Targeting ferroptosis and cuproptosis in gastrointestinal cancers: molecular mechanisms, metabolic vulnerabilities, and therapeutic interventions

Ferroptosis is an iron-dependent form of regulated cell death driven by lipid peroxidation, which is morphologically, biochemically, and genetically distinct from other cell death modalities such as apoptosis, necrosis, and autophagy [18, 28]. This process is characterized by an imbalance between cellular oxidative and antioxidant systems, ultimately resulting in severe peroxidative damage to membrane phospholipids [29]. Its complex molecular regulatory network can be understood as a dynamic equilibrium between a "pro-death" substrate supply system and a "pro-survival" core detoxification system [30]. On one hand, polyunsaturated fatty acids (PUFAs) in cell membranes and the labile iron pool provide the necessary chemical foundation for ferroptosis. On the other hand, a robust intracellular antioxidant network, particularly the defense axis centered on glutathione peroxidase 4 (GPX4), constitutes the critical line of defense against ferroptosis [19]. When this defense is breached, the initiation of ferroptosis becomes inevitable. The core molecular mechanism of ferroptosis is illustrated in Fig. 1 of this review.

The initiation and execution of ferroptosis are highly dependent on two key elements of its "pro-death arm": oxidizable lipid substrates and the iron ions that catalyze the oxidative reactions [31]. First, PUFAs embedded in membrane phospholipids, especially arachidonic acid (AA) and adrenic acid (AdA), are the primary substrates for lipid peroxidation [31]. Acyl-CoA synthetase long-chain family member 4 (ACSL4) acts as the "initiator" in this process, responsible for activating these free PUFAs into their corresponding acyl-CoAs [32]. Subsequently, lysophosphatidylcholine acyltransferase 3 (LPCAT3) efficiently incorporates these activated PUFAs into membrane phospholipid molecules, a process that significantly increases the membrane's susceptibility to peroxidative damage [32]. Lipid peroxidation can be initiated through both enzymatic and non-enzymatic pathways. The enzymatic pathway is primarily mediated by iron-dependent lipoxygenases (LOXs) [30]. A pivotal study found that the scaffold protein phosphatidylethanolamine-binding protein 1 (PEBP1) can form a complex with 15-lipoxygenase (15-LO). This binding alters the substrate specificity of 15-LO, enabling it to directly catalyze the peroxidation of phosphatidylethanolamine (PE) in the membrane, thereby generating hydroperoxy-phosphatidylethanolamine (hydroperoxy-PE) and triggering the ferroptosis signal [33]. Second, central to the "iron-dependent" nature of ferroptosis, ferrous ions (Fe²⁺) in the labile iron pool are the key catalysts for the Fenton reaction, which generates highly reactive hydroxyl radicals. These radicals, in turn, initiate and propagate the chain reaction of lipid peroxidation [17, 18]. Cells tightly regulate the level of the labile iron pool through various mechanisms, among which the selective autophagy process known as ferritinophagy is a key upstream event promoting ferroptosis [34]. In this process, the specific cargo receptor, nuclear receptor coactivator 4 (NCOA4), recognizes and binds to the intracellular iron-storage protein complex, ferritin, thereby targeting it for degradation in the lysosome. This action releases a large amount of iron ions, greatly expanding the labile iron pool and providing an ample catalyst for the Fenton reaction [34].

In contrast, the "pro-survival arm" comprises multiple precise and hierarchical antioxidant systems designed to neutralize or repair damage caused by lipid peroxidation. The most classic and central defense axis is the System Xc⁻–glutathione (GSH)–GPX4 pathway [19, 30]. System Xc⁻, composed of the SLC7A11 and SLC3A2 subunits, is responsible for importing extracellular cystine, which is then reduced to cysteine intracellularly, providing the key raw material for GSH biosynthesis [35]. As the most abundant hydrophilic antioxidant in the cell, GSH is an essential reducing cofactor for the catalytic activity of GPX4 [19, 36]. GPX4 is a unique selenoenzyme that crucially utilizes GSH to efficiently reduce lipid hydroperoxides (PLOOH) formed on membrane phospholipids to their non-toxic corresponding lipid alcohols (PLOH), thereby directly terminating the lipid peroxidation chain reaction [37, 38]. However, the cell's defense system is not limited to this pathway. Recent studies have revealed several parallel, GSH-independent defense pathways. One important pathway is the ferroptosis suppressor protein 1 (FSP1)–coenzyme Q10 (CoQ10) axis [20, 39]. FSP1 is a membrane-localized, NAD(P)H-dependent oxidoreductase that reduces the oxidized form of CoQ10 (ubiquinone) to its reduced form, CoQH₂ (ubiquinol) [39]. As a potent lipophilic radical-trapping antioxidant, ubiquinol can directly scavenge lipid peroxy radicals within the membrane, thus inhibiting the initiation and propagation of ferroptosis independently of the GSH/GPX4 system [20]. Another parallel defense pathway is mediated by GTP cyclohydrolase 1 (GCH1) and its product, tetrahydrobiopterin (BH4) [40]. The expression of GCH1 leads to the synthesis of BH4, which not only possesses intrinsic antioxidant activity but also contributes to a robust anti-ferroptotic barrier by promoting the endogenous production of CoQ10 and regulating lipid remodeling [40].

