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Cancer Discovery Nov 2021Cancer cells must overcome anoikis (detachment-induced death) to successfully metastasize. Using proteomic screens, we found that distinct oncoproteins upregulate IL1...
Cancer cells must overcome anoikis (detachment-induced death) to successfully metastasize. Using proteomic screens, we found that distinct oncoproteins upregulate IL1 receptor accessory protein (IL1RAP) to suppress anoikis. IL1RAP is directly induced by oncogenic fusions of Ewing sarcoma, a highly metastatic childhood sarcoma. IL1RAP inactivation triggers anoikis and impedes metastatic dissemination of Ewing sarcoma cells. Mechanistically, IL1RAP binds the cell-surface system X transporter to enhance exogenous cystine uptake, thereby replenishing cysteine and the glutathione antioxidant. Under cystine depletion, IL1RAP induces cystathionine gamma lyase (CTH) to activate the transsulfuration pathway for cysteine synthesis. Therefore, IL1RAP maintains cyst(e)ine and glutathione pools, which are vital for redox homeostasis and anoikis resistance. IL1RAP is minimally expressed in pediatric and adult normal tissues, and human anti-IL1RAP antibodies induce potent antibody-dependent cellular cytotoxicity of Ewing sarcoma cells. Therefore, we define IL1RAP as a new cell-surface target in Ewing sarcoma, which is potentially exploitable for immunotherapy. SIGNIFICANCE: Here, we identify cell-surface protein IL1RAP as a key driver of metastasis in Ewing sarcoma, a highly aggressive childhood sarcoma. Minimal expression in pediatric and adult normal tissues nominates IL1RAP as a promising target for immunotherapy...
Topics: Adult; Anoikis; Cell Line, Tumor; Child; Humans; Interleukin-1 Receptor Accessory Protein; Proteomics; Receptors, Interleukin-1; Sarcoma, Ewing
PubMed: 34021002
DOI: 10.1158/2159-8290.CD-20-1690 -
British Journal of Cancer Jun 2022Lymph node (LN) metastasis confers gastric cancer (GC) progression, poor survival and cancer-related death. Aberrant activation of Wnt/β-catenin promotes...
BACKGROUND
Lymph node (LN) metastasis confers gastric cancer (GC) progression, poor survival and cancer-related death. Aberrant activation of Wnt/β-catenin promotes epithelial-mesenchymal transition (EMT) and LN metastasis, whereas the constitutive activation mutation of Wnt/β-catenin is rare in GC, suggesting that the underlying mechanisms enhancing Wnt/β-catenin activation need to be further investigated and understood.
METHODS
Bioinformatics analyses and immunohistochemistry (IHC) were used to identify and detect LN metastasis-related genes in GC. Cellular functional assays and footpad inoculation mouse model illustrate the biological function of CCT5. Co-immunoprecipitation assays, western blot and qPCR elucidate the interaction between CCT5 and E-cadherin, and the regulation on β-catenin activity.
RESULTS
CCT5 is upregulated in LN metastatic GCs and correlates with poor prognosis. In vitro assays prove that CCT5 markedly promotes GC cell proliferation, anti-anoikis, invasion and lymphatic tube formation. Moreover, CCT5 enhances xenograft GC growth and popliteal lymph node metastasis in vivo. Furthermore, CCT5 binds the cytoplasmic domain of E-cadherin and abrogates the interaction between E-cadherin and β-catenin, thereby releasing β-catenin to the nucleus and enhancing Wnt/β-catenin signalling activity and EMT.
CONCLUSION
CCT5 promotes GC progression and LN metastasis by enhancing wnt/β-catenin activation, suggesting a great potential of CCT5 as a biomarker for GC diagnosis and therapy.
Topics: Animals; Cell Line, Tumor; Cell Movement; Cell Proliferation; Chaperonin Containing TCP-1; Epithelial-Mesenchymal Transition; Heterografts; Humans; Lymphatic Metastasis; Mice; Neoplasm Metastasis; Stomach Neoplasms; Wnt Signaling Pathway; beta Catenin
PubMed: 35194191
DOI: 10.1038/s41416-022-01747-0 -
Nature Mar 2023Most human cells require anchorage for survival. Cell-substrate adhesion activates diverse signalling pathways, without which cells undergo anoikis-a form of programmed...
