A new isoxazolyl-urea derivative induces apoptosis, paraptosis, and ferroptosis by modulating MAPKs in pancreatic cancer cells.

MAPK pathway regulates the major events including cell division, cell death, migration, invasion, and angiogenesis. Small molecules that modulate the MAPK pathway have been demonstrated to impart cytotoxicity in cancer cells. Herein, the synthesis of a new isoxazolyl-urea derivative (QR-4) has been described and its effect on the growth of pancreatic cancer cells has been investigated. QR-4 reduced the cell viability in a panel of pancreatic cancer cells with minimal effect on normal hepatocytes. QR-4 induced the cleavage of PARP and procaspase-3, reduced the expression of antiapoptotic proteins, increased SubG1 cells, and annexin V/PI-stained cells indicating the induction of apoptosis. QR-4 also triggered paraptosis as witnessed by the reduction of mitochondrial membrane potential, decrease in the expression of Alix, increase in the levels of ATF4 and CHOP, and enhanced ER stress. QR-4 also modulated ferroptosis-related events such as elevation in iron levels, alteration in GSH/GSSG ratio, and increase in the expression of TFRC with a parallel decrease in the expression of GPX4 and SLC7A11. The mechanistic approach revealed that QR-4 increases the phosphorylation of all three forms of MAPKs (JNK, p38, and ERK). Independent application of specific inhibitors of these MAPKs resulted in a partial reversal of QR-4-induced effects. Overall, these reports suggest that a new isoxazolyl-urea imparts cell death via apoptosis, paraptosis, and ferroptosis by regulating the MAPK pathway in pancreatic cancer cells.

A Two-Pronged Nanostrategy of Iron Metabolism Disruption to Synergize Tumor Therapy by Triggering the Paraptosis-Apoptosis Hybrid Pathway.

Iron metabolism has emerged as a promising target for cancer therapy; however, the innate metabolic compensatory capacity of cancer cells significantly limits the effectiveness of iron metabolism therapy. Herein, bioactive gallium sulfide nanodots (GaSx), with dual functions of "reprogramming" and "interfering" iron metabolic pathways, were successfully developed for tumor iron metabolism therapy. The constructed GaSx nanodots ingeniously harness hydrogen sulfide (H2S) gas, which is released in response to the tumor microenvironment, to reprogram the inherent transferrin receptor 1 (TfR1)-ferroportin 1 (FPN1) iron metabolism axis in cancer cells. Concurrently, the gallium ions (Ga3+) derived from GaSx act as a biochemical "Trojan horse", mimicking the role of iron and displacing it from essential biomolecular binding sites, thereby influencing the fate of cancer cells. By leveraging the dual mechanisms of Ga3+-mediated iron disruption and H2S-facilitated reprogramming of iron metabolic pathways, GaSx prompted the initiation of a paraptosis-apoptosis hybrid pathway in cancer cells, leading to marked suppression of tumor proliferation. Importantly, the dysregulation of iron metabolism induced by GaSx notably increased tumor cell susceptibility to both chemotherapy and immune checkpoint blockade (ICB) therapy. This study underscores the therapeutic promise of gas-based interventions and metal ion interference strategies for the tumor metabolism treatment.

Isoxazolyl-urea derivative evokes apoptosis and paraptosis by abrogating the Wnt/ß-catenin axis in colon cancer cells.