At the mitochondrial level, this organelle plays an indispensable and complex role in the regulatory network of ferroptosis. It acts as both a signal amplifier and possesses its own independent defense systems; its dysfunction and oxidative stress are key components of the ferroptotic mechanism [41, 42]. Although the final execution site of ferroptosis is the rupture of the plasma membrane, the functional state of mitochondria profoundly influences a cell's susceptibility to ferroptosis [43, 44]. First, mitochondria are the primary sites of reactive oxygen species (ROS) in the cell [41]. During the operation of the mitochondrial electron transport chain (mtETC), superoxide anions and other ROS inevitably "leak" out, significantly exacerbating the cell's overall oxidative stress level. This synergizes with Fenton reaction-driven lipid peroxidation, potently amplifying the ferroptosis signal [29, 45]. Second, mitochondria are the crossroads of several key metabolic pathways, including the tricarboxylic acid (TCA) cycle, fatty acid β-oxidation, and the biosynthesis of iron-sulfur (Fe-S) clusters [38]. These metabolic activities are directly linked to the core elements of ferroptosis. For example, the metabolic state of the TCA cycle can impact the cellular redox balance. At the same time, dysfunction of mitochondria as the center of Fe-S cluster synthesis can directly affect iron metabolism and distribution, potentially leading to an expansion of the labile iron pool and thus promoting ferroptosis [46]. More importantly, mitochondria harbor specific ferroptosis defense mechanisms. GPX4 has its own mitochondrial isoform (mGPX4), which is specifically responsible for eliminating lipid peroxides in the inner mitochondrial membrane, thereby protecting mitochondrial structure and functional integrity [47]. Furthermore, a mitochondrial defense mechanism independent of the classic pathways revolves around dihydroorotate dehydrogenase (DHODH), which is localized to the inner mitochondrial membrane [48]. While playing its role in the de novo pyrimidine synthesis pathway, DHODH can also utilize its redox activity to reduce ubiquinone to ubiquinol on the mitochondrial membrane. This activity forms a potent local antioxidant barrier that effectively inhibits mitochondrial lipid peroxidation and protects the cell against ferroptosis [48].

Cuproptosis is a recently elucidated and unique form of regulated cell death triggered by copper overload, with a core mechanism distinct from known pathways such as apoptosis, ferroptosis, or necroptosis [21, 49, 50]. Unlike apoptosis, cuproptosis is independent of caspase executioners and originates from severe biochemical disturbances within respiring mitochondria [21]. The fundamental cause is that excess copper ions directly target and disrupt key proteins in the tricarboxylic acid (TCA) cycle, triggering a cascade of events that ultimately lead to cell death from unbearable proteotoxic stress and metabolic collapse [22, 51]. This discovery underscores the importance of intracellular metal homeostasis and offers a novel molecular understanding of the biological toxicity of copper [52]. The key steps of this mitochondria-centric cell death pathway are summarized in Fig. 2 of this review.

The molecular mechanism of cuproptosis is centered on the mitochondrion, and its initiation and execution involve a precise and interconnected signaling chain [21, 53]. Intracellular copper concentration is strictly regulated by copper transporters, with SLC31A1 (also known as CTR1) being the primary copper importer. At the same time, ATP7A and ATP7B are responsible for pumping excess copper out of the cell to maintain copper homeostasis [54, 55]. When copper homeostasis is disrupted and a large influx of copper ions enters the cell, the cuproptosis pathway is activated. In this process, ferredoxin 1 (FDX1) plays a crucial upstream regulatory role [56]. As a mitochondrial [2Fe-2S] protein, FDX1 performs a dual key function: it not only efficiently reduces cupric ions (Cu²⁺) to the more reactive cuprous ions (Cu⁺) but is also essential for the process of protein lipoylation [52, 56]. Protein lipoylation, the covalent attachment of lipoic acid to lysine residues of specific proteins, is a critical prerequisite for cuproptosis. It generates the direct targets for copper ions, paving the way for the subsequent lethal binding events [57].

