Role of PANoptosis in cancer: Molecular mechanisms and therapeutic insights

Role of PANoptosis in cancer: Molecular mechanisms and therapeutic insights | Authorea try { document.documentElement.classList.add('js'); } catch (e) { } var _gaq = _gaq || []; _gaq.push(['_setAccount', 'G-8VDV14Y67G']); _gaq.push(['_trackPageview']); (function() { var ga = document.createElement('script'); ga.type = 'text/javascript'; ga.async = true; ga.src = ('https:' == document.location.protocol ? 'https://ssl' : 'http://www') + '.google-analytics.com/ga.js'; var s = document.getElementsByTagName('script')[0]; s.parentNode.insertBefore(ga, s); })(); Skip to main content Preprints Collections Wiley Open Research IET Open Research Ecological Society of Japan All Collections About About Authorea FAQs Contact Us Quick Search anywhere Search for preprint articles, keywords, etc. Search Search ADVANCED SEARCH SCROLL This is a preprint and has not been peer reviewed. Data may be preliminary. 15 November 2025 V1 Latest version Share on Role of PANoptosis in cancer: Molecular mechanisms and therapeutic insights Authors : Yi Zhang [email protected] , Xue-Li Wang , Yan-Wen Wang , Ti Chu , Lei Cao , Yong-Qi Fan , Yu-Hang Chen , Wei-Rong Si , Qi-Ying Jiang , and Dongdong Wu 0000-0001-6739-8437 Authors Info & Affiliations https://doi.org/10.22541/au.176320286.60816100/v1 229 views 109 downloads Contents Abstract Supplementary Material Information & Authors Metrics & Citations View Options References Figures Tables Media Share Abstract Abnormal cell death plays a crucial role in cancer development. Inducing tumor cell death is an important strategy for cancer treatment. Against this backdrop, with the in-depth study of the regulatory mechanisms of cell death, the discovery of interactions existing among pyroptosis, apoptosis, and necroptosis has resulted in the emergence of the concept of PANoptosis. This review article summarizes the discovery and molecular mechanisms of PANoptosis, as well as its roles in cancer development, tumor microenvironment regulation, and tumor metastasis, and explores its application prospects in cancer treatment. PANoptosis can promote anti-tumor immunity, but it has a dual-edged role in tumor metastasis. Targeted therapies against its key signaling molecules and combination therapies have shown promising results. For example, inhibiting the activity of ZBP1 can reduce the side effects of chemotherapy, and the combination of multiple molecules with radiotherapy or immunotherapy can enhance the therapeutic effect. However, in-depth research is still needed on the interaction and regulation mechanisms of its signaling pathways, as well as the activation mechanisms and functions of some molecules. Moreover, relevant targeted therapies and combination therapy regimens require more clinical trials for verification. In summary, PANoptosis provides a new perspective for tumor research and is expected to promote the development of precision medicine for tumors. 1. Introduction In the process of cancer progression, irregularities in cell death occur., such as the obstruction of apoptosis, will disrupt the balance between cell division and death, leading to excessive cell proliferation. At the same time, tumor cells evade death and continue to proliferate by inhibiting the cell death pathways, thus promoting the development of cancer [1, 2]. In cancer treatment, inducing the death of tumor cells is an important therapeutic strategy, which includes activating different pathways such as apoptosis, pyroptosis, autophagy, and ferroptosis [3, 4]. With the revelation of more complex regulatory mechanisms controlling cell death, the concept of PANoptosis has emerged [5]. For a long time, the pathways of pyroptosis, apoptosis, and necroptosis have been considered to operate in parallel with little overlap [6]. However, the interactions between pyroptosis and apoptosis, between apoptosis and necroptosis, and between pyroptosis and necroptosis have now been clearly defined. Moreover, an expanding group of investigators have elucidated the interactions among these three cell death modalities in terms of their mechanisms, and these findings have contributed to the establishment of the concept of PANoptosis [7]. 2. Discovery and Molecular Mechanism of PANoptosis 2.1 Discovery of PANoptosis As early as 2016, investigates had revealed the crucial role of Z-DNA binding protein 1 (ZBP1) as a key innate immune sensor during influenza A virus (IAV) infection. It triggers cell death and an inflammatory response through the RIPK1-RIPK3-Caspase-8 axis [8]. The absence of ZBP1 endows cells with resistance to IAV-induced cell death, reduces the inflammatory response and epithelial damage, and safeguards mice from death caused by IAV infection. ZBP1 interacts with the nucleoprotein (NP) and polymerase basic protein 1 (PB1) proteins of IAV, Initiates the activation process of the NOD-like receptor protein 3 (NLRP3) inflammasome and enables the discharge of inflammatory cytokines including interleukin-1β (IL-1β) and interleukin-18 (IL-18), and coordinates multiple cell death pathways, including pyroptosis (P), apoptosis(A), and necroptosis(N), these pathways are key characteristics of PANoptosis [9, 10]. After ZBP1 senses IAV infection and activates the NLRP3 inflammasome and PANoptosis, the deletion or functional inactivation of TAK1 leads to the activation of multiple forms of cell death. The inhibitory phosphorylation of receptor-interacting protein kinase 1 (RIPK1) by TAK1 is crucial for restricting its activation and preventing the spontaneous activation of PANoptosis [11, 12]. Additionally, it has been discovered that the inhibition of TAK1 can directly activate cysteine-aspartic protease 8 (Caspase-8), drive the cleavage of gasdermin D (GSDMD), and induce pyroptosis, independent of Cysteine-aspartic protease 1 (Caspase-1) (Fig.1). These findings provide an important theoretical foundation and potential therapeutic targets for future research [13, 14]. 2.2 Molecular Mechanism of PANoptosis 2.2.1 Activation Mechanism of the Inflammasome The inflammasome is a multimeric complex composed of various proteins and serves as a crucial platform for initiating PANoptosis [15]. Taking the NLRP3 inflammasome as an example, comprising the NLRP3 protein, the adaptor protein ASC (apoptosis-associated speck-like protein with a CARD), and the Caspase-1 precursor [16] . The NLRP3 inflammasome becomes active when cells are subjected to numerous stimuli. These stimuli include lipopolysaccharide (LPS) from bacteria, the leakage of intracellular potassium ions, and the release of mitochondrial DNA (mtDNA) due to mitochondrial damage, the NLRP3 inflammasome can be activated [17]. Firstly, pathogen-associated molecular patterns (PAMPs) like LPS can activate the Nuclear factor kappa-light-chain-enhancer of activated B (NF-κB) signaling pathway through Toll-like receptors (TLRs), leading to an upregulation of the expression of inflammation-related proteins such as NLRP3 and pro-IL-1β [18, 19]. After that, particular danger signals within the cell, for instance the efflux of potassium ions, trigger a change in the conformation of the NLRP3 protein, thus bringing about its oligomerization. The oligomerized NLRP3 interacts with the PYD-PYD and CARD-CARD domains of ASC, it attracts the precursor of Caspase-1 to assemble and form an active NLRP3 inflammasome complex [20, 21]. In addition to the NLRP3 inflammasome, the activation of the NLRP1 inflammasome differs from that of NLRP3. The NLRP1 inflammasome contains a unique FIIND domain. When infected by certain pathogens, the FIIND domain can be cleaved by specific proteases, thus activating the NLRP1 inflammasome. The absent in melanoma 2 (AIM2) inflammasome, on the other hand, recognizes double-stranded DNA in the cytoplasm through its HIN domain, following that, it calls in ASC and the Caspase-1 precursor, leading to their assembly and the formation of an active inflammasome [22, 23]. 