Iron accumulation, glutathione depletion, and lipid peroxidation must occur simultaneously during ferroptosis and are mutually amplifying events

doi:10.1016/j.mehy.2017.02.017

Medical Hypotheses

Volume 101, April 2017, Pages 69-74

https://doi.org/10.1016/j.mehy.2017.02.017 Get rights and content

Abstract

Ferroptosis is a recently discovered form of regulated necrosis that involves iron-dependent lipid peroxidation. How cells die once ferroptosis is triggered remains unclear. Ferroptosis is hypothesized to require three critical events: (1) accumulation of redox-active iron, (2) glutathione depletion, and (3) lipid peroxidation. It is proposed that these three events must unfold simultaneously because stopping any critical event also stops ferroptosis. These events are hypothesized to amplify in severity through positive feedback loops. The cause of death in ferroptosis is therefore the synergistic combination of antioxidant depletion, iron toxicity, and membrane denaturation. The relevance of these feedback loops for cancer and neurodegenerative therapies is discussed.

Graphical abstract

Introduction

Ferroptosis is an iron-dependent mode of regulated necrosis that is biochemically, genetically, and morphologically distinct from apoptosis, autophagy, and other forms of necrosis, and is therefore a new way that cells can die [1], [2]. The glutamate/cystine antiporter (named X_C⁻) supplies extracellular cystine in exchange for intracellular glutamate, a process required for the biosynthesis of the endogenous antioxidant glutathione. The antitumor molecules erastin and sorafenib trigger ferroptosis by inhibiting X_C⁻, resulting in glutathione depletion and oxidative damage [3], [4]. Ferroptosis may also be induced by small molecules RSL3 and ML162 by inhibiting glutathione peroxidase 4 (Gpx4), a lipid repair enzyme essential for life [5]. Iron is suggested to be involved in ferroptosis because death is prevented by co-treatment with the iron chelator deferoxamine [1]. Ferroptosis is regulated by several genes, for example, iron metabolism genes TFRC and IREB2 [1], glutaminolysis-regulating genes SLC38A1 and GLS2 [6], the pentose phosphate pathway gene G6PD [1], and autophagy-regulating genes ULK1 and BECN1 [7]. Interest in ferroptosis as a natural tumor-suppressing process has been spurred by the discovery that tumor-suppressor proteins RB1 and p53 can activate ferroptosis [8], [9], [10], [11]. Ferroptosis is observed in both in vitro and in vivo models, and research is underway expanding the repertoire of ferroptosis-inducing and -suppressing molecules [12].

How death occurs once ferroptosis is triggered remains unclear. Firstly, it is hypothesized that for ferroptosis to occur three critical events are required: (1) Accumulation of ‘free’ iron, which causes oxidative stress through Fenton catalysis, (2) depletion of the antioxidant glutathione, resulting in oxidative stress, and (3) accumulation of lipid oxidative damage, leading to cell membrane denaturation. Secondly, it is hypothesized that each event must unfold simultaneously for ferroptosis to occur because experimental evidence suggests that stopping any of these events also stops ferroptosis. Thirdly, it is hypothesized that these three critical events, once triggered, can be mutually exacerbated through positive feedback loops and are therefore amplifiable events. Death from ferroptosis is therefore proposed to be the synergistically lethal combination of iron toxicity, antioxidant depletion, and membrane damage. These three hypotheses are summarized in Fig. 1.

Section snippets

Hypothesis 1A – Relevance of glutathione depletion to ferroptosis

Glutathione is a tripeptide (Glu-Cyc-Gly) and an important cellular antioxidant that protects lipids, proteins, and DNA from oxidative damage [13]. Glutathione donates electrons via glutathione peroxidase by dimerizing to glutathione disulfide. Enzymatic reduction of glutathione disulfide restores glutathione. Glutathione depletion is associated with Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, and Friedreich’s ataxia [14]. Glutathione depletion

