorotate dehydrogenase–dihydroubiquinone (DHODH–CoQH₂) system,⁵⁸ and the O-acyltransferase domain containing 1/2–monounsaturated fatty acids (MBOAT1/2–MUFA) system.⁵⁹ Many key components of the ferroptosis pathway, e.g., the principal proteins and enzymes engaged in the induction and inhibition of ferroptosis, are transcriptionally controlled by NF-E2 p45-related factor 2 (Nrf2), the transcription factor encoded by NFE2L2.^{60,61,62,63,64} Nrf2 and Kelch-like ECH-associated protein 1 (KEAP1), which is the principal negative regulator of Nrf2 and an E3 ligase adaptor, are critical for maintaining metabolic, redox and protein homeostasis.^65,66 Nrf2 is involved in regulating the transcription of enzymes responsible for GSH biosynthesis, such as glutamate-cysteine ligase catalytic subunit (GCLC), glutamate-cysteine ligase modifier subunit, glutathione disulfide reductase, and GSH synthetase (GSS), which support GPX4-mediated suppression of ferroptosis by increasing and maintaining the GSH level. Other downstream targets of Nrf2 include SLC7A11, GPX4, and NAD(P)H-quinone oxidoreductase 1, as well as iron metabolism proteins, such as ferritin light chain (FTL), ferritin heavy chain 1 (FTH1), ferroportin-1 (FPN1) and heme oxygenase-1 (HO-1), all of which are directly relevant to ferroptosis.^{60,61,62,63,64,67,68,69}

Fig. 1

The diagram depicting key milestones in the field of ferroptosis research

Fig. 2

Core mechanisms of ferroptosis. The core of ferroptosis initiation is iron-dependent lipid peroxidation of polyunsaturated fatty acid (PUFA)-containing phospholipids (PUFA-PLs). When the ferroptosis-promoting factors (or Ferroptosis prerequisites) exceeding the buffering capability of cellular antioxidant systems (or ferroptosis defense systems), lethal accumulation of lipid peroxides on cellular membranes lead to membrane rupture, resulting in ferroptosis-related cell death. The ferroptosis-promoting factors consist of PUFA-PL synthesis and peroxidation, iron metabolism among others. Cells have evolved at least four ferroptosis defense systems,which includes GPX4/xCT system, the FSP1/CoQH₂ system, the DHODH /CoQH₂ system, and the GCH1/BH₄ system, with different subcellular localizations to detoxify lipid peroxides and thus protect cells against ferroptosis. The cytosolic GPX4 (GPX4^cyto) cooperates with FSP1 on the plasma membrane (and other non-mitochondrial membranes) and mitochondrial GPX4 (GPX4^mito) cooperates with DHODH in the mitochondria to neutralize lipid peroxides. ACSL4 and LPCAT3 mediate the synthesis of PUFA-PLs, which are susceptible to LPO through both non-enzymatic and enzymatic mechanisms. Iron initiates the non-enzymatic Fenton reaction and acts as an essential cofactor for ALOXs and POR, which promote LPO. When ferroptosis-promoting factors significantly exceed the detoxification capabilities of ferroptosis defense systems, an excessive and lethal accumulation of lipid peroxides on cellular membranes result in membrane rupture and trigger ferroptosis-mediated cell death

Ferroptosis-promoting factors

PUFA-PL synthesis and peroxidation

The core mechanism of ferroptosis is membrane LPO, a radical-induced chain reaction consisting of a series of chemical reactions between molecular oxygen (O₂), oxidizable lipids, and iron, leading to the incorporation of O₂ into lipids.^70,71 Because of their susceptibility to peroxidation, PUFA-PLs are the substrates for LPO during ferroptosis.^70,72 The critical mediators of the synthesis of PUFA-PLs include acyl-coenzyme A (CoA) synthetase long-chain family member 4 (ACSL4) and lysophosphatidylcholine acyltransferase 3 (LPCAT3).^73,74 ACSL4 ligates the free long-chain PUFAs adrenic acid (AdA) and arachidonic acid (AA) to CoA to generate AA-CoA and ADA-CoA (i.e., PUFA-CoAs), respectively.^74,75 Subsequently, these PUFA-CoAs are re-esterified and incorporated into PLs by LPCAT3 to produce PUFA-PLs (such as AA-phosphatidylethanolamine [PE] and ADA-PE).^73,75,76 PKCβII-mediated phosphorylation of ACSL4 can further activate ACSL4.⁷⁷ Acetyl-CoA carboxylase mediates the synthesis of PUFAs from the basic building block acetyl-CoA.⁷² Nonenzymatic autoxidation through the iron-mediated Fenton reaction is the primary driver of LPO of PUFA-PLs.^70,78,79 The enzymatic reactions mediated by arachidonate lipoxygenase (ALOX) or Cytochrome P450 oxidoreductase are also involved in facilitating LPO.^{80,81,82,83,84} final step of ferroptosis is the formation of pores in plasma or organelle membranes driven by LPO or its secondary products (4-hydroxynonenal and malondialdehyde), which eventually results in cell death. During recent decades, the involvement of ferroptosis in diseases has attracted great interest, not only in the cancer research community.^85,86

Iron homeostasis

Free iron is involved in the core mechanism of ferroptosis in at least two different ways.: inducing accumulation of lethal lipid peroxides and initiating ferroptosis by catalyzing the nonenzymatic Fenton reaction for direct peroxidation of PUFA-PLs^70,79 or functioning as an essential cofactor for POR and ALOX, both of which are enzymes that participate in LPO.^52,80,81,87 Mammalian cells contain a relatively stable pool of intracellular iron—the labile iron pool (LIP)—and maintain this pool by orchestrating the regulation of iron uptake, utilization, storage (ferritin: FTH1/FTL) and export (FPN, the iron export transporter).^88,89 Deregulation of iron metabolism processes can suppress or promote ferroptosis as a result of a decrease or increase in intracellular LIP, respectively. Intracellular iron is mostly stored in the ferritin protein complex, which is composed of 24 subunits of FTL and FTH1.⁹⁰ Poly rC-binding protein 1 binds to and incorporates ferrous iron (Fe²⁺) into ferritin, and this Fe²⁺ is further oxidized to ferric iron (Fe³⁺) by FTH1 inside the ferritin cage, resulting in inert deposits of Fe³⁺ that are unavailable for intracellular use or ROS production.⁹¹ Ferritinophagy, a nuclear receptor coactivator 4 (NCOA4)-mediated autophagy-like process, can degrade ferritin, release iron stored in ferritin into the LIP and facilitate the availability of iron in cells, thereby boosting LPO-driven ferroptosis.^{72,92,93,94,95} Through its function as a selective autophagy receptor, NCOA4 transports intracellular ferritin to autophagic lysosomes and releases free iron through binding FTH1.⁹⁶ Inhibition of cytosolic glutamate oxaloacetate transaminase 1, which enhances ferritinophagy, can increase the LIP and promote ferroptosis.⁹⁷ Conversely, inhibition of ferritinophagy mediated by NCOA4 decreases the LIP and suppresses ferroptosis.^93,94

Ferroptosis defense systems

SLC7A11–GSH–GPX4 axis

There are four antiferroptosis defense systems (or cellular antioxidant systems) that directly neutralize lipid peroxides with distinctive subcellular localizations. Related to the metabolism of amino acids, the SLC7A11–GSH–GPX4 axis is a well-defined and major antiferroptosis defense system.^6,53 SLC7A11 (also named xCT) and solute carrier family 3 member 2 constitute system Xc⁻.^98,99 xCT is a transporter subunit and mediates the antiporter activity of system Xc⁻ through the import of extracellular cystine and export of intracellular glutamate.^98,100 Through a reduction reaction mediated by nicotinamide adenine dinucleotide phosphate (NADPH) consumption in the cytosol, extracellular cystine taken up via SLC7A11 is rapidly reduced to cysteine, which then serves as the rate-limiting precursor for GSH biosynthesis.⁹⁹ GSH is the major cofactor for GPX4-mediated detoxification of lipid peroxides.⁹⁹ Inhibition of SLC7A11 activity or depletion of cystine promotes ferroptosis in various cancer cells.⁹⁹ GPX4 is a member of the GPX protein family with enzymatic lipid repair activity,^101,102 and it has been identified as a key ferroptosis inhibitor.^4,103,104 GPX4 can promote the production of nontoxic lipid PL alcohols from PL hydroperoxides (L-OOH) and simultaneously oxidize two GSH molecules to yield oxidized GSH (GSSG).^46,105 Accumulating studies have revealed the critical role of GPX4 in inhibiting ferroptosis through its genetic or pharmacological manipulation.^106,107 GPX4 is regulated by epigenetic modifications and PTMs.¹⁰⁸ Small molecule inhibitors of GPX4 could be optimized for use as anticancer agents.¹⁰⁹

The FSP1–CoQH₂ system

Several nonperoxidase mechanisms function in parallel with GPX4 to inhibit LPO and ferroptosis. FSP1, localized on the plasma membrane, is also known as apoptosis-inducing factor mitochondria-associated 2. In 2019, FSP1 was identified as the second main protein inhibiting ferroptosis independent of GPX4 through the production of coenzyme Q₁₀ (CoQ₁₀, also known as ubiquinone), reduced forms of endogenous electron carriers and vitamin K, all of which possess significant antioxidant (RTA) activity.^56,57,110 FSP1 functions as an NAD(P)H-dependent oxidoreductase to reduce CoQ₁₀ to regenerate CoQ₁₀-H₂, the reduced form of CoQ₁₀, which can trap lipid peroxyl radicals to hinder LPO and halt ferroptosis.^56,57 FSP1 also inhibits ferroptosis independently of its oxidoreductase function by activating ESCRT-III-dependent membrane repair, which halts ferroptosis.^111,112 Small molecule inhibitors of FSP1 could also be optimized as anticancer agents.⁵⁷

The GCH1–BH₄ system

A study in 2020 revealed that the GCH1–BH4 system is another critical GPX4-independent inhibitor of ferroptosis that acts by suppressing LPO.^54,55 GCH1, which mediates the rate-limiting reaction generating the endogenous metabolite BH₄, was discovered as a suppressor by Kraft⁵⁴ and as an enhancer of ferroptosis by Birsoy in 2020.⁵⁵ As a different RTA, BH4 can be regenerated following its RTA reactions by dihydrofolate reductase (DHFR). Inhibition of DHFR synergizes with inhibition of GPX4 to induce ferroptosis.⁵⁵ Moreover, BH4 enhances CoQ₁₀ synthesis by converting phenylalanine into tyrosine, which can be further converted to 4-OH-benzoate, the precursor of CoQ₁₀.⁵⁴ GCH1-mediated BH4 synthesis reprograms lipid metabolism and inhibits ferroptosis by selectively preventing two polyunsaturated fatty acyl tails from depleting PLs.⁵⁴

The DHODH–CoQH₂ system

A newly identified GPX4-independent mitochondria-localized ferroptosis defense system, the DHODH–CoQH₂ system can compensate for GPX4 loss and detoxify mitochondrial lipid peroxides.⁵⁸ DHODH is localized to the inner mitochondrial membrane, where it catalyzes de novo pyrimidine synthesis through which CoQ10 can be reduced to CoQH₂ at the rate-limiting fourth step. The function of CoQH₂ is analogous to that of FSP1 in extra-mitochondrial membranes.⁵⁸ After acute inactivation of GPX4, DHODH-mediated flux is significantly increased, leading to increased generation of CoQH₂, which neutralizes LPO and inhibits ferroptosis in mitochondria.⁵⁸ Inactivation of both mitochondrial DHODH and GPX4 causes robust ferroptosis by unleashing potent LPO reactions in mitochondria.⁶ Low expression of DHODH or high expression of GCH1 renders cells more sensitive or resistant to ferroptosis, respectively.

MBOAT1/2–MUFA system

The MBOAT1/2-PE-MUFA system is a newly identified ferroptosis defense mechanism independent of GPX4 and FSP1.⁵⁹ Jiang and colleagues identified new PL-modifying enzymes, MBOAT1 and MBOAT2, which function as ferroptosis suppressors.⁵⁹ By functioning as a lyso-PL acyltransferase (LPLAT), membrane-bound MBOAT2 inhibits ferroptosis by selectively incorporating MUFAs into lysophosphatidylethanolamine (lyso-PE), thereby correspondingly increasing the abundance of cellular PE-MUFAs and decreasing the abundance of cellular PE-PUFAs. PE-PUFAs are the preferred substrate for LPO and determine ferroptosis sensitivity.^73,74 The sex hormone receptors, i.e., the estrogen receptor (ER) and androgen receptor (AR), directly transcriptionally upregulate MBOAT1 and MBOAT2, respectively. AR or ER antagonists boost the antitumor activity of ferroptosis inducers in AR⁺ prostate cancers and ER⁺ breast cancers with or without resistance to single-agent hormone therapies.⁵⁹

Epigenetic and posttranslational modifications

Epigenetic modification, a dynamic and reversible process, regulates gene expression without changing the DNA sequence.^9,10 There are four major mechanisms of epigenetic regulation: DNA methylation, chromatin structure regulation, histone PTM, and ncRNA regulation.^9,10,113 The common well-studied epigenetic regulatory mechanisms are DNA methylation, histone modification, and ncRNA regulation.¹¹ The histone subunit in the nucleosome contains a characteristic tail possessing specific amino acids for covalent PTMs, such as ubiquitination, phosphorylation, methylation, acetylation, SUMOylation, acylation, glycosylation, hydroxylation, serotonylation, glycation, and ADP-ribosylation.^{114,115,116,117} Epigenetic regulation of gene expression is mediated by various classes of proteins, most of which have enzymatic activities. Four classes of epigenetic regulators, i.e., “writers”, “erasers”, “readers”, and “remodelers”, constitute the molecular component of the epigenetic regulators of DNA and histone modifications and chromatin structure.^113,118 The writers and erasers add and remove epigenetic marks, respectively. The readers recognize specific epigenetic marks to mediate downstream effects, while the remodelers modulate the chromatin state.¹⁰ Approximately 1000 epigenetic regulators form one of the largest protein groups in mammals. Cancer develops as a result of progressive accumulation of cell-intrinsic genetic and epigenetic changes, which are key characteristics of most cancers.^119,120 Epigenetic mechanisms regulate cancer biology in multiple ways, including driving tumorigenesis and invasion and modulating the immune response. Furthermore, modulation of the epigenome exposes cancer cells to immune-mediated attack, increasing cancer cell sensitivity to immunotherapy in various solid tumors.^121,122 As covalent or enzymatic modifications of synthesized proteins, PTMs modify or introduce functional groups, such as phosphoryl, methyl, acetyl and glycosyl groups, to dynamically modulate protein localization, activity and molecular interactions.^12,13 PTMs include phosphorylation, ubiquitination, SUMOylation, acetylation, methylation, ADP-ribosylation, palmitoylation, neddylation, glycosylation, prenylation, cholesterylation, myristoylation, glutathionylation, sulfhydration, citrullination, S-nitrosylation, and several novel PTMs.¹³ PTMs are usually reversible. Epigenetic modifications and PTMs are strongly correlated with the occurrence and genesis of many diseases. Epigenetic modifications and PTMs transcriptionally and posttranscriptionally regulate gene expression, respectively, and posttranscriptionally modulate protein activity, function and degradation.¹²³ These modifications are required for the maintenance of tissue-specific expression of genes and proteins and for normal cellular development. Dysregulation of epigenetic modifications and PTMs causes aberrant expression of gene and protein signatures and transformation into malignant phenotypes, which induces disease onset and progression.^123,124,125 Accumulating evidence has revealed that dysregulated epigenetic regulation contributes to tumor drug resistance, NSDs, CVDs, liver diseases, lung diseases, and kidney diseases.