2.2.2 The Pyroptosis Pathway Upon the inflammasome activating Caspase-1, the activated form of Caspase-1 will selectively cleave GSDMD [18]. GSDMD is a protein containing an N-terminal domain and a C-terminal domain. After Caspase-1 cleaves GSDMD [24], the N-terminal domain with membrane-pore-forming activity is released [25, 26]. The N-terminal domains can oligomerize and enter the cell membrane, resulting in the formation of pores with an approximate diameter of 10 to 14 nanometers. After the formation of pores in the cell membrane, the ion balance within the cell is disrupted. A large amount of water enters the cell, leading to cell swelling and rupture, which are typical morphological changes of pyroptosis [27]. In the meantime, cell-contained inflammatory cytokines, for example, IL-1β and IL-18, which originally exist in an inactive precursor form, are cleaved into their active, mature forms under the action of Caspase-1. These active cytokines are then released extracellularly through the pores formed by GSDMD, triggering a robust inflammatory response [28, 29]. 2.2.3 The Apoptosis Pathways 2.2.3.1 The Intrinsic Apoptosis Pathway Once cells face internal stress signals, including oxidative stress and DNA damage, the outer-membrane permeability of mitochondria is modified. For instance, during oxidative stress, the synthesis of reactive oxygen species (ROS) in mitochondria elevates, leading to a decrease in the mitochondrial membrane potential [30]. At this time, the proteins of the B-cell lymphoma-2 (Bcl-2) family located on the inner mitochondrial membrane have a significant regulatory function [31]. Pro-apoptotic proteins like Bcl-2-associated X protein (Bax) and Bcl-2 homologous antagonist/killer (Bak) are capable of forming oligomers on the outer mitochondrial membrane. This leads to the opening of the mitochondrial membrane permeability transition pore (MPTP), this enables cytochrome c to be discharged from the mitochondria into the cytoplasm [32]. The cytochrome c that gets into the cytoplasm combines with apoptotic protease activating factor-1 (Apaf-1), and simultaneously binds to adenosine triphosphate (ATP) to form a heptameric apoptosome. The apoptosome recruits and activates the precursor of Caspase-9, leading to its self-cleavage and activation [33, 34]. The activated Caspase-9 then activates downstream effector caspases, such as Caspase-3, Caspase-6, and Caspase-7. These effector caspases can cleave a variety of intracellular substrates, such as poly (ADP-ribose) polymerase (PARP), etc. Ultimately, this results in apoptosis, manifested by morphological changes such as nuclear pyknosis, chromatin condensation, and membrane blebbing [35]. ¿p#1 2.2.3.2 Extrinsic Apoptosis Pathway When death receptors such as Fas cell surface death receptor (Fas) and tumor necrosis factor receptor 1 (TNFR1) located on the cell surface attach to their respective ligands, a sequence of signal transduction events is triggered [36]. Take Fas as an example. Once Fas ligand (FasL) binds to Fas, the intracellular death domain (DD) of Fas aggregates, attracting the adapter protein FADD, which contains a death effector domain (DED) [37]. Through DED-DED interactions, FADD recruits the precursor of Caspase-8, resulting in the assembly of a death-inducing signaling complex (DISC) [38, 39]. Within the DISC, the precursor of Caspase-8 undergoes self-cleavage and activation. The activated Caspase-8 is capable of directly activating downstream effector caspases, such as Caspase-3, thereby triggering apoptosis. In certain scenarios, Caspase-8 can also cleave the Bid protein, converting it into tBid. tBid can then translocate to the mitochondria, activating the intrinsic apoptosis pathway and further amplifying the apoptotic signal [40, 41]. 2.2.4 The Necroptosis Pathway Necroptosis is primarily triggered by elements within the tumor necrosis factor (TNF) superfamily. As an illustration, in the case of TNFR1 activation, when TNF attaches to TNFR1, the intracellular portion of TNFR1 attracts the tumor necrosis factor receptor-associated death domain protein (TRADD, TNF receptor-associated death domain). Following this, TRADD recruits RIPK1, giving rise to the formation of complex I [42, 43]. Within complex I, RIPK1 undergoes phosphorylation. Subsequently, RIPK1 dissociates from complex I and associates with receptor-interacting protein kinase 3 (RIPK3) and Fas-associated death domain protein (FADD), constructing complex IIb, and it is also named the necrosome [44, 45]. In the necrosome, RIPK3 interacts with RIPK1 via its receptor-interacting protein homotypic interaction motif (RHIM) domain and is activated through phosphorylation by RIPK1 [46]. The activated RIPK3 goes on to phosphorylate the mixed lineage kinase domain-like protein (MLKL) further. Once phosphorylated, MLKL forms oligomers and migrates from the cytoplasm to the cell membrane. The oligomerized MLKL embeds into the cell membrane, disturbing the membrane integrity, thereby causing the outflow of intracellular substances (Fig 2). As a result, the cell experiences necroptosis and displays morphological alterations like cell swelling and cell membrane rupture [47, 48]. ¿p#1 2.2.5 Regulation of Molecular Interactions in PANoptosis Protein kinase R (PKR) plays a crucial role during viral infection. Once a cell is infected by a virus, double-stranded RNA (dsRNA) is produced, which can activate PKR [49]. The activated PKR undergoes autophosphorylation and subsequently phosphorylates eukaryotic translation initiation factor 2α (eIF2α) [50]. The phosphorylation of eIF2α hinders the initiation of protein synthesis, thereby suppressing the synthesis of viral proteins. Meanwhile, PKR is also capable of triggering the NF-κB signaling pathway. This activation facilitates the expression of inflammatory cytokines including IL-6 and TNF-α, and PKR takes part in the PANoptosis process [51, 52]. The mitogen-activated protein kinase (MAPK) group, comprising extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK, also contributes to the regulation of PANoptosis [41, 53]. For instance, under inflammatory stimuli, JNK and p38 MAPK can be activated. They can phosphorylate and activate transcription factors such as c-Jun and ATF2, regulate the expression of related genes, promote apoptosis and inflammatory responses, and thus participate in the process of PANoptosis (Fig.3). In contrast, in certain situations, ERK can inhibit the activity of apoptosis-related proteins through phosphorylation, exerting a negative regulatory effect on PANoptosis [54, 55]. 3. Functions of PANoptosis in Cancer 3.1 Enhancement of Antitumor Immunity PANoptosis is a programmed cell death process that integrates multiple forms of cell death, including apoptosis, pyroptosis, and necroptosis. In the realm of cancer, activating the PANoptosis signaling pathway can drive cancer cells to undergo a series of intricate biochemical and morphological transformations. These changes disrupt the normal structure and function of cancer cells, ultimately leading to their demise [56, 57] (Fig.1). For instance, specific stimuli can trigger the activation of relevant intracellular proteins, setting off an intracellular cascade reaction. This reaction gives rise to events such as cell membrane rupture, organelle damage, and DNA fragmentation, all of which culminate in inducing the death of cancer cells [57, 58]. When undergoing PANoptosis, cancer cells discharge numerous damage-associated molecular patterns (DAMPs). Examples of these DAMPs are high -mobility group box 1 protein (HMGB1) and ATP. Pattern recognition receptors (PRRs) present on the surface of immune cells are capable of identifying these DAMPs, consequently triggering the activation of innate immune cells, including dendritic