RESUMO
Type I and II interferons (IFNs) stimulate pro-inflammatory programs that are critical for immune activation, but also induce immune-suppressive feedback circuits that impede control of cancer growth. Here, we sought to determine how these opposing programs are differentially induced. We demonstrated that the transcription factor interferon regulatory factor 2 (IRF2) was expressed by many immune cells in the tumor in response to sustained IFN signaling. CD8+ T cell-specific deletion of IRF2 prevented acquisition of the T cell exhaustion program within the tumor and instead enabled sustained effector functions that promoted long-term tumor control and increased responsiveness to immune checkpoint and adoptive cell therapies. The long-term tumor control by IRF2-deficient CD8+ T cells required continuous integration of both IFN-I and IFN-II signals. Thus, IRF2 is a foundational feedback molecule that redirects IFN signals to suppress T cell responses and represents a potential target to enhance cancer control.
Assuntos
Interferon Tipo I , Neoplasias , Humanos , Fator Regulador 2 de Interferon/genética , Linfócitos T CD8-Positivos , Fatores de Transcrição , Exaustão das Células T , Neoplasias/patologiaRESUMO
Extensive cellular heterogeneity exists within specific immune-cell subtypes classified as a single lineage, but its molecular underpinnings are rarely characterized at a genomic scale. Here, we use single-cell RNA-seq to investigate the molecular mechanisms governing heterogeneity and pathogenicity of Th17 cells isolated from the central nervous system (CNS) and lymph nodes (LN) at the peak of autoimmune encephalomyelitis (EAE) or differentiated in vitro under either pathogenic or non-pathogenic polarization conditions. Computational analysis relates a spectrum of cellular states in vivo to in-vitro-differentiated Th17 cells and unveils genes governing pathogenicity and disease susceptibility. Using knockout mice, we validate four new genes: Gpr65, Plzp, Toso, and Cd5l (in a companion paper). Cellular heterogeneity thus informs Th17 function in autoimmunity and can identify targets for selective suppression of pathogenic Th17 cells while potentially sparing non-pathogenic tissue-protective ones.
Assuntos
Encefalomielite Autoimune Experimental/patologia , Análise de Sequência de RNA , Análise de Célula Única , Células Th17/metabolismo , Células Th17/patologia , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Transporte/metabolismo , Sistema Nervoso Central/patologia , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Perfilação da Expressão Gênica , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Linfonodos/patologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Glicoproteína Mielina-Oligodendrócito/metabolismo , Fragmentos de Peptídeos/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica , Receptores Acoplados a Proteínas G/metabolismo , Receptores Imunológicos/metabolismo , Receptores Depuradores , Células Th17/imunologiaRESUMO
The molecular mechanisms whereby CD8+ T cells become "exhausted" in the tumor microenvironment remain unclear. Programmed death ligand-1 (PD-L1) is upregulated on tumor cells and PD-1-PD-L1 blockade has significant efficacy in human tumors; however, most patients do not respond, suggesting additional mechanisms underlying T cell exhaustion. B7 superfamily member 1 (B7S1), also called B7-H4, B7x, or VTCN1, negatively regulates T cell activation. Here we show increased B7S1 expression on myeloid cells from human hepatocellular carcinoma correlated with CD8+ T cell dysfunction. B7S1 inhibition suppressed development of murine tumors. Putative B7S1 receptor was co-expressed with PD-1 but not T cell immunoglobulin and mucin-domain containing-3 (Tim-3) at an activated state of early tumor-infiltrating CD8+ T cells, and B7S1 promoted T cell exhaustion, possibly through Eomes overexpression. Combinatorial blockade of B7S1 and PD-1 synergistically enhanced anti-tumor immune responses. Collectively, B7S1 initiates dysfunction of tumor-infiltrating CD8+ T cells and may be targeted for cancer immunotherapy.