Most human cells require anchorage for survival. Cell-substrate adhesion activates diverse signalling pathways, without which cells undergo anoikis-a form of programmed cell death. Acquisition of anoikis resistance is a pivotal step in cancer disease progression, as metastasizing cells often lose firm attachment to surrounding tissue. In these poorly attached states, cells adopt rounded morphologies and form small hemispherical plasma membrane protrusions called blebs. Bleb function has been thoroughly investigated in the context of amoeboid migration, but it has been examined far less in other scenarios. Here we show by three-dimensional imaging and manipulation of cell morphological states that blebbing triggers the formation of plasma membrane-proximal signalling hubs that confer anoikis resistance. Specifically, in melanoma cells, blebbing generates plasma membrane contours that recruit curvature-sensing septin proteins as scaffolds for constitutively active mutant NRAS and effectors. These signalling hubs activate ERK and PI3K-well-established promoters of pro-survival pathways. Inhibition of blebs or septins has little effect on the survival of well-adhered cells, but in detached cells it causes NRAS mislocalization, reduced MAPK and PI3K activity, and ultimately, death. This unveils a morphological requirement for mutant NRAS to operate as an effective oncoprotein. Furthermore, whereas some BRAF-mutated melanoma cells do not rely on this survival pathway in a basal state, inhibition of BRAF and MEK strongly sensitizes them to both bleb and septin inhibition. Moreover, fibroblasts engineered to sustain blebbing acquire the same anoikis resistance as cancer cells even without harbouring oncogenic mutations. Thus, blebs are potent signalling organelles capable of integrating myriad cellular information flows into concerted cellular responses, in this case granting robust anoikis resistance.
Topics: Humans; Anoikis; Cell Survival; Melanoma; Phosphatidylinositol 3-Kinases; Septins; Signal Transduction; Cell Surface Extensions; Carcinogenesis; Cell Adhesion; Extracellular Signal-Regulated MAP Kinases; Fibroblasts; Mutation; Cell Shape; Imaging, Three-Dimensional; Mitogen-Activated Protein Kinase Kinases
PubMed: 36859545
DOI: 10.1038/s41586-023-05758-6 -
Journal of Experimental & Clinical... Jan 2022The development of lethal cancer metastasis depends on the dynamic interactions between cancer cells and the tumor microenvironment, both of which are embedded in the...
BACKGROUND
The development of lethal cancer metastasis depends on the dynamic interactions between cancer cells and the tumor microenvironment, both of which are embedded in the extracellular matrix (ECM). The acquisition of resistance to detachment-induced apoptosis, also known as anoikis, is a critical step in the metastatic cascade. Thus, a more in-depth and systematic analysis is needed to identify the key drivers of anoikis resistance.
METHODS
Genome-wide CRISPR/Cas9 knockout screen was used to identify critical drivers of anoikis resistance using SKOV3 cell line and found protein-L-isoaspartate (D-aspartate) O-methyltransferase (PCMT1) as a candidate. Quantitative real-time PCR (qRT-PCR) and immune-histochemistry (IHC) were used to measure differentially expressed PCMT1 in primary tissues and metastatic cancer tissues. PCMT1 knockdown/knockout and overexpression were performed to investigate the functional role of PCMT1 in vitro and in vivo. The expression and regulation of PCMT1 and integrin-FAK-Src pathway were evaluated using immunoprecipitation followed by mass spectrometry (IP-MS), western blot analysis and live cell imaging.
RESULTS
We found that PCMT1 enhanced cell migration, adhesion, and spheroid formation in vitro. Interestingly, PCMT1 was released from ovarian cancer cells, and interacted with the ECM protein LAMB3, which binds to integrin and activates FAK-Src signaling to promote cancer progression. Strikingly, treatment with an antibody against extracellular PCMT1 effectively reduced ovarian cancer cell invasion and adhesion. Our in vivo results indicated that overexpression of PCMT1 led to increased ascites formation and distant metastasis, whereas knockout of PCMT1 had the opposite effect. Importantly, PCMT1 was highly expressed in late-stage metastatic tumors compared to early-stage primary tumors.