Deregulated activation of the Wnt/ß-catenin pathway is observed in many types of human malignancies including colon cancer. Abrogation of the Wnt/ß-catenin pathway has been demonstrated as an effective way of inducing cancer cell death. Herein, a new isoxazolyl-urea (QR-5) was synthesized and examined its efficacy on the viability of colon cancer cell lines. QR-5 displayed selective cytotoxicity towards colon cancer cells over normal counterparts. QR-5 induced apoptosis as evidenced by elevation in sub-G1 cells, decrease in Bcl-2, MMP-9, COX-2, VEGF and cleavage of PARP and caspase-3. QR-5 reduced the mitochondrial membrane potential, decreased the expression of Alix and elevated the expression of ATF4 and CHOP indicating the induction of paraptosis. The inhibitor of apoptosis (Z-DEVD-FMK) and paraptosis (CHX) could not restore Alix expression and PARP cleavage in QR-5 treated cells, respectively suggesting the complementation between the two cell death pathways. QR-5 suppressed the expression of Wnt/ß-catenin pathway proteins which was also evidenced by the downregulation of nuclear and cytoplasmic ß-catenin. The dependency of QR-5 on ß-catenin for inducing apoptosis and paraptosis was demonstrated by knockdown experiments using ß-catenin specific siRNA. Overall, QR-5 induces apoptosis as well as paraptosis by mitigating the Wnt/ß-catenin axis in colon cancer cells.

Fangchinoline targets human renal cell carcinoma cells through modulation of apoptotic and non­apoptotic cell deaths.

The process of apoptosis is one of the essential processes involved in maintenance of homeostasis in the human body. It can aid to remove misfolded proteins or cellular organelles. This sequence is especially necessary in cancer cells. However, specifically targeting already apoptotic pathways can induce drug resistance in cancer cells and hence drugs can induce cell death by alternative mechanism. We investigated whether fangchinoline (FCN) can target renal carcinoma cells by inducing multiple cell death mechanisms. Both paraptosis, autophagy, and apoptosis were induced by FCN through stimulation of diverse molecular signaling pathways. FCN induced ROS production with GSH/GSSG imbalance, and ER stress. In addition, formation of autophagosome and autophagy related markers were stimulated by FCN. Moreover, FCN induced cell cycle arrest and PARP cleavage. Except for blocking protein synthesis, these three cell death pathways were found to be complementarily working together with each other. FCN also exhibited synergistic effects with paclitaxel in inducing programmed cell death in RCC cells. Our data indicates that FCN could induce apoptotic cell death and non-apoptotic cell death pathways and can be con-tribute to development of novel cancer prevention or therapy.

Targeted delivery of FAK siRNA by engineered exosomes to reverse cetuximab resistance via activating paraptosis in colon cancer.

BACKGROUND: Cetuximab is extensively used in the treatment of metastatic colorectal cancer (mCRC). However, resistance poses a significant challenge to successful therapy. Recently, paraptosis, a non-classical programmed cell death, has garnered increased attention for its potential application value in antitumor treatments. We aimed to identify the essential pathways and signaling molecules involved in paraptosis inhibition and select them as therapeutic targets in cetuximab resistance. Additionally, engineered exosome technology is used as a drug delivery system with both targeted and effector properties. RESULTS: By comparing the differential expression of paraptosis-related genes between drug-resistant colon cancer cells and sensitive cells, it was observed that the paraptosis level induced by cetuximab was significantly downregulated in drug-resistant cells. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis identified the focal adhesion kinase (FAK) signaling pathway as a key pathway involved in the suppression of paraptosis. The biological function of FAK in cetuximab-resistant cells was investigated through cell morphology observation, CCK-8 assay, colony formation assay, RT-qPCR, Western Blot, and loss-of-function experiments. The results showed that the FAK signaling pathway was significantly upregulated in cetuximab-resistant colon cancer cells, and siRNA interference targeting FAK could notably inhibit cell proliferation while upregulating the paraptosis level. Based on this, engineered colon cancer cells targeted and FAK siRNA loaded exosomes (CT-Exo-siFAK1) were constructed. In vitro experiments, CT-Exo-siFAK1 could effectively activate paraptosis and inhibit the proliferation of drug-resistant colon cancer cells. In vivo experiments also confirmed that CT-Exo-siFAK1 significantly suppressed tumor growth and metastasis while upregulating the paraptosis level. CONCLUSION: This study suggests that FAK signaling pathway-mediated inhibition of paraptosis levels is crucial in the sensitivity of cetuximab targeted therapy in colon cancer, and the use of engineered exosomes to deliver FAK siRNA may be an effective strategy to reverse cetuximab resistance.