cells and macrophages [59, 60]. Once activated, dendritic cells are able to engulf, process, and present tumor antigens to T lymphocytes. This action activates the T cell-mediated adaptive immune response, significantly enhancing the body’s capacity to identify and eliminate cancer cells, and thus establishing an effective antitumor immune defense [61]. ¿p#1 3.2 Involvement in the Regulation of the Tumor Microenvironment The tumor microenvironment serves as a crucial foundation for the growth, proliferation, and metastasis of cancer cells. PANoptosis can influence the tumor microenvironment in multiple ways. On one hand, the cytokines and chemokines released during the process of PANoptosis can regulate the recruitment and activation of immune cells, altering the composition and function of immune cells within the tumor microenvironment. This, in turn, affects tumor immune surveillance and immune escape. On the other hand, these released substances may also impact processes such as tumor angiogenesis and extracellular matrix remodeling, thus having an impact on the growth, invasive ability, and metastatic potential of cancer cells [58, 62]. PANoptosis can induce immunogenic cell death, releasing DAMPs that activate the immune system and enhance the antitumor immune response [63, 64]. For example, targeted therapies aimed at PANoptosis can boost antitumor immunity. In the tumor microenvironment, the death of tumor and immune cells mediated by PANoptosis has the potential to regulate the tumor microenvironment (TME), affecting the effectiveness of tumor immunotherapy [65]. The expression and activity of PANoptosis-related genes can change the immunogenicity of cancer cells and influence the functional state of immune cells, thereby regulating the immune balance within the tumor microenvironment [66]. For example, researchers have disclosed a substantial association between PANoptosis and the infiltration degrees of immune cells within the tumor microenvironment. These immune cells consist of dendritic cells (DCs) and natural killer cells (NKs), CD8+ T cells, and CD4+ T cells. Additionally, the expression profiles of PANoptosis-related genes are associated with the infiltration levels of immune cells in gastric cancer patients, suggesting that PANoptosis can stimulate a robust antitumor immune response [67, 68]. 3.3 Role in Tumor Metastasis PANoptosis plays a dual-sided role in tumor metastasis: (1) Upon activation, PANoptosis can recruit immune cells by releasing inflammasome-related cytokines like IL-1β and IL-18, thus strengthening the body’s antitumor immune response. This, in turn, inhibits the migration and invasion of cancer cells, impeding tumor metastasis [69]. Meanwhile, the cell death induced by PANoptosis directly reduces the number of cancer cells with metastatic potential [70]. (2) DAMPs released by PANoptosis, such as HMGB1, may create a pro-inflammatory microenvironment. This microenvironment provides a favorable condition for the survival and proliferation of cancer cells, along with the epithelial-mesenchymal transition (EMT). It heightens the migration and invasion capacities of cancer cells, thereby facilitating tumor metastasis. Moreover, the chronic inflammation triggered by persistent PANoptosis may induce angiogenesis, providing a pathway for the distant metastasis of cancer cells [71, 72] (Fig.4). 4. Application Prospects of PANoptosis in Cancer Treatment 4.1 Targeted Therapy of Key Signaling Molecules 4.1.1 ZBP1 ZBP1 holds a crucial position in the PANoptosis triggered by DNA damage. When DNA sustains damage, it can prompt ZBP1 to construct a panoptosome complex in tandem with RIPK3 and Caspase-6. This complex synergistically activates PANoptosis, and this process is independent of the classical p53 signaling pathway [73, 74]. Meanwhile, DNA damage will activate the transcription of endogenous retroviruses (ERVs), and the generated dsRNA can bind to ZBP1 and act as a ligand triggering PANoptosis [75]. In mouse models, DNA-damaging chemotherapeutic drugs can activate ZBP1-mediated PANoptosis, leading to chemotherapy side effects such as intestinal and pulmonary inflammation, tissue damage, and a reduction in immune cells. However, in ZBP1-deficient mice, these side effects are significantly alleviated. This indicates that by inhibiting the activity of ZBP1, it may be possible to reduce the toxic effects of chemotherapy on normal tissues without compromising the therapeutic efficacy [76]. The herpes simplex virus type 1-based oncolytic virus (oHSV) can trigger ZBP1-mediated PANoptosis, this leads to augmented anti-tumor immune effects. oHSV upregulates the expression of interferon-stimulated genes, giving rise to the build-up of endogenous Z-RNA and the subsequent activation of ZBP1. The outer membrane vesicles of Fusobacterium nucleatum (Fn-OMV) are able to elevate the expression of PANoptosis execution proteins. When Fn-OMV and oHSV are combined, they exhibit powerful anti-tumor immunogenicity [77]. Through transcriptomic analysis of various cancers, it has been found that the prognostic significance of ZBP1 expression varies among different cancers. In low-grade glioma (LGG), the prognostic significance of ZBP1 expression is negative. In contrast, in cutaneous melanoma (SKCM), high ZBP1 expression indicates a positive prognosis. This implies that personalized treatment plans can be formulated for patients with different cancers based on the expression level of ZBP1, and the treatment outcomes can be predicted [78]. 4.1.2 RIPK3 RIPK3 is a crucial regulatory factor in necroptosis, a form of programmed cell death. It can interact with RIPK1 and further activate downstream substrates such as MLKL, thereby initiating necroptosis [79, 80]. In cancer treatment, inducing necroptosis in cancer cells is an effective therapeutic strategy. For instance, activating RIPK3 through specific drugs or gene therapies can drive cancer cells towards necroptosis, thus inhibiting tumor growth [80]. Necroptosis mediated by RIPK3 releases a variety of cellular contents, which in turn alters the TME [81]. DAMPs released by necrotic cells can attract immune cells such as macrophages and dendritic cells to infiltrate the tumor site. For example, HMGB1, a typical DAMP, can activate pattern recognition receptors on the surface of immune cells and initiate an immune response upon release [82, 83]. Meanwhile, RIPK3-related necroptosis may also influence angiogenesis in the tumor microenvironment. Abnormal angiogenesis of tumor blood vessels is crucial for tumor growth and metastasis. Necrosis of cancer cells caused by RIPK3 activation may inhibit the disorderly growth of tumor blood vessels by affecting the discharge of angiogenesis-linked factors, for example, vascular endothelial growth factor (VEGF), cutting off the nutrient supply to the tumor, and thereby suppressing tumor development [84, 85]. Drug resistance of some cancer patients to chemotherapy and targeted therapy is a major challenge in clinical treatment. Research has found that RIPK3 may play a key role in this process [86]. In certain drug-resistant cancer cells, the RIPK3 signaling pathway is abnormally regulated. For example, in some breast cancer cells that have developed resistance to chemotherapeutic drugs, the expression of RIPK3 is downregulated, making it difficult to induce necroptosis, allowing the cancer cells to survive and continue to proliferate [87]. Conversely, by upregulating the expression of RIPK3 or activating its downstream signals, there is hope to restore the sensitivity of drug-resistant cancer cells to therapeutic drugs. The signaling pathway involving RIPK3 does not exist in isolation. It has complex interactions with other signaling pathways related to cell death and survival. For example, it may have cross-regulation with the apoptosis-related signaling pathway, and this interaction affects