Epigenetic and posttranslational modifications regulating ferroptosis in diseases

Epigenetic and posttranslational modifications regulating ferroptosis in cancer

Ubiquitination-mediated regulation of ferroptosis in cancer

Ubiquitination is a key and highly conserved PTM and plays a vital role in modulating the degradation of various protein substrates.^126,127 Deubiquitinases (DUBs) can remove ubiquitin chains to reverse ubiquitination, leading to termination of ubiquitination and preservation of substrate protein expression.¹²⁷ The interaction between ubiquitination by ubiquitinases and DUBs plays an important role in controlling almost all aspects of biological activities. Emerging studies have revealed that deubiquitination/ubiquitination are involved in regulating ferroptosis in cancer. Specific regulators can modulate ferroptosis by regulating the ubiquitination of ferroptosis-related factors (Table 1 and Fig. 3).

Table 1 Posttranslational modification of ferroptosis by ubiquitination in cancer

Fig. 3

Posttranslational modification of ferroptosis by ubiquitination in cancer. ALL acute lymphoblastic leukemia; BAP1 tumor suppressor BRCA1-associated protein 1, BC breast cancer, CCA cholangiocarcinoma, CISD2 CDGSH iron sulfur domain 2, ccRCC clear cell renal cell carcinoma, CRC colorectal cancer, ESCC esophageal squamous cell carcinoma, GBC gallbladder cancer, GBM glioblastoma, GC gastric cancer, HCC hepatocellular carcinoma, HNRNPA2B1 heterogeneous nuclear ribonucleoprotein A2/B1, OC ovarian cancer, QSOX1 quiescin sulfhydryl oxidase 1, TEAD4 TEA domain family member 4, USP ubiquitin specific peptidase, RND1 Rho family GTPase 1, SOCS2 suppressor of cytokine signaling 2, βTrCP beta-transducin repeat containing E3 ubiqutin protein ligase

Hepatocellular carcinoma (HCC): Suppressor of cytokine signaling 2-mediated ubiquitination of SLC7A11 enhances ferroptosis and radiosensitization in HCC.¹²⁸ PCDHB14 functions as a novel tumor suppressor by enhancing ferroptosis through promotion of RNF182-dependent ubiquitination of p65 and blockade of its binding to the promoter of SLC7A11, thereby downregulating SLC7A11 in HCC.¹²⁹ P53 binds to PCDHB14 to induce its expression.¹²⁹ Quiescin sulfhydryl oxidase 1 (QSOX1) promotes ferroptosis induced by sorafenib in HCC by driving ubiquitin-mediated degradation of EGFR, leading to suppression of Nrf2 activation, suggesting that QSOX1 is an inducer of ferroptosis.¹³⁰ Tribbles homolog 2 (TRIB2) functions as one of the key molecules that stabilizes GPX4 and attenuates oxidative stress-induced cell damage.¹³¹ A recent study revealed that TRIB2 inhibits and desensitizes ferroptosis through βTrCP-mediated ubiquitination of TfRC, thereby leading to a decline in the LIP in liver cancer cells.¹³² The largest RNA-binding protein, high-density lipoprotein-binding protein (HDLBP), is an important transporter that protects cells against cholesterol overaccumulation. Elevated expression of HDLBP stabilizes lncFAL to decrease ferroptosis vulnerability by diminishing TRIM69-mediated FSP1 degradation in HCC cells.¹³³ Inhibition of FSP1 enhances the antitumor activity of ferroptosis inducers, supporting the potential utility of targeting FSP1 as a therapeutic approach for HCC patients with high HDLBP or lncFAL expression.¹³³

Gastric cancer (GC): Erastin and RSL3 upregulate the cation channel transient receptor potential melastatin-2 (TRPM2) in GC cell lines. TRPM2 knockdown induces ferroptosis in GC cells, as evidenced by the reductions in the GSH content and GPX activity and increased concentrations of Fe²⁺, ROS and lipid peroxides. Silencing TRPM2 increases the sensitivity of GC cells to RSL3- and erastin-induced ferroptosis by destabilizing the HIF-1α and Nrf2 proteins, suggesting that TRPM2 functions as a negative regulator of erastin- and RSL3-induced ferroptosis¹³⁴ and indicating that the combination of TRPM2-targeted drugs with chemotherapeutics potentiates the effectiveness of treatment and improves the outcome of patients. Increased expression of the long noncoding RNA (lncRNA) BDNF-AS inhibits ferroptosis by recruiting WDR5 to transcriptionally upregulate FBXW7, thereby mediating the ubiquitin-dependent degradation of VDAC3 and promoting the progression of GC.¹³⁵ Increased expression of CST1 promotes the progression and metastasis of GC via inhibition of ferroptosis by reducing GPX4 ubiquitination and degradation by recruiting OTUB1.¹³⁶ Paclitaxel and cisplatin (CDDP) increase the secretion of cancer-associated fibroblast-derived miR-522 by activating the USP7/hnRNPA1 axis, thereby suppressing ALOX15 expression and ferroptosis and ultimately leading to reduced chemosensitivity.¹³⁷

Colorectal cancer (CRC): Increased expression of LINC00239 is associated with poorer prognosis in patients with CRC.¹³⁸ Overexpression of LINC00239 inhibits erastin- and RSL3-mediated antitumor activity by inhibiting ferroptosis. LINC00239 interacts with and binds to the Kelch domain of Keap1, thereby inhibiting the ubiquitination of Nrf2 to stabilize it, suggesting that LINC00239 functions as an oncogene by inhibiting ferroptosis by binding to Keap1 and stabilizing Nrf2 in CRC cells.¹³⁸ Loss of ribonucleotide reductase subunit M1 (RRM1) destabilizes p53 and increases the sensitivity of different types of cancer cells to ferroptosis by inhibiting the expression of GPX4. Silencing RRM1 promotes the interaction of MDM2 and P53 while decreasing the binding of USP11 to p53, thereby stimulating the ubiquitination of p53, in turn leading to decreased expression of p21, which eventually induces ferroptosis through inhibition of GPX4 and results in cancer cell death.¹³⁹

Cholangiocarcinoma (CCA): FBXO31 functions as a tumor suppressor, and its expression increases the sensitivity of stem cell-like cells to cisplatin by enhancing ferroptosis through promotion of the proteasomal degradation of GPX4 in CCA cells.¹⁴⁰ The expression of a component of the linear ubiquitin chain activation complex, shank-associated RH domain interacting protein (SHARPIN), was found to be increased, promoting cell proliferation through ferroptosis inhibition mediated by promoting the ubiquitination and degradation of P53, thereby upregulating SLC7A11/GPX in CCA cells.¹⁴¹ Blocking SHARPIN-mediated inhibition of ferroptosis via the P53/SLC7A11/GPX4 axis and targeting SHARPIN might be promising treatment approaches for CCA.

Non-small cell lung carcinoma (NSCLC): Inducing ferroptosis is a good treatment approach for LUAD patients with late-stage and/or therapy-resistant tumors. Erastin promotes the nuclear translocation of USP7 by increasing its interaction with p53, which erases the monoubiquitination of lysine 120 on histone H2B (H2Bub1) in the SLC7A11 gene regulatory region and inactivates SCL7A11 expression, eventually leading to ferroptosis in NSCLC cells.¹⁴² TRIM6 functions as an oncogene by promoting SLC1A5 ubiquitination and degradation, thereby inhibiting glutamine import, glutaminolysis, LPO, and ferroptosis.¹⁴³ USP35 is upregulated in NSCLC. USP35 knockdown promotes ferroptosis and increases the sensitivity of lung cancer cells to paclitaxel and cisplatin.¹⁴⁴ Conversely, overexpression of USP35 facilitates lung cancer cell growth and tumor progression by reducing Erastin/RSL3-triggered ferroptosis. USP35 directly interacts with the FPN protein to maintain its stability, suggesting that USP35 functions as an oncogene by inhibiting ferroptosis through stabilization of FPN.¹⁴⁴ USP11 deubiquitinates and stabilizes Nrf2. Elevated USP11 expression promotes cancer cell proliferation by inhibiting ferroptosis through deubiquitination and stabilization of Nrf2 in NSCLC cells.¹⁴⁵

Glioblastoma: Overexpression of TRIM7 inhibits NCOA4-mediated ferritinophagy and ferroptosis through directly binding and ubiquitinating NCOA4 in human glioblastoma cells. Ablation of TRIM7 increases the sensitivity of human glioblastoma cells to temozolomide, suggesting that TRIM7 functions as a negative regulator of ferroptosis.¹⁴⁶ Downregulated expression of Rho family GTPase 1 (RND1) predicts a better prognosis in patients with glioblastoma multiforme (GBM). RND1 induces ferroptosis by interacting with and deubiquitinating P53, thereby inhibiting SLC7A11 in GBM cells.¹⁴⁷ Circular RNAs (circRNAs) regulate ferroptosis through several mechanisms in GBM. Downregulation of circLRFN5 promotes malignancy by inhibiting ferroptosis in GBM cells. CircLRFN5 binds to the paired related homeobox 2 (PRRX2) protein and promotes its ubiquitin-mediated degradation, thereby transcriptionally upregulating the expression of the ferroptosis suppressor GCH1 in glioma stem cells (GSCs), leading to ferroptosis induction.¹⁴⁸

Clear cell renal cell carcinoma (ccRCC): As a tumor suppressor, the H2A DUB BRCA1-associated protein 1 (BAP1) suppresses tumorigenesis by inducing ferroptosis through suppression of SLC7A11 in ccRCC. BAP1, which encodes a nuclear DUB to reduce histone 2A ubiquitination (H2Aub) on chromatin, reduces the H2Aub level in the SLC7A11 promoter and suppresses SLC7A11 expression in a deubiquitination-dependent manner, leading to inhibition of cystine uptake, LPO and ferroptosis.¹⁴⁹ BAP1 inhibits the progression of tumors partially by inducing ferroptosis through suppression of SLC7A11 expression, and cancer-associated BAP1 mutants lose their ability to suppress SLC7A11 and promote ferroptosis. BAP1 promotes Erastin-induced ferroptosis by repressing SLC7A11 expression. BAP1 and PRC1 (a major H2Aub ligase) coordinately suppress SLC7A11 expression by regulating the level of H2Aub in the SLC7A11 promoter.¹⁵⁰

Gynecologic neoplasms: An increased expression level of estrogen receptor 1 (ESR1) promotes cancer by inhibiting ferroptosis in breast cancer cells through NEDD4L-mediated ubiquitination and degradation of CD71.¹⁵⁰ Silencing ESR1 significantly promotes ionizing radiation-mediated ferroptosis and increases the CD71 protein level.¹⁵⁰ The results suggest that in breast cancer, ESR1 is an inhibitor of ferroptosis, while CD71 is an inducer of ferroptosis.¹⁵⁰ Methylenetetrahydrofolate reductase (MTHFR), a key enzyme for folic acid metabolism, inhibits TRC8-mediated HMOX1 ubiquitination, thereby blocking ferroptosis and promoting the growth of ovarian cancer (OC) cells.¹⁵¹ The E3 ubiquitin ligase 3-hydroxy-3-methylglutaryl reductase (HRD1) exhibits decreased degradation in ovarian cancer tissues and functions as a tumor suppressor. HRD1 was found to inhibit the proliferation and colony formation of ovarian cancer cells by inducing ferroptosis through facilitation of ubiquitination-dependent SLC7A11 degradation. This finding suggests that HRD1 exerts antitumor effects by promoting ferroptosis in ovarian cancer cells by increasing SLC7A11 degradation.¹⁵²

Other tumors: In bladder cancer (BCa), an important serine metabolism enzyme, phosphoglycerate dehydrogenase (PHGDH), is highly expressed. PHGDH interacts with the RNA-binding protein poly(rC)-binding Protein 2 and inhibits its ubiquitin-mediated degradation, which in turn upregulates SLC7A11 and inhibits ferroptosis, thereby promoting malignant progression.¹⁵³ The PHGDH inhibitor NCT-502 enhances ferroptosis and halts tumor progression in BCa.¹⁵³ This finding indicates that inhibition of PHGDH could be a therapeutic strategy for BCa. PAQR3, a member of the Progestin and AdipoQ Receptor (PAQR) family, is a newly discovered tumor suppressor, and its expression is decreased in acute lymphoblastic leukemia (ALL). PAQR3 suppresses cell proliferation and aggravates ferroptosis by increasing ubiquitin-dependent degradation of Nrf2 in ALL cells.¹⁵⁴ Activation of autophagy was found to sensitize ALL cells to Erastin-induced ferroptosis by inhibiting ubiquitination-dependent degradation of VDAC3 mediated by the E3 ligase FBXW7,¹⁵⁵ indicating that autophagy activation combined with ferroptosis induction is a potential therapeutic strategy for ALL. FOXM1 and Nedd4 regulate VDAC2/3 during ferroptosis in melanoma cells.¹⁵⁶ Erastin induces FOXM1 expression to activate the transcription of Nedd4, which degrades VDAC2/3 and suppresses ferroptosis. Ablation of Nedd4 inhibits the degradation of VDAC2/3 proteins, increasing the sensitivity of cancer cells to Erastin-induced ferroptosis.¹⁵⁶ These results suggest that Nedd4 regulates ferroptosis and highlight Nedd4 as a target for overcoming Erastin-induced resistance in melanoma cells. Silencing CDGSH iron sulfur domain 2, an iron-sulfur protein with a [2Fe-2S] cluster that is critical for cell proliferation and iron homeostasis, increases the degradation of p62 (an autophagy adaptor), leading to increased Keap1-mediated ubiquitination of Nrf2 and its subsequent degradation, thereby promoting ferroptosis in fibrosarcoma cells.¹⁵⁷

Phosphorylation-mediated regulation of ferroptosis in cancer

Histone phosphorylation is a histone modification whose modulation is catalyzed by many protein kinases and phosphatases, such as protein phosphatase 1, mitogen- and stress-activated kinases, and Aurora B, and is achieved through the addition of phosphate groups to threonine, serine or tyrosine residues in histone tails.¹⁵⁸ Histone phosphorylation frequently occurs early after the formation of a DNA double-strand break and mediates the recruitment of DNA damage repair proteins.¹⁵⁹ Histone phosphorylation has been revealed to be associated with transcriptional activation. As an important epigenetic PTM, phosphorylation is strongly associated with tumorigenesis.^160,161 Emerging studies suggest that phosphorylation regulates ferroptosis in cancer (Table 2 and Fig. 4).