CDK7/CDK9 mediates transcriptional activation to prime paraptosis in cancer cells.

BACKGROUND: Paraptosis is a programmed cell death characterized by cytoplasmic vacuolation, which has been explored as an alternative method for cancer treatment and is associated with cancer resistance. However, the mechanisms underlying the progression of paraptosis in cancer cells remain largely unknown. METHODS: Paraptosis-inducing agents, CPYPP, cyclosporin A, and curcumin, were utilized to investigate the underlying mechanism of paraptosis. Next-generation sequencing and liquid chromatography-mass spectrometry analysis revealed significant changes in gene and protein expressions. Pharmacological and genetic approaches were employed to elucidate the transcriptional events related to paraptosis. Xenograft mouse models were employed to evaluate the potential of paraptosis as an anti-cancer strategy. RESULTS: CPYPP, cyclosporin A, and curcumin induced cytoplasmic vacuolization and triggered paraptosis in cancer cells. The paraptotic program involved reactive oxygen species (ROS) provocation and the activation of proteostatic dynamics, leading to transcriptional activation associated with redox homeostasis and proteostasis. Both pharmacological and genetic approaches suggested that cyclin-dependent kinase (CDK) 7/9 drive paraptotic progression in a mutually-dependent manner with heat shock proteins (HSPs). Proteostatic stress, such as accumulated cysteine-thiols, HSPs, ubiquitin-proteasome system, endoplasmic reticulum stress, and unfolded protein response, as well as ROS provocation primarily within the nucleus, enforced CDK7/CDK9-Rpb1 (RNAPII subunit B1) activation by potentiating its interaction with HSPs and protein kinase R in a forward loop, amplifying transcriptional regulation and thereby exacerbating proteotoxicity leading to initiate paraptosis. The xenograft mouse models of MDA-MB-231 breast cancer and docetaxel-resistant OECM-1 head and neck cancer cells further confirmed the induction of paraptosis against tumor growth. CONCLUSIONS: We propose a novel regulatory paradigm in which the activation of CDK7/CDK9-Rpb1 by nuclear proteostatic stress mediates transcriptional regulation to prime cancer cell paraptosis.

Docosahexaenoic Acid Coordinating with Sodium Selenite Promotes Paraptosis in Colorectal Cancer Cells by Disrupting the Redox Homeostasis and Activating the MAPK Pathway.

Tumor cells are characterized by a delicate balance between elevated oxidative stress and enhanced antioxidant capacity. This intricate equilibrium, maintained within a threshold known as redox homeostasis, offers a unique perspective for cancer treatment by modulating reactive oxygen species (ROS) levels beyond cellular tolerability, thereby disrupting this balance. However, currently used chemotherapy drugs require larger doses to increase ROS levels beyond the redox homeostasis threshold, which may cause serious side effects. How to disrupt redox homeostasis in cancer cells more effectively remains a challenge. In this study, we found that sodium selenite and docosahexaenoic acid (DHA), a polyunsaturated fatty acid extracted from marine fish, synergistically induced cytotoxic effects in colorectal cancer (CRC) cells. Physiological doses of DHA simultaneously upregulated oxidation and antioxidant levels within the threshold range without affecting cell viability. However, it rendered the cells more susceptible to reaching the upper limit of the threshold of redox homeostasis, facilitating the elevation of ROS levels beyond the threshold by combining with low doses of sodium selenite, thereby disrupting redox homeostasis and inducing MAPK-mediated paraptosis. This study highlights the synergistic anticancer effects of sodium selenite and DHA, which induce paraptosis by disrupting redox homeostasis in tumor cells. These findings offer a novel strategy for more targeted and less toxic cancer therapies for colorectal cancer treatment.