the response of cancer cells to different treatment methods. Studying this interaction mechanism is helpful for the development of combination treatment regimens and improving the efficacy of cancer treatment [88, 89]. As found in some investigates, regulating the synergistic effect between RIPK3 and the apoptosis pathway can enhance the sensitivity of cancer cells to chemotherapeutic drugs and increase the success rate of cancer treatment [71, 90]. ¿p#1 4.1.3 NLRP3 The NLRP3 inflammasome assumes a two-fold function in cancer treatment. Firstly, once activated, the NLRP3 inflammasome stimulates the activation of Caspase-1, resulting in the liberation of inflammatory cytokines like IL-1β and IL-18. These cytokines draw in immune cells, thereby generating an inflammatory microenvironment that enables the proliferation, invasion, metastasis, and angiogenesis of cancer cells [91, 92]. IL-1β is capable of triggering the NF-κB signaling pathway, thus augmenting the proliferative potential and anti-apoptotic capability of cancer cells. Additionally, NLRP3 has the ability to modulate the expression of cell cycle-associated proteins, which contributes to the proliferation of cancer cells. The immunosuppressive cells within the inflammatory microenvironment and the released inflammatory cytokines interfere with the function of antigen-presenting cells, which makes it possible for cancer cells to escape the immune system [93]; on the other hand, after the activation of the NLRP3 inflammasome, the activated Caspase-1 cuts GSDMD, producing the GSDMD-N terminal domain. This leads to the induction of pyroptosis in cancer cells. As a result, danger signals are released, which attract immune cells and boost the anti-tumor immune response [94, 95]. Inflammatory cytokines, for example, IL-1β and IL-18, which are discharged by the NLRP3 inflammasome, have the ability to activate immune cells such as natural killer (NK) cells and cytotoxic T lymphocytes (CTLs). Additionally, IL-18 facilitates the differentiation of Th1 cells and the release of IFN-γ. This, in turn, strengthens the immune system’s capacity to assault cancer cells from various perspectives [96]. Furthermore, the NLRP3 inflammasome can regulate the expression of cell cycle-related proteins, causing cancer cells to arrest at specific cell cycle stages, inhibiting their proliferation rate, and restricting tumor growth, thus providing support for cancer treatment [97]. For example, 5-fluorouracil (5-FU), one of the main drugs for treating gastrointestinal cancers, can selectively reduce the number of myeloid-derived suppressor cells (MDSCs) and enhance the generation of interferon-γ (IFN-γ) by tumor-specific CD8+ T cells [98, 99]. Nevertheless, although 5-FU eradicates immunosuppressive MDSCs, it simultaneously triggers the activation of the NLRP3 inflammasome in apoptotic MDSCs. This activation results in the release of IL-1β, the recruitment of T helper 17 cells, and the production of IL-17. Subsequently, IL-17 accelerates tumor growth through promoting angiogenesis [100]. The anti-tumor immune response and tumor suppression effects induced by chemotherapy are reliant on PTEN in myeloid cells. PTEN directly associates with NLRP3 and dephosphorylates it. This dephosphorylation event allows NLRP3 to interact with ASC, facilitating the assembly and activation of the inflammasome. As a consequence, Caspase-1 is activated, and IL-1β or IL-18 is secreted, thereby initiating the innate anti-tumor immunity [101, 102]. Clinical investigates have shown that in cancer patients, the expression of PTEN in myeloid cells is associated with the chemotherapy-induced production of IL-1β and anti-tumor immunity. This research indicates that NLRP3 plays a crucial role in PTEN-mediated anti-tumor immunity during chemotherapy, and activating the NLRP3 inflammasome can help enhance the effectiveness of chemotherapy [100]. 4.1.4 IRF1 Interferon regulatory factor 1 (IRF1) is an upstream regulator of PANoptosis, capable of inducing cell death. It functions in bone marrow and epithelial cells and exerts a protective effect against chemically induced and spontaneous colorectal cancer [103]. Investigates have revealed that mice lacking IRF1 are highly susceptible to colorectal cancer. In murine models of colorectal cancer that are elicited by azoxymethane (AOM) and dextran sulfate sodium (DSS), as well as in spontaneous colorectal cancer mouse models, IRF1 exerts its protective effect by regulating PANoptosis [104]. If IRF1 is absent, it will lead to defects in pyroptosis, apoptosis, and necroptosis (PANoptosis) of colonic cells. Consequently, cell death is weakened, increasing the risk of tumorigenesis [105]. IRF1 plays a pivotal role in the development and recruitment of immune cells, and is also essential for the proper functioning of cytotoxic T cells and natural killer cells. Cytotoxic T cells and natural killer cells are vital immune cells within the body’s immune system, having the capacity to directly assail cancer cells. Through the regulation of related gene expression, IRF1 facilitates the maturation and activation of these immune cells. As a result, it strengthens their ability to identify and eliminate cancer cells. This process significantly contributes to the body’s elimination of cancer cells, thus having an impact on anti-tumor immunity [106, 107]. Nevertheless, within cancer cells, IRF1 has the potential to repress the host’s IFN-I and IRF1-reliant anti-tumor immunity. It also leads to an elevation in the levels of diverse immune checkpoints, like IDO-1 and PD-L1. These immune checkpoint proteins are capable of impeding the activity of T cells. As a consequence, cancer cells are able to elude the recognition and assault of the body’s immune system. Simultaneously, IRF1 can modify the expression of proteins associated with antigen presentation, which further aids cancer cells in attaining immune escape [108, 109]. ¿p#1 4.1.5 AIM2 AIM2 can recognize double-stranded DNA in the cytoplasm, recruit ASC and Caspase-1 to form an inflammasome. Once Caspase-1 is activated, it cleaves the forerunners of IL-1β and IL-18, causing them to attain maturity and expediting their emission. This sets off an inflammatory reaction, rousing the immune system to fight against cancer cells [110, 111]. The inflammasome activated by AIM2 not only triggers an immune response but also assumes a crucial part in triggering PANoptosis within cells. Caspase-1 activated by the inflammasome can activate the downstream GSDMD. Upon cleavage of GSDMD, its N-terminal domain punches holes in the cell membrane, increasing the membrane’s permeability. As a result, the cell swells and eventually ruptures, a process known as pyroptosis [112, 113]. Simultaneously, Caspase-1 can also activate apoptosis-related caspases such as caspase-3, triggering apoptosis and directly killing cancer cells, thereby inhibiting tumor growth. Moreover, inflammatory cytokines released during the process of PANoptosis, like IL-1β and IL-18, is capable of drawing more immune cells to the tumor location. This enhances the surveillance and killing capabilities of immune cells against cancer cells, further suppressing tumor development [114]. 