Ferroptosis-Targeting Drugs In Cancer Therapy

Table 1. Cell morphology, biochemical features, and key regulators of ferroptosis, apoptosis, necroptosis.

Beyond these findings, ferroptosis has recently gained much importance in cancer treatment, and emerging evidence shows that ferroptosis influences a growing number of oncogenic pathways. For example, P53 regulation promotes tumour cell ferroptosis but also decreases tumour metastases

to blood, lung, and liver. Viswanathan and colleagues contended the contribution of ferroptosis to drug resistant phenomenon and cancer immunotherapeutic efficacy.

Basis of ferroptosis

Figure 1. The complicated interplay of iron, cysteine and lipid metabolism takes an important role in ferroptosis

 Mechanisms of Ferroptosis. Excess irons are regarded as an important factor for ferroptosis. The circulated iron (Fe³⁺) combined with transferrin (TF) enters cells mediated by transferrin receptor (TFR). Under the catalysis of iron oxide reductase STEAP3, Fe³⁺ can be deoxidized to Fe²⁺ and ultimately, releasing it into labile iron pool (LIP) mediated by DMT1. LIP consists of iron from endosomal uptake of circulated iron and ferritin degradation (ferritinophagy). System Xc-mediate the uptake of cystine (Cys2). Cys2, glutamate (Glu) and glycine (Gly) are materials of glutathione (GSH), which is an important antioxidant in cells. Transsulfurylation pathway may also increase the level of cysteine transformed from methionine (Met). Cysteine can be imported directly by alanine/serine/cysteine transporter (system ASC) under reducing conditions. The PE-PUFAs can be oxidized to PE-PUFAs-OOH by lipoxygenases (LOXs), leading to ferroptosis. GPX4 roles as a protector to transfer PE-PUFAs-OOH to PE-OH. CoQ10, coenzyme Q10; DMT1, divalent metal transporter 1; FPN, ferroprotein; Gln, glutamine; HAMP, hepcidin antimicrobial peptide; HMGCR, 3-hydroxy-3-methylglutaryl-CoA reductase; IREB2, iron-responsive element binding protein 2; NCOA4, Nuclear receptor coactivator 4; STEAP3: six-transmembrane epithelial antigen of the prostate 3.

The most prominent character of ferroptosis is ROS generation, mainly caused by iron metabolism disorders . The endosomal uptake of circulated iron(Fe³⁺) is mediated by its binding to transferrin (TF) and transferrin receptor 1 (TFR1). Iron Fe³⁺ is deoxidized to iron Fe²⁺, under the catalysis of iron oxide reductase named six-transmembrane epithelial antigen of the prostate 3 (STEAP3) and ultimately release into labile iron pool (LIP), due to Fe²⁺’s characteristics of high solubility and transfer electron capability. Increasing formation of LIP may trigger the Fenton reaction [the process of ROS generation mediated by interaction between Fe²⁺ and hydrogen peroxide (H₂O₂)], which may result in iron poisonous . Compared with RAS un-mutated ferroptosis-insensitive cells, RAS-mutated ferroptosis-sensitive cells increased the expression of TFR1 and decreased the expression of ferritin light chain (FTL) and ferritin heavy chain 1 (FTH1) in the iron-storage protein subunits. Apart from the mechanisms discussed above, there are several other pathways involved in ferroptosis (Figure 2). In mevalonate (MVA) pathway, the activity of FIN56-targeted protein squalene synthase (SQS) reduces the idebenone level in cells, therefore decreasing the cellular anti-oxidation activity. Nuclear factor (erythroid derived 2) like 2 (Nrf2) is a regulator in the iron metabolism, and Nrf2 activation was reported capable of inhibiting ferroptosis in hepatocellular carcinoma cells. The famous tumour suppressor P53 also plays an essential role in ferroptosis regulation. Other pathways such as the sulphur-transfer pathway, heat shock factor-1 (HSF1)-heat shock protein beta-1 (HSPB1) pathway, and mucin 1 C-terminal (MUC1-C)/system Xc⁻ (xCT) also contribute to ferroptosis regulation (Figure 2) . Further, evaluation of ferroptosis-related molecules provides diverse approaches to monitor the ferroptosis process in-vitro and in-vivo (Tables 2,3).                                 