Assuntos
Antígeno B7-H1/metabolismo , Linfócitos T CD8-Positivos/imunologia , Carcinoma Hepatocelular/imunologia , Neoplasias Hepáticas/imunologia , Linfócitos do Interstício Tumoral/imunologia , Células Mieloides/imunologia , Inibidor 1 da Ativação de Células T com Domínio V-Set/metabolismo , Animais , Carcinoma Hepatocelular/patologia , Modelos Animais de Doenças , Humanos , Neoplasias Hepáticas/patologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas com Domínio T/metabolismo , Inibidor 1 da Ativação de Células T com Domínio V-Set/genéticaRESUMO
We developed a highly sensitive assay for detecting protein-protein interaction using chimeric receptors comprising two molecules of interest in the extracellular domain and interferon alpha and beta receptor subunit 1 or 2 (IFNAR1/2) in the intracellular domain. This intracellular IFNAR1/2 reconstitution system (IFNARRS) proved markedly more sensitive than the NanoBiT system, currently considered one of the best detection systems for protein interaction. Employing chimeric receptors with extracellular domains from the IFNγ or IL-2 receptor and the intracellular domains of IFNAR1/2, the IFNARRS system effectively identifies low IFNγ or IL-2 levels. Cells stably expressing these chimeric receptors responded to IFNγ secreted by activated T cells following various stimuli, including a specific peptide-antigen. The activation signals were further enhanced by the expression of relevant genes, such as costimulators, via IFN-stimulated response elements in the promoters. Besides IFNγ or IL-2, the IFNARRS system demonstrated the capability to detect other cytokines by using the corresponding extracellular domains from these target cytokine receptors.
Assuntos
Interferon gama , Interleucina-2 , Receptor de Interferon alfa e beta , Linfócitos T , Humanos , Receptor de Interferon alfa e beta/metabolismo , Receptor de Interferon alfa e beta/genética , Linfócitos T/metabolismo , Linfócitos T/imunologia , Interleucina-2/metabolismo , Interferon gama/metabolismo , Receptores de Interleucina-2/metabolismo , Receptores de Interleucina-2/genética , Ligação Proteica , Ativação Linfocitária , Células HEK293RESUMO
Despite extensive analysis of pRB phosphorylation in vitro, how this modification influences development and homeostasis in vivo is unclear. Here, we show that homozygous Rb∆K4 and Rb∆K7 knock-in mice, in which either four or all seven phosphorylation sites in the C-terminal region of pRb, respectively, have been abolished by Ser/Thr-to-Ala substitutions, undergo normal embryogenesis and early development, notwithstanding suppressed phosphorylation of additional upstream sites. Whereas Rb∆K4 mice exhibit telomere attrition but no other abnormalities, Rb∆K7 mice are smaller and display additional hallmarks of premature aging including infertility, kyphosis, and diabetes, indicating an accumulative effect of blocking pRb phosphorylation. Diabetes in Rb∆K7 mice is insulin-sensitive and associated with failure of quiescent pancreatic ß-cells to re-enter the cell cycle in response to mitogens, resulting in induction of DNA damage response (DDR), senescence-associated secretory phenotype (SASP), and reduced pancreatic islet mass and circulating insulin level. Pre-treatment with the epigenetic regulator vitamin C reduces DDR, increases cell cycle re-entry, improves islet morphology, and attenuates diabetes. These results have direct implications for cell cycle regulation, CDK-inhibitor therapeutics, diabetes, and longevity.