α-Hederin induces paraptosis by targeting GPCRs to activate Ca2+/MAPK signaling pathway in colorectal cancer.

BACKGROUND: Non-apoptotic cell death is presently emerging as a potential direction to overcome the apoptosis resistance of cancer cells. In the current study, a natural plant agent α-hederin (α-hed) induces caspase-independent paraptotic modes of cell death. PURPOSE: The present study is aimed to investigate the role of α-hed induces paraptosis and the associated mechanism of it. METHODS: The cell proliferation was detected by CCK-8. The cytoplasm organelles were observed under electron microscope. Calcium (Ca2+) level was detected by flow cytometry. Swiss Target Prediction tool analyzed the potential molecule targets of α-hed. Molecular docking methods were used to evaluate binding abilities of α-hed with targets. The expressions of genes and proteins were analyzed by RT-qPCR, western blotting, immunofluorescence, and immunohistochemistry. Xenograft models in nude mice were established to evaluate the anticancer effects in vivo. RESULTS: α-hed exerted significant cytotoxicity against a panel of CRC cell lines by inhibiting proliferation. Besides, it induced cytoplasmic vacuolation in all CRC cells. Electron microscopy images showed the aberrant dilation of endoplasmic reticulum and mitochondria. Both mRNA and protein expressions of Alg-2 interacting proteinX (Alix), the marker of paraptosis, were inhibited by α-hed. Besides, both Swiss prediction and molecular docking showed that the structure of α-hed could tightly target to GPCRs. GPCRs were reported to activate the phospholipase C (PLC)-ß3/ inositol 1,4,5-trisphosphate receptor (IP3R)/ Ca2+/ protein kinase C alpha (PKCα) pathway, and we then found all proteins and mRNA expressions of PLCß3, IP3R, and PKCα were increased by α-hed. After blocking the GPCR signaling, α-hed could not elevate Ca2+ level and showed less CRC cell cytotoxicity. MAPK cascade is the symbol of paraptosis, and we then demonstrated that α-hed activated MAPK cascade by elevating Ca2+ flux. Since non-apoptotic cell death is presently emerging as a potential direction to overcome chemo-drug resistance, we then found α-hed also induced paraptosis in 5-fluorouracil-resistant (5-FU-R) CRC cells, and it reduced the growth of 5-FU-R CRC xenografts. CONCLUSIONS: Collectively, our findings proved α-hed as a promising candidate for inducing non-apoptotic cell death, paraptosis. It may overcome the resistance of apoptotic-based chemo-resistance in CRC.

Necrosis Links Neurodegeneration and Neuroinflammation in Neurodegenerative Disease.

The mechanisms of neuronal cell death in neurodegenerative disease remain incompletely understood, although recent studies have made significant advances. Apoptosis was previously considered to be the only mechanism of neuronal cell death in neurodegenerative diseases. However, recent findings have challenged this dogma, identifying new subtypes of necrotic neuronal cell death. The present review provides an updated summary of necrosis subtypes and discusses their potential roles in neurodegenerative cell death. Among numerous necrosis subtypes, including necroptosis, paraptosis, ferroptosis, and pyroptosis, transcriptional repression-induced atypical cell death (TRIAD) has been identified as a potential mechanism of neuronal cell death. TRIAD is induced by functional deficiency of TEAD-YAP and self-amplifies via the release of HMGB1. TRIAD is a feasible potential mechanism of neuronal cell death in Alzheimer's disease and other neurodegenerative diseases. In addition to induction of cell death, HMGB1 released during TRIAD activates brain inflammatory responses, which is a potential link between neurodegeneration and neuroinflammation.

Prognostic and immunological implications of paraptosis-related genes in lung adenocarcinoma: Comprehensive analysis and functional verification of hub gene.