4.1.6 ASC ASC primarily participates in the assembly of inflammasomes during PANoptosis. When cells recognize PAMPs or DAMPs, for instance, after AIM2 recognizes double-stranded DNA in the cytoplasm, it will recruit ASC [115, 116]. ASC interacts with the PYD (pyrin domain) of AIM2 through its own PYD. Then, by utilizing its CARD (caspase recruitment domain), ASC recruits Caspase-1 to form an inflammasome [117, 118]. The activated Caspase-1 can cleave the precursors of IL-1β and IL-18, releasing mature inflammatory cytokines, which initiates an inflammatory response and activates immune cells to combat cancer cells [114]. ASC is not merely capable of triggering PANoptosis modalities, like pyroptosis, in cancer cells by activating the signaling pathway associated with the inflammasome. This action, in turn, inhibits tumor growth. Additionally, ASC has the ability to modulate the recruitment and activation of immune cells within the tumor microenvironment. It releases inflammatory cytokines to attract immune cells like macrophages and NK cells to the tumor site, enhancing the killing ability of immune cells against cancer cells. Additionally, it remodels the tumor microenvironment, inhibiting the proliferation, migration, and invasion capabilities of cancer cells [119, 120]. ¿p#1 4.1.7 FADD The sufficient expression of FADD is crucial for inducing apoptosis in cancer cells. TAT-FADD, formed by chemically conjugating the FADD protein with a cell-permeable trans-activator of transcription peptide (TAT), can be internalized into cancer cells via the caveolae-mediated endocytosis pathway and exert its function in the cytosol. It rapidly assembles the components of the DISC, triggering apoptosis signals and thus increasing the death rate of cancer cells [121, 122]. Investigates have shown that the apoptosis-inducing ability of TAT-FADD is comparable to that of conventional apoptosis inducers. There are numerous inflammatory signals present in the tumor microenvironment, which can promote tumor growth and metastasis. FADD can regulate the activation of NF-κB, reducing the constitutive activation of NF-κB and the expression of related downstream anti-apoptotic genes, such as Bcl-2, cFLIPL, RIP1, and cIAP2. Moreover, FADD can also inhibit the activation of the canonical NLRP3 inflammasome and restrict the processing and secretion of pro-inflammatory IL-1β. In this way, it modifies the tumor microenvironment and suppresses tumor development [123]. In the DISC, multiple procaspase-8 molecules aggregate and activate through auto-cleavage, generating active Caspase-8. The activated Caspase-8 can further cleave and activate downstream effector caspases, such as Caspase-3 and Caspase-7. These effector caspases act on numerous intracellular substrates, including cytoskeletal proteins and DNA repair enzymes. Ultimately, this leads to the appearance of morphological and biochemical characteristics of apoptosis, such as cell membrane shrinkage, chromatin condensation, and DNA fragmentation [124, 125]. When cells are stimulated by internal stressors such as DNA damage, oxidative stress, and hypoxia, the outer membrane permeability of mitochondria changes, releasing cytochrome c into the cytoplasm. Cytochrome c binds with apoptotic protease Apaf-1) and ATP to form an apoptosome [126, 127]. Within the apoptosome, the conformation of Apaf-1 changes. It recruits and activates the procaspase-9 through its caspase recruitment domain (CARD). The activated Caspase-9 then activates downstream Caspase-3 and Caspase-7, triggering the cell apoptosis program [128]. 4.1.8 Caspases Inflammatory caspases (Caspase-1, Caspase-4, Caspase-5 in humans, and Caspase-11 in mice) are the core initiating enzymes of pyroptosis. They cleave proteins of the gasdermin family, generating fragments with membrane-pore-forming activity in cells. This leads to cell swelling and rupture, releasing inflammatory factors, and playing a crucial role in inflammatory responses and immune defenses [129, 130]. When cells are stimulated by PAMPs or DAMPs, PRRs within the cells recognize these signals. Through a series of signal transduction processes, inflammasomes are activated [131, 132]. An inflammasome is a multi-protein complex that can recruit and activate inflammatory caspases such as Caspase-1. The activated Caspase-1 is capable of cleaving proteins of the gasdermin family, such as GSDMD. The N-terminal domain of the cleaved GSDMD has membrane-pore-forming activity, which creates pores in the cell membrane. This results in an imbalance of ions inside the cell, a change in osmotic pressure, cell swelling, and eventually rupture, releasing cellular contents including inflammatory cytokines like IL-1β and IL-18, and triggering an inflammatory response [133]. Necroptosis is a programmed necrosis process, in which Caspase-8 plays a complex regulatory role. Under normal circumstances, Caspase-8 can negatively regulate necroptosis by inhibiting the activities of receptor-interacting protein kinase 1 (RIPK1) and RIPK3 [134]. When cells are stimulated by tumor necrosis factor (TNF) and other factors, and the activity of Caspase-8 is inhibited (for example, inhibited by a virus-encoded Caspase-8 inhibitor, or in some pathological conditions where Caspase-8 is abnormally expressed), RIPK1 and RIPK3 will interact and phosphorylate, forming a necrosome [135, 136]. The necrosome further recruits and activates MLKL. MLKL is phosphorylated and oligomerized, then translocates to the cell membrane, creating pores in the cell membrane, leading to cell membrane rupture and necroptosis of the cell [137, 138]. When abnormal situations occur in cells, such as DNA damage and activation of proto-oncogenes, the apoptosis signaling pathways within the cells are activated. Initiator caspases, such as Caspase-8 and Caspase-9, are able to sense these apoptosis signals and get activated. The activated initiator caspases further activate effector caspases, such as Caspase-3 and Caspase-7 [139]. These effector caspases will cleave important proteins within the cell, disrupting the structure and function of the cell, inducing apoptosis, and thus eliminating cells that may become cancerous, maintaining the homeostasis of cells in the body. Numerous investigates have shown that in various types of cancer, the deletion or mutation of caspase genes leads to a decrease in caspase activity, blocking cell apoptosis and allowing cancer cells to escape the body’s clearance mechanism, thereby promoting tumorigenesis and tumor development [133]. 4.1.9 MLKL MLKL is a key execution protein for necroptosis and PANoptosis. When cancer cells are exposed to specific stimuli such as chemotherapeutic drugs and immune attacks, MLKL will be activated, triggering necroptosis and PANoptosis within cancer cells, thereby restricting the proliferation and spread of cancer cells [140, 141]. For example, in leukemia cell lines, the attack of specific immune cells can activate MLKL, triggering the cell death program and inhibiting the growth of cancer cells. This process is of great significance for maintaining the body’s tumor immune surveillance and preventing the unrestricted proliferation of cancer cells. Cell death mediated by MLKL can release danger signal molecules, which activate the immune system and attract immune cells such as macrophages and NK cells to recognize and kill cancer cells, thus exerting an anti-tumor effect. With the involvement of PANoptosis, the effect of immune activation may be further enhanced. For instance, in a melanoma model, MLKL-mediated tumor cell death promotes the aggregation of immune cells, effectively inhibiting tumor growth [141, 142]. In the necroptosis signaling pathway, when death receptors such as the tumor necrosis factor receptor (TNFR) bind to their corresponding ligands, they will recruit the adapter proteins TRADD and TRAF2. This leads to the activation of RIPK1, and subsequently, RIPK3 binds to RIPK1 to form a ”necrosome”. After activation, RIPK3 further phosphorylates MLKL. Phosphorylated MLKL oligomerizes and translocates to the plasma membrane, inserting into the plasma membrane to form pores, which disrupts the integrity of the cell membrane, causing the leakage of intracellular substances and ultimately triggering necroptosis [143, 144]. When cells undergo MLKL-mediated necroptosis, a large number of DAMPs are released, such as HMGB1, ATP, etc. These DAMPs can activate the pattern recognition receptors of surrounding cells, thereby inducing the production and release of inflammatory cytokines such as