Assuntos
Envelhecimento/fisiologia , Ácido Ascórbico/farmacologia , Diabetes Mellitus Experimental/prevenção & controle , Proteína do Retinoblastoma/metabolismo , Animais , Senescência Celular/efeitos dos fármacos , Quinase 2 Dependente de Ciclina/antagonistas & inibidores , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patologia , Fator de Transcrição E2F1/metabolismo , Desenvolvimento Embrionário/genética , Feminino , Fibroblastos/efeitos dos fármacos , Técnicas de Introdução de Genes , Células Secretoras de Insulina/patologia , Camundongos , Fosforilação , Gravidez , Proteína do Retinoblastoma/genética , Telômero/genéticaRESUMO
Activated T cells produce reactive oxygen species (ROS), which trigger the antioxidative glutathione (GSH) response necessary to buffer rising ROS and prevent cellular damage. We report that GSH is essential for T cell effector functions through its regulation of metabolic activity. Conditional gene targeting of the catalytic subunit of glutamate cysteine ligase (Gclc) blocked GSH production specifically in murine T cells. Gclc-deficient T cells initially underwent normal activation but could not meet their increased energy and biosynthetic requirements. GSH deficiency compromised the activation of mammalian target of rapamycin-1 (mTOR) and expression of NFAT and Myc transcription factors, abrogating the energy utilization and Myc-dependent metabolic reprogramming that allows activated T cells to switch to glycolysis and glutaminolysis. In vivo, T-cell-specific ablation of murine Gclc prevented autoimmune disease but blocked antiviral defense. The antioxidative GSH pathway thus plays an unexpected role in metabolic integration and reprogramming during inflammatory T cell responses.
Assuntos
Glutamato-Cisteína Ligase/deficiência , Glutationa/metabolismo , Inflamação/metabolismo , Linfócitos T/metabolismo , Animais , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/metabolismo , Metabolismo Energético/genética , Glutamato-Cisteína Ligase/genética , Glutamina/metabolismo , Glicólise , Immunoblotting , Inflamação/genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fatores de Transcrição NFATC/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/genética , Serina-Treonina Quinases TOR/metabolismoRESUMO
Protein-tyrosine phosphatases (PTPs), along with protein-tyrosine kinases, play key roles in cellular signaling. All Class I PTPs contain an essential active site cysteinyl residue, which executes a nucleophilic attack on substrate phosphotyrosyl residues. The high reactivity of the catalytic cysteine also predisposes PTPs to oxidation by reactive oxygen species, such as H(2)O(2). Reversible PTP oxidation is emerging as an important cellular regulatory mechanism and might contribute to diseases such as cancer. We exploited these unique features of PTP enzymology to develop proteomic methods, broadly applicable to cell and tissue samples, that enable the comprehensive identification and quantification of expressed classical PTPs (PTPome) and the oxidized subset of the PTPome (oxPTPome). We find that mouse and human cells and tissues, including cancer cells, display distinctive PTPomes and oxPTPomes, revealing additional levels of complexity in the regulation of protein-tyrosine phosphorylation in normal and malignant cells.
Assuntos
Proteínas Tirosina Fosfatases/análise , Proteômica/métodos , Animais , Linhagem Celular , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/metabolismo , Oxirredução , RatosRESUMO
Understanding the factors that impede immune responses to persistent viruses is essential in designing therapies for HIV infection. Mice infected with LCMV clone-13 have persistent high-level viremia and a dysfunctional immune response. Interleukin-7, a cytokine that is critical for immune development and homeostasis, was used here to promote immunity toward clone-13, enabling elucidation of the inhibitory pathways underlying impaired antiviral immune response. Mechanistically, IL-7 downregulated a critical repressor of cytokine signaling, Socs3, resulting in amplified cytokine production, increased T cell effector function and numbers, and viral clearance. IL-7 enhanced thymic output to expand the naive T cell pool, including T cells that were not LCMV specific. Additionally, IL-7 promoted production of cytoprotective IL-22 that abrogated liver pathology. The IL-7-mediated effects were dependent on endogenous IL-6. These attributes of IL-7 have profound implications for its use as a therapeutic in the treatment of chronic viral diseases.