CONCLUSIONS
Through systematically identifying the drivers of anoikis resistance, we uncovered the contribution of PCMT1 to focal adhesion (FA) dynamics as well as cancer metastasis. Our study suggested that PCMT1 has the potential to be a therapeutic target in metastatic ovarian cancer.
Topics: Animals; CRISPR-Cas Systems; Cell Line, Tumor; Female; Humans; Mice; Mice, Nude; Middle Aged; Neoplasm Metastasis; Ovarian Neoplasms; Protein D-Aspartate-L-Isoaspartate Methyltransferase
PubMed: 35033172
DOI: 10.1186/s13046-022-02242-3 -
Theranostics 2020Peritoneal metastasis predicts poor prognosis of gastric cancer (GC) patients, and the underlying mechanisms are poorly understood. The 2-DIGE, MALDI-TOF/TOF MS and...
Peritoneal metastasis predicts poor prognosis of gastric cancer (GC) patients, and the underlying mechanisms are poorly understood. The 2-DIGE, MALDI-TOF/TOF MS and single-cell transcriptome were used to detect differentially expressed proteins among normal gastric mucosa, primary GC and peritoneal metastatic tissues. Lentiviruses carrying shRNA and transcription activator-like effector nuclease technology were used to knock down myosin heavy chain 9 (MYH9) expression in GC cell lines. Immunofluorescence, immune transmission electron microscopy, chromatin fractionation, co-immunoprecipitation, and assays for chromatin immunoprecipitation, dual luciferase reporter, agarose-oligonucleotide pull-down, flow cytometry and cell anoikis were performed to uncover nuclear MYH9-induced β-catenin () transcription . Nude mice and conditional transgenic mice were used to investigate the findings . We observed that MYH9 was upregulated in metastatic GC tissues and was associated with a poor prognosis of GC patients. Mechanistically, we confirmed that MYH9 was mainly localized in the GC cell nuclei by four potential nuclear localization signals. Nuclear MYH9 bound to the promoter through its DNA-binding domain, and interacted with myosin light chain 9, β-actin and RNA polymerase II to promote transcription, which conferred resistance to anoikis in GC cells and . Staurosporine reduced nuclear MYH9 S1943 phosphorylation to inhibit transcription, Wnt/β-catenin signaling activation and GC progression in both orthotropic xenograft GC nude mouse and transgenic GC mouse models. This study identified that nuclear MYH9-induced CTNNB1 expression promotes GC metastasis, which could be inhibited by staurosporine, indicating a novel therapy for GC peritoneal metastasis.
Topics: Animals; Anoikis; Cell Line, Tumor; Cell Movement; Cell Nucleus; Cell Proliferation; Chemotherapy, Adjuvant; Female; Gastrectomy; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Humans; Lung Neoplasms; Male; Mice; Mice, Transgenic; Myosin Heavy Chains; Phosphorylation; Promoter Regions, Genetic; Staurosporine; Stomach; Stomach Neoplasms; Transcriptional Activation; Up-Regulation; Wnt Signaling Pathway; Xenograft Model Antitumor Assays; beta Catenin
PubMed: 32685004
DOI: 10.7150/thno.46001 -
Proceedings of the National Academy of... Jun 2023Although recent studies demonstrate active mitochondrial metabolism in cancers, the precise mechanisms through which mitochondrial factors contribute to cancer...