BACKGROUND: Lung adenocarcinoma (LUAD) poses significant clinical challenges due to its inherent heterogeneity and variable response to treatment. Recent research has specifically focused on elucidating the role of Paraptosis-related genes (PRGs) in the progression of cancer and the prognosis of patients. METHODS: We conducted a comprehensive analysis of the differential expression of PRGs in LUAD. Additionally, univariate Cox regression analysis was utilized to determine the prognostic significance of these genes. Furthermore, consensus clustering was employed to differentiate molecular subtypes within LUAD, while immune heterogeneity was assessed. To evaluate treatment outcomes, the expression of immune checkpoint inhibitors was examined, and the sensitivity of LUAD patients to chemotherapy drugs was assessed. Moreover, machine learning algorithms were employed to construct a Paraptosis-related risk score with prognostic and immunological indicators. Finally, to validate the findings, in vitro experiments were performed to verify the regulatory effect of key PRGs on Paraptosis. RESULTS: Our analysis identified 24 PRGs that exhibited differential expression, with CDKN3, TP53, and PHB emerging as the most prominently upregulated genes in tumor tissues. Among these genes, seven were identified as prognostic markers, with HSPB8 being the sole protective factor. Notably, our analysis also revealed the existence of two distinct molecular subtypes within LUAD, each characterized by unique prognoses and immune responses. Specifically, Subtype B displayed a poorer prognosis but demonstrated increased sensitivity to both chemotherapy and immunotherapy. In addition, our development of a Paraptosis-Associated Risk Score yielded a significant prognostic value in predicting patient outcomes. Furthermore, we found regulatory effect of CDKN3 on Paraptosis in two cell lines. CONCLUSIONS: Our study highlights the importance of PRGs in LUAD, particularly in prognosis and treatment response. The identified molecular subtypes and Paraptosis-Associated Risk Score offer valuable insights for personalized treatment strategies.

An anti-tumor protein PFAP specifically interacts with cholesterol-enriched membrane domains of A549 cells and induces paraptosis and endoplasmic reticulum stress.

Nowadays, non-small cell lung cancer (NSCLC) is still one of the most life-threatening diseases in the world. In previous studies, a fungal protein PFAP with anti-NSCLC properties was isolated and identified from Pleurotus ferulae lanzi. In this study, the amino acid sequence of PFAP was analyzed and found to be highly homologous to the aegerolysin family. PFAP, like other members of the aegerolysin family, specifically recognizes lipid raft domains rich in cholesterol and sphingomyelin, which is probably its specific anti-tumor mechanism. Previous studies have shown that PFAP can induce AMPK-mediated autophagy and G1-phase cell cycle arrest in A549 lung cancer cells. This study further revealed that PFAP can also induce paraptosis and endoplasmic reticulum stress (ERS) in A549 cells in vitro by targeting AMPK. PFAP induces multi-pathway death of A549 cells, and thus demonstrates its potential value for developing new drugs for NSCLC.

Cannabidiol activates MAPK pathway to induce apoptosis, paraptosis, and autophagy in colorectal cancer cells.

Mitogen-activated protein kinase (MAPK) activation by natural compounds is known to be involved in the induction of apoptosis, paraptosis, and autophagy. Cannabidiol (CBD), a bioactive compound found in Cannabis sativa, is endowed with many pharmacological activities. We investigated the cytotoxic effect of CBD in a panel of colorectal cancer (CRC) cells (HT-29, SW480, HCT-116, and HCT-15). CBD induced significant cytotoxicity as evidenced by the results of MTT  assay, live-dead assay, and flow cytometric analysis. Since CBD displayed cytotoxicity against CRC cells, we examined the effect of CBD on apoptosis, paraptosis, and autophagy. CBD decreased the expression of antiapoptotic proteins and increased the Annexin-V-positive as well as TUNEL-positive cells suggesting that CBD induces apoptosis. CBD increased the expression of ATF4 (activating transcription factor 4) and CHOP (CCAAT/enhancer-binding protein homologous protein), elevated endoplasmic reticulum stress, and enhanced reactive oxygen species levels indicating that CBD also promotes paraptosis. CBD also induced the expression of Atg7, phospho-Beclin-1, and LC3 suggesting that CBD also accelerates autophagy. Since, the MAPK pathway is a common cascade that is involved in the regulation of apoptosis, paraptosis, and autophagy, we investigated the effect of CBD on the activation of JNK, p38, and ERK pathways. CBD activated all the forms of MAPK proteins and pharmacological inhibition of these proteins reverted the observed effects. Our findings implied that CBD could induce CRC cell death by activating apoptosis, paraptosis, and autophagy through the activation of the MAPK pathway.