The core execution event of cuproptosis is the direct binding of cuprous ions (Cu⁺) to lipoylated protein components of the TCA cycle [21, 22]. The most critical target protein is dihydrolipoamide S-acetyltransferase (DLAT), which is the E2 subunit of the pyruvate dehydrogenase (PDH) complex [21, 58]. Excess Cu⁺ binds to the lipoic acid moieties on DLAT, causing its irreversible oligomerization and aggregation [21, 59]. This protein aggregation directly induces severe mitochondrial dysfunction. First, it induces intense proteotoxic stress, as a large number of misfolded and aggregated proteins accumulate in the mitochondrial matrix, overwhelming the mitochondrion's own processing capacity [51, 60]. Second, this process further induces the loss and degradation of iron-sulfur (Fe-S) cluster proteins [21, 61]. Because Fe-S cluster proteins are essential cofactors for many key enzymes in the mitochondrial electron transport chain (ETC) complexes and the TCA cycle, their loss severely impairs mitochondrial respiration. It blocks ATP production, leading to a cellular energy crisis [41, 62]. Even more fatally, the collapse of the ETC exacerbates electron leakage, leading to an explosive production of ROS and plunging the cell into a state of severe oxidative stress [62]. Therefore, FDX1-mediated copper ion reduction and protein lipoylation serve as the "upstream switch" for cuproptosis, while the aggregation of lipoylated proteins, such as DLAT, the loss of Fe-S cluster proteins, and the ensuing lethal mitochondrial dysfunction act as its "downstream executioner" [53, 63].

Although ferroptosis and cuproptosis are two forms of regulated cell death driven by different metal ions and possessing distinct execution mechanisms, they do not exist in isolation. Instead, they are intertwined within the cell through multiple molecular nodes, forming a complex regulatory network [25, 26]. These two pathways exhibit significant crosstalk at the levels of cellular metabolism, organelle functional homeostasis, and upstream signaling [61, 64]. Their interaction is mainly manifested in shared metabolic hubs, a common platform for organellar stress, and overlapping upstream regulatory pathways. These points of intersection make it possible to simultaneously target both death modalities, an approach that is expected to produce a synergistic killing effect [65–67].

The most central and direct node of crosstalk between these two death pathways is the cellular glutathione (GSH)/cysteine metabolic axis [25, 36]. GSH plays a critical inhibitory role in both pathways: it is both the essential cofactor for GPX4 to function and inhibit lipid peroxidation, and an important buffer that chelates excess copper ions to mitigate their toxicity [36, 68]. Therefore, any factor that leads to GSH depletion will simultaneously weaken the cell's resistance to both forms of cell death [36, 69]. Mitochondria serve as the second key platform for ferroptosis–cuproptosis crosstalk, acting as a "reactor" where death signals from both pathways are mutually amplified and transmitted [41, 43, 70]. This crosstalk constitutes a positive feedback loop rather than a simple summation. The core event of cuproptosis is the aggregation of lipoylated proteins, such as DLAT, which directly leads to the stalling of the TCA cycle and the degradation of Fe-S cluster proteins [21]. The degradation of Fe-S cluster proteins, in turn, directly releases their iron ions, expanding the labile iron pool, and causes the collapse of ETC function, leading to massive ROS production and providing a strong oxidative drive for lipid peroxidation [61, 71]. Conversely, the massive lipid peroxidation that occurs during ferroptosis severely damages mitochondrial membrane integrity, leading to mitochondrial dysfunction. This damage makes the mitochondria more sensitive to the toxic effects of copper ions [26].

These two death pathways also share some upstream master regulatory transcription factors, such as NRF2 and p53, whose activity status can simultaneously determine the cell's sensitivity to both death modalities [72, 73]. NRF2 is the central regulator of the cellular antioxidant stress response [74]. Regarding ferroptosis, NRF2 directly regulates the expression of SLC7A11, GPX4, and ferritin (FTH1). For cuproptosis, NRF2's target genes also include enzymes required for GSH synthesis and metallothioneins (MTs), which can directly chelate copper ions. Thus, NRF2 activation confers dual resistance to both ferroptosis and cuproptosis. In contrast, p53 primarily plays a pro-death role [73]. p53 can promote ferroptosis by inhibiting the transcription of SLC7A11, and the downregulation of SLC7A11 also reduces GSH synthesis, thereby indirectly weakening the cell's defense against cuproptosis [75]. Furthermore, autophagy plays a complex dual role in the crosstalk between these two death pathways [54, 76]. Ferritinophagy is a key upstream initiator of ferroptosis [34], while mitophagy, by clearing damaged mitochondria, can mitigate oxidative stress and thus may inhibit the progression of both ferroptosis and cuproptosis [43]. This complex interplay lays the foundation for a synergistic "dual-strike" strategy, which is illustrated in Fig. 3 of this review.