Table 2 Posttranslational modification of ferroptosis by phosphorylation in cancer

Fig. 4

Posttranslational modification of ferroptosis by phosphorylation and acetylation in cancer. ATF4 Activating transcription factor 4, ADCY10 adenylyl cyclase 10; β-OHB β-hydroxy-butyric acid; βTrCP beta-transducin repeat containing E3 ubiqutin protein ligase, CBP/p300 histone acetyltransferases CBP and p300, CCA cholangiocarcinoma, CRC colorectal cancer, CHAC1 glutathione specific gamma-glutamylcyclotransferase 1, CISD2 CDGSH iron sulfur domain 2, CKB creatine kinase B, DDR2 discoidin domain receptor tyrosine kinase 2, eIF2α alpha subunit of eukaryotic initiation factor 2, FSP1 ferroptosis suppressor protein 1, GC gastric cancer, GCN2 general control nonderepressible 2, GSS glutathione synthetase, HCC hepatocellular carcinoma, HMGCL ketogenesis-related hydroxy-methyl-glutaryl-CoA lyase, HSPB1 heat shock protein beta-1, HSF1 heat shock factor 1, IGF1R insulin-like growth factor 1 receptor, LUAD lung adenocarcinoma, NAT10 N-acetyltransferase 10, PKC protein kinase C, MESH1 metazoan SpoT homolog 1, RRM2 ribonucleotide reductase regulatory subunit M2, RND1 Rho family GTPase 1, TNBC, triple-negative breast cancer, YAP Yes-associated protein

Colorectal cancer: AMP-activated protein kinase (AMPK)-mediated phosphorylation of BECN1 enhances ferroptosis through binding to SLC7A11 to directly block system Xc^–.¹⁶² Silencing BECN1 inhibits ferroptosis induced by the system Xc^- inhibitors erastin-, sulfasalazine-, and sorafenib. Phosphorylation of BECN1 (Ser90/93/96) induced by AMPK is required for BECN1-SLC7A11 complex formation and LPO. Inhibition of PRKAA/AMPKα reduces Erastin-mediated BECN1 phosphorylation at S93/96, BECN1-SLC7A11 complex formation, and subsequent ferroptosis. Activation of the BECN1 pathway increases ferroptosis in CRC cells.¹⁶² KIF20A expression is increased in oxaliplatin-resistant CRC cell lines. Silencing KIF20A increases the sensitivity of cancer cells to oxaliplatin and suppresses NUAK1, thereby upregulating the expression of PP1β, which subsequently decreases the phosphorylation of downstream GSK3β to suppress the activation of Nrf2 and the expression of GPX4, abolishing oxaliplatin resistance in these cells.¹⁶³

Hepatocellular carcinoma: AKT activated by insulin-like growth factor 1 receptor (IGF1R) signaling phosphorylates creatine kinase B (CKB) at T133, reducing its metabolic activity and increasing its binding to and phosphorylation of GPX4 at S104, which prevents HSC70 binding to GPX4, thereby abrogating degradation of GPX4 by chaperone-mediated autophagy, inhibiting ferroptosis and promoting tumor growth in mice.¹⁶⁴ Elevated expression of ribonucleotide reductase regulatory subunit M2 (RRM2) inhibits ferroptosis in HCC cells. Ferroptotic stress induces phosphorylation of RRM2 at T33, thus promoting the RRM2-GSS interaction, which results in the translocation of RRM2 and GSS to the proteasome for simultaneous degradation.¹⁶⁵

Gastric cancer: A recent study showed that Metazoan SpoT Homolog 1 (MESH1) is the first NADPH phosphatase regulating ferroptosis in the cytosol.¹⁶⁶ Silencing MESH1 dramatically protects cells against ferroptosis. In GC, MESH1 knockdown upregulates the protein expression of ATF3 and ATF4, increases eIF2α phosphorylation, and induces the mRNA expression of XBPs, ATF3, and CHOP. Concurrent silencing of ATF4 restores the sensitivity of MESH1-depleted RCC4 cells to ferroptosis. Concurrent knockdown of NADK abolishes ATF3 induction.¹⁶⁷

Breast cancer: PKCβII phosphorylates ACSL4 to boost LPO to induce ferroptosis.⁷⁷ PKCβII was found to function as a critical contributor to ferroptosis by sensing initial lipid peroxides and amplifying ferroptosis-associated LPO by phosphorylating and activating ACSL4. Inhibiting the PKCβII-ACSL4 pathway attenuates ferroptosis in vitro and impedes immunotherapy-induced ferroptosis in vivo. Murine recurrent breast tumor cells are highly sensitive to ferroptosis. The receptor for collagen I, discoidin domain receptor tyrosine kinase 2 (DDR2), is upregulated in human mesenchymal breast cancer cells and ferroptosis-sensitive recurrent tumor cells. Upregulation of DDR2 increases the susceptibility of recurrent breast tumors to ferroptosis through the Hippo pathway.¹⁶⁸ Erastin induces upregulation and phosphorylation of DDR2. Epithelial–mesenchymal transition (EMT)-driven DDR2 upregulation maintains a growth advantage but results in ferroptosis susceptibility mediated by YAP/TAZ in recurrent tumors.¹⁶⁸ Silencing DDR2 reduces the clonogenic proliferation of recurrent tumor cells. These results reveal an important role of EMT-driven DDR2 upregulation in maintaining the growth advantage but endowing YAP/TAZ-mediated ferroptosis susceptibility in recurrent tumors, highlighting potential therapeutic strategies to eradicate recurrent breast cancer cells with mesenchymal features.¹⁶⁸ Cystine starvation activates GCN2 to increase the phosphorylation of eIF2α and the expression of the ATF4 protein and its target gene CHAC1. Silencing CHAC1 rescues cystine depletion-induced ferroptosis in human triple-negative breast cancer (TNBC) cells.¹⁶⁹

Non-small cell lung carcinoma: Inhibition of system XC⁻ increases endogenous glutamate accumulation, which enhances adenylyl cyclase (ADCY)-mediated Ca²⁺-dependent cAMP production to stimulate protein kinase A (PKA)-associated phosphorylation and suppress glutamine-fructose-6-phosphate transaminase (GFPT1), thereby suppressing YAP expression and failing to sustain ferritinophagy-triggered transcriptional compensation of FTH1, leading to increases in the LIP and ferroptosis sensitivity.¹⁷⁰

Osteosarcoma/prostate adenocarcinoma: Heat shock protein beta-1 (HSPB1) functions as an inhibitor of ferroptosis in cancer. Erastin enhances heat shock factor 1 (HSF1)-dependent HSPB1 expression in cancer cells. Silencing HSPB1 and HSF1 promotes but HSPB1 overexpression inhibits erastin-induced ferroptosis. Protein kinase C (PKC)-mediated HSPB1 phosphorylation inhibits ferroptosis. Inhibition of HSPB1 phosphorylation and HSF1-HSPB1 signaling boosts the anticancer activity of erastin in vivo.¹⁷¹

Acetylation-mediated regulation of ferroptosis in cancer

Protein acetylation is required for key cellular processes related to physiology and diseases, such as transcriptional activity, protein stability, enzyme activity, protein‒protein interactions, subcellular localization, and protein‒DNA interactions.¹⁷² Histone acetylation was first found to modulate gene transcription as early as the 1960s.¹⁷³ Since the first acetylation modification of a nonhistone protein, p53, was found in the 1980s, various nonhistone proteins have been identified as targets for acetylation.¹⁷² Emerging studies suggest that acetylation regulates ferroptosis in cancer (Table 3 and Fig. 4).

Table 3 Posttranslational modification of ferroptosis by acetylation in cancer

Non-small cell lung carcinoma: Regulation of ferroptosis by P53 was first reported in 2015, and the associated study revealed SLC7A11 as a direct target gene of P53 for suppression.¹⁷⁴ The acetylation-defective mutant p53^3KR was found to be unable to induce cell cycle arrest, senescence and apoptosis but retained the full ability to inhibit SLC7A11 expression and induce ferroptosis. Gu and colleagues revealed the role of acetylation in modulating P53-mediated ferroptosis and tumor suppression in NSCLC.¹⁷⁵ Expression of P53^3KR efficiently inhibited tumor growth, which was restored by the overexpression of SLC7A11 in vivo, suggesting the important role of SLC7A11 inhibition in p53-mediated tumor suppression. However, p53^4KR (K98R + 3KR) lost the ability to suppress SLC7A11, thus inducing ferroptosis and tumor suppression.¹⁷⁵ These results revealed the role of acetylation in regulating p53-mediated ferroptosis and tumor suppression. Most recent studies have shown that GINS4, a regulator of initiation and elongation during DNA replication, negatively regulates ferroptosis in lung adenocarcinoma (LUAD). Ablation of GINS4 facilitates ferroptosis.¹⁷⁶ GINS4 suppresses P53-mediated ferroptosis by stabilizing p53 via activation of snail, which antagonizes the acetylation of P53 at K351. These results indicate that GINS4 is a potential oncogene that destabilizes p53 and then inhibits ferroptosis, thus constituting a potential therapeutic target for LUAD.¹⁷⁶

Hepatocellular carcinoma: Dipeptidyl peptidase 4 (DPP4) is a key protein that maintains intracellular iron accumulation and LPO. Ketogenesis-related hydroxy-methyl-glutaryl-CoA lyase (HMGCL) negatively regulates cell proliferation and metastasis in HCC. HMGCL increases β-hydroxybutyric acid (β-OHB)-mediated acetylation of DPP4 at histone 3 lysine 9 (H3K9) and promotes its expression, leading to increased vulnerability of HCC cells to erastin- and sorafenib-induced ferroptosis.¹⁷⁷ This observation suggests that HMGCL functions as a tumor suppressor by increasing ferroptosis susceptibility driven by β-OHB-mediated acetylation of DPP4.

Colorectal cancer: NAT10 negatively regulates tumorigenesis and metastasis in CRC. Upregulation of N-acetyltransferase 10 (NAT10) promotes cancer progression by inhibiting ferroptosis via N4 acetylation and stabilization of FSP1 mRNA in CRC cells.¹⁷⁸ FSP1 mRNA undergoes N4-acetylcytidine (ac4C) modification, leading to inhibition of ferroptosis. This observation reveals that NAT10-mediated N4 acetylation of FSP1 mRNA terminates ferroptosis in colon cancer cells.

Glioma: KAT6B, a histone acetyltransferase, promotes glioma progression by inhibiting ferroptosis through epigenetic induction of STAT3.¹⁷⁹ KAT6B expression is increased in glioma. KAT6B reverses erastin-induced ferroptosis in glioma cells, indicating that KAT6B functions as an inhibitor of ferroptosis. A mechanistic study showed that ablation of KAT6B represses the expression of STAT3. Silencing KAT6B inhibits the enrichment of RNA polymerase II (RNA pol II) and histone H3 lysine 23 acetylation (H3K23ac) on the STAT3 promoter, while loss of STAT3 reverses KAT6B-induced inhibition of ferroptosis in glioma cells.

Osteosarcoma: K139 has been identified as a novel acetylation site in human p53 accounting for p53-mediated mTOR suppression.¹⁸⁰ The p53-4KR mutant retains the ability to inhibit mTOR activity, which is completely abolished in the p53-5KR (K136R + K98R + K117R + K161R + K162R) mutant. The 5KR mutation series in p53 abolishes its remaining tumor suppressor function. Treatment with an mTOR inhibitor was found to suppress early-onset tumor formation in P535KR/5KR mice, which was similar to that observed in p53-null mice. This finding reveals a role of p53-mediated mTOR regulation in tumor suppression.¹⁸⁰

Methylation-mediated regulation of ferroptosis in cancer

First discovered in 1959, protein methylation is an important PTM that regulates the functions of both nonhistone and histone proteins.^181,182 Histon methylation was identified in 1964.¹⁸³ Currently, accumulating discoveries have revealed much of the biology of protein methylation.¹⁸⁴ Protein methylation occurs mainly on side chains of arginine (Arg) and lysine (Lys) residues.¹⁸⁵ The protein Arg methyltransferases (PRMTs) that use S-adenosylmethionine (SAM) as the methyl donor induce mono- or dimethylation of Arg on its side chains,^185,186 whereas Lys residues may undergo mono-, di- or trimethylation (me1, me2 or me3, respectively) in a SAM-dependent manner.¹⁸⁷ Circumstantial evidence has shown that dysregulation of protein methylation is involved in tumorigenesis.^188,189 Emerging studies have suggested that methylation regulates ferroptosis in cancer (Table 4 and Fig. 5).