Apoptosis and Paraptosis Induced by Disulfiram-Loaded Ca2+/Cu2+ Dual-Ions Nano Trap for Breast Cancer Treatment.

Regarded as one of the hallmarks of tumorigenesis and tumor progression, the evasion of apoptotic cell death would also account for treatment resistance or failure during cancer therapy. In this study, a Ca2+/Cu2+ dual-ion "nano trap" to effectively avoid cell apoptosis evasion by synchronously inducing paraptosis together with apoptosis was successfully designed and fabricated for breast cancer treatment. In brief, disulfiram (DSF)-loaded amorphous calcium carbonate nanoparticles (NPs) were fabricated via a gas diffusion method. Further on, the Cu2+-tannic acid metal phenolic network was embedded onto the NPs surface by self-assembling, followed by mDSPE-PEG/lipid capping to form the DSF-loaded Ca2+/Cu2+ dual-ions "nano trap". The as-prepared nanotrap would undergo acid-triggered biodegradation upon being endocytosed by tumor cells within the lysosome for Ca2+, Cu2+, and DSF releasing simultaneously. The released Ca2+ could cause mitochondrial calcium overload and lead to hydrogen peroxide (H2O2) overexpression. Meanwhile, Ca2+/reactive oxygen species-associated mitochondrial dysfunction would lead to paraptosis cell death. Most importantly, cell paraptosis could be further induced and strengthened by the toxic dithiocarbamate (DTC)-copper complexes formed by the Cu2+ combined with the DTC, the metabolic products of DSF. Additionally, the released Cu2+ will be reduced by intracellular glutathione to generate Cu+, which can catalyze the H2O2 to produce a toxic hydroxyl radical by a Cu+-mediated Fenton-like reaction for inducing cell apoptosis. Both in vitro cellular assays and in vivo antitumor evaluations confirmed the cancer therapeutic efficiency by the dual ion nano trap. This study can broaden the cognition scope of dual-ion-mediated paraptosis together with apoptosis via a multifunctional nanoplatform.

The Mechanisms of Regulated Cell Death: Structural and Functional Proteomic Pathways Induced or Inhibited by a Specific Protein-A Narrative Review.

Billions of cells die in us every hour, and our tissues do not shrink because there is a natural regulation where Cell Death (CD) is balanced with cell division. The process in which cells eliminate themselves in a controlled manner is called Programmed Cell Death (PCD). The PCD plays an important role during embryonic development, in maintaining homeostasis of the body's tissues, and in the elimination of damaged cells, under a wide range of physiological and developmental stimuli. A multitude of protein mediators of PCD have been identified and signals have been found to utilize common pathways elucidating the proteins involved. This narrative review focuses on caspase-dependent and caspase-independent PCD pathways. Included are studies of caspase-dependent PCD such as Anoikis, Catastrophe Mitotic, Pyroptosis, Emperitosis, Parthanatos and Cornification, and Caspase-Independent PCD as Wallerian Degeneration, Ferroptosis, Paraptosis, Entosis, Methuosis, and Extracellular Trap Abnormal Condition (ETosis), as well as neutrophil extracellular trap abnormal condition (NETosis) and Eosinophil Extracellular Trap Abnormal Condition (EETosis). Understanding PCD from those reported in this review could shed substantial light on the processes of biological homeostasis. In addition, identifying specific proteins involved in these processes is mandatory to identify molecular biomarkers, as well as therapeutic targets. This knowledge could provide the ability to modulate the PCD response and could lead to new therapeutic interventions in a wide range of diseases.