IL-1β and interleukin-6 (IL-6). Meanwhile, after MLKL is activated, It has the potential to further facilitate the expression and emission of inflammatory cytokines via activating signaling pathways including NF-κB, amplifying the inflammatory response [145]. During endoplasmic reticulum stress, the accumulation of unfolded or misfolded proteins activates the PERK-eIF2α signaling pathway. Once this pathway is activated, on the one hand, it inhibits the overall protein synthesis, and on the other hand, it promotes the expression of proteins related to necroptosis such as MLKL. As a result, the cell undergoes PANoptosis, and MLKL functions to facilitate the cell death process [146, 147]. MLKL also has a certain connection with cell death or survival pathways such as autophagy. When cells face different stress conditions, they coordinate with each other to jointly determine the fate of the cell. For example, autophagy may inhibit MLKL-mediated necroptosis to a certain extent, or provide some signal support for it [147, 148]. ¿p#1 4.1.10 The GSDM Protein Family The gasdermin protein family members typically comprise an N-terminal domain and a C-terminal domain. These two domains are linked by a linker peptide in the middle part. Under normal conditions, the activity of the N-terminal domain is inhibited by the C-terminal domain. Once cells are stimulated, specific proteases cleave the gasdermin proteins, thereby activating them. For example, GSDMD can be cleaved and activated by Caspase-1 in the inflammasome, releasing the active GSDMD-NT (N-terminal domain); gasdermin E (GSDME) can be cleaved and activated by caspase-3 [149, 150]. For instance, in the case of GSDMD, the activated GSDMD-NT creates pores on the cell membrane. As a result, potassium ions and other substances flow out of the cell, disrupting the ionic balance within the cell. As a result, the cell swells, the cell membrane ruptures, and cellular contents such as inflammatory cytokines like IL-1β are released. These inflammatory cytokines can recruit immune cells and trigger an inflammatory response [151, 152]. Upon stimulation by chemotherapeutic drugs and the like, GSDME is cleaved by caspase-3 to generate GSDME-NT, thereby inducing pyroptosis [153, 154]. In addition, gasdermin A (GSDMA) may also form pores in the cell membrane through its N-terminal domain, mediating cell death. However, its specific activation mechanism and functions in different cell types still require further investigation [149, 155]. GSDME can induce pyroptosis in normal tissues, thus eliminating abnormal cells and playing an inhibitory role in the occurrence and development of tumors. In various tumors such as colorectal cancer, the promoter region of the GSDME gene is often methylated, leading to the suppression of GSDME expression. This allows cancer cells to avoid pyroptosis and consequently keep proliferating and developing. Gasdermin-mediated pyroptosis has the capacity to discharge inflammatory cytokines and DAMPs, like IL-1β and HMGB1. These substances are able to trigger the activation of the immune system. They attract immune cells including macrophages, natural killer cells, and T cells to the tumor location, and boost the anti-tumor immune response. Simultaneously, cancer cells experiencing pyroptosis reveal a greater number of tumor-associated antigens. This, in turn, activates immune cells such as T cells, equipping them to more effectively identify and eliminate cancer cells [156, 157]. The activation of GSDMD can trigger pyroptosis and immune activation. Through high-throughput screening, researchers have discovered the small-molecule compound DMB. It can bind to the N-terminal domain of GSDMD to promote its activation, effectively inducing pyroptosis of cancer cells and inhibiting tumor growth in multiple tumor cell lines and mouse models. At the same time, it releases tumor-associated antigens and inflammatory cytokines to activate the immune system. Moreover, toxicity tests have confirmed that DMB shows no obvious toxicity at effective doses, providing new strategies and potential targets for tumor treatment [158, 159]. ¿p#1 4.2 Application of Key Signaling Molecules in Combination Therapies ¿p#1 4.2.1 ZBP1 During radiotherapy, cancer cells generate endogenous Z-RNA, a process that activates ZBP1 [160]. Moreover, oHSV can also upregulate the expression of ZBP1 in cancer cells, inducing ZBP1-mediated PANoptosis to exert an anti-tumor effect [77, 161]. When radiotherapy is combined with oHSV, on one hand, radiotherapy can directly kill cancer cells. On the other hand, oHSV, by promoting a higher expression of ZBP1, induces PANoptosis in cancer cells. The synergistic effect of these two approaches significantly enhances the killing efficiency against cancer cells [162, 163]. Simultaneously, the ZBP1 activated by radiotherapy can also trigger necroptosis by activating the downstream molecule MLKL. During necroptosis, cytoplasmic DNA accumulates, which in turn activates the cGAS-STING signaling pathway, forming a positive feedback loop with radiotherapy. This loop further intensifies the inflammatory response, thereby enhancing the anti-tumor effect of radiotherapy [164, 165]. In immunotherapy, especially in immune checkpoint blockade (ICB) therapy, the ZBP1-mediated PANoptosis triggered by oHSV can release tumor-associated antigens and inflammatory signals. These released substances enhance the ability of the immune system to recognize and eliminate cancer cells. The combination of oHSV and ICB therapy is capable of remarkably enhancing the effectiveness of the ICB treatment [166, 167]. Furthermore, exploring other drugs or agents that can enhance ZBP1-mediated PANoptosis and combining them with immunotherapeutic drugs could activate both innate and adaptive immunity, comprehensively strengthening the anti-tumor immune response [168]. 4.2.2 The GSDM Protein Family ROS generated by radiotherapy can disrupt the redox balance of cancer cells. Some nanomaterials, such as Au@AgBiS₂, possess unique photothermal and radiosensitizing properties. During radiotherapy, Au@AgBiS₂ absorbs the energy from radiation, generating an increased amount of ROS. The high concentration of ROS activates caspase 3. Once activated, caspase 3 cleaves GSDMD, resulting in the oligomerization of its N-terminal domain and its insertion into the cell membrane, thereby forming pores. This causes the discharge of cellular contents and, in turn, initiates pyroptosis within the cells [169, 170]. By developing more nanomaterials with similar properties and combining them with radiotherapy, the pyroptosis rate of cancer cells can be significantly increased. This enhancement not only strengthens the ability of radiotherapy to kill cancer cells but also improves the overall therapeutic efficacy of radiotherapy [171, 172]. The small-molecule agonist of GSDMD, DMB, can directly bind to GSDMD, inducing a conformational change and activating it. The activated GSDMD creates pores in the tumor cell membrane, triggering pyroptosis, and causing cancer cells to release tumor-associated antigens. The anti-PD-1 monoclonal antibody can block the PD-1/PD-L1 signaling pathway, thereby relieving the immunosuppression of T cells [173]. When DMB is combined with the anti-PD-1 monoclonal antibody, DMB induces pyroptosis in cancer cells, and the released antigens activate T cells. Meanwhile, the anti-PD-1 monoclonal antibody enhances the activity and proliferation ability of T cells. Through their synergistic effect, the recognition and killing ability of immune cells against cancer cells are enhanced, thus producing a synergistic anti-tumor effect [152, 174]. During radiotherapy, it is possible to combine with nanomaterials loaded with relevant substances activated by ultrasound, such as nanoparticles loaded with chemotherapeutic drugs or immunomodulators. The cavitation effect