Table 4 Epigenetic modification of ferroptosis by methylation in cancer

Fig. 5

Epigenetic modification of ferroptosis by methylation, glcNAcylation, and SUMOylation in cancer. ALG3 Alpha 1,3-mannosyltransferase, CBS cystathionine -beta-synthase, CREB, cAMP response element-binding protein, CYP2E1 cytochrome P450 family two subfamily E member 1, EGFR epidermal growth factor receptor, ELOVL5 elongation of very long-chain fatty acid protein 5, EP300 E1A binding protein P300, FADS2 fatty acid desaturase 2, FASN fatty acid synthase, FADS1 fatty acid desaturase 1, FSP1 ferroptosis suppressor protein 1, FTH ferritin heavy chain, GALNT14 N-Acetylgalactosaminyltransferase-14, GBM glioblastoma, HMGCL ketogenesis-related hydroxy-methyl-glutaryl-CoA lyase, KDM3B histone lysine demethylase 3B, KDM4A histone lysine demethylase 4 A, β-OHB β-hydroxy-butyric acid, MM multiple myeloma, NAT10 N-acetyltransferase 10, PSAT1 phosphoserine aminotransferases 1, SDH succinate dehydrogenase, SENP1 small ubiquitin-like modifier (SUMO)-specific protease 1, TFRC transferrin receptor

Cancers of the digestive system

Increased expression of ELOVL5 and FADS1 enhances the sensitivity of mesenchymal-type GC cells to ferroptosis. ELOVL5 and FADS1 are silenced by DNA methylation in intestinal-type GCs, rendering cells resistant to ferroptosis. AA and AdA derived from linoleic acid are not generated in intestinal-type GCs. AA supplementation restores the ferroptosis sensitivity of intestinal-type GCs.¹⁹⁰ In CRC, increased methylation levels of solute carrier family 2 member 1 (SLC2A1) greatly inhibit autophagy and ferroptosis, resulting in immunosuppression and a poor prognosis in patients.¹⁹¹ Aberrant methylation of the promoter inactivates PCDHB14 in HCC patients. PCDHB14 functions as a tumor suppressor to enhance cell cycle arrest, inhibit cell proliferation, and induce ferroptosis. PCDHB14 inhibits HCC progression by enhancing RNF182-mediated degradation of p65 and promotes cell sensitivity to ferroptosis in HCC by suppressing SLC7A11.¹²⁹

Non-small cell lung carcinoma

There are low levels of DNA methylation in upstream regions of GPX4 and an increased level of H3K4me3. H3K27ac leads to increased expression of GPX4. Inhibiting the expression of GPX4 induces ferroptosis in cancer cells and boosts the anticancer effect of cisplatin.¹⁹² cAMP response element-binding protein (CREB) is upregulated in LUAD. Silencing CREB decreases the viability and inhibits the growth of cancer cells by promoting ferroptosis. CREB suppresses ferroptosis by binding the promoter region of GPX4, and E1A binding protein P300 (EP300) enhances this binding.¹⁹³ A lysine monomethylase, SET7, directly interacts with the DUB OTUB1 to catalyze its methylation at lysine 122, inhibiting the binding of OTUB1 to the E2 ubiquitin-conjugating enzyme UBC13. SET7-mediated methylation of OTUB1 promotes ferroptosis through relieving OTUB1-mediated suppression of ferroptosis, highlighting that SET7 inhibitor treatment might enhance OTUB1 function as a therapeutic approach.¹⁹⁴

Clear cell renal cell carcinoma

CX3CL1 overexpression attenuates the proliferation and metastasis of ccRCC cells by increasing their sensitivity to ferroptosis. The expression of CX3CL1 is associated with its DNA methylation level in ccRCC.¹⁹⁵ Decreased expression of succinate dehydrogenase (SDH), which is responsible for oxidative phosphorylation (OXPHOS) and flux through the tricarboxylic acid (TCA) cycle, is correlated with ccRCC progression. SDH deficiency enhances tumorigenesis by inhibiting ferroptosis in ccRCC cells. High mutation rates and increased methylation in the SDH promoter lead to SDH deficiency, thereby promoting tumorigenesis through inhibition of ferroptosis.¹⁹⁶

The promoter of the gene coding for FSP1 is hypermethylated in ALL cells, silencing the expression of FSP1 and generating selective dependency on GSH-centered antiferroptosis defenses. Expression of FSP1 in trans increases the resistance of leukemic cells to compounds targeting the GSH-dependent antiferroptosis pathway. FSP1 overexpression promotes ALL tumor growth.

Acute lymphoblastic leukemia

ALL cells are selectively sensitive to compounds that block the GSH-dependent ferroptosis defense system. The promoter of FSP1 has been found to be hypermethylated in ALL cell lines and patient biopsies. Silencing FSP1 produces selective dependency on the GSH-centered ferroptosis defense system. Overexpression of FSP1 enhances the resistance of leukemic cells to compounds that target the GSH-dependent antiferroptosis pathway, revealing metabolic vulnerability in ALL.¹⁹⁷

Ferroptosis induction leads to changes in histone modification and DNA methylation associated with cellular senescence.

Multiple myeloma

RSL3-induced ferroptosis leads to changes in histone modification and DNA methylation related to cellular senescence.¹⁹⁸ MM1 multiple myeloma cells are sensitive to ferroptosis induced by RSL3 and epigenetic reprogramming. Enrichment of CpG probes in genes associated with cell cycle progression and senescence was found in ferroptotic MM cells, suggesting that ferroptotic cell death is associated with an epigenomic stress response that might increase the therapeutic applicability of ferroptotic compounds.¹⁹⁸

Fibrosarcoma

The H3K9 demethylase KDM3B functions as a potential epigenetic regulator of ferroptosis. KDM3B can inhibit erastin-triggered ferroptosis. Overexpression of KDM3B reduces H3K9 methylation and inhibits ferroptosis by upregulating SLC7A11 in cooperation with the transcription factor ATF4.¹⁹⁹

Osteosarcoma

Upregulated KDM4A expression was found to be associated with poorer prognosis. Knockdown of KDM4A promotes ferroptosis through regulation of SLC7A11 transcription by controlling the demethylation of H3K9me3 in the SLC7A11 promoter.²⁰⁰

Glycosylation-mediated regulation of ferroptosis in cancer

O-GlcNAcylation, the attachment of O-linked N-acetylglucosamine (O-GlcNAc) moieties to threonine or serine residues of proteins in the nucleus, cytosol or mitochondria, is an important PTM that links nutrient flux to gene transcription in tumorigenesis.^201,202 O-GlcNAcylation is dynamically and finely modulated by O-GlcNAcase (OGA) and O-GlcNAc transferase (OGT) proteins. Abnormal O-GlcNAcylation has been identified as a common characteristic in cancers due to deregulated cellular nutrient flux.^203,204 Emerging studies suggest that GlcNAcylation regulates ferroptosis in cancer (Table 5 and Fig. 5).

Table 5 Posttranslational modification of Ferroptosis by GlcNAcylation in cancer

Cancers of the digestive system: GlcNAcylated c-Jun represses ferroptosis by antagonizing the synthesis of GSH in HCC. Erastin inhibits malignant phenotypes by inhibiting the O-GlcNAcylation, protein expression, transcriptional activity and nuclear accumulation of c-Jun in HCC. An overabundance of O-GlcNAcylated c-Jun conversely inhibits ferroptosis by increasing the synthesis of GSH via increased transcription of PSAT1 and CBS.²⁰⁵ This observation indicates that O-GlcNAcylated c-Jun is at the core of ferroptosis and that targeting c-Jun O-GlcNAcylation might be a potential therapeutic approach for HCC.²⁰⁵ O-GlcNAcylation stabilizes and enhances the expression of YAP, which plays a pivotal role in controlling ferroptosis. O-GlcNAcylated YAP mediates increased ferroptosis sensitivity through a transcriptional increase in TfRC expression in HCC cells.²⁰⁶ Knockdown or mutation of YAP abolishes the O-GlcNAcylation-mediated increase in sensitivity to ferroptosis. The related study provided the first evidence that O-GlcNAcylation can increase the sensitivity of HCC cells to ferroptosis via YAP/TFRC, highlighting new therapeutic strategies for HCC.²⁰⁶ In pancreatic cancer, O-GlcNAcylation of zinc finger E-box-binding homeobox 1 (ZEB1) enhances ferroptosis in mesenchymal pancreatic cancer cells. High glucose exposure increases the O-GlcNAcylation of ZEB1, transcriptionally inducing fatty acid synthase (FASN) and fatty acid desaturase 2 (FADS2) expression, thereby resulting in ferroptosis in mesenchymal pancreatic cancer cells.²⁰⁷ These results indicate that glycolipid metabolism and O-GlcNAcylation play a novel role in increasing ferroptosis susceptibility in mesenchymal cancer cells, which reveals a new molecular mechanism of ferroptosis and suggests a therapeutic strategy for refractory pancreatic cancers.

Gynecologic neoplasms: Increased expression of GALNT14 is found in ovarian cancer. Silencing GALNT14 eliminates ovarian cancer cells by promoting apoptosis and ferroptosis by reducing the O-glycosylation of EGFR and promoting its degradation, thereby suppressing mTOR pathway activity.²⁰⁸ Combination treatment with an mTOR inhibitor and cisplatin induced apoptosis and ferroptosis, suggesting that the combination of cisplatin with an mTOR inhibitor might be a promising strategy to combat cisplatin resistance in ovarian cancer.²⁰⁸ Alpha 1,3-mannosyltransferase (ALG3) is involved in protein glycosylation critical for the assembly of lipid-linked oligosaccharides and in N-linked glycosylation of proteins at the luminal side of the endoplasmic reticulum (ER). Inhibition of ALG3 leads to N-linked glycosylation deficiency-mediated ferroptosis to boost the efficacy of anti-PD1 immunotherapy.²⁰⁹ Ablation of ALG3 was found to attenuate tumor growth in a cytotoxic T-cell-dependent manner in mice.²⁰⁹ Moreover, ALG3 inhibition and treatment with tunicamycin (an N-linked glycosylation inhibitor) synergize with anti-PD1 therapy to inhibit tumor growth in mouse models.²⁰⁹ Inhibition of ALG3 induces impairment of posttranslational N-linked glycosylation, resulting in sterol-regulated element-binding protein (SREBP1)-dependent lipogenesis and excessive lipid accumulation, which induces immunogenic ferroptosis in cancer cells and leads to the formation of a proinflammatory microenvironment, thereby boosting antitumor immune responses.

Osteosarcoma: Protein O-GlcNAcylation orchestrates both mitophagy and ferritinophagy to support ferroptosis in osteosarcoma cells.²¹⁰ RSL3 modulates ferroptosis by inducing a biphasic change in protein O-GlcNAcylation. Inhibition of O-GlcNAcylation enhances ferritinophagy, leading to an increased labile iron content, thus rendering the cell more sensitive to ferroptosis. De-O-GlcNAcylation of FTH at S179 facilitates its interaction with NCOA4, resulting in labile iron accumulation to support ferroptosis.²¹⁰ These results reveal links between dynamic O-GlcNAcylation and both iron metabolism and ferroptosis initiation, highlighting a potential therapeutic regimen for cancers.

SUMOylation-mediated regulation of ferroptosis in cancer

A PTM involving conjugation of small ubiquitin-like modifier (SUMO) proteins to substrate proteins, SUMOylation is involved in various cellular processes and is a vital cellular mechanism in stress responses.²¹¹ SUMOylation is also a reversible and dynamic process. SUMOylation occurs through an enzymatic cascade that consists of a dimeric SUMO-activating E1 enzyme (SAE1 and SAE2/UBA2), a single E2 enzyme (ubiquitin-conjugating enzyme 9, UBC9), and a limited set of E3 ligases.²¹² SUMO-specific proteases (SENPs) cooperate with SUMO molecules to modulate the SUMOylation status of a substrate protein by specifically de-SUMOylating the substrate protein. Aberrantly overactivated SUMOylation has been detected in many cancers and is involved in EMT, tumorigenesis, metastasis, drug resistance, and antitumor immunity.^211,213 Emerging studies suggest that SUMOylation regulates ferroptosis in cancer (Table 6 and Fig. 5). Aberrantly increased SENP1 expression predicts a poor prognosis in patients with lung cancer. Inhibition of SENP1 inhibits the proliferation and growth of lung cancer cells. Overexpression of SENP1 inhibits Erastin- or cisplatin-induced ferroptosis. Elevated SENP1 expression mediates A20 SUMOylation to reduce the expression of ACSL4 and induce the expression of SLC7A11, thereby inhibiting ferroptosis in lung cancers.²¹⁴ SENP1 functions as a ferroptosis suppressor, as determined through a novel network analysis of how A20 SUMOylation links SLC7A11 and ACSL4 in lung cancer cells. Inhibition of SENP1 enhances ferroptosis. Elevated HSP27 expression inhibits ferroptosis by inducing SUMOylation of ACSL4 to reduce its stability in GBM cells.²¹⁵

Table 6 Posttranslational modification of Ferroptosis by SUMOylation in cancer

N₆-methyladenosine (m⁶A) modification-mediated regulation of ferroptosis in cancer

Accumulating studies have shown that m⁶A modification plays a vital role in epigenetic regulation in organisms and in the pathogenesis of malignant diseases. Aberrant m⁶A modification, a dynamic and reversible posttranscriptional RNA modification mediated by methyltransferases (writers), demethylases (erasers), and m⁶A binding proteins (readers), is associated with the development, progression, occurrence, and prognosis of cancer. Emerging studies suggest that m⁶A modification regulates ferroptosis in cancer (Table 7 and Fig. 6).

Table 7 Epigenetic modification of Ferroptosis by m⁶A in cancer

Fig. 6

Epigenetic modification of ferroptosis by m⁶A in cancer. ALKBH5 AlkB homolog 5, ACSL3 acyl-CoA synthetase long-chain family member 3, BC breast cancer, CBS cystathionine-beta-synthase, FSP1 ferroptosis suppressor protein 1, FTH1 ferritin heavy chain 1, FTO fat mass and obesity-associated protein, GBM glioblastoma, GC gastric cancer, HCC hepatocellular carcinoma, GPX4 glutathione peroxidase 4, HDLBP high-density lipoprotein-binding protein, HPSCC hypopharyngeal squamous cell carcinoma, IGF2BP1 Insulin-like growth factor 2 mRNA binding protein 1, IGF2BP2 Insulin-like growth factor 2 mRNA binding protein 2, METTL3 methyltransferase-like protein 3, METTL14 methyltransferase -like 14, METTL16 methyltransferase-like 16, NKAP NF-κB activating protein, NPC nasopharyngeal carcinoma, NSCLC non-small cell lung carcinoma, SFPQ splicing factor proline and glutamine-rich, SLC3A2 solute carrier family 3 member 2, YTHDC2 YTH domain containing 2, YTHDF1 YTH N6-methyladenosine RNA binding protein 1, YTHDF2 YTH N6-methyladenosine RNA binding protein 2, YTHDC2 YTH domain containing 2

Non-small cell lung carcinoma: Exosomal miR-4443 promotes cisplatin resistance by enhancing METLL3-mediated m⁶A modification of FSP1, thereby inhibiting ferroptosis.²¹⁶ Downregulation of the m⁶A reader YT521-B homology domain containing 2 (YTHDC2) is related to poor clinical outcomes in patients with LUAD. YTHDC2 was found to inhibit tumorigenesis in a mouse model of spontaneous LUAD. YTHDC2 inhibits cystine uptake and blocks the downstream antioxidant system. Lung tumorigenesis is rescued by supplementation with downstream cystine antioxidants in mice with pulmonary YTHDC2 overexpression. YTHDC2 inhibits LUAD tumorigenesis by suppressing the transcription of SLC7A11 mRNA and promoting its decay in an m⁶A-dependent manner,²¹⁷ suggesting that increased cystine uptake through suppression of YTHDC2 is critical for LUAD tumorigenesis and that blocking this process may be a therapeutic approach. This observation was corroborated by other studies, which showed that YTHDC2 promoted m⁶A methylation and subsequently destabilized HOXA13 mRNA to suppress SLC3A2 expression, thereby inducing ferroptosis and inhibiting LUAD tumorigenesis.²¹⁸ The upregulated reader protein IGF2BP3 recognizes and binds target mRNAs encoding antiferroptotic factors that can undergo METTL3-mediated m⁶A methylation, leading to suppression of ferroptosis and stimulation of tumorigenesis.²¹⁹ The writer METTL3 promotes tumor growth and inhibits ferroptosis by stabilizing the m⁶A modification of SLC7A11 in LUAD.²²⁰