Paraptosis: a non-classical paradigm of cell death for cancer therapy.

Due to the sustained proliferative potential of cancer cells, inducing cell death is a potential strategy for cancer therapy. Paraptosis is a mode of cell death characterized by endoplasmic reticulum (ER) and/or mitochondrial swelling and cytoplasmic vacuolization, which is less investigated. Considerable evidence shows that paraptosis can be triggered by various chemical compounds, particularly in cancer cells, thus highlighting the potential application of this non-classical mode of cell death in cancer therapy. Despite these findings, there remain significant gaps in our understanding of the role of paraptosis in cancer. In this review, we summarize the current knowledge on chemical compound-induced paraptosis. The ER and mitochondria are the two major responding organelles in chemical compound-induced paraptosis, which can be triggered by the reduction of protein degradation, disruption of sulfhydryl homeostasis, overload of mitochondrial Ca2+, and increased generation of reactive oxygen species. We also discuss the stumbling blocks to the development of this field and the direction for further research. The rational use of paraptosis might help us develop a new paradigm for cancer therapy.

Aloperine targets lysosomes to inhibit late autophagy and induces cell death through apoptosis and paraptosis in glioblastoma.

Glioblastoma (GBM) is an aggressive intracranial tumour, and current chemotherapy regimens have limited efficacy. Aloperine (ALO), a natural alkaline compound, has shown potential as an antitumor agent. However, the effect of ALO against GBM remains unclear. This study aimed to investigate the function of ALO in treating GBM. U87, A172, and GL261 cell lines were used for in vitro experiments, and GL261 was also used to establish in vivo models. The results showed that ALO inhibited the proliferation of GBM cells by cell cycle arrest and apoptosis. Furthermore, autophagy was found to play a critical role, suggested by observation of autophagosomes under the transmission electron microscopy. It was discovered for the first time that ALO targeted lysosomes directly in glioma cells, tested by fluo-rescence-labelled ALO and organelle-localizing probes. In addition, ALO inhibited late autophagy and induced paraptosis in GBM, verified by classical gene expression changes in qPCR and western blotting. Also, ALO inhibited tumour growth and acted synergistically with temozolomide in intracranial glioma mice models in vivo. Our findings suggest that ALO targets lysosomes to inhibit late autophagy in GBM, inducing cell cycle arrest, paraptosis, and apoptosis. ALO may therefore be a promising therapeutic agent for the treatment of GBM.

Ferritinophagy-mediated apoptosis and paraptosis induction involved MAPK and PI3K/AKT pathway in mechanism of an iron chelator.

Melanoma cells were more resistant to ferroptosis with still poor therapy outcomes. Sensitizing melanoma cell to the ferroptosis inducer was a crucial strategy for treatment of melanoma. In the present study, 2,2'-di-pyridylketone hydrazone dithiocarbamate s-butyric acid (DpdtbA) displayed superior inhibitory activity than ferroptosis inducer Erastin in melanoma cells, which prompt us to explore the underlying mechanism. The analyses from flow cytometry and Western blot showed that the growth inhibition of DpdtbA against SK-MEL-28 and A375 cells involved apoptosis induction and G1 phase arrest. Surprisingly, the cytoplasmic vacuoles were found upon the treatment; transmission electron microscopy and endoplasmic reticulum (ER) staining revealed that the cytoplasmic vacuoles were in ER; while down-regulation of alix and requirement of protein synthesis suggested there was an occurrence of paraptosis. However, both NAC and 3-MA could significantly attenuate the cytoplasmic vacuolization and growth inhibition, hinting that both ROS and autophagy involved the paraptosis induction. The additional evidence revealed that there was an occurrence of continuous ferritinophagy, which was responsible for the ROS production. Downregulation of NCOA4 clearly attenuated the apoptosis and paraptosis induction. In addition, activation of MAPK involved regulation of paraptosis, but only ERK and JNK had role in the formation of cytoplasmic vacuoles and growth inhibition. Furthermore, a ROS dependent regulation of PI3K/AKT pathway was also involved. Taken together, our result firstly demonstrated that a continuous ferritinophagy contributed to the apoptosis and paraptosis induction, highlighting that the lysosomal labile iron pool had a crucial role in control of melanoma cell fate.