of ultrasound can enhance the penetration and uptake of nanomaterials in tumor tissues. Taking nasopharyngeal carcinoma as an example, radiotherapy activates caspase 3 through the intrinsic mitochondrial apoptosis pathway. The activated caspase 3 cleaves gasdermin E (GSDME), inducing GSDME-dependent pyroptosis [175, 176]. By designing specific drug carriers, such as pH-responsive nanomicelles, drugs can be released in the acidic environment within cancer cells, targeting and activating GSDME. This further enhances the pyroptosis of cancer cells induced by radiotherapy, significantly increasing the lethality of radiotherapy [177]. In tumors, exogenous GSDME can be expressed by using gene editing techniques, such as the CRISPR/Cas9 system, to introduce the GSDME gene into cancer cells. Cancer cells with overexpressed GSDME are more prone to pyroptosis when attacked by immune cells [178, 179]. When combining the gene therapy vector with overexpressed GSDME with chimeric antigen receptor T-cell (CAR-T) therapy, CAR-T cells can specifically recognize and bind to the antigens on the surface of cancer cells, activating GSDME and triggering pyroptosis in cancer cells. In liver cancer, targeting GSDME can promote the polarization of macrophages into the M1 phenotype. M1 macrophages secrete pro-inflammatory cytokines, enhancing the anti-tumor immune response, which provides a new perspective for the combination of GSDME and immunotherapy [180]. ¿p#1 4.2.3 MLKL Radiotherapy induces DNA damage in cancer cells, which activates ZBP1. ZBP1 then activates MLKL through RIPK1 and RIPK3. Drugs capable of inhibiting the negative regulators of MLKL, such as small-molecule inhibitors, can be developed. These inhibitors block the binding between the negative regulators and MLKL, making MLKL more readily activated [181, 182]. Once activated, MLKL undergoes phosphorylation and oligomerization, translocates to the cell membrane, and disrupts the integrity of the cell membrane, leading to necroptosis of the cell. By enhancing the activation of the ZBP1-MLKL pathway induced by radiotherapy, the sensitivity of cancer cells to radiotherapy can be increased. This promotes the death of more cancer cells and improves the therapeutic effect of radiotherapy [146, 168]. The cell death mediated by the activation of MLKL releases immunogenic substances, such as heat shock proteins and ATP. Drugs or agents that can activate MLKL, such as small-molecule agonists, can be combined with tumor vaccines [183]. The small-molecule agonists activate MLKL, causing cancer cells to release more antigens. Tumor vaccines, which contain tumor-associated antigens, can stimulate the body’s immune response. When these two are combined, the immune response elicited by the tumor vaccine is enhanced, activating more T cells and B cells and improving the efficacy of immunotherapy [184, 185]. ¿p#1 5. Conclusion and Prospect Abnormal cell death holds a pivotal position in the development of cancer. The impairment of apoptosis and other factors give rise to an imbalance between cell proliferation and death. This imbalance enables cancer cells to avoid death and keep proliferating continuously. Consequently, inducing the death of cancer cells stands as a central strategy in cancer treatment. Against this background, as research on the regulatory mechanisms of cell death delves deeper, interactions among pyroptosis, apoptosis, and necroptosis have been unearthed, leading to the emergence of the concept of PANoptosis. From the perspective of molecular mechanisms, ZBP1, acting as the top-level sensor of PANoptosis, upon detecting DNA damage, initiates the assembly of the PANoptosome complex. It does this by recognizing double-stranded RNA originating from endogenous retroviruses, thereby commencing PANoptosis. Besides, molecules like NLRC4, NLRP3, IRF1, AIM2, RIPK1, and RIPK3 are also implicated. These molecules either function as sensors to activate inflammasomes and facilitate cell death or act as upstream regulators to modulate related processes. Adaptors ASC and FADD link sensors with catalytic effectors, facilitating the formation of multiprotein complexes. Catalytic effectors, such as the Caspase protein family, MLKL, and the gasdermin protein family, are in charge of carrying out the final stages of cell death. They cleave relevant proteins, trigger inflammatory reactions, or modify the permeability of the cell membrane, driving the cell towards death. Concerning its function in cancer, PANoptosis, on one hand, bolsters anti-tumor immunity. When cancer cells experience PANoptosis, they release danger-associated molecular patterns. These patterns activate innate immune cells and then trigger an adaptive immune response, strengthening the body’s capacity to eliminate cancer cells. On the other hand, it is involved in the regulation of the tumor microenvironment. The cytokines and chemokines released affect the recruitment and activation of immune cells, tumor angiogenesis, and extracellular matrix remodeling. Furthermore, its role in tumor metastasis is two-sided. It can inhibit tumor metastasis by activating the anti-tumor immune system. However, it may also promote tumor metastasis by generating a pro-inflammatory microenvironment. In terms of the application prospects in cancer treatment, targeted therapy against key signaling molecules holds great significance. For example, inhibiting the activity of ZBP1 can reduce the side effects of chemotherapy, and personalized treatment plans can be formulated based on its expression level. Activating RIPK3 can induce programmed necrosis in cancer cells, and regulating its synergistic effect with the apoptosis pathway can improve the effectiveness of cancer treatment. The NLRP3 inflammasome has a dual role in cancer treatment, and it needs to be rationally regulated according to specific circumstances. Molecules such as IRF1, AIM2, ASC, and FADD also play roles in cancer treatment by regulating the function of immune cells, activating inflammasomes, inducing apoptosis, and other means. Meanwhile, key signaling molecules have demonstrated promising results in combination therapies. For instance, ZBP1, the GSDM protein family, MLKL, etc., when combined with radiotherapy, immunotherapy, nanomaterials, etc., can enhance the killing effect on cancer cells or improve the efficacy of immunotherapy. Currently, although certain progress has been made in the research of PANoptosis in tumors, there are still many unknown aspects that await exploration. At the basic research level, in-depth analysis is required for the complex interactive regulatory mechanisms among various signaling pathways of PANoptosis, especially the differences in different tumor types and development stages. For example, further clarifying the precise regulatory mechanisms of key molecules such as ZBP1 and RIPK3 within a specific tumor microenvironment will contribute to the more precise design of targeted therapeutic strategies. Meanwhile, the activation mechanisms of some members of the gasdermin protein family and their functions in different cell types, such as GSDMA, still need further investigation. This will provide more potential targets for tumor treatment. ¿p#1 Abbreviations IAV: Influenza A virus; ZBP1: Z-DNA binding protein 1; RIPK1: Receptor-interacting protein kinase 1; RIPK3: Receptor-interacting protein kinase 3; Caspase-8: Cysteine-aspartic protease 8; NP: Nucleoprotein; PB1: Polymerase basic protein 1; NLRP3: NOD-like receptor protein 3; ASC: Apoptosis-associated speck-like protein containing a CARD; Caspase-1: Cysteine-aspartic protease 1; LPS: Lipopolysaccharide; mtDNA: Mitochondrial DNA; PAMPs: Pathogen-associated molecular patterns; TLRs: Toll-like receptors; NF-κB: Nuclear factor kappa-light-chain-enhancer of activated B cells; GSDMD: Gasdermin D; IL-1β: Interleukin-1β; IL-18: Interleukin-18; ROS: Reactive