Figure 2:ferroptosis regulation

Ferroptosis modulation in tumour. Small molecules such as elastin, sorafenib, glutamate, and sulfasalazine induce ferroptosis by inhibiting system Xc- and impeding cysteine uptake, which could result in a subsequent decline of glutathione and a decrease of cells’ anti-oxidative ability. mucin 1 C-terminal (MUC1-C) binds with CD44v to promote stability of the system Xc⁻. The cysteine level can also be supplemented by cellular methionine via the sulphur-transfer pathways. GPX4 can prevent ferroptosis by suppressing cellular lipid peroxides and the mevalonate (MVA) pathway is crucial for its maturation and the products of it (IPP and CoQ10) can promote synthesis of GPX4. Treatment FIN56 modulates squalene synthase (SQS) to reduce CoQ10. Ferroptosis inducer RSL3 can suppress GPX4 directly to regulate ferroptosis. The p62-Keap1-Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway is able to regulate Nrf2-targeted genes such as hemi oxygenase-1 (HO-1), ferritin heavy chain 1 (FTH1), and NAD(P)H: quinone oxidoreductase 1 (NQO1) against ferroptosis. CISD1, PHKG2, and IREB2 are important in regulating iron metabolism and ferroptosis. Iron chelators can inhibit ferroptosis. The HSPB1 also impedes ferroptosis by inhibiting increase of intracellular iron. In addition, p53 also regulate ferroptosis through inhibiting SLC7A11 and promoting lipid peroxides production. BSO, buthionine sulfoximine; FTH1, ferritin heavy chain 1; HSP, heat-shock protein; HO-1, hemi oxygenase-1; MUC1-C, mucin 1 C-terminal; MVA, mevalonate; NQO1, NAD(P)H: quinone oxidoreductase 1; Nrf2, nuclear factor (erythroid-derived 2)-like 2; SQS, squalene synthase.

Table 2. Ferroptosis inducers.

Table 3. Ferroptosis inhibitors.

Hall marks of ferroptosis in cancer

Role of ferroptosis in tumour suppression:

Tumour suppressor P53 inactivation is very common in cancers. The anti-tumour activity of P53 was thought to drive cell senescence, cell cycle arrest and apoptostraditionally. These years, P53 has been explored to be essential in some other activities to suppress tumour progression (53,54). In the study of Jiang’s group, the acetylated defective mutant TP533KR lost its function to induce cell senescence, cell cycle arrest and apoptosis, while the function of ferroptosis-induction was still kept. The cancer progression was depressed through the inhibition of Cys2 uptake and elevation of tumours' sensitivity to ferroptosis by repressing the SLC7A11 expression (55). Additionally, evidence has shown a high release of mobility group box 1 (HMGB1) in ferroptosis. We may conjecture that ferroptotic tumour cells might be immunogenic (56). Release of damage-associated molecular patterns (DAMPs) can trigger Toll-like receptor 4 (TLR4) signals in ferroptotic cell death. Such phenomena have been observed in the attraction of neutrophils and dendritic cells (DCs), thus activating the innate immune system.

Role of ferroptosis in tumour promotion and tumour evasion

Ferroptotic cancer cells can release oxidized lipid mediators, which might regulate antitumor immunity (Figure 3). Eicosanoids such as 5-hydroxyeicosatetraenoic acid (5-HETE), 11-HETE and 15-HETE released from ferroptotic cells can induce GPX4 depletion and affect anti-tumour immunity. GPX4 inactivation was reported associated with ferroptosis promotion of T cells. The lower GPX4 activity is, the more pro-inflammatory lipid mediators such as 5-HETE and leukotriene B4 (LTB4) produced, whereas LTB4, a kind of pro-inflammatory leukotriene, is crucial to carcinogenesis. Beyond free eicosanoids, esterified eicosanoids also role in immune response. Oxidized phosphatidylcholine was reported to inhibit DC maturation through Nrf2 activation and suppress the differentiation of T helper 17 (TH17) cells.                                

Figure 3. Regulate antitumor immunity

Role of ferroptosis in cancer. AA, arachidonic acid; AdA, adrenaline; ACSL4, acyl-CoA synthetase long-chain family 4; cDC1, type 1 dendritic cell; CAF, cancer-associated fibroblast; DMT1, divalent metal transporter 1; DAMP, damage-associated molecular pattern; HETE, hydroxy eicosatetraenoic acid; HMGB1, high mobility group box 1; FPN, ferroportin; LF, lactoferrin; LIP, labile iron pool; LCN, lipocalin; LPCAT3, lysophosphatidylcholine acyltransferase 3; NTBI, non-transferrin-bound iron; NCOA4, nuclear receptor coactivator 4; NK cell, natural killer cell; PE, phosphatidylethanolamine; PTGS2, prostaglandin-endoperoxide synthase 2; SCARA5, scavenger receptor A member 5; TF, transferrin; TFR1, transferrin receptor 1; TAM, tumour-associated macrophage; TME, tumour microenvironment.