Ultimately, a crucial functional intersection of these two death modalities is their ability to induce immunogenic cell death (ICD), thereby converting an intrinsic cellular death signal into an extrinsic anti-tumor immune response [77]. When tumor cells undergo ferroptosis or cuproptosis, they release a series of damage-associated molecular patterns (DAMPs), such as high mobility group box 1 (HMGB1), ATP, and calreticulin (CRT) [78]. These DAMPs act as "danger signals" that can be recognized by the immune system, leading to the recruitment and activation of dendritic cells (DCs), promoting their maturation and antigen presentation [79, 80]. Mature DCs then activate cytotoxic T lymphocytes (CTLs), ultimately triggering a specific immune attack against the tumor [44]. Therefore, inducing ferroptosis or cuproptosis can not only directly kill tumor cells but also, through an "in situ vaccine" effect, transform immunologically "cold" tumors into "hot" tumors infiltrated by immune cells [81]. This process, which links cell death to anti-tumor immunity, is depicted in Fig. 4 of this review. To more clearly distinguish between these death modalities, a comparison of the key features of ferroptosis, cuproptosis, and other relevant RCD pathways is provided in Table 1 [52, 82–84].

To meet the demands of uncontrolled proliferation and adapt to the harsh tumor microenvironment, gastrointestinal (GI) tumor cells systematically remodel their metabolic networks [23, 85]. These metabolic adaptations, mediated by oncogenic signaling or microenvironmental pressures, confer a survival advantage but also create therapeutically exploitable 'metabolic vulnerabilities' [29, 86]. A dependency on specific metabolic pathways characterizes such vulnerabilities. Disruption of these pathways compromises cellular homeostasis and sensitizes cells to regulated cell death, including ferroptosis or cuproptosis [17]. This section will conceptually elaborate on the common vulnerability axes in GI tumors related to lipid, amino acid, metal ion, and energy metabolism, as well as their antioxidant defense systems, laying the theoretical groundwork for the subsequent in-depth discussion of specific molecular mechanisms in each cancer type.

The clinical application of therapies targeting ferroptosis and cuproptosis requires a context-specific understanding that extends beyond common metabolic vulnerabilities. The efficacy of inducing these death pathways is determined by the distinct molecular landscapes, oncogenic drivers, and tumor microenvironments of each GI cancer type. This section outlines the specific regulatory networks of ferroptosis and cuproptosis in major gastrointestinal malignancies, aiming to identify disease-specific pathways and therapeutic targets. The anatomical sequence of the gastrointestinal tract organizes the content.

Small-molecule compounds are the cornerstone tools for exploring and inducing ferroptosis and cuproptosis. They precisely regulate the initiation and execution of cell death by targeting key proteins in the core pathways.

The induction of ferroptosis is primarily achieved through two classic routes: inhibiting the cystine/glutamate antiporter System Xc⁻ or directly inhibiting glutathione peroxidase 4 (GPX4) [137]. Erastin, the first discovered ferroptosis inducer, blocks the function of System Xc⁻, leading to impaired cellular cystine uptake, which in turn depletes glutathione (GSH). This ultimately inactivates GPX4, causing the accumulation of lipid peroxides [36, 138]. Some clinically approved drugs have also been shown to exhibit a similar mechanism of action. For example, the anti-inflammatory drug sulfasalazine (SSZ) effectively inhibits System Xc⁻. In colorectal cancer (CRC) models, it not only induces ferroptosis but also enhances the cytotoxicity of cisplatin by depleting GSH [35]. As a clinically long-used anti-inflammatory drug, the favorable safety profile of sulfasalazine makes it a promising "old drug, new use" candidate. Clinical trials are currently evaluating its application in cancer therapy. Another class of ferroptosis inducers directly targets GPX4, with RSL3 (RAS-selective lethal 3) being the most representative. It covalently binds to the active site of GPX4, directly inactivating it and thereby efficiently inducing ferroptosis [139]. Furthermore, numerous natural products have been discovered to possess ferroptosis-inducing potential. For instance, β-Lapachone induces ferroptosis in CRC cells by activating the JNK pathway, which promotes NCOA4-mediated ferritinophagy and increases the intracellular labile iron pool [140]. The metabolite γ-linolenic acid from Lactobacillus plantarum triggers ferroptosis in CRC cells by damaging mitochondria [101]. A more detailed summary of these small-molecule compounds, which regulate cell death by targeting core pathways, along with their mechanisms and applications in gastrointestinal cancers, is provided in Table 2.

The induction of cuproptosis primarily relies on the excessive accumulation of intracellular copper ions, particularly their toxic effects within the mitochondria [103]. Elesclomol is a potent copper ionophore that forms a complex with copper ions, facilitating their transport across membranes and specific accumulation in the mitochondria [153]. Excess copper ions in the mitochondria directly bind to lipoylated proteins of the tricarboxylic acid (TCA) cycle, such as the E2 subunit of the pyruvate dehydrogenase complex (DLAT), causing these proteins to aggregate and lose function while also disrupting the stability of iron-sulfur cluster proteins. This triggers proteotoxic stress, ultimately leading to cell death [49].