Rifampicin-induced ER stress and excessive cytoplasmic vacuolization instigate hepatotoxicity via alternate programmed cell death paraptosis in vitro and in vivo.

AIMS: Rifampicin-induced hepatotoxicity is a primary cause of drug-induced liver injury (DILI), posing a significant challenge to its continued clinical application. Moreover, the mechanism underlying rifampicin-induced hepatotoxicity remains unclear. MAIN METHODS: Human hepatocyte line-17 (HHL-17) cells were treated with an increasing dose of rifampicin for 24 h, and male Wistar rats were given rifampicin [150 mg/kg body weight (bw)] orally for 28 days. Viability assay, protein expression, and cell death assays were analyzed in vitro. Moreover, liver serum markers, body/organ weight, H&E staining, protein expression, etc., were assayed in vivo. KEY FINDINGS: Rifampicin induced a dose-dependent hepatotoxicity in HHL-17 cells (IC50; 600 µM), and increased the serum levels of liver injury markers, e.g., alanine transaminase (ALT) and aspartate transaminase (AST) in rats. Rifampicin-induced cell death was non-apoptotic and non-necroptotic both in vitro and in vivo. Further, excessive cellular vacuolization and reduced expression of Alix protein confirmed the induction of paraptosis both in vitro and in vivo. In addition, a significant increase in the endoplasmic reticulum (ER) stress markers (e.g., BiP, CHOP, and total polyubiquitinated proteins) was detected, demonstrating the induction of ER stress and altered protein homeostasis. Interestingly, rifampicin-induced hepatotoxicity was associated with the inhibition of autophagy and enhanced reactive oxygen species (ROS) generation in HHL-17 cells. Furthermore, inhibition of protein synthesis by cycloheximide (CHX) suppressed paraptosis by alleviating rifampicin-induced ER stress and ROS generation. SIGNIFICANCE: Rifampicin-induced hepatotoxicity involves ER stress-driven paraptosis as a novel mechanism of its toxicity that may be targeted to protect liver cells from rifampicin toxicity.

Cyclometalated iridium(III) complexes induce immunogenic cell death in HepG2 cells via paraptosis.

Immunotherapy has been shown to provide superior antitumor efficacy by activating the innate immune system to recognize, attack and eliminate tumor cells without seriously harming normal cells. Herein, we designed and synthesized three new cyclometalated iridium(III) complexes (Ir1, Ir2, Ir3) then evaluated their antitumor activity. When co-incubated with HepG2 cells, the complex Ir1 localized in the lysosome, where it induced paraptosis and endoplasmic reticulum stress (ER stress). Notably, Ir1 also induced immunogenic cell death (ICD), promoted dendritic cell maturation that enhanced effector T cell chemotaxis to tumor tissues, down-regulated proportions of immunosuppressive regulatory T cells within tumor tissues and triggered activation of antitumor immunity throughout the body. To date, Ir1 is the first reported iridium(III) complex-based paraptosis inducer to successfully induce tumor cell ICD. Furthermore, Ir1 induced ICD of HepG2 cells without affecting cell cycle or reactive oxygen species levels.

Corrigendum: Paraptosis: a unique cell death mode for targeting cancer.

[This corrects the article DOI: 10.3389/fphar.2023.1159409.].