Hepatocellular carcinoma: Elevated expression of HDLBP, the largest RNA-binding protein, inhibits ferroptosis in HCC cells. HDLBP binds to and stabilizes lncFAL. The splicing of lncFAL is enhanced by YTHDF2 in an m⁶A-dependent manner. lncFAL reduces vulnerability to ferroptosis by directly binding to FSP1 and competitively abolishing Trim69-dependent polyubiquitination and degradation of FSP1.¹³³ Upregulation of IGF2BP3 is strongly associated with early recurrence, tumor invasion, and poor prognosis in HCC. Silencing IGF2BP3 significantly promotes sorafenib-induced ferroptosis in HCC cells. Moreover, Nrf2 mRNA was identified as an important target of IGF2BP3, which stabilizes Nrf2 mRNA via m⁶A modification. Upregulation of IGF2BP3 inhibits sorafenib-induced ferroptosis by promoting Nrf2 mRNA stability in an m⁶A-dependent manner,²²¹ suggesting a new anticancer strategy aimed at improving the efficacy of sorafenib by inhibiting IGF2BP3. METTL3-mediated m⁶A modification of SLC7A11 promotes ferroptosis resistance in hepatoblastoma cells. METTL3/IGF2BP1-mediated m⁶A modification stabilizes SLC7A11 mRNA and upregulates SLC7A11 expression by inhibiting the deadenylation process in hepatoblastoma cells.²²²

Thyroid cancer: ALKBH5 inhibits the progression of thyroid cancer by enhancing ferroptosis through Nrf2 inactivation mediated by decreasing the m⁶A level in TIAM1.²²³ FTO inhibits the progression of thyroid cancer by inducing ferroptosis through m⁶A-mediated downregulation of SLC7A11.²²⁴

Breast cancer: METTL16 promotes the progression of breast cancer by inhibiting ferroptosis by epigenetically increasing GPX4 expression via m⁶A modification.²²⁵ The m⁶A modification level is reduced due to downregulation of METTL14 in anti-HER2 therapy-resistant breast cancer. This decrease in the m6A level prevents YTHDC2-mediated degradation of FGFR4 mRNA, thus leading to the accumulation of FGFR4 in resistant breast cancer cells. FGFR4 phosphorylates GSK-3β and activates β-catenin/TCF4 signaling to increase the transcription of the SLC7A11 and FPN1 genes. Upregulation of SLC7A11 and FPN1 accelerates GSH synthesis and Fe²⁺ efflux, which confer anti-HER2 resistance by attenuating ferroptosis in breast cancer cells. Roblitinib, a highly selective inhibitor of FGFR4, combats anti-HER2 resistance by inducing ferroptosis in refractory HER2-positive breast cancer.²²⁶

Other tumors: In GBM, NKAP inhibits ferroptosis by recruiting SFPQ to promote SLC7A11 mRNA splicing in a manner dependent on METTL3-mediated m⁶A modification.²²⁷ Upregulation of FTO promotes radioresistance by repressing ferroptosis through promotion of OTUB1 expression in nasopharyngeal carcinoma (NPC).²²⁸ ALKBH5 inhibits ferroptosis by activating Nrf2 in an m⁶A-IGF2BP2-dependent manner in hypopharyngeal squamous cell carcinoma (HPSCC).²²⁹ HIF-1α mediates lncRNA-CBSLR expression to recruit the YTHDF2 protein and CBS mRNA to form the CBSLR/YTHDF2/CBS complex, resulting in m6A-dependent destabilization of CBS mRNA and leading to decreased methylation of the ACSL4 protein and an increase in its ubiquitin–proteasome-dependent degradation, thereby inhibiting ferroptosis in GC cells.²³⁰

Noncoding RNA-induced modulation of ferroptosis in cancer

ncRNAs, encompassing microRNAs (miRNAs), lncRNAs, and circRNAs, do not encode proteins. However, ncRNAs are considered master regulators of various cellular processes, particularly in cancers, where they are implicated in all hallmarks of cancer. Recent studies have demonstrated that ncRNAs, particularly miRNAs, lncRNAs, and circRNAs, are involved in the biological process of ferroptosis by regulating the molecular mechanism of ferroptosis in tumor cells. We refer readers to some recent excellent reviews for a detailed discussion of the roles of ncRNAs in regulating ferroptosis in cancer.^{231,232,233,234}

Epigenetic and posttranslational modifications regulating ferroptosis in central nervous system (CNS) diseases

Accumulating evidence supports neuronal ferroptosis as a critical factor in traumatic brain injury (TBI), multiple sclerosis (MS), spinal cord injury (SCI), neurodegenerative diseases such as Alzheimer’s disease (AD) and Parkinson’s disease (PD), and stroke, including spontaneous intracerebral hemorrhage (ICH), acute ischemic stroke (AIS), and subarachnoid hemorrhage (SAH); in addition, pharmacological inhibition of ferroptosis is supported as a therapeutic strategy for these diseases.¹⁹ Emerging evidence has revealed the role of epigenetic modifications and PTMs in regulating ferroptosis in CNS diseases. Below, we summarize the epigenetic modifications and PTMs regulating ferroptosis in CNS diseases (Table 8 and Fig. 7).

Table 8 Epigenetic and posttranslational modification of Ferroptosis in neurological diseases

Fig. 7

Epigenetic and posttranslational modification of ferroptosis in CNS disease and CVD. AIS acute ischemic stroke, BAP1 BRCA1-associated protein 1, BMSC-Exos bone marrow mesenchymal stem cells (BMSCs)-derived exosomes, CVD cardiovascular diseases, DIC doxorubicin-induced cardiomyopathy, HFD High-fat diet, Egr-1 transcription factor early growth response-1, FSP1 ferroptosis suppressor protein 1, FTH1 ferritin heavy chain 1, METTL14 methyltransferase-like 14, ME2 malic enzyme 2, MIRI myocardial ischemia/reperfusion injury, MPP⁺ 1-methyl-4-phenylpyridinium, MS multiple sclerosis, MSCs-exo mesenchymal stem cells-derived exosomes, NAMPT nicotinamide phosphoribosyltransferase, NHLRC1 NHL repeat-containing 1, Ptgs2 prostaglandin-endoperoxide synthase-2, TBI traumatic brain injury, SIC sepsis-induced cardiomyopathy, SPATA2 Spermatogenesis-associated protein 2, TfR1 transferrin receptor 1, TMEM43 transmembrane protein 43

Parkinson’s disease

PD is the second most common neurodegenerative disease and is a progressive condition pathologically characterized by the degeneration of nigrostriatal dopaminergic neurons located in the substantia nigra pars compacta (SNpc) of the brainstem and the presence of Lewy bodies, which consist mainly of misfolded α-synuclein, Parkin, ubiquitin, PTEN-induced kinase-1 (PINK1), and other proteins, in the surviving neurons.^{235,236,237,238} PD is characterized by motor and nonmotor symptoms and affects more than 2% of the population above 65 years of age.^237,239 Accumulating evidence indicates that ferroptosis plays a role in the genesis of PD.^{19,87,240,241,242,243,244,245,246,247,248,249,250,251,252,253,254,255,256,257,258,259,260,261,262} However, data revealing the roles of epigenetic regulation of ferroptosis by ncRNAs in PD are limited. Upregulation of the lncRNA NEAT1 was observed in 1-methyl-4-phenylpyridinium (MPP⁺)-treated SK-N-SH cells. Silencing lncRNA NEAT1 was found to increase cell viability through ferroptosis inhibition by sponging and suppressing miR-150-5p.⁴¹ Overexpression of miR-150-5p upregulates SLC7A11 expression by directly binding to BAP1 to suppress ferroptosis. BAP1 overexpression or miR-150-5p inhibition mitigate sh-NEAT1-mediated inhibition of ferroptosis.⁴¹ Together, these observations indicate that upregulation of NEAT1 promotes MPP⁺-induced ferroptosis by regulating the miR-150-5p/BAP1/SLC7A11 pathway in SK-N-SH cells.⁴¹ miR-335 was found to promote ferroptosis through the degradation of FTH1 in vivo and in vitro in 6-hydroxydopamine (6-OHDA)-induced models of PD.²⁶³ Midbrain dopamine oxidation promotes ferroptosis in dopaminergic neurons by facilitating NEDD4-mediated ubiquitination of GPX4.²⁴⁰

Acute ischemic stroke

Caused by arterial occlusion, AIS is the most common type of stroke, represents a significant threat to human life and is among the most frequent causes of disability and death worldwide.^264,265 Emerging evidence has shown that ferroptosis plays a role in the genesis of neuronal injury after ischemic stroke.^{19,266,267,268,269,270,271,272,273,274,275,276,277,278,279,280,281,282,283,284,285,286,287,288,289,290,291,292} Recently, accumulating evidence has revealed the roles of epigenetic regulation of ferroptosis via ncRNA expression, methylation, ubiquitination, and acetylation in AIS.^{293,294,295,296,297,298,299,300,301,302} Upregulation of the circRNA Carm1 inhibits ferroptosis through its binding to microRNA-3098-3p to downregulate ACSL4 in OGD/R-treated HT22 cells.²⁹³ Upregulation of LncRNA PVT1 promotes ferroptosis through miR-214-mediated suppression of TFR1 and P53 expression.²⁹⁴ miR-27a promotes ferroptosis by inhibiting Nrf2 expression during ischemic stroke.²⁹⁵ miR-27a aggravates cerebral ischemia/reperfusion injury (IRI) by promoting ferroptosis through inhibition of SLC7A11.²⁹⁶ miR-760-3p in exosomes from adipose-derived stem cells (ADSC-Exos) inhibits ferroptosis by targeting CHAC1 in neurons.²⁹⁷ GATA6 suppresses neuronal autophagy and ferroptosis through a miR-193b/ATG7 axis-dependent mechanism.²⁹⁸ LncRNA Meg3 promotes oxygen-glucose deprivation (OGD)-mediated hyperglycemic reperfusion-induced ferroptosis by upregulating p53, thereby inhibiting GPX4 expression in rat brain microvascular endothelial cells (RBMVECs).²⁹⁹ ELAVL1 suppresses ferroptosis-induced cerebral IRI and subsequent brain damage by inhibiting PINK1 expression through stabilization of DNMT3B mRNA.³⁰⁰ USP14 stabilizes NCOA4 to promote ferritinophagy-mediated ferroptosis in ischemic stroke.³⁰¹ HDAC9 promotes neuronal ferroptosis through deacetylation and deubiquitination, dependently increasing HIF-1 expression and thus increasing the transcription of TfR1 and decreasing the Sp1 protein level by deacetylation and ubiquitination, leading to downregulation of GPX4 in in vitro and in vivo models of stroke.³⁰²

Intracerebral hemorrhage

ICHs are defined by brain injury resulting from acute extravasation of blood into the brain parenchyma from a ruptured cerebral blood vessel.³⁰³ Occurring spontaneously and accounting for 80% of hemorrhagic strokes and 10-15% of all strokes,³⁰³ ICH is an acute subtype of cerebral stroke. Emerging evidence has shown that ferroptosis plays a role in the genesis of secondary brain injury after ICH (SBI-ICH).^{19,304,305,306,307,308,309,310,311,312,313,314,315,316,317,318,319,320,321,322,323,324,325,326,327,328} Data revealing the roles of epigenetic regulation of ferroptosis by ncRNAs in ICH are limited. LncRNA H19 protects against ICH-related injuries by regulating the microRNA-106b-5p/ACSL4 axis.³²⁹ miR-137 inhibits ferroptosis in oxyhemoglobin (oxyHb)-treated SH-SY5Y cells via the COX2/PGE2 pathway.³³⁰ Downregulation of miR-124 induces FPN expression and attenuates iron accumulation.³³¹ Acupuncture may alleviate neuronal cell death, inflammation, and ferroptosis after ICH by downregulating miR-23a-3p.³³²

Traumatic brain injury

Structural and physiological disruption of brain function caused by external forces results in TBI, which is a major cause of disability and death in patients worldwide and a key injury mechanism involving the death of nerve cells.^{333,334,335,336,337} Emerging evidence indicates that ferroptosis plays a vital role in secondary brain injury and worsens long-term outcomes after TBI.^{338,339,340,341,342,343,344,345,346,347,348,349,350,351,352,353} Emerging studies have revealed the roles of epigenetic regulation of ferroptosis by ncRNAs in TBI.³⁵⁴ Upregulated circPtpn14 (mmu_circ_0000130) promotes ferroptosis by sponging miR-351-5p, which targets the 5-LOX mRNA for degradation, thereby upregulating the expression of 5-LOX after TBI. Decreased miR-212-5p expression has been found in TBI. Overexpression of miR-212-5p attenuates cell death by inhibiting ferroptosis through repression of Ptgs2 after TBI.³⁵⁴

Spinal cord injury

SCI is a severely disabling neurological condition causing primary damage to the spinal cord via compression and laceration, followed by secondary damage consisting of inflammation and ischemia, culminating in substantial loss of tissue.^{355,356,357,358,359} Ferroptosis plays a key role in secondary SCI, and this role is closely related to inflammation, immunity, and chronic injuries.^{339,340,360,361,362} Emerging studies have revealed the roles of epigenetic regulation of ferroptosis by ncRNAs and ubiquitination in SCI.³⁵⁴ miR-672-3p enhances functional recovery by inhibiting ferroptosis via upregulation of FSP1 in contusive SCI.³⁶³Mesenchymal stem cell (MSC) exosomal lncGm36569 attenuates neuronal dysfunction through ferroptosis inhibition by sponging miR-5627-5p to induce FSP1 upregulation.³⁶⁴ Silencing miR-6315 attenuates neuronal dysfunction by inhibiting ferroptosis via upregulation of GPX4.³⁶⁵ Downregulated USP7 expression and upregulated HMOX-1 expression were found in SCI rat models. USP7 overexpression alleviates SCI through deubiquitination of HMOX-1 and promotion of its expression, thereby reducing ferroptosis.³⁶⁶

Multiple sclerosis

A CNS disorder characterized by inflammation, demyelination, gliosis and neuroaxonal degeneration, MS is highly heterogeneous and affects over 2.8 million people worldwide.^{367,368,369,370,371} Recently, ferroptosis was shown to play a key role in the genesis of MS, and inhibition of ferroptosis was found to attenuate disease progression in an experimental autoimmune encephalitis (EAE) mouse model.^{372,373,374,375,376,377} Emerging studies have revealed the roles of epigenetic regulation of ferroptosis by ncRNAs and methylation in MS. The histone methyltransferase G9a promotes neurodegeneration by inducing ferroptosis by catalyzing the writing of the repressive mark H3K9me2, thereby suppressing the expression of GCLC, CBS, and GPX4.³⁷⁸ In addition, bone marrow mesenchymal stem cell-derived exosomes (BMSC-Exos) containing miR-367-3p were found to attenuate the severity of EAE by inhibiting ferroptosis through repression of EZH2 expression, leading to overexpression of SLC7A11.³⁷²

Epigenetic and posttranslational modifications regulating ferroptosis in cardiovascular diseases

Accumulating evidence supports ferroptosis as a critical factor in CVDs, including myocardial IRI (MIRI), doxorubicin (DOX)-induced cardiomyopathy (DIC), cardiac fibrosis (CF), heart failure, high-fat diet (HFD)-induced cardiac injury, cardiac hypertrophy, aortic dissection (AD), and septic cardiomyopathy (SCM). Below, we summarize the epigenetic regulation of ferroptosis in CVDs (Table 9 and Fig. 7).