Among small molecules, the combination of disulfiram (DSF) and copper (DSF-Cu) exemplifies a dual-induction strategy, demonstrating a powerful potential to trigger both ferroptosis and cuproptosis [154]. DSF itself is a copper ionophore that efficiently delivers copper ions into the cell, thereby initiating the cuproptosis program [153]. More critically, DSF and its metabolites can effectively deplete the cell's GSH reserves [138]. GSH depletion acts as a central hub connecting the two death modalities in this process. On one hand, the decline in GSH levels weakens the cell's ability to chelate and detoxify copper ions, greatly enhancing the effect of cuproptosis [153]. On the other hand, GSH depletion directly leads to the inactivation of GPX4, rendering the cell unable to eliminate lipid peroxides and thus initiating the ferroptosis program [61]. This mechanistic synergy, which involves simultaneously striking two key cellular pathways with a single drug combination, not only enhances anti-tumor efficacy but also offers a new approach to overcoming the adaptive resistance that may arise from targeting a single pathway [115].

Although small-molecule compounds have shown great potential in mechanistic studies and preliminary applications, their limitations, including solubility, targeting specificity, and bioavailability, have become increasingly apparent. To overcome these obstacles, nanotechnology has provided revolutionary solutions for achieving efficient and precise drug delivery.

Nanotechnology provides platforms for the delivery of ferroptosis and cuproptosis inducers. Through specific engineering, nanodelivery systems can address the limitations of small-molecule drugs, including issues with solubility, bioavailability, and targeting [155]. The clinical translation of these nanoplatforms, however, is contingent on a critical evaluation of their design and performance, as existing systems face several challenges.

In terms of targeting specificity, although the enhanced permeability and retention (EPR) effect provides a theoretical basis for passive targeting of nanomedicines, its significant heterogeneity across different tumor types, and even within the same tumor, significantly limits its universality and efficiency [156]. The dense extracellular matrix (ECM) and high interstitial fluid pressure, characteristic features of solid tumors (especially pancreatic cancer), constitute physical barriers that hinder the effective penetration and uniform distribution of nanoparticles [157]. To overcome this limitation, active targeting strategies have emerged, involving the modification of nanocarrier surfaces with specific ligands to enhance binding to tumor cells. However, the heterogeneous expression of target receptors poses a challenge that active targeting strategies must address. Regarding drug-loading capacity, different platforms vary significantly. Platforms with high porosity, such as metal–organic frameworks (MOFs), exhibit excellent drug-loading capabilities [158], whereas liposomes and polymer micelles may face challenges with loading efficiency [155]. The controllability of the release mechanism is key to determining the efficacy of nanomedicines. An ideal release mechanism should respond to specific signals in the tumor microenvironment (TME), such as low pH or high GSH concentration, to achieve "smart" drug release [159], but precise regulation of release kinetics remains challenging. Biocompatibility and potential toxicity are core considerations for the clinical translation of all nanomaterials. While many organic materials (such as lipids and polymers) have good biocompatibility, the long-term safety of inorganic materials (especially nanozymes or MOFs containing heavy metals) requires careful evaluation, as metal ion leakage can cause systemic toxicity [160]. Finally, the influence of the TME itself on the efficacy of nanotherapeutics cannot be ignored. For example, high levels of GSH in the TME not only serve as a key defense line for tumor cells against oxidative stress and ferroptosis but can also directly chelate and inactivate specific metal ions (such as Cu²⁺), thereby weakening the effectiveness of therapeutic strategies based on metal-catalyzed reactions [161]. Therefore, a successful nanomedicine design must consider the complexity of the TME as both a barrier and a trigger to be exploited.

To address the aforementioned challenges, the field has rapidly evolved from simple drug carriers to intelligent, multifunctional theranostic platforms. These next-generation systems not only aim to deliver payloads but also to actively sense and respond to the unique biochemical landscape of the TME, act as catalysts to amplify therapeutic effects in situ, and even integrate with biological systems to create complex bio-hybrid therapies. This section will systematically elaborate on the design strategies and latest advancements in nanodelivery systems for inducing ferroptosis, cuproptosis, and their synergistic effects.

Inducing ferroptosis and cuproptosis can not only directly kill tumor cells but also produce powerful synergistic effects with traditional anticancer treatments, such as immunotherapy, radiotherapy, and chemotherapy, by modulating the tumor microenvironment and cellular signaling pathways [192]. This synergy is primarily mediated by the induction of immunogenic cell death (ICD), which can convert immunologically non-responsive tumors into tumors with active immune cell infiltration [193].

