RESUMEN
Metabolic dysfunction-associated steatohepatitis-driven hepatocellular carcinoma (MASH-HCC) is a global clinical challenge for which there is a limited understanding of disease pathogenesis and a subsequent lack of therapeutic interventions. We previously identified that tumor necrosis factor-alpha (TNF-α) upregulated apoptosis antagonizing transcription factor (AATF) in MASH. Here, we investigated the effect of TNF-α converting enzyme (TACE) inhibition as a promising targeted therapy against AATF-mediated steatohepatitis to hepatocarcinogenesis. A preclinical murine model that recapitulates human MASH-HCC was used in the study. C57Bl/6 mice were fed with chow diet normal water (CD) or western diet sugar water (WD) along with a low dose of carbon tetrachloride (CCl4; 0.2 µL·g-1, weekly) for 24 weeks. TACE activity, TNF-α levels, and AATF expression were measured. The mice were treated with the TACE inhibitor Marimastat for 12 weeks, followed by analyses of liver injury, fibrosis, inflammation, and oncogenic signaling. In vitro experiments using stable clones of AATF control and AATF knockdown were also conducted. We found that AATF expression was upregulated in WD/CCl4 mice, which developed severe MASH at 12 weeks and advanced fibrosis with HCC at 24 weeks. WD/CCl4 mice showed increased TACE activity with reduced hepatic expression of sirtuin 1 (Sirt1) and tissue inhibitor of metalloproteinase 3 (