Table 9 Epigenetic and posttranslational modification of ferroptosis in cardiovascular diseases

Myocardial ischemia/reperfusion injury

Myocardial reperfusion strategies and reoxygenation are the preferred and effective treatments for acute myocardial infarction (AMI).^379,380 However, reperfusion inevitably causes cardiomyocyte death, increases the infarct size, and aggravates the condition; these events are collectively referred to as MIRI.³⁸¹ MIRI leads to energy metabolism disruption and oxidative stress, among other issues.³⁸² A recent novel study revealed a role of ferroptosis in MIRI in murine models, establishing a correlation between cardiac cell death and ferroptosis in vivo.^383,384 Thereafter, emerging studies have attempted to elucidate the pathophysiological role of ferroptosis in the genesis of MIRI.^{92,382,383,385,386,387,388,389,390,391,392,393,394,395} There are also emerging studies that have revealed the roles of epigenetic regulation of ferroptosis by ncRNAs, ubiquitination, and phosphorylation in MIRI^{385,396,397,398,399,400,401,402,403,404,405} (Table 9). CircRNA FEACR alleviates MIRI through inhibition of ferroptosis by interacting with nicotinamide phosphoribosyltransferase (NAMPT), which upregulates NAMPT-dependent sirtuin1 (Sirt1) expression, thereby reducing the forkhead box protein O1 (FOXO1) acetylation level to promote its transcriptional activity, eventually upregulating the transcription of FTH1.³⁹⁶ Upregulation of LncRNA SEMA5A-IT1 protects cardiomyocytes against hypoxia/reoxygenation (H/R) injury partially through inhibition of ferroptosis by sponging miR-143-3p to upregulate SLC7A11.³⁹⁷ miR-210-3p attenuates myocardial cell injury induced by H/R through inhibition of ferroptosis by downregulating TfR1.³⁹⁸ miR-190a-5p promotes ferroptosis through inhibition of GLS2 expression.³⁹⁹ MiR-199a-5p promotes cardiomyocyte death in OGD/reperfusion (OGD/R)-treated H9c2 cells by inducing ferroptosis via downregulation of GPX4 through inhibition of the Akt/eNOS signaling pathway.⁴⁰⁰ Upregulated expression of miR-214-3p promotes MIRI by inducing ferroptosis through suppression of ME2.⁴⁰¹ Egr-1-mediated upregulation of miR-15a-5p promotes MIRI by inducing ferroptosis through suppression of GPX4.⁴⁰² Upregulated expression of frataxin alleviates cardiomyocyte ferroptosis. NHL repeat-containing 1 (NHLRC1) is an E3 ligase that mediates ubiquitin-mediated degradation of frataxin.⁴⁰³ USP7 promotes MIRI by inducing ferroptosis via activation of the p53/TfR1 pathway.³⁸⁵ USP22 alleviates MIRI by inhibiting ferroptosis via the SIRT1-p53/SLC7A11 axis.⁴⁰⁴ The expression of SIRT1, USP22, and SLC7A11 is inhibited by IRI, whereas increased expression of p53 is found in affected myocardial tissues. Conversely, overexpression of USP22, SIRT1, or SLC7A11 inhibits IRI and increases the viability of cardiomyocytes by inhibiting ferroptosis.⁴⁰⁴ Upregulated expression of HIP-55 alleviates cardiomyocyte ferroptosis and MI injury.⁴⁰⁵ HIP-55 is phosphorylated by AKT at S269/T291, leading to negatively regulation of MAP4K1 by AKT to attenuate MI injury in a site-specific manner, as evidenced by the finding that S269A/T291A-mutated HIP-55 (HIP-55AA) decreased the interaction between HIP-55 and MAP4K1, resulting in loss of ferroptosis inhibition.⁴⁰⁵

Doxorubicin-induced cardiomyopathy

Anthracycline-based chemotherapy can lead to the progressive development of cardiomyopathy. Although DOX is an effective chemotherapeutic agent prescribed to treat breast, ovarian, and gastrointestinal cancers, it causes cardiotoxicity, resulting in DCM, which leads to congestive heart failure.^25,406 The exact mechanisms of DOX-induced cardiotoxicity remain poorly understood. However, emerging studies have attempted to elucidate the pathophysiological role of ferroptosis in the genesis of DCM.^{25,383,407,408,409} In addition, emerging studies have revealed the roles of epigenetic regulation of ferroptosis by m⁶A modification, other methylation modifications, phosphorylation, and ubiquitination in DCM^{410,411,412,413,414,415} (Table 9). METTL14 promotes ferroptosis in DOX-induced cardiomyocytes by mediating IGF2BP1 m6A-dependent stabilization of KCNQ1OT1 to sponge miR-7-5p, thereby increasing the level of the transferrin receptor.⁴¹⁰ Upregulated expression of PRMT4 aggravates DIC by promoting ferroptosis via an interaction with Nrf2 to promote its enzymatic methylation, thereby suppressing GPX4 expression.⁴¹¹ Activation of AMPK α2 attenuates DOX-induced cardiotoxicity by inhibiting ferroptosis.⁴¹² TRIM21 ablation protects against DIC through inhibition of ferroptosis by enhancing sequestration of Keap1 by P62.⁴¹³ MITOL knockdown reduced susceptibility to DOX in cultured cardiomyocytes stressed with DOX.⁴¹⁴ Overexpressed SPATA2 recruits CYLD to promote ferritinophagy by decreasing NCOA4 ubiquitination and ferritin expression and promote ferroptosis by increasing ACSL4 expression.⁴¹⁵

Cardiac fibrosis

Knockout of SIRT3 results in increased p53 acetylation and ferroptosis through downregulation of GPX4 in mouse hearts. p53 acetylation-induced ferroptosis is partially involved in SIRT3-mediated CF in myofibroblasts.⁴¹⁶ MiR-375-3p promotes CF by accelerating ferroptosis in cardiomyocytes by downregulating GPX4.⁴¹⁷

Heart failure

circSnx12 functions as an endogenous sponge to bind miR-224-5p, and FTH1 has miRNA binding sites in its 3’UTR.⁴¹⁸ Downregulated expression of PGAM5 promotes ferroptosis in models of heart failure by decreasing Keap1 ubiquitination, thereby reducing the activation of Nrf2.⁴¹⁹

HFD-induced cardiac injury

The IL-6/STAT3 axis exacerbates cardiac injury by increasing NCOA4-mediated ferritinophagy and ferroptosis in HFD-fed mice.⁴²⁰

Cardiac hypertrophy

CD147 promotes cardiac remodeling and dysfunction by promoting ferroptosis in a glycosylation-dependent manner by binding to TRAF2 to activate the downstream TRAF2-TAK1 axis.⁴²¹

Aortic dissection

Upregulated expression of METTL3 facilitates ferroptosis in human aortic smooth muscle cells by promoting the degradation of SLC7A11 and FSP1 mRNA.⁴²²

Septic cardiomyopathy

Seventy percent of patients with sepsis have SCM, which is the leading cause of sepsis-related mortality and morbidity.^423,424 Ferroptosis was recently found to play a key role in the genesis of SCM, and inhibition of ferroptosis attenuates SCM progression.^{425,426,427,428,429} Thus, ferroptosis is involved in SCM. Emerging studies have revealed the roles of epigenetic regulation of ferroptosis in SCM^{410,411,412,413,414,415} (Table 9). TMEM43 knockdown promotes lipopolysaccharide (LPS)-induced cardiac injury and dysfunction in mice by inducing ferroptosis by increasing the levels of P53 and ferritin and decreasing the levels of GPX4 and SLC7A11.⁴²⁵

Epigenetic and posttranslational modifications regulating ferroptosis in liver diseases

Accumulating studies have shown that ferroptosis plays a role in the genesis of liver diseases,^29,30 and pharmacological induction and inhibition of ferroptosis show significant potential utility for the treatment of hepatic disorders.^{430,431,432,433,434} Accumulating evidence indicates epigenetic modifications regulate ferroptosis in liver diseases, including acute liver injury (ALI), nonalcoholic fatty liver disease (NAFLD), liver fibrosis, hepatic ischemia/reperfusion injury, and toxin-mediated hepatic toxicity. In this section, we summarize the epigenetic regulation of ferroptosis by ubiquitination, phosphorylation, and acetylation in liver diseases (Table 10 and Fig. 8).

Table 10 Epigenetic and posttranslational modification of ferroptosis in liver diseases

Fig. 8

Epigenetic and posttranslational modification of ferroptosis in Liver diseases and Lung diseases. AUF1 AU-rich element (ARE)-binding factor 1, BRD7 bromodomain-containing protein 7, COPD chronic obstructive pulmonary disease, GCN5L1 mitochondrial general control of amino acid synthesis 5 like 1, IRI ischemia/reperfusion injury, LOXs lipoxygenases, NETs neutrophil extracellular traps, Ptgs2 prostaglandin-endoperoxide synthase 2, STAT3 signal transducer and activator of transcription 3, TfR1 transferrin receptor 1, TMEM16A transmembrane member 16A, TMEM16A transmembrane member 16A

Acute liver injury

ALI is the primary cause of liver diseases and is associated with high morbidity and mortality, causing 3.5% of deaths worldwide.⁴³⁵ Various hepatotoxic factors, including lipid deposition, viruses, and drugs, can induce ALI. Emerging studies have suggested that ferroptosis plays a role in the genesis of ALI^{436,437,438,439,440,441,442,443,444} and have revealed the roles of epigenetic regulation of ferroptosis by ubiquitination in ALI^{410,411,412,413,414,415} (Table 10). Mesenchymal stem cells (MSCs) and MSC-derived exosomes (MSC-Exos) alleviate injury through ferroptosis inhibition by decreasing the mRNA levels of Ptgs2 and LOXs while increasing the protein level of SLC7A11 in CCl₄-induced ALI. MSC-Exo-induced expression of the SLC7A11 protein was found to be accompanied by increases in CD44 and OTUB1 expression. OTUB1-mediated deubiquitination rescued CCl₄-triggered ubiquitination of SLC7A11, thus stabilizing SLC7A11 to activate system X_C⁻ and thereby preventing hepatocyte ferroptosis.⁴⁴⁵ The overexpressed ubiquitin E3 ligase HUWE1 inhibits ferroptosis by targeting TfR1 for ubiquitination and proteasomal degradation in the context of CCl₄-induced liver injury, suggesting that HUWE1 functions as an inhibitor to mitigate ALI by antagonizing both aberrant iron accumulation and ferroptosis.⁴⁴⁶

Liver fibrosis

Liver fibrosis is a pathological state and abnormal repair response to chronic liver injury that is characterized by diffuse and progressive excessive deposition of extracellular matrix (ECM) in the liver. Liver fibrosis typically results from chronic liver damage resulting from infection, toxin or drug exposure, NAFLD, cholestasis, or autoimmune insults.^447,448 Liver fibrosis has become one of the leading causes of liver disease globally and often progresses to liver cirrhosis and HCC. Accumulating evidence has shown that ferroptosis is involved in the pathogenesis of liver fibrosis.^{449,450,451,452,453,454,455,456,457} Recent studies have also shown the roles of epigenetic regulation of ferroptosis by phosphorylation⁴⁵⁸ and ubiquitination^459,460,461 in liver fibrosis. Upregulated expression of ELAVL1 induces liver fibrosis by promoting ferritinophagy in hepatic stellate cells (HSCs).⁴⁵⁹ Downregulation of ZFP36 mediated by the ubiquitin ligase FBXW7/CDC4 promotes activation of ferritinophagy and induction of ferroptosis in human HSCs.⁴⁵⁸ BRD7 upregulation induced by xCT inhibition, GPX4 inhibition, and GSH depletion triggers mitochondrial translocation of p53 via direct binding to the N-terminal transactivation domain, thus increasing mitochondrial iron and LPO and eventually causing ferroptosis.⁴⁶⁰ TRIM26 mediates the ubiquitination of SLC7A11 to promote ferroptosis in HSCs, thereby suppressing liver fibrosis.⁴⁶¹

Nonalcoholic fatty liver disease

GCN5L1 expression has been found to be increased in human patients and mice with nonalcoholic steatohepatitis (NASH). GCN5L1 acetylates CypD to enhance its binding to ATP5B, resulting in the release of mitochondrial ROS (mtROS) into the cytoplasm, which promotes ferroptosis in hepatocytes and induces the accumulation of high mobility group box 1 in the microenvironment, thereby inducing the formation of neutrophil extracellular traps (NETs).⁴⁶²

Hepatic ischemia/reperfusion injury

TMEM16A ubiquitinates and degrades GPX4 to induce ferroptosis. TMEM16A deficiency alleviates hepatic IRI by suppressing GPX4-mediated ferroptosis.⁴⁶³

Toxin-mediated hepatic toxicity

Ethyl carbamate induces ferroptosis by inhibiting the activation of Nrf2 through suppression of its phosphorylation and nuclear translocation, thereby inhibiting SLC7A11 expression and leading to GSH depletion.⁴⁶⁴ Glyphosate triggers ferroptosis in hepatocytes by suppressing Nrf2 via blockade of Nrf2 phosphorylation and nuclear translocation, resulting in GSH depletion and inhibition of GPX4.⁴⁶⁵

Epigenetic and posttranslational modifications regulating ferroptosis in lung diseases

Accumulating studies have shown epigenetic regulation of ferroptosis in the genesis of lung diseases, including chronic obstructive pulmonary disease (COPD), acute lung injury, pulmonary fibrosis, and sepsis-induced acute lung injury. In this section, we summarize the epigenetic regulation of ferroptosis in these lung diseases (Table 11 and Fig. 8).