Iron Metabolism

Iron is an indispensable and most abundant trace element in the body, and it participates in many important physiological and biochemical functions in the body. Iron is the main raw material for the synthesis of haemoglobin and myoglobin. It not only participates in the biosynthesis of DNA and ATP but is also an important electron transport chain in mitochondria and a cofactor of metalloproteinases . Under normal conditions, the body maintains the homeostasis of iron through food sources of iron and the “iron cycle” (a process in which aging red blood cells release iron ions under the action of heme oxygenase, and macrophages re-engulf and recycle iron). The hepcidin synthesized and secreted by the liver directly regulates the level of serum iron, while the regulation of iron homeostasis in the body’s cells is mainly played by the iron responsive element, the hepcidin system. The trivalent iron ion (Fe3+) in the peripheral circulation combines with transferrin to form a complex and then binds to the transferrin receptor on the cell membrane and enters the endosomes in the cell.

Regulation of lipid metabolism pathway

Accumulations of PLOOH is the hallmark of ferroptosis. Lipid metabolism is closely related to cell vulnerability to ferroptosis. PLOOH can accumulate in both enzymatic and nonenzymatic ways. Dull et  al.’s study revealed that ACSL4 is an essential component for ferroptosis execution. ACSL4 functions in converting long chain-PUFAs to acyl-CoA, subsequently participating in producing PLOOH in an enzymatic way. Ye et  al. Ribosylation factor 6 (ARF6) overcame gemcitabine Resistance in pancreatic cancer by inducing ferroptosis. In addition, several LOXs have been reported to Be able to oxygenate PUFAs directly in a nonenzymatic Way, thereby mediating ferroptosis . Zhang et  al. Showed that arachidonate lipoxygenase 15 (ALOX15), A LOX member, was closely linked with the suppression of ferroptosis in gastric cancer. Downregulation of microRNA-522, which promotes ALOX15 expression, provides novel methods to enhance cisplatin/paclitaxel sensitivity in gastric cancer by inducing ferroptosis.

Therapeutic strategies targeting ferroptosis

Potential compounds to target ferroptosis based cancer therapy:

Ferroptosis can be induced by suppressing antioxidant defence components such as systems Xc–, GSH, and GPX4, or by precisely controlling various endogenous elements like intracellular iron concentration and PUFA-containing phospholipids.74 Up till now, the potential effectiveness of a variety of naturally occurring substances and clinically utilized medications in inducing ferroptosis has been investigated. While some of these drugs are already FDA-approved, others are still in the preclinical and clinical trial stages (Table 1). Here, we are reminded of the substances already on the market for various purposes that could be converted to ferroptosis-based cancer therapy.

Potential compounds to target intracellular iron levels

One of the key characteristics of ferroptosis is iron overload. The following is a description of the medications and substances that cause ferroptosis by raising the amount of iron within cells: salinomycin, artesunate, neratinib, lapatinib+siramesine, and ruscogenin are some of these agents.

Salinomycin

A polyether ionophore molecule called salinomycin was discovered from the bacterium Streptomycin albus. It exhibits a broad spectrum of antibacterial activity against viruses, gram-positive bacteria, fungi, and parasites. Salinomycin has antitumor effects that significantly reduce the growth of breast tumours in the mice xenograft model,76 according to a 2009 study by Gupta et al.Studies have shown that salinomycin dramatically reduces the ability of docetaxel-resistant prostate cancer cells to form colonies.77 Additionally, the resistance of ovarian cancer cells to chemotherapy medicines based on platinum is substantially eliminated by salinomycin and its derivatives. By focusing on the hypoxia-inducible factor 1-α (HIF-1α)/vascular endothelial growth factor (VEGF) signaling pathway, salinomycin significantly inhibits the angiogenesis and development of breast cancer cells.

80Tumor cells treated with salinomycin have been shown to undergo apoptosis and autophagy caused by endoplasmic reticulum (ER) stress.81-82 Additionally, it has been discovered that salinomycin increases intracellular ROS levels and downregulates NRF2 expression, hence improving the radiosensitivity of nasopharyngeal cancer cells.83Salinomycin treatment for colon cancer cells may cause ROS production and mitochondrial dysfunction. Researchers have discovered that administering salinomycin and AM5 to cancer stem cells prevents iron transport from the lysosome lumen to the cytosol. This, in turn, causes cytosolic iron depletion, which is shown by ferritin degradation and an increase in IREB2 and TfR.85-86By causing lipid peroxide buildup and inducing ferroptosis, delivery of salinomycin-loaded gold nanoparticles (SalAuNPs) to breast cancer stem cells can efficiently kill tumour cells.