In synergy with immunotherapy, dying tumor cells undergoing ferroptosis or cuproptosis release a series of damage-associated molecular patterns (DAMPs), including ATP, calreticulin (CRT), and high-mobility group box 1 (HMGB1) [194, 195]. These DAMPs act as "danger signals" that are recognized by antigen-presenting cells (APCs), particularly dendritic cells (DCs), thereby promoting DC maturation and their antigen-presenting capacity [196]. Mature DCs then migrate to lymph nodes, where they effectively present tumor antigens to naive T cells, activating and expanding tumor-specific cytotoxic T lymphocytes (CTLs) [197]. These activated CTLs subsequently infiltrate the tumor tissue and specifically kill tumor cells. Therefore, initiating ICD by inducing ferroptosis or cuproptosis can effectively enhance the efficacy of immune checkpoint blockade (ICB), such as anti-PD-1/PD-L1 antibodies [198]. For example, in a colorectal cancer model, ferroptosis induced by the natural product Icariin (ICA) activated CD8 + T cells to secrete IFN-γ, thereby synergistically enhancing the effect of anti-PD-1 therapy [148]. Similarly, in hepatocellular carcinoma, a nanoplatform that induces cuproptosis also promoted the infiltration of CTLs, showing significant anti-tumor synergy when combined with anti-PD-1 therapy [196].

In synergy with radiotherapy (RT), the amplification of oxidative stress is the key mechanism [199]. Radiotherapy itself generates a large amount of ROS in cells through ionizing radiation, which damages DNA and induces cell death. However, tumor cells often possess robust antioxidant systems that enable them to withstand this damage [108]. Ferroptosis or cuproptosis inducers severely weaken the antioxidant defenses of tumor cells by inhibiting GPX4 or depleting GSH, allowing the ROS generated by radiotherapy to cause more lasting and extensive damage, thereby significantly enhancing the killing effect of RT [112]. Studies have shown that radiotherapy itself can induce ferroptosis by downregulating the expression of SLC7A11. This effect can be amplified by IFN-γ, providing a new molecular link for the synergy between radiotherapy and immunotherapy [200]. Furthermore, radiotherapy can directly induce cuproptosis by upregulating the expression of copper transporters (like CTR1) and increasing mitochondrial copper ion concentration. This provides a direct molecular basis for the combined use of radiotherapy and cuproptosis inducers (like Elesclomol) and has been confirmed to have a sensitizing effect in an esophageal squamous cell carcinoma model [201].

Synergy with chemotherapy mainly focuses on overcoming drug resistance. The resistance mechanisms to many chemotherapy drugs are closely related to the enhancement of the antioxidant capacity of tumor cells [103]. For instance, in colorectal cancer, cells that have developed resistance to platinum-based drugs, such as cisplatin or oxaliplatin, often exhibit higher levels of GSH and more vigorous GPX4 activity [35, 202]. By using ferroptosis inducers such as SSZ or RSL3, this resistant phenotype can be effectively reversed, thereby restoring the sensitivity of tumor cells to chemotherapy drugs [35]. In gastric cancer, a traditional Chinese medicine formula can reverse cisplatin resistance by inducing ferroptosis via the AKT/GSK3β/NRF2/GPX4 axis [93]. In pancreatic cancer, knocking out the ACOT8 gene promotes ferroptosis and increases the tumor's chemosensitivity to gemcitabine [118]. This strategy, by targeting the metabolic reprogramming that tumor cells undergo to adapt to chemotherapy stress, offers a new avenue for treating recurrent and refractory tumors.

The clinical translation of therapies targeting ferroptosis and cuproptosis in gastrointestinal (GI) malignancies is in early stages, despite extensive preclinical evidence. Current clinical trials are proceeding through two main strategies: repurposing existing drugs with established safety profiles and developing novel agents specifically designed to modulate these cell death pathways. This approach combines the use of established pharmaceuticals with investment in new, mechanism-driven therapeutics.