Assuntos
Interleucina-7/uso terapêutico , Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/fisiologia , Animais , Antígenos de Diferenciação/metabolismo , Regulação para Baixo , Fatores de Transcrição Forkhead/metabolismo , Humanos , Interleucina-6/imunologia , Interleucina-7/imunologia , Camundongos , Receptor de Morte Celular Programada 1 , Proteínas Recombinantes/metabolismo , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/metabolismo , Linfócitos T/imunologiaRESUMO
Mutations in IDH1, IDH2, and TET2 are recurrently observed in myeloid neoplasms. IDH1 and IDH2 encode isocitrate dehydrogenase isoforms, which normally catalyze the conversion of isocitrate to α-ketoglutarate (α-KG). Oncogenic IDH1/2 mutations confer neomorphic activity, leading to the production of D-2-hydroxyglutarate (D-2-HG), a potent inhibitor of α-KG-dependent enzymes which include the TET methylcytosine dioxygenases. Given their mutual exclusivity in myeloid neoplasms, IDH1, IDH2, and TET2 mutations may converge on a common oncogenic mechanism. Contrary to this expectation, we observed that they have distinct, and even opposite, effects on hematopoietic stem and progenitor cells in genetically engineered mice. Epigenetic and single-cell transcriptomic analyses revealed that Idh2R172K and Tet2 loss-of-function have divergent consequences on the expression and activity of key hematopoietic and leukemogenic regulators. Notably, chromatin accessibility and transcriptional deregulation in Idh2R172K cells were partially disconnected from DNA methylation alterations. These results highlight unanticipated divergent effects of IDH1/2 and TET2 mutations, providing support for the optimization of genotype-specific therapies.
Assuntos
Proteínas de Ligação a DNA , Dioxigenases , Isocitrato Desidrogenase , Células-Tronco , Animais , Camundongos , Dioxigenases/genética , Proteínas de Ligação a DNA/genética , Isocitrato Desidrogenase/genética , Isocitrato Desidrogenase/metabolismo , Ácidos Cetoglutáricos/metabolismo , Mutação , Neoplasias , Células-Tronco/metabolismoRESUMO
Asbestos is the main cause of malignant mesothelioma. Previous studies have linked asbestos-induced mesothelioma to the release of HMGB1 from the nucleus to the cytoplasm, and from the cytoplasm to the extracellular space. In the cytoplasm, HMGB1 induces autophagy impairing asbestos-induced cell death. Extracellularly, HMGB1 stimulates the secretion of TNFα. Jointly, these two cytokines kick-start a chronic inflammatory process that over time promotes mesothelioma development. Whether the main source of extracellular HMGB1 were the mesothelial cells, the inflammatory cells, or both was unsolved. This information is critical to identify the targets and design preventive/therapeutic strategies to interfere with asbestos-induced mesothelioma. To address this issue, we developed the conditional mesothelial HMGB1-knockout (Hmgb1ΔpMeso) and the conditional myelomonocytic-lineage HMGB1-knockout (Hmgb1ΔMylc) mouse models. We establish here that HMGB1 is mainly produced and released by the mesothelial cells during the early phases of inflammation following asbestos exposure. The release of HMGB1 from mesothelial cells leads to atypical mesothelial hyperplasia, and in some animals, this evolves over the years into mesothelioma. We found that Hmgb1ΔpMeso, whose mesothelial cells cannot produce HMGB1, show a greatly reduced inflammatory response to asbestos, and their mesothelial cells express and secrete significantly reduced levels of TNFα. Moreover, the tissue microenvironment in areas of asbestos deposits displays an increased fraction of M1-polarized macrophages compared to M2 macrophages. Supporting the biological significance of these findings, Hmgb1ΔpMeso mice showed a delayed and reduced incidence of mesothelioma and an increased mesothelioma-specific survival. Altogether, our study provides a biological explanation for HMGB1 as a driver of asbestos-induced mesothelioma.