oxygen species; Bcl-2: B-cell lymphoma-2; Bax: Bcl-2-associated X protein; Bak: Bcl-2 homologous antagonist/killer; MPTP: Mitochondrial permeability transition pore; Apaf-1: Apoptotic protease activating factor-1; PARP: Poly (ADP-ribose) polymerase; Fas: Fas cell surface death receptor; TNFR1: Tumor necrosis factor receptor 1; FasL: Fas ligand; DD: Death domain; FADD: Fas-associated death domain protein; DED: Death effector domain; DISC: Death-inducing signaling complex; TRADD: Tumor necrosis factor receptor-associated death domain; MLKL: Mixed lineage kinase domain-like protein; PKR: Protein kinase R; dsRNA: Double-stranded RNA; eIF2α: Eukaryotic translation initiation factor 2α; MAPK: Mitogen-activated protein kinase; ERK: Extracellular signal-regulated kinase; JNK: c-Jun N-terminal kinase; p38 MAPK: p38 mitogen-activated protein kinase; DAMPs: Damage-associated molecular patterns; HMGB1: High-mobility group box 1 protein; ATP: Adenosine triphosphate; PRRs: Pattern recognition receptors; TME: Tumor microenvironment; ERVs: Endogenous retroviruses; oHSV: Oncolytic herpes simplex virus type 1; Fn-OMV: Outer membrane vesicles of Fusobacterium nucleatum; LGG: Low-grade glioma; SKCM: Cutaneous melanoma; VEGF: Vascular endothelial growth factor; IRF1: Interferon regulatory factor 1; AIM2: Absent in melanoma 2; TAT: Trans-activator of transcription peptide; ICB: Immune checkpoint blockade; CAR-T: Chimeric antigen receptor T-cell; GSDME: Gasdermin E; GSDMA: Gasdermin A. Conflicts of interest The authors declare that have no competing interests. ¿p#1 Author Contributions Conceptualization: W.R.S., Q.Y.J., D.D.W.; Data curation: Y.Z.; Funding acquisition: Q.Y.J., D.D.W.; Writing-original draft: Y.Z., X.L.W., Y.W.W., T.C., L.C., Y.Q.F., Y.H.C.; Visualization and supervision: W.R.S., Q.Y.J., D.D.W.; Writing-review and editing: Z.Y., D.D.W. Acknowledgments This work was supported by grants from the Foundation of Science & Technology Department of Henan Province, China (No. 252102310338), the Natural Science Foundation of Education Department of Henan Province, China (No. 24B310001), and Henan University School of Stomatology Research Grant for Young Scholar, China (No. HUSSYS2024001). References1. Koo, N., A.K. Sharma, and S. Narayan, Therapeutics Targeting p53-MDM2 Interaction to Induce Cancer Cell Death. Int J Mol Sci, 2022. 23 (9).2. Tesfay, L., et al., Stearoyl-CoA Desaturase 1 Protects Ovarian Cancer Cells from Ferroptotic Cell Death. 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ZBP1 acts as an initiating signaling molecule, triggering three distinct cell death pathways. The first pathway is the apoptosis pathway: ZBP1 activates Bcl-2 and Caspase-8, and Caspase-8 further activates Caspase-3, ultimately leading to apoptosis. The second pathway is the necroptosis pathway: ZBP1 promotes the interaction between RIPK1 and RIPK3, activating MLKL, and the activation of MLKL induces necroptosis. The third pathway is the pyroptosis pathway: ZBP1 activates the NLRP3 inflammasome, and NLRP3 recruits and activates Caspase-1 through the adaptor protein ASC. Activated Caspase-1 leads to pyroptosis. Regardless of which of the above pathways leads to cell death, the end result is the activation of anti-tumor immunity. ZBP1: Z-DNA binding protein 1; Bcl-2: B-cell lymphoma-2; Caspase-8: Cysteine-aspartic protease 8; Caspase-3: Cysteine-aspartic protease 3; RIPK1: Receptor-interacting protein kinase 1; RIPK3: Receptor-interacting protein kinase 3; MLKL: Mixed lineage kinase domain-like protein; NLRP3: NOD-like receptor protein 3; ASC: Apoptosis-associated speck-like protein containing a CARD; Caspase-1: Cysteine-aspartic protease 1. Figure 2 . The binding of LPS to TLR4 or K⁺ efflux leads to the generation of reactive oxygen species (ROS), which activates the NLRP3 inflammasome, resulting in the activation of Caspase-1. This subsequently cleaves GSDMD, ultimately leading to pyroptosis. FasL binding to Fas forms the DISC, which activates Caspase-8. Meanwhile, ROS stimulation can induce the release of apoptosis-related substances from mitochondria, activating Caspase-9. Both of these pathways ultimately lead to apoptosis. TNF binding to TNFR1 induces the phosphorylation of RIPK1, which subsequently activates RIPK3 and MLKL, resulting in necroptosis. LPS: Lipopolysaccharide; TLR4: Toll-likereceptor4; ROS: Reactive oxygen species; NLRP3: NOD-like receptor protein 3; Caspase-1: Cysteine-asparticprotease1; GSDMD: Gasdermin D; FasL: Fas ligand; Fas: Fas cell surface death receptor; DISC: Death-inducing signaling complex; Caspase-8: Cysteine-asparticprotease8; Caspase-9: Cysteine-asparticprotease9; TNF: Tumor necrosis factor; TNFR1: Tumornecrosisfactorreceptor1; RIPK1: Receptor-interactingproteinkinase1; RIPK3: Receptor-interactingproteinkinase3; MLKL: Mixed lineage kinase domain-like protein. Figure 3 . After viral infection of cells, dsDNA is produced. dsDNA activates PKR, leading to its phosphorylation. Phosphorylated PKR has two directions of action: on the one hand, it activates NF-κB, and on the other hand, it phosphorylates eIF2α. NF-κB and phosphorylated eIF2α ultimately upregulate the expression of IL-6 and TNF-α, thereby triggering inflammatory responses and apoptosis. In the inflammatory signaling pathway, inflammatory stimuli activate and phosphorylate JNK and ERK, respectively. Phosphorylated JNK activates the transcription factor c-JUN, which also participates in the process of apoptosis and inflammatory response. In contrast, phosphorylated ERK plays a negative regulatory role in apoptosis and inflammatory response. PKR: Protein kinase R; NF-κB: Nuclear factor kappa-light-chain-enhancer of activated B cells; eIF2α: Eukaryotic translation initiation factor 2α; IL-6: Interleukin-6; TNF-α: Tumor necrosis factor-α; JNK: c-Jun N-terminal kinase; ERK: Extracellular signal-regulated kinase; c-JUN: c-Jun protein. Figure 4 . When cancer cells undergo pan-apoptosis, they release cytokines and DAMPs, which can be recognized by immune cells. Upon activation, immune cells can produce three effects: First, they activate adaptive immunity to initiate a specific immune response to attack cancer cells; second, they promote tumor angiogenesis to provide nutrients for tumor growth; and third, they facilitate tumor metastasis by altering the tumor microenvironment, aiding the spread of cancer cells to other locations. DAMPs: Damage-associated molecular patterns. Supplementary Material File (figure 1.docx) Download 215.13 KB File (figure 2_2.docx) Download 403.29 KB File (figure 3.docx) Download 273.48 KB File (figure 4.docx) Download 274.56 KB Information & Authors Information Version history V1 Version 1 15 November 2025 Copyright This work is licensed under a Non Exclusive No Reuse License. Authors Affiliations Yi Zhang [email protected] Henan University School of Stomatology View all articles by this author Xue-Li Wang Henan University School of Stomatology View all articles by this author Yan-Wen Wang Henan University School of Stomatology View all articles by this author Ti Chu Henan University School of Stomatology View all articles by this author Lei Cao Henan University School of Stomatology View all articles by this author Yong-Qi Fan Henan University School of Stomatology View all articles by this author Yu-Hang Chen Henan University School of Stomatology View all articles by this author Wei-Rong Si Henan University School of Basic Medical Sciences View all articles by this author Qi-Ying Jiang Henan University School of Stomatology View all articles by this author Dongdong Wu 0000-0001-6739-8437 Henan University School of Stomatology View all articles by this author Metrics & Citations Metrics Article Usage 229 views 109 downloads .FvxKWukQNSOunydq8rnd { width: 100px; } Citations Download citation Yi Zhang, Xue-Li Wang, Yan-Wen Wang, et al. 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