The drug repurposing strategy has provided the earliest clinical insights. Sorafenib, a multikinase inhibitor approved for hepatocellular carcinoma (HCC), was retrospectively found to induce ferroptosis by inhibiting System Xc⁻. This dual mechanism has prompted further investigation, such as in trial NCT02989870↗, which explored its synergy with stereotactic body radiation therapy (SBRT) in HCC, and NCT05068752↗, which is evaluating its combination with vemurafenib in KRAS-mutant pancreatic cancer. Similarly, sulfasalazine, a long-standing anti-inflammatory drug and a canonical System Xc⁻ inhibitor, is being formally tested in combination with standard chemotherapy for metastatic colorectal cancer (NCT06134388↗). Another notable example is the antimalarial drug artesunate, which induces ferroptosis by generating reactive oxygen species (ROS). The ongoing NeoART trial (NCT02633098↗) is strategically significant as it assesses artesunate in the neoadjuvant setting for colorectal cancer, aiming to improve surgical outcomes and reduce recurrence by targeting the disease at an earlier stage. In the realm of cuproptosis, the combination of the alcohol-aversion drug disulfiram with copper has been the cornerstone of clinical investigation. The Phase 1 trial NCT00742911↗, the foundational trial, established the safety and tolerability of this combination in patients with liver metastases but observed no objective responses, highlighting the central challenge of translating this strategy into potent clinical efficacy. Subsequent studies, such as NCT03714555↗ in metastatic pancreatic cancer, have continued to explore this combination in specific GI tumor contexts.

To overcome the potential limitations in potency and specificity of repurposed drugs, the field is increasingly focusing on novel and targeted therapeutic strategies. These approaches aim to engage the core machinery of ferroptosis and cuproptosis more directly and powerfully. One prominent example is eprenetapopt (APR-246), a novel small molecule that not only reactivates mutant p53 but also potently depletes cellular glutathione (GSH), thereby inducing ferroptosis robustly. Its investigation in the NCT04383938↗ trial, which combines eprenetapopt with the immune checkpoint inhibitor pembrolizumab and includes an expansion cohort for advanced gastric cancer, exemplifies a modern strategy to test the synergy between ferroptosis-induced immunogenic cell death and immunotherapy. A more direct approach to modulating the GSH axis is demonstrated in trial NCT04205357↗, which combines buthionine sulfoximine, a specific inhibitor of GSH synthesis, with gemcitabine in patients with pancreatic ductal adenocarcinoma. Perhaps the most forward-looking developments are in the realm of purpose-built nanomedicine. These platforms are engineered to address the fundamental challenge of delivering therapeutic metal ions, such as iron, specifically to tumors while minimizing systemic toxicity. A landmark study in this area is the recently completed Phase 1 trial NCT06048367↗, which evaluated the safety and preliminary efficacy of intratumoral injection of Carbon Nanoparticle-Loaded Iron in patients with advanced solid tumors, including various GI cancers. This trial represents a critical first-in-human validation of a nanomedicine specifically designed to induce ferroptosis, marking a pivotal step from preclinical concept to clinical reality. The current landscape of representative clinical trials investigating these strategies in gastrointestinal cancers is summarized in Table 4.

Patient stratification based on the expression profiles of cuproptosis- and ferroptosis-related genes is being used to investigate the heterogeneity of GI tumors. Unsupervised consensus clustering of cuproptosis-related genes (CRGs) has been employed to identify previously unrecognized tumor subtypes. These subtypes are not randomly distributed but show consistent and significant associations with distinct clinical outcomes and TME characteristics [203–205].

In gastric cancer (GC), clustering analysis based on CRG expression profiles has successfully identified multiple molecular subtypes. Crucially, these subtypes are closely related to prognosis; for instance, studies have found that subtypes characterized by high expression of CRGs paradoxically have a better prognosis [206, 207]. This finding challenges the simple assumption that an active cell death pathway is always beneficial for tumor suppression, suggesting that the context and specific mechanisms of cuproptosis activation are critical. Similarly, in colorectal cancer (CRC), two distinct cuproptosis-related molecular subtypes have been identified, which show significant differences in clinicopathological features, overall survival, and TME activity [205, 208]. One study directly linked three cuproptosis patterns to three distinct immune infiltration phenotypes, namely immune-desert, immune-inflamed, and immune-excluded, thereby tightly connecting cuproptosis regulation with the tumor's immune microenvironment [208].

Even in pancreatic ductal adenocarcinoma (PDAC), known for its extreme heterogeneity and therapeutic resistance, CRG-based clustering analysis has identified three unique cuproptosis subtypes. These subtypes exhibit significant differences in patient prognosis and TME cell infiltration, highlighting the immense potential of cuproptosis-based stratification strategies even in the most challenging gastrointestinal malignancies [204]. Research in cholangiocarcinoma (CCA) has also found two CRG-related clusters with opposing survival times, further validating the general applicability of this method across multiple gastrointestinal tumors [209].

A recurring and compelling theme in these findings is the "prognostic paradox" of cuproptosis activity. Intuitively, high expression of CRGs might imply a greater potential for cuproptosis, which should lead to more tumor cell death and a better prognosis. However, multiple studies consistently show that high expression levels of CRGs are associated with better patient survival [206, 207]. There may be a more profound biological logic behind this phenomenon. High CRG expression may not represent a continuously active cell death process but rather reflect a specific metabolic state that is highly dependent on mitochondrial respiration. This metabolic phenotype itself may be less aggressive or have a weaker immune escape capability, leading to a better clinical outcome independent of the actual cuproptosis induction process. Another possibility is that this "primed-for-death" state makes the tumor extremely sensitive to endogenous copper fluctuations or therapeutic interventions, ultimately leading to its effective control.