Table 11 Epigenetic and posttranslational modification of ferroptosis in lung diseases

Acute lung injury

A common and critical illness caused by both pulmonary and extrapulmonary factors, acute lung injury (ALI) and its most severe form, acute respiratory distress syndrome (ARDS), result in high morbidity and mortality and have no effective treatments.^466,467 Accumulating evidence has revealed that ferroptosis is involved in the pathogenesis of ALI.^42,468 Recent studies have also shown the roles of epigenetic regulation of ferroptosis by phosphorylation^469,470 and acetylation^471,472 in acute lung injury. Signal transducer and activator of transcription 6 (STAT6) alleviates acute lung injury through ferroptosis inhibition by competitively binding the critical acetyltransferase for p53 acetylation, i.e., CREB-binding protein, to suppress p53 acetylation and transcriptionally restore the expression of SLC7A11.⁴⁷¹ Decreased SIRT1 expression triggers heat stress-induced lung epithelial cell injury through ferroptosis induction by increasing the acetylation of P53, which transcriptionally inhibits SLC7A11.⁴⁷² mtROS-initiated dysfunction of mitochondria-associated endoplasmic reticulum membranes (MAMs) is partially implicated in arsenic-induced ferroptosis and ALI.⁴⁶⁹ Nrf2 was found to activate and cooperate with STAT3 to upregulate SLC7A11 to inhibit ferroptosis in an intestinal ischemia/reperfusion-induced acute lung injury (IIR-ALI) model.⁴⁷⁰

Sepsis-associated acute lung injury (SALI)

Accumulating evidence has demonstrated that ferroptosis is involved in the pathogenesis of SALI.^{43,473,474,475,476,477,478,479} Recent studies have also shown the roles of epigenetic regulation of ferroptosis by phosphorylation and acetylation in SALI. NETs induce ferroptosis through METTL3-induced m⁶A modification of GPX4 in the pathogenesis of sepsis-associated ALI.⁴⁷³ miR-125b-5p in ADSC-Exos alleviates sepsis-induced ferroptosis in mouse pulmonary microvascular endothelial cells (MPVECs) by upregulating Keap1/Nrf2/GPX4 expression, hence ameliorating sepsis-induced acute lung injury.⁴⁸⁰ Inhibition of MUC1 aggravates lung injury by triggering ferroptosis via a reduction in the phosphorylation level of GSK3β and an increase in Keap1 expression, thereby inhibiting the activation of Nrf2 to decrease GPX4 expression.⁴⁷⁴ FBXW7 mediates the protein degradation of AU-rich element (ARE)-binding factor 1 (AUF1). AUF1 alleviates sepsis-induced acute lung injury through ferroptosis inhibition by upregulating Nrf2 and downregulating ATF3.⁴⁷⁵ ncRNAs are also involved in the modulation of SALI by regulating ferroptosis. Septic conditions were found to upregulate the expression of circEXOC5 in both in vivo and in vitro sepsis models. CircEXOC5 exacerbates SALI through ferroptosis induction by stabilizing ACSL4 mRNA via PTBP1 binding.⁴⁷⁹ Silencing circEXOC5 inhibits lung injury by alleviating ferroptosis, as evidenced by upregulated GPX4 protein expression, decreased ROS levels, and decreased ACSL4 expression.⁴⁷⁹

Chronic obstructive pulmonary disease

NF-κB/RelA-mediated PRMT7 upregulation induces monomethylation of histones at enhancers and can regulate Rap1a expression, which is crucial for MAPK signaling downstream of G-protein coupled receptor activation, integrin activation, and the consequent adhesion and migration abilities of monocytes. ALOX5-mediated release of LTB4 in inflammatory macrophages upregulates ACSL4 in AT2 cells, thereby promoting tissue injury through increasing susceptibility to cigarette smoke-induced ferroptosis.⁴⁸¹ Nrf2 promoter hypermethylation-induced inhibition of Nrf2 expression induces ferroptosis by inhibiting GPX4 in human bronchial epithelial (HBE) cells challenged with cigarette smoke extract.⁴⁸² Cigarette smoke-induced USP14 downregulation aggravates bronchial epithelial cell ferroptosis through proteasomal degradation of MFG-E8.⁴⁸³

Epigenetic and posttranslational modifications regulating ferroptosis in kidney diseases

Accumulating studies have shown that ferroptosis plays a role in the genesis of kidney diseases.³⁵ Accumulating evidence has shown epigenetic regulation of ferroptosis in kidney diseases, including acute kidney injury (AKI),^484,485 diabetic nephropathy (DN),⁴⁸⁶ renal fibrosis,⁴⁸⁷ renal IRI (RIRI),^{488,489,490,491,492} sepsis-induced acute kidney injury (SAKI),⁴⁹³ toxin-mediated kidney toxicity,⁴⁹⁴ and crystal nephropathies.⁴⁹⁵ Below, we summarize the epigenetic regulation of ferroptosis by phosphorylation, ncRNAs, deacetylation, and ubiquitination in kidney diseases (Table 12 and Fig. 9).

Table 12 Epigenetic and posttranslational modification of ferroptosis in kidney diseases

Fig. 9

Epigenetic and posttranslational modification of ferroptosis in Kidney disease. AKI Acute kidney injury, ALK activin receptor-like kinase, CHAC1 Cation transport regulator-like protein 1, DNMT1 DNA methyltransferase 1, FMR1 FMRP translational regulator 1, IRI ischemia/reperfusion injury, SAKI sepsis-induced acute kidney injury, TBK1 TANK-binding kinase 1, TIGAR TP53-induced glycolysis and apoptosis regulator

Acute kidney injury

AKI is caused by sudden loss of excretory kidney function, leading to increased serum creatinine, decreased urine output, or both.^496,497 The various etiologies of AKI include kidney ischemia, exposure to nephrotoxins, dehydration and sepsis.⁴⁹⁸ AKI develops in ~10–15% of inpatients. Kidney damage or dysfunction can occur over a longer period or follow AKI along a continuum with acute and chronic kidney disease.⁴⁹⁶ Ferroptosis is closely associated with the development of AKI.^{44,45,499,500,501} Accumulating evidence has revealed epigenetic regulation of ferroptosis by phosphorylation and ncRNAs in AKI. Translocation of P66Shc to mitochondria promotes cisplatin-induced AKI by promoting ferroptosis.⁴⁸⁴ MicroRNA-214-3p aggravates cisplatin-induced AKI through ferroptosis induction by targeting GPX4.⁴⁸⁵

Diabetic nephropathy

DN, also called diabetic kidney disease (DKD), results from microvascular damage sustained as a result of diabetes, leading to proximal tubule injury.⁵⁰² DKD is the leading cause of kidney failure worldwide, affecting approximately half of patients with type 2 diabetes and one-third of patients with type 1 diabetes. The characteristics of DKD are the accumulation of ECM, hypertrophy and fibrosis in kidney glomerular and tubular cells.⁵⁰³

Increasing evidence has shown that ferroptosis plays a vital role in the genesis of DKD.^{504,505,506,507,508,509} A recent study also showed the role of epigenetic regulation of ferroptosis by ncRNAs in DKD. Downregulated expression of mmu_circRNA_0000309 results in a decrease in its competitive sponging of miR-188-3p, subsequently promoting GPX4 expression, thereby blocking ferroptosis-dependent mitochondrial damage and podocyte apoptosis. In addition, overexpression of GPX4 counteracts mmu_circRNA_0000309 silencing-mediated ferroptosis in germacrone-exposed MPC5 cells.⁴⁸⁶

Renal fibrosis

Renal fibrosis is a common final outcome of a wide variety of chronic kidney disease (CKD) and is characterized by excessive deposition of ECM that leads to tissue scarring.⁵¹⁰ Evidence has shown that ferroptosis is involved in the pathogenesis of kidney fibrosis.^{432,487,511,512,513,514,515,516} A recent study also showed the role of epigenetic regulation of ferroptosis by deacetylation in kidney fibrosis. Sirtuin 1-mediated p53 deacetylation was found to attenuate calcium oxalate deposition-induced renal fibrosis by suppressing ferroptosis.⁴⁸⁷

Renal ischemia/reperfusion injury

RIRI is the main cause of AKI and contributes to rapid renal dysfunction, morbidity and mortality in patients with a wide range of injuries.⁵¹⁷ Evidence has demonstrated that ferroptosis is involved in the pathogenesis of RIRI.^{518,519,520,521,522,523} Increasing evidence shows the roles of epigenetic regulation of ferroptosis by ncRNAs and ubiquitination in RIRI. Upregulated expression of miR-20a-5p attenuates IRI and postischemic renal fibrosis through ferroptosis inhibition by repressing ACSL4.⁴⁸⁸ Upregulated expression of miR-182-5p and miR-378a-3p leads to ferroptosis in the context of renal injury by downregulating GPX4 and SLC7A11, respectively, in the rat kidneys subjected to ischemia/reperfusion.⁴⁸⁹ Increased USP14 expression has been found in H/R-exposed HK-2 cells and in kidney tissues of mice subjected to I/R. Inhibiting USP14 suppresses H/R-induced ferroptosis in HK-2 cells.⁴⁹⁰ TRIM21 aggravates ischemia/reperfusion-induced AKI by promoting ferroptosis via ubiquitination of GPX4.⁴⁹¹ Inhibiting USP7 attenuates RIRI through ferroptosis inhibition by decreasing the ubiquitination of TBK1 and promoting DNMT1-mediated methylation of FMR1.⁴⁹²

Sepsis-induced acute kidney injury

SAKI is a common critical illness characterized by rapid sepsis-associated deterioration of renal function; it is one of the leading causes of death worldwide and is a common and life-threatening complication in hospitalized and critically ill patients.^{524,525,526,527} SAKI leads to a greatly increased risk of CKD, cardiovascular events and death. Novel findings suggest that ferroptosis plays a pathological role in SAKI.^528,529,530 Recent evidence has shown the role of epigenetic regulation of ferroptosis by ncRNAs in SAKI, demonstrating that LPS increases the phosphatidylcholine content and the activity of LPCAT3 and decreases the expression of miR-124-3p.1 in human renal tubular epithelial cells in a SAKI model.⁵³¹ miR-124-3p.1 overexpression in turn suppresses cell injury by inhibiting LPCAT3-related ferroptosis, suggesting that miR-124-3p.1 is a ferroptosis inhibitor.⁵³¹

Modulation of epigenetic and posttranslational modifications regulating ferroptosis for disease therapy

Modulation of epigenetic modifications and PTMs regulating ferroptosis for cancer therapy

Epigenetic modifications modulate multiple aspects of cancer biology, and during the last decade, this observation has increased the interest in developing epigenetic strategies to impact tumor growth and drug resistance, thereby combating cancers.^121,122,532 Unlike genetic mutations, dysregulated epigenetic mechanisms and PTMs can feasibly be targeted by small molecule compounds to treat cancers (Table 13).

Table 13 Small molecule compounds target dysregulated epigenetic and posttranslational mechanisms to induce ferroptosis in cancer

Hepatocellular carcinoma

Tiliroside, a potent TBK1 inhibitor, increases the sensitivity of HCC to sorafenib by targeting TBK1 to induce ferroptosis by decreasing the phosphorylation of p62 and the affinity of p62 for Keap1 and promoting Keap1-mediated Nrf2 ubiquitination and degradation in HCC.⁵³³ Corosolic acid can increase the sensitivity of HCC to ferroptosis by inhibiting GSH synthesis via HERPUD1, which decreases MDM2 ubiquitination and promotes the ubiquitination of GSS, thereby inhibiting GSH synthesis to enhance ferroptosis in HCC.⁵³⁴ DHA triggers ferroptosis by enhancing the formation of the PEBP1/15-LO complex and promoting cell membrane LPO. DHA promotes PEBP1 protein expression through inhibition of its ubiquitination and degradation.⁵³⁵ Polyphyllin VI induces ferroptosis by inhibiting STAT3 phosphorylation, which inhibits GPX4 expression in HCC.⁵³⁶ Anisomycin activates p38MAPK to induce ferritinophagy through the phosphorylation of histone H3 on serine 10 (p-H3S10) in HCC.⁵³⁷

Non-small cell lung carcinoma

Bufotalin induces ferroptosis by facilitating the ubiquitination and degradation of GPX4 in NSCLC.⁵³⁸ Sanguinarine triggers ferroptosis by destabilizing GPX4 by promoting its ubiquitination and degradation mediated by its E3 ligase STUB1 in NSCLC.⁵³⁹ Selenite induces ferroptosis by activating the p38-ATF4-DDIT3 axis. Selenite also alters the cellular DNA methylation machinery by upregulating TET1 and downregulating DNMT1. Low-dose selenite treatment enhances the antitumor activity of osimertinib in EGFR-mutant H1975 NSCLC cells and of adagrasib in KRAS-mutant H358 NSCLC cells.⁵⁴⁰

Breast cancer

Eupaformosanin significantly decreases the viability of TNBC cells by inducing ferroptosis via ubiquitination of mutant p53.⁵⁴¹ DMOCPTL induces ferroptosis through ubiquitination of GPX4 in breast cancer cells.⁵⁴² NN3 kills p53-positive breast cancer cells by promoting ferroptosis through downregulation of the SLC7A11 pathway by triggering the ubiquitination and proteasome-dependent degradation of PARP1.⁵⁴³ Ketamine induces ferroptosis through GPX4 repression by decreasing the occupancy of KAT5 in its promoter region, thus inhibiting the enrichment of H3K27ac and RNA pol II, in breast cancer cells.⁵⁴⁴

Glioblastoma/glioma

ALZ003 induces FBXL2-mediated AR ubiquitination and degradation. ALZ003 significantly inhibits glioblastoma cell survival by inducing ferroptosis.⁵⁴⁵ Paeoniflorin inhibits glioma growth by inducing ferroptosis through upregulation of NEDD4L to mediate STAT3 ubiquitination-mediated repression of Nrf2 and GPX4 in glioma.⁵⁴⁶ Myrislignan induces ferroptosis through the Slug-SLC7A11 axis by inactivating NF-κB signaling through blockade of p65 phosphorylation in GBM cells.⁵⁴⁷

Pancreatic cancer

QD394 induces ferroptosis by inhibiting STAT3 phosphorylation, thereby inhibiting GPX4 expression in pancreatic cancer.⁵⁴⁸

Esophageal squamous cell carcinoma (ESCC)

Allicin may induce ferritinophagy by increasing AMPK phosphorylation and decreasing mTOR in ESCC.⁵⁴⁹

Ovarian cancer

Eriodictyol induces ferroptosis by downregulating Nrf2 phosphorylation, thereby decreasing the protein levels of SLC7A11 and GPX4 in ovarian cancer.⁵⁵⁰

Thyroid cancer

RSL3 activates ferroptosis by suppressing mTOR-triggered autophagy. Inhibiting GPX4 mirrored the effect of RSL3 on mTOR pathway suppression in thyroid cancer.⁵⁵¹

Modulation of epigenetic and posttranslational modifications regulating ferroptosis for CNS disease therapy

A therapeutic regimen for pharmacologically inhibiting ferroptosis to treat disease has been convincingly established in CNS diseases, especially in neurodegenerative diseases and stroke.¹⁹ Small molecule compounds also feasibly target dysregulated epigenetic mechanisms and PTMs in CNS diseases (Table 14).