Assuntos
Amianto , Proteína HMGB1 , Mesotelioma Maligno , Mesotelioma , Animais , Camundongos , Fator de Necrose Tumoral alfa/genética , Proteína HMGB1/genética , Mesotelioma/induzido quimicamente , Mesotelioma/genética , Amianto/toxicidade , Inflamação , Microambiente TumoralRESUMO
The E3 ligase ARIH2 has an unusual structure and mechanism of elongating ubiquitin chains. To understand its physiological role, we generated gene-targeted mice deficient in ARIH2. ARIH2 deficiency resulted in the embryonic death of C57BL/6 mice. On a mixed genetic background, the lethality was attenuated, with some mice surviving beyond weaning and then succumbing to an aggressive multiorgan inflammatory response. We found that in dendritic cells (DCs), ARIH2 caused degradation of the inhibitor IκBß in the nucleus, which abrogated its ability to sequester, protect and transcriptionally coactivate the transcription factor subunit p65 in the nucleus. Loss of ARIH2 caused dysregulated activation of the transcription factor NF-κB in DCs, which led to lethal activation of the immune system in ARIH2-sufficent mice reconstituted with ARIH2-deficient hematopoietic stem cells. Our data have therapeutic implications for targeting ARIH2 function.
Assuntos
Células Dendríticas/imunologia , Desenvolvimento Embrionário/imunologia , Insuficiência de Múltiplos Órgãos/imunologia , Ubiquitina-Proteína Ligases/fisiologia , Animais , Células Cultivadas , Modelos Animais de Doenças , Desenvolvimento Embrionário/genética , Hematopoese/genética , Humanos , Sistema Imunitário/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Terapia de Alvo Molecular , Insuficiência de Múltiplos Órgãos/genética , NF-kappa B/metabolismo , Ativação Transcricional/imunologia , Ubiquitina-Proteína Ligases/genética , Ubiquitinação/genética , Ubiquitinação/imunologiaRESUMO
Acute myeloid leukemia (AML) with TP53 mutation is one of the most lethal cancers and portends an extremely poor prognosis. Based on in silico analyses of druggable genes and differential gene expression in TP53-mutated AML, we identified pololike kinase 4 (PLK4) as a novel therapeutic target and examined its expression, regulation, pathogenetic mechanisms, and therapeutic potential in TP53-mutated AML. PLK4 expression was suppressed by activated p53 signaling in TP53 wild-type AML and was increased in TP53-mutated AML cell lines and primary samples. Short-term PLK4 inhibition induced DNA damage and apoptosis in TP53 wild-type AML. Prolonged PLK4 inhibition suppressed the growth of TP53-mutated AML and was associated with DNA damage, apoptosis, senescence, polyploidy, and defective cytokinesis. A hitherto undescribed PLK4/PRMT5/EZH2/H3K27me3 axis was demonstrated in both TP53 wild-type and mutated AML, resulting in histone modification through PLK4-induced PRMT5 phosphorylation. In TP53-mutated AML, combined effects of histone modification and polyploidy activated the cGAS-STING pathway, leading to secretion of cytokines and chemokines and activation of macrophages and T cells upon coculture with AML cells. In vivo, PLK4 inhibition also induced cytokine and chemokine expression in mouse recipients, and its combination with anti-CD47 antibody, which inhibited the "don't-eat-me" signal in macrophages, synergistically reduced leukemic burden and prolonged animal survival. The study shed important light on the pathogenetic role of PLK4 and might lead to novel therapeutic strategies in TP53-mutated AML.