Although recent studies demonstrate active mitochondrial metabolism in cancers, the precise mechanisms through which mitochondrial factors contribute to cancer metastasis remain elusive. Through a customized mitochondrion RNAi screen, we identified succinyl-CoA ligase ADP-forming subunit beta (SUCLA2) as a critical anoikis resistance and metastasis driver in human cancers. Mechanistically, SUCLA2, but not the alpha subunit of its enzyme complex, relocates from mitochondria to the cytosol upon cell detachment where SUCLA2 then binds to and promotes the formation of stress granules. SUCLA2-mediated stress granules facilitate the protein translation of antioxidant enzymes including catalase, which mitigates oxidative stress and renders cancer cells resistant to anoikis. We provide clinical evidence that SUCLA2 expression correlates with catalase levels as well as metastatic potential in lung and breast cancer patients. These findings not only implicate SUCLA2 as an anticancer target, but also provide insight into a unique, noncanonical function of SUCLA2 that cancer cells co-opt to metastasize.
Topics: Humans; Catalase; Stress Granules; Succinate-CoA Ligases; Oxidation-Reduction; Neoplasms
PubMed: 37253003
DOI: 10.1073/pnas.2217332120 -
Cancer Gene Therapy Nov 2022Transmembrane ATPases are membrane-bound enzyme complexes and ion transporters that can be divided into F-, V-, and A-ATPases according to their structure. The... (Review)
Review
Transmembrane ATPases are membrane-bound enzyme complexes and ion transporters that can be divided into F-, V-, and A-ATPases according to their structure. The V-ATPases, also known as H-ATPases, are large multi-subunit protein complexes composed of a peripheral domain (V1) responsible for the hydrolysis of ATP and a membrane-integrated domain (V0) that transports protons across plasma membrane or organelle membrane. V-ATPases play a fundamental role in maintaining pH homeostasis through lysosomal acidification and are involved in modulating various physiological and pathological processes, such as macropinocytosis, autophagy, cell invasion, and cell death (e.g., apoptosis, anoikis, alkaliptosis, ferroptosis, and lysosome-dependent cell death). In addition to participating in embryonic development, V-ATPase pathways, when dysfunctional, are implicated in human diseases, such as neurodegenerative diseases, osteopetrosis, distal renal tubular acidosis, and cancer. In this review, we summarize the structure and regulation of isoforms of V-ATPase subunits and discuss their context-dependent roles in cancer biology and cell death. Updated knowledge about V-ATPases may enable us to design new anticancer drugs or strategies.
Topics: Humans; Vacuolar Proton-Translocating ATPases; Cell Membrane; Neoplasms; Cell Death
PubMed: 35504950
DOI: 10.1038/s41417-022-00477-y -
Cell Communication and Signaling : CCS Sep 2023Metastatic cancer cells can develop anoikis resistance in the absence of substrate attachment and survive to fight tumors. Anoikis is mediated by endogenous... (Review)
Review
Metastatic cancer cells can develop anoikis resistance in the absence of substrate attachment and survive to fight tumors. Anoikis is mediated by endogenous mitochondria-dependent and exogenous death receptor pathways, and studies have shown that caspase-8-dependent external pathways appear to be more important than the activity of the intrinsic pathways. This paper reviews the regulation of anoikis by external pathways mediated by death receptors. Different death receptors bind to different ligands to activate downstream caspases. The possible mechanisms of Fas-associated death domain (FADD) recruitment by Fas and TNF receptor 1 associated-death domain (TRADD) recruitment by tumor necrosis factor receptor 1 (TNFR1), and DR4- and DR5-associated FADD to induce downstream caspase activation and regulate anoikis were reviewed. This review highlights the possible mechanism of the death receptor pathway mediation of anoikis and provides new insights and research directions for studying tumor metastasis mechanisms. Video Abstract.
Topics: Anoikis; Proteolysis; Caspases; Mitochondria; Protein Processing, Post-Translational
PubMed: 37667281
DOI: 10.1186/s12964-023-01247-5 -
Thoracic Cancer Jan 2023Lung adenocarcinoma (LUAD) is the most prevalent histotype of non-small cell lung cancer. Anoikis, an alternative form of programmed cell death, plays a pivotal role in...
BACKGROUND
Lung adenocarcinoma (LUAD) is the most prevalent histotype of non-small cell lung cancer. Anoikis, an alternative form of programmed cell death, plays a pivotal role in cancer invasion and metastasis, preventing the detached cancer cells from readhering to other substrates for abnormal proliferation. The aim of this study was to conduct a comprehensive analyses of the prognostic implications of anoikis-related genes (ARGs) in LUAD.