Testing the hypothesis

As the amplification of requisite events is the most important implication of this work, approaches to experimentally validate this positive feedback loop will be the subject of focus. Oxidative stress causes lipid peroxides to accumulate. This is predicted to result in the permeabilization of the lysosomal membrane and subsequent leakage of iron into the cytosol [39]. Notably, the iron chelator deferioxamine, used in the majority of published studies on ferroptosis, is eventually localized

Relevance to cancer therapy

Metabolic remodeling is imperative to cancer survival [58]. Such remodeling includes heightened glutathione levels afforded by stabilizing X_C⁻ expression [59] as well as increased intracellular iron intake due to the metabolic demands of rapidly replicating cells [60], [61]. Nonetheless, cancer cells typically suffer chronic oxidative stress due to dysfunctional metabolism and are hypersensitive to free radicals [62], [63], [64]. The hypothesis predicts that sabotaging antioxidant defences or

Conclusion

Ferroptosis is a novel cell death mechanism that can be triggered by disruption of the membrane repair enzyme Gpx4 or the X_C⁻ antiporter required for glutathione biosynthesis. A hypothesis has been presented explaining ferroptosis as the consequence of three critical events that must operate concurrently: (1) Iron accumulation, (2) glutathione depletion, and (3) lipid membrane oxidation. Once ferroptosis is triggered, these events can amplify through positive feedback mechanisms, expediting

Conflict of interest statement

There are no conflicts of interest to declare.

Funding

National Sciences and Engineering Research Council of Canada (No. 460639-2014).

Acknowledgements

This work was supported by the National Sciences and Engineering Research Council of Canada (NSERC) (No. 460639-2014). Anonymous reviewers are also acknowledged for their insightful contributions during manuscript development.

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Iron accumulation, glutathione depletion, and lipid peroxidation must occur simultaneously during ferroptosis and are mutually amplifying events

Abstract

Graphical abstract

Introduction

Section snippets

Hypothesis 1A – Relevance of glutathione depletion to ferroptosis

Testing the hypothesis

Relevance to cancer therapy

Conclusion

Conflict of interest statement

Funding

Acknowledgements

Cell

Mol Cell

Cancer Lett

Cell Rep

Biochim Biophys Acta

Free Radical Biol Med

J Biol Chem

J Biol Chem

Arch Biochem Biophys

Free Radical Biol Med

Cell Metab

Biochem Biophys Res Commun

Cell Metab

Free Radical Biol Med

Neoplasia

J Biol Chem

J Biol Chem

Cancer Cell

Crit Rev Oncol Hematol

FEBS Lett

Cancer Cell

Biochem Pharmacol

Adv Drug Deliv Rev

Mutat Res

Biomaterials

Free Radical Biol Med

Biomaterials

Cell

Ferroptosis: an iron-dependent form of nonapoptotic cell death

Cell

Mechanisms of ferroptosis

Cell Mol Life Sci

Pharmacological inhibition of cystine-glutamate exchange induces endoplasmic reticulum stress and ferroptosis

eLife

Iron-dependent cell death of hepatocellular carcinoma cells exposed to sorafenib

Int J Cancer

Ferroptosis is an autophagic cell death process

Cell Res

Ferroptosis as a p53-mediated activity during tumour suppression

Nature

Activation of SAT1 engages polyamine metabolism with p53-mediated ferroptotic responses

Proc Natl Acad Sci USA

Synchronized renal tubular cell death involves ferroptosis

Proc Natl Acad Sci USA

Glutathione in cellular redox homeostasis: association with the excitatory amino acid carrier 1 (EAAC1)

Molecules

Impaired glutathione synthesis in neurodegeneration

Int J Mol Sci

Fenton chemistry in biology and medicine

Pure Appl Chem

Redox active metal-induced oxidative stress in biological systems

Transition Met Chem

Iron homeostasis and management of oxidative stress response in bacteria

Metallomics

Lethal oxidative damage and mutagenesis are generated by iron in ∆fur mutants of Escherichia coli: protective role of superoxide dismutase

J Bacteriol

Iron-dependent formation of reactive oxygen species and glutathione depletion after accumulation of magnetic iron oxide nanoparticles by oligodendroglial cells

J Nanopart Res