Assuntos
Histonas , Leucemia Mieloide Aguda , Animais , Camundongos , Histonas/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Mutação , Metilação , Nucleotidiltransferases/metabolismo , Leucemia Mieloide Aguda/patologia , Imunidade , PoliploidiaRESUMO
Current vaccine efforts to combat SARS-CoV-2 are focused on the whole spike protein administered as mRNA, viral vector, or protein subunit. However, the SARS-CoV-2 receptor-binding domain (RBD) is the immunodominant portion of the spike protein, accounting for 90% of serum neutralizing activity. In this study, we constructed several versions of RBD and together with aluminum hydroxide or DDA (dimethyldioctadecylammonium bromide)/TDB (d-(+)-trehalose 6,6'-dibehenate) adjuvant evaluated immunogenicity in mice. We generated human angiotensin-converting enzyme 2 knock-in mice to evaluate vaccine efficacy in vivo following viral challenge. We found that 1) subdomain (SD)1 was essential for the RBD to elicit maximal immunogenicity; 2) RBDSD1 produced in mammalian HEK cells elicited better immunogenicity than did protein produced in insect or yeast cells; 3) RBDSD1 combined with the CD4 Th1 adjuvant DDA/TDB produced higher neutralizing Ab responses and stronger CD4 T cell responses than did aluminum hydroxide; 4) addition of monomeric human Fc receptor to RBDSD1 (RBDSD1Fc) significantly enhanced immunogenicity and neutralizing Ab titers; 5) the Beta version of RBDSD1Fc provided a broad range of cross-neutralization to multiple antigenic variants of concern, including Omicron; and 6) the Beta version of RBDSD1Fc with DDA/TDB provided complete protection against virus challenge in the knock-in mouse model. Thus, we have identified an optimized RBD-based subunit vaccine suitable for clinical trials.
Assuntos
COVID-19 , Vacinas Virais , Humanos , Animais , Camundongos , SARS-CoV-2 , Vacinas contra COVID-19 , Hidróxido de Alumínio , Glicoproteína da Espícula de Coronavírus , Vacinas de Subunidades Antigênicas , Anticorpos Antivirais , Anticorpos Neutralizantes , MamíferosRESUMO
PI3K and PTEN lipid phosphatase control the level of cellular phosphatidylinositol (3,4,5)-trisphosphate, an activator of AKT kinases that promotes cell growth and survival. Mutations activating AKT are commonly observed in human cancers. We report here that ENTPD5, an endoplasmic reticulum (ER) enzyme, is upregulated in cell lines and primary human tumor samples with active AKT. ENTPD5 hydrolyzes UDP to UMP to promote protein N-glycosylation and folding in ER. Knockdown of ENTPD5 in PTEN null cells causes ER stress and loss of growth factor receptors. ENTPD5, together with cytidine monophosphate kinase-1 and adenylate kinase-1, constitute an ATP hydrolysis cycle that converts ATP to AMP, resulting in a compensatory increase in aerobic glycolysis known as the Warburg effect. The growth of PTEN null cells is inhibited both in vitro and in mouse xenograft tumor models. ENTPD5 is therefore an integral part of the PI3K/PTEN regulatory loop and a potential target for anticancer therapy.
Assuntos
Proliferação de Células , Retículo Endoplasmático/metabolismo , Glicosilação , Proteínas Oncogênicas/metabolismo , Trifosfato de Adenosina/metabolismo , Aerobiose , Animais , Linhagem Celular Tumoral , Glicólise , Guanosina Monofosfato/metabolismo , Humanos , Camundongos , Transplante de Neoplasias , Proteína Oncogênica v-akt/metabolismo , Proteínas Oncogênicas/genética , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Pirofosfatases , Transplante Heterólogo , Uridina Monofosfato/metabolismoRESUMO