METHODS
ARGs were selected from The Cancer Genome Atlas (TCGA) database and Genecards dataset using differential expression analysis. The signature incorporating ARGs was identified using univariate Cox regression analysis and LASSO regression analysis. Furthermore, a nomogram containing the signature and clinical information was developed through univariate and multivariate Cox regression analysis. Kaplan-Meier survival analysis and receiver operating characteristic (ROC) curves were applied to evaluate the predictive validity of these risk models. Finally, functional analysis of the selected ARGs in signature and analysis of immune landscape were also conducted.
RESULTS
A 16-gene signature was integrated to stratify LUAD patients into different survival risk groups. The prognostic risk score generated from the signature and TNM stage were identified as independent prognostic factors and utilized to develop a nomogram. Both the signature and the nomogram showed satisfactory prediction performance in predicting overall survival (OS) of LUAD patients. The ARGs were enriched in several biological functions and signaling pathways. Finally, differences of immune landscape were investigated among the high- and low-risk groups stratified by the signature.
CONCLUSIONS
This study revealed potential relationships between ARGs and prognosis of LUAD. The prognostic predictors identified in present study could be utilized as potential biomarkers for clinical applications.
Topics: Humans; Anoikis; Carcinoma, Non-Small-Cell Lung; Tumor Microenvironment; Lung Neoplasms; Prognosis; Adenocarcinoma of Lung
PubMed: 36507553
DOI: 10.1111/1759-7714.14766 -
Frontiers in Immunology 2023Stroke is the second leading cause of death and the third leading cause of disability worldwide, with ischemic stroke (IS) being the most prevalent. A substantial number...
BACKGROUND
Stroke is the second leading cause of death and the third leading cause of disability worldwide, with ischemic stroke (IS) being the most prevalent. A substantial number of irreversible brain cell death occur in the short term, leading to impairment or death in IS. Limiting the loss of brain cells is the primary therapy target and a significant clinical issue for IS therapy. Our study aims to establish the gender specificity pattern from immune cell infiltration and four kinds of cell-death perspectives to improve IS diagnosis and therapy.
METHODS
Combining and standardizing two IS datasets (GSE16561 and GSE22255) from the GEO database, we used the CIBERSORT algorithm to investigate and compare the immune cell infiltration in different groups and genders. Then, ferroptosis-related differently expressed genes (FRDEGs), pyroptosis-related DEGs (PRDEGs), anoikis-related DEGs (ARDEGs), and cuproptosis-related DEGs (CRDEGs) between the IS patient group and the healthy control group were identified in men and women, respectively. Machine learning (ML) was finally used to generate the disease prediction model for cell death-related DEGs (CDRDEGs) and to screen biomarkers related to cell death involved in IS.
RESULTS
Significant changes were observed in 4 types of immune cells in male IS patients and 10 types in female IS patients compared with healthy controls. In total, 10 FRDEGs, 11 PRDEGs, 3 ARDEGs, and 1 CRDEG were present in male IS patients, while 6 FRDEGs, 16 PRDEGs, 4 ARDEGs, and 1 CRDEG existed in female IS patients. ML techniques indicated that the best diagnostic model for both male and female patients was the support vector machine (SVM) for CDRDEG genes. SVM's feature importance analysis demonstrated that SLC2A3, MMP9, C5AR1, ACSL1, and NLRP3 were the top five feature-important CDRDEGs in male IS patients. Meanwhile, the PDK4, SCL40A1, FAR1, CD163, and CD96 displayed their overwhelming influence on female IS patients.
CONCLUSION
These findings contribute to a better knowledge of immune cell infiltration and their corresponding molecular mechanisms of cell death and offer distinct clinically relevant biological targets for IS patients of different genders.
Topics: Female; Humans; Male; Ischemic Stroke; Cell Death; Biomarkers; Brain; Stroke; Brain Death
PubMed: 37435058
DOI: 10.3389/fimmu.2023.1164742