Table 14 Targeting dysregulated epigenetic and posttranslational modification by small molecule compounds to treat CNS, cardiovascular, liver, lung, and kidney diseases

Intracerebral hemorrhage

Paeonol, a naturally occurring phenolic agent extracted from Cortex Moutan, exerts neuroprotective effects in patients with many CNS diseases.^{552,553,554,555,556,557,558,559,560,561,562,563,564,565,566,567,568,569,570,571,572,573} Paeonol inhibits ferroptosis in neurons in ICH by downregulating HOTAIR-dependent UPF1-mediated degradation of ACSL4.³¹² Isorhynchophylline (IRN), a tetracyclic oxindole alkaloid extracted from Uncaria rhynchophylla, has diverse biological activities, such as antioxidant, neuroprotective and anti-inflammatory activities.^{574,575,576,577,578,579,580,581,582,583} Isorhynchophylline attenuates cell damage by inhibiting ferroptosis via upregulation of miR-122-5p and SLC7A11 mRNA expression and inhibiting p53 expression in ferric ammonium citrate (FAC)-treated hippocampal HT-22 cells. The protective effects of IRN against FAC-induced ferroptosis were weakened by miR-122-5p knockdown.³¹³

Parkinson’s disease

(-)Clausenamide (Clau), a scavenger of lipid peroxidation products, is an alkaloid that is isolated from the plant Clausena lansium (Lour.) and exhibits neuroprotective activities both in vivo and in vitro.^{574,575,576,577,578,579,580,581,582,583} Clau prevents the nuclear translocation of ALOX5 by directly interacting with Ser663 of ALOX5, the PKCα phosphorylation site.²⁴⁰

Traumatic brain injury

Melatonin, an indole hormone that contributes to neuroprotection, exerts a neuroprotective effect by inhibiting ferroptosis.^{584,585,586,587} A recent study showed that melatonin ameliorates neurological deficits by inhibiting neuroinflammation and ferroptosis via MT2/IL-33/ferritin H signaling in TBI.⁵⁸⁸ Melatonin improves brain function in mice after TBI by inhibiting ferroptosis and ER stress by reducing the expression of circPtpn14 (mmu_circ_0000130), which sponges miR-351-5p to downregulate 5-LOX.⁵⁸⁹

Multiple sclerosis

The G9a inhibitor UNC0642 reduces inflammation-induced neuronal loss and improves clinical outcomes through restoring the expression of antiferroptotic genes. The neuronal expression of antiferroptotic genes was found to be decreased in MS brain tissue but was increased by G9a inhibition in human neuronal cultures.³⁷⁸

Modulation of epigenetic modifications regulating ferroptosis for CVD therapy

Increasing evidence has suggested that ferroptosis may represent a therapeutic approach for CVD. Some small molecule compounds can target dysregulated epigenetic mechanisms in liver diseases (Table 14).

Aortic dissection

BRD4770 attenuates aortic dilation by inhibiting the inflammatory response and ferroptosis.⁵⁹⁰ Liproxstatin-1 inhibits BAPN-induced aortic aneurysm and dissection (AAD) in mice. Inhibition of METTL3 expression or ferroptosis is an effective intervention strategy for AD.⁴²² Ferrostatin-1 (Fer-1) abolishes the detrimental role of PRMT4-mediated ferroptosis in DIC.⁴¹¹

Disseminated intravascular coagulation (DIC)

Metformin (MET) improves cardiac function by inhibiting ferroptosis through activation of AMPKα2 phosphorylation.⁴¹² Ferrostatin-1 suppresses the exacerbation of DOX-induced myocardial damage in MITOL-knockout hearts.⁴¹⁴

Cardiac fibrosis

C646 attenuates ferroptosis by inhibiting p53 acetylation in H9c2 myofibroblasts.⁴¹⁶ Treatment of SIRT3-cKO mice with ferrostatin-1 was found to lead to significant reductions in ferroptosis and CF.⁴¹⁶ Ferrostatin-1 increases the antioxidant capacity of cardiac fibroblasts, reduces GPX4-mediated ferroptosis and alleviates I/R-induced CF.⁴¹⁷

HFD-induced cardiac injury

Celastrol induces ferroptosis resistance via AKT/GSK3β signaling in HFD-induced cardiac injury.⁵⁹¹ Piperlongumine inhibits ferritinophagy-mediated ferroptosis in cardiomyocytes by reducing the phosphorylated STAT3 level.⁴²⁰

Myocardial ischemia/reperfusion injury

Inhibition of miR-190a-5p upregulates GLS2, resulting in decreased ferroptosis, and this effect could be blocked by the GLS2 inhibitor compound 968.³⁹⁹

Modulation of epigenetic modification and posttranslational modifications regulating ferroptosis for liver disease therapy

Emerging evidence has suggested that ferroptosis may represent a novel therapeutic approach for liver diseases. Dysregulated epigenetic mechanisms and PTMs can also be feasibly targeted by small molecule compounds in liver diseases (Table 14).

Liver fibrosis

Berberine attenuates liver fibrosis in mice by inducing ROS-mediated HSC ferroptosis.⁵⁹² Recombinant FGF21 inhibits liver fibrosis by inhibiting hepatocyte ferroptosis through promotion of HO-1 ubiquitination and degradation and Nrf2 activation.⁵⁹³ Artemether alleviates liver fibrosis in vivo and in vitro by inducing HSC ferroptosis through inhibition of IRP2 ubiquitination, thereby inducing an increase in the iron content in HSCs.⁵⁹⁴

Nonalcoholic fatty liver disease

Urolithin C ameliorates NAFLD by regulating the AMPK-ferroptosis axis, maintaining the intestinal mucosal barrier and counteracting gut dysbiosis.⁵⁹⁵

Toxin-mediated hepatic toxicity

Ferrostatin-1 and tert-butylhydroquinone (tBHQ) reverse ethyl carbamate-induced liver dysfunction and inflammation by inhibiting ferroptosis in BALB/c mice.⁴⁶⁴ Ferrostatin-1 and tBHQ were also found to attenuate liver damage resulting from glyphosate-triggered ferroptosis in a mouse model.⁴⁶⁵

Acute liver failure

AGK2 alleviates thioacetamide-induced acute liver failure by inhibiting ferroptosis by regulating the MFN2-PERK pathway.⁵⁹⁶

Modulation of epigenetic and posttranslational modifications regulating ferroptosis for lung disease therapy

Emerging evidence has suggested that targeting ferroptosis may represent a novel therapeutic approach for lung diseases. Dysregulated epigenetic mechanisms and PTMs can also be feasibly targeted by small molecule compounds in lung diseases (Table 14).

Acute lung injury

MitoQ pretreatment counteracts arsenic-induced pulmonary ferroptosis and ALI.⁴⁶⁹ Ferrostatin-1 mitigates arsenic-triggered mitochondrial damage and ferroptosis in MLE-12 cells.⁴⁶⁹ PERK inhibitors mitigate arsenic-induced ferroptosis in MLE-12 cells.⁴⁶⁹ Ferrostatin-1 alleviates lung injury and pulmonary edema.⁴⁷⁰

Sepsis-induced acute lung injury

Obacunone alleviates LPS-induced acute lung injury by inhibiting ferroptosis through inhibition of the ubiquitin-mediated proteasomal degradation of Nrf2, thereby upregulating GPX4 and SLC7A11.⁵⁹⁷

Pulmonary epithelial senescence

SIRT3 activation by melatonin alleviates PM2.5-induced senescence and ferroptosis in mice by deacetylating p53 at lysine 320 (K320).⁵⁹⁸

Chronic obstructive pulmonary disease

hMFG-E8 ameliorates ferroptosis induced by cigarette smoke extract in BEAS-2B cells and HBE cells.⁴⁸³

Modulation of epigenetic and posttranslational modifications regulating ferroptosis for kidney disease therapy

An increasing body of evidence indicates that targeting ferroptosis may represent a therapeutic approach for kidney disease. Dysregulated epigenetic mechanisms and PTMs can also be feasibly targeted by small molecule compounds in kidney diseases (Table 14).

Diabetic nephropathy

Germacrone ameliorates DN by inhibiting ferroptosis through upregulation of mmu_circRNA_0000309, which sponges miR-188-3p, subsequently upregulating GPX4 expression.⁴⁸⁶

Renal IRI

The small molecule USP14 inhibitors IU1 and NAC ameliorate renal injury in I/R mice.⁴⁹⁰ The JAK2 inhibitor fedratinib downregulates TRIM21 expression and reduces damage both in vivo and in vitro, and these effects are correlated with upregulation of GPX4.⁴⁹¹

Toxin-mediated kidney toxicity

SB431542 and SB505124, ALK4/5 kinase inhibitors, suppress cadmium-induced HK-2 cell death.⁴⁹⁴ SIS3, a selective inhibitor of TGFβ1-dependent Smad3 phosphorylation, attenuates cadmium-induced cell death.⁴⁹⁴

Folic acid-induced kidney injury

Roxadustat (FG-4592) ameliorates folic acid-induced kidney injury by repressing ferroptosis via Akt/GSK-3β-mediated Nrf2 activation.⁵⁹⁹

Acute kidney injury

Baicalein ameliorates polymyxin B-induced acute renal injury by inhibiting ferroptosis through a reduction in p53 K382 acetylation via upregulation of SIRT1 expression.⁶⁰⁰ Dihydromyricetin attenuates cisplatin-induced AKI by inhibiting ferroptosis.⁶⁰¹ Myo-inositol ameliorates cisplatin-induced AKI by inhibiting ferroptosis through the promotion of CHIP-mediated ubiquitination of NOX4.⁶⁰²

Diabetic kidney disease

Dapagliflozin ameliorates tubular injury by inhibiting ferroptosis through stabilization of SLC40A1 by reducing its ubiquitin-mediated degradation.⁶⁰³ Ginkgolide B alleviates DKD by inhibiting ferroptosis through inhibition of GPX4 ubiquitination.⁶⁰⁴ Schisandrin A attenuates ferroptosis and pyroptosis in DKD through attenuation of mitochondrial damage via AdipoR1 ubiquitination.⁶⁰⁵

Adriamycin-induced renal damage

Astragaloside IV attenuates adriamycin-induced renal damage by inhibiting ferroptosis through activation of PI3K/Akt and Nrf2.⁶⁰⁶

Concluding remarks and future perspectives

Epigenetic modifications and PTMs are essential for the maintenance of physical homeostasis and are implicated in the pathology of a variety of diseases. Accumulating evidence indicates that epigenetic modifications and PTMs play vital roles in regulating ferroptosis by controlling the expression of ferroptosis-associated genes in cancer, NSDs, CVDs, liver diseases, lung diseases, and kidney diseases. The present review highlights the crucial roles of epigenetic modifications and PTMs regulating ferroptosis in tumorigenesis and the genesis of NSDs, CVDs, liver diseases, lung diseases, and kidney diseases and clarified the regulatory interactions between epigenetic modifications and PTMs regulating ferroptosis in the genesis of these diseases. However, epigenetic modification-mediated regulation of ferroptosis in cancers is an emerging field and is still in its infancy. It is crucial to determine whether other epigenetic regulators play a role in regulating ferroptosis in cancers. First, in addition to ubiquitination, phosphorylation, acetylation, SUMOylation, O-GlcNAcylation, methylation, and ncRNA regulation, whether other epigenetic modifications, including ISGylation and lactylation, are involved in regulating ferroptosis remains unclear. Second, specific epigenetic modifications and PTMs regulate ferroptosis in specific cancers, NSDs, CVDs, liver diseases, lung diseases, and kidney diseases, and whether these epigenetic regulatory mechanisms are actually specific in other types of cancers remains an open question for future investigation. Third, epigenetic modifications and PTMs of targets that induce or inhibit ferroptosis have been identified, and dysregulated epigenetic modifications and PTMs can be feasibly targeted by small molecule compounds. Epigenetic drugs have exhibited viable therapeutic potential for NSDs,^{607,608,609,610} CVDs,^{611,612,613,614,615} liver diseases,^616,617 lung diseases,^618,619,620 and kidney diseases^621,622,623 in preclinical and clinical trials. Epigenetic drugs targeting epigenetic regulators through modulation of epigenetic mechanisms have been applied and clinically translated for the treatment of hematological malignancies, greatly contributing to the development of antitumor drugs.^118,124,624 Epigenetic drugs were found to exhibit viable therapeutic potential for solid tumors in preclinical and clinical trials.^118,625 Combinations of epigenetic drugs with other therapies are being tested in preclinical research and clinical trials for the treatment of solid tumors.^626,627 According to previous studies on clinical epigenetic drugs for hematological malignancies and studies that have revealed key roles of ferroptosis in mediating antitumor activity, we believe that epigenetic drugs targeting epigenetic regulators and ferroptosis could erase the roadmap to cancer. However, much remains to be done before the practical application of these treatment modalities, and inevitable challenges remain and need further clinical investigation. Herein, we highlight in detail the current efforts to translate this knowledge into clinical benefit for patients. Fourth, whether epigenetic modifications affect multiple ferroptosis regulators and how these different epigenetic modifications and PTMs cooperate with diverse signaling pathways to control the susceptibility of cancer cells to ferroptosis remain unclear. The epigenetic modification network of ferroptosis needs extensive investigation. Fifth, although ferroptosis has recently been confirmed to play a vital role in tumorigenesis and in the genesis of NSDs, CVDs, liver diseases, lung diseases, and kidney diseases, the detailed mechanisms related to ferroptosis and the molecular pathways involved remain open topics for future investigation. Moreover, the mechanisms by which epigenetic modifications and PTM events control the expression of ferroptosis-related genes in cancers, NSDs, CVDs, liver diseases, lung diseases, and kidney diseases remain incompletely elucidated. Determining whether mechanisms of novel epigenetic modifications and PTMs are actually specific in different diseases and cell types needs more systematic and comprehensive investigation.

In conclusion, epigenetic modifications and PTMs play fundamental roles in diseases, such as cancers, NSDs, CVDs, liver diseases, lung diseases, and kidney diseases. More studies on the effects of epigenetic modifications and PTMs on ferroptosis will ensure a better understanding of the pathogenesis of these diseases and identify novel paradigms for their treatment.