Furthermore, molecular subtyping has become a bridge connecting cell death and tumor immunology. The consistent association between cuproptosis-defined subtypes and the TME landscape [205, 208] implies that the regulation of copper metabolism is not an isolated cellular process; it is fundamentally linked to the mechanisms that shape tumor-immune interactions. The molecular machinery that regulates cuproptosis likely determines the tumor's potential for immunogenic cell death (ICD). A subtype with high expression of key cuproptosis execution proteins (such as FDX1, DLAT) may be more prone to ICD when stressed, thereby shaping an "immune-inflamed" microenvironment over time. Conversely, a subtype that has evolved to suppress the expression of these genes would be resistant to ICD, ultimately forming an "immune-desert" phenotype. Therefore, cuproptosis subtyping is not only a prognostic tool but also a functional interpretation of the tumor's immunogenic potential.

Multi-gene prognostic signature models are being constructed as an extension of molecular subtyping. These models utilize machine learning algorithms to generate a quantitative risk score from gene expression data, enabling the prediction of patient outcomes. A common framework involves an initial screen for prognosis-related genes using univariate Cox regression, followed by variable selection with the least absolute shrinkage and selection operator (LASSO) regression, and final model construction through multivariate Cox analysis [210–212]. This rigorous statistical approach aims to prevent model overfitting and enhance its generalizability.

Signature models based on protein-coding genes have been validated in various gastrointestinal tumors. In cholangiocarcinoma (CCA), a 4-gene signature (ATP7A, FDX1, DBT, LIAS) was established as an independent prognostic factor [212], while another study constructed a signature model containing 10 genes [213]. In colorectal cancer (CRC), multiple models have also been developed, including a 5-gene signature [214] and a 12-gene signature [215]. In gastric cancer (GC), a 7-gene prognostic model was established using the random forest method [216].

A significant and recent development is the shift in research focus towards constructing models based on CRLs. Due to their high stability and tissue specificity, lncRNAs may offer superior potential as biomarkers. In colon adenocarcinoma (COAD), multiple studies have successfully constructed prognostic signatures based on CRLs, identifying models containing 5, 13, and 15 lncRNAs, respectively [211, 217, 218]. These models consistently demonstrate high predictive accuracy (area under the receiver operating characteristic curve, AUC values typically > 0.75) and can serve as independent prognostic factors[211, 219]. In pancreatic cancer (PAAD), a 5-lncRNA signature model was validated in both the TCGA and ICGC cohorts [220]. Other studies have further confirmed the value of CRLs in prognostic assessment [221]. In esophageal cancer (ESCA), a 6-CRL model was constructed to predict prognosis [222].

Some of the most innovative models integrate genes from multiple cell death pathways or other related biological processes. In esophageal squamous cell carcinoma (ESCC), a prognostic model integrating cuproptosis, ferroptosis, and immune-related genes has been constructed, thereby creating a more comprehensive signature that reflects the tumor's cell death and immune landscape [223, 224]. In colorectal cancer, one study developed a signature model based on cuproptosis-related RNA methylation regulators, directly linking RNA epigenetics with cuproptosis and prognosis [225].

Research has increasingly shifted from protein-coding genes to their regulators, such as cuproptosis-related lncRNAs (CRLs). This focus is based on the biological function of lncRNAs as upstream regulators that can control the expression of multiple protein-coding genes. Consequently, signature models based on lncRNAs may offer greater robustness compared to those based on downstream effector genes.

Although the various signature models in different cancer types contain different genes, they all functionally converge on a "tumor vulnerability index." A high-risk score always predicts a poor prognosis [211, 214, 226]. These risk models are not merely measuring the expression of random genes but are quantifying a core biological property of the tumor. A "high-risk" score may reflect that the cell has successfully suppressed endogenous cell death pathways (such as cuproptosis and ferroptosis). This suppression is a prerequisite for the tumor to achieve aggressive behaviors (such as proliferation, invasion, and therapeutic resistance). Therefore, the risk score can be seen as a proxy for the overall resilience and adaptability of the tumor, quantifying the success of cancer cells in evading death by reprogramming their metabolism, which is intrinsically linked to a more malignant phenotype and poorer prognosis. To systematically present these findings, the following table summarizes representative cuproptosis- and ferroptosis-related prognostic models in gastrointestinal tumors.

To visually summarize the clinical translational blueprint for targeting these emerging cell death pathways in gastrointestinal cancers, we present Fig. 5.

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The author declares no competing financial interests.

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