Potential compounds to target system Xc-, GSH, And GPX4

Sorafenib

NEXAVAR, commonly known as sorafenib, is an oral bioavailable multitarget kinase inhibitor that is being used to treat patients with thyroid, liver, and advanced renal cell carcinoma.119 By targeting Raf serine/threonine kinases and various cell surface receptor tyrosine kinases, such as VEGFR1-3, PDGFRβ, cKitprotein, FMSlike tyrosine kinase 3 (FLT3), and platelet derived growth factor receptor β (PDGFRβ)derived growth factor receptor β (PDG-FRβ), sorafenib is able to exert its antineoplastic activity.120 It has been discovered that sorafenib not only causes apoptosis but also activates autophagy in cancer cells.121-122 Additionally, it has been discovered by researchers that sorafenib's anti-cancer effect can also be mediated by ferroptosis induction, which is separate from its conventional kinase inhibitory function.123-124 By inhibiting system Xc and subsequently depleting

GSH, sorafenib mechanistically causes ferroptosis.125 Furthermore, research indicates that sorafenib-mediated ferroptosis is also modulated by the expression of a few genes, such as NRF2, retinoblastoma (RB), and MT-1G.126-127 In hepatocellular carcinoma cells, sorafenib-induced ferroptotic cell death is negatively regulated by MT-1G, a transcriptional NRF2 target gene, which inhibits GSH depletion-mediated lipid peroxidation.126 Significantly, sorafenib's anticancer effectiveness is enhanced by decreasing MT-1G both in vitro and in vivo.

Potential compounds to target HMG-CoA Reductase

Statins (Fluvastatin, Pravastatin, Lovastatin, Simvastatin)

Known for their ability to decrease cholesterol, statins are frequently administered to patients with hypercholesterolemia. Mechanistically, statins inhibit HMG-CoA reductase, an essential enzyme involved in the mevalonate pathway-mediated production of cholesterol, IPP, and CoQ10.186 Numerous accounts exist on encouraging attempts to treat cancer with statins.186 Statins have been shown to promote apoptosis in tumour cells and mediate cell cycle G1/S-phase arrest. According to a paper, statins work in concert to enhance sorafenib's antitumor activity in vitro.188 Furthermore, in comparison to control groups, the combination of simvastatin with lipid nano emulsions paclitaxel considerably reduces the tumour growth and metastatic rate of melanoma-bearing animal models. Accordingly, statins may decrease intracellular GPX4 levels and increase lipid peroxidation in a manner that is dose- and time-dependent, according to Viswanathan et al.

Potential compounds to target SCD1 and ACSL4

MF-438, CAY10566, and A939572

SCD1, an enzyme linked to the endoplasmic reticulum, is essential for the transformation of saturated fatty acids (SFAs) into monounsaturated fatty acids (MUFAs). It's interesting to note that a rise in cellular MUFA concentration and overexpression of SCD1 have been seen in a number of cancer types. SCD1 may be a viable target for antitumor therapy since, as a wealth of data over the last ten years has demonstrated, it zl. lays a remarkable role in encouraging tumour growth and metastasis.191 According to Pisanu et al., pharmacological targeting of SCD1 with MF-438 dramatically increases the cisplatin susceptibility of lung cancer stem cells.192 More research is required to determine the exact underlying mechanism by which SCD1 inhibition inhibits tumour growth. SCD1 inhibition induced by MF-438 and CAY1-0566 inhibits tumour cell growth and chinitiates apoptosis.193 Moreover, administering CAY1-0566 to hepatocellular carcinoma cells induces autophagy via AMP-activated protein kinase (AMPK).194 By suppressing YAP/TAZ activity, MF-438 has been demonstrated to eliminate lung cancer cells' capacity to form spheres.195 Further more, A939572-inhibiting SCD1 prevents tumour cell migration that is driven by cancer-associated fibroblasts (CAF).196 The ferroptosis is maintained by suppressing SCD1, which lowers MUFA and CoQ10 levels. According to Tesfay and colleagues, administering SCD1 inhibitors to ovarian cancer cells causes an increase in lipid peroxidation and ferroptosis-mediated cell death, which is prevented when Fer-1 and oleic acid are present. Furthermore, erastin, RSL3, and SCD1 inhibitors work in concert to suppress tumour growth in vivo and in vitro.

Nanoparticle inducers of ferroptosis in cancer

Nanotechnology applications have attracted much attention with specific physicochemical properties recently (Table 4).

Table 4