Deregulation of cell cycle is a typical feature of cancer cells. Normal cells rely on the strictly coordinated spindle assembly checkpoint (SAC) to maintain the genome integrity and survive. However, cancer cells could bypass this checkpoint mechanism. In this study, we showed the clinical relevance of threonine tyrosine kinase (TTK) protein kinase, a central regulator of the SAC, in hepatocellular carcinoma (HCC) and its potential as therapeutic target. Here, we reported that a newly developed, orally active small molecule inhibitor targeting TTK (CFI-402257) effectively suppressed HCC growth and induced highly aneuploid HCC cells, DNA damage, and micronuclei formation. We identified that CFI-402257 also induced cytosolic DNA, senescence-like response, and activated DDX41-STING cytosolic DNA sensing pathway to produce senescence-associated secretory phenotypes (SASPs) in HCC cells. These SASPs subsequently led to recruitment of different subsets of immune cells (natural killer cells, CD4+ T cells, and CD8+ T cells) for tumor clearance. Our mass cytometry data illustrated the dynamic changes in the tumor-infiltrating immune populations after treatment with CFI-402257. Further, CFI-402257 improved survival in HCC-bearing mice treated with anti-PD-1, suggesting the possibility of combination treatment with immune checkpoint inhibitors in HCC patients. In summary, our study characterized CFI-402257 as a potential therapeutic for HCC, both used as a single agent and in combination therapy.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Inibidores de Proteínas Quinases , Pirazóis , Pirimidinas , Animais , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Células Matadoras Naturais/metabolismo , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Camundongos , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Serina-Treonina Quinases , Proteínas Tirosina Quinases/metabolismo , Pirazóis/uso terapêutico , Pirimidinas/uso terapêuticoRESUMO
BACKGROUND AND AIMS: Prognosis of HCC remains poor due to lack of effective therapies. Immune checkpoint inhibitors (ICIs) have delayed response and are only effective in a subset of patients. Treatments that could effectively shrink the tumors within a short period of time are idealistic to be employed together with ICIs for durable tumor suppressive effects. HCC acquires increased tolerance to aneuploidy. The rapid division of HCC cells relies on centrosome duplication. In this study, we found that polo-like kinase 4 (PLK4), a centrosome duplication regulator, represents a therapeutic vulnerability in HCC. APPROACH AND RESULTS: An orally available PLK4 inhibitor, CFI-400945, potently suppressed proliferating HCC cells by perturbing centrosome duplication. CFI-400945 induced endoreplication without stopping DNA replication, causing severe aneuploidy, DNA damage, micronuclei formation, cytosolic DNA accumulation, and senescence. The cytosolic DNA accumulation elicited the DEAD box helicase 41-stimulator of interferon genes-interferon regulatory factor 3/7-NF-κß cytosolic DNA sensing pathway, thereby driving the transcription of senescence-associated secretory phenotypes, which recruit immune cells. CFI-400945 was evaluated in liver-specific p53/phosphatase and tensin homolog knockout mouse HCC models established by hydrodynamic tail vein injection. Tumor-infiltrated immune cells were analyzed. CFI-400945 significantly impeded HCC growth and increased infiltration of cluster of differentiation 4-positive (CD4 + ), CD8 + T cells, macrophages, and natural killer cells. Combination therapy of CFI-400945 with anti-programmed death-1 showed a tendency to improve HCC survival. CONCLUSIONS: We show that by targeting a centrosome regulator, PLK4, to activate the cytosolic DNA sensing-mediated immune response, CFI-400945 effectively restrained tumor progression through cell cycle inhibition and inducing antitumor immunity to achieve a durable suppressive effect even in late-stage mouse HCC.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Animais , Camundongos , Aneuploidia , Carcinoma Hepatocelular/patologia , Ciclo Celular , Linhagem Celular Tumoral , Neoplasias Hepáticas/patologia , Proteínas Serina-Treonina Quinases/metabolismoRESUMO
Passenger mutations specific to particular mouse strains can distort experimental outcomes. In this issue of Immunity, Vanden Berghe et al. (2015) demonstrate that passenger mutations are frequent in most genetically engineered congenic mice and persist even after extensive backcrossing.
Assuntos
Variação Genética/genética , Genoma/genética , Camundongos Endogâmicos C57BL/genética , AnimaisRESUMO
A hallmark of cancer, including pancreatic ductal adenocarcinoma (PDA), is a massive stromal and inflammatory reaction. Many efforts have been made to identify the anti- or protumoral role of cytokines and immune subpopulations within the stroma. Here, we investigated the role of interleukin-17A (IL17A) and its effect on tumor fibroblasts and the tumor microenvironment. We used a spontaneous PDA mouse model (KPC) crossed to IL17A knockout mice to show an extensive desmoplastic reaction, without impaired immune infiltration. Macrophages, especially CD80+ and T cells, were more abundant at the earlier time point. In T cells, a decrease in FoxP3+ cells and an increase in CD8+ T cells were observed in KPC/IL17A-/- mice. Fibroblasts isolated from IL17A+/+ and IL17A-/- KPC mice revealed very different messenger RNA (mRNA) and protein profiles. IL17A-/- fibroblasts displayed the ability to restrain tumor cell invasion by producing factors involved in extracellular matrix remodeling, increasing T cell recruitment, and producing higher levels of cytokines and chemokines favoring T helper 1 cell recruitment and activation and lower levels of those recruiting myeloid/granulocytic immune cells. Single-cell quantitative PCR on isolated fibroblasts confirmed a very divergent profile of IL17A-proficient and -deficient cells. All these features can be ascribed to increased levels of IL17F observed in the sera of IL17A-/- mice, and to the higher expression of its cognate receptor (IL17RC) specifically in IL17A-/- cancer-associated fibroblasts (CAFs). In addition to the known effects on neoplastic cell transformation, the IL17 cytokine family uniquely affects fibroblasts, representing a suitable candidate target for combinatorial immune-based therapies in PDA.
Assuntos
Adenocarcinoma/genética , Carcinoma Ductal Pancreático/genética , Interleucina-17/genética , Receptores de Interleucina/genética , Adenocarcinoma/patologia , Animais , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Fibroblastos Associados a Câncer/metabolismo , Fibroblastos Associados a Câncer/patologia , Carcinogênese/genética , Carcinoma Ductal Pancreático/patologia , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/genética , Humanos , Camundongos , Camundongos Knockout , Microambiente Tumoral/genéticaRESUMO
Ribosome biogenesis in the nucleolus is an important process that consumes 80% of a cell's intracellular energy supply. Disruption of this process results in nucleolar stress, triggering the activation of molecular systems that respond to this stress to maintain homeostasis. Although nucleolar stress was originally thought to be caused solely by abnormalities of ribosomal RNA (rRNA) and ribosomal proteins (RPs), an accumulating body of more current evidence suggests that many other factors, including the DNA damage response and oncogenic stress, are also involved in nucleolar stress response signaling. Cells reacting to nucleolar stress undergo cell cycle arrest or programmed death, mainly driven by activation of the tumor suppressor p53. This observation has nominated nucleolar stress as a promising target for cancer therapy. However, paradoxically, some RP mutations have also been implicated in cancer initiation and progression, necessitating caution. In this article, we summarize recent findings on the molecular mechanisms of nucleolar stress and the human ribosomal diseases and cancers that arise in its wake.
Assuntos
Neoplasias , Proteínas Ribossômicas , Humanos , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo , Ribossomos/genética , Ribossomos/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , RNA Ribossômico/genética , RNA Ribossômico/metabolismo , Pontos de Checagem do Ciclo Celular/genética , Neoplasias/genética , Neoplasias/metabolismoRESUMO
Bladder cancer (BlC) is the fourth most common cancer in males worldwide, but few systemic chemotherapy options for its effective treatment exist. The development of new molecularly-targeted agents against BlC is therefore an urgent issue. The Hippo signaling pathway, with its upstream LATS kinases and downstream transcriptional co-activators YAP1 and TAZ, plays a pivotal role in diverse cell functions, including cell proliferation. Recent studies have shown that overexpression of YAP1 occurs in advanced BlCs and is associated with poor patient prognosis. Accessing data from our previous screening of a chemical library of compounds targeting the Hippo pathway, we identified DMPCA (N-(3,4-dimethoxyphenethyl)-6-methyl-2,3,4,9-tetrahydro-1H-carbazol-1-amine) as an agent able to induce the phosphorylation of LATS1 and YAP1/TAZ in BlC cells, thereby suppressing their viability both in vitro and in mouse xenografts. Our data indicate that DMPCA has a potent anti-tumor effect, and raise the possibility that this agent may represent a new and effective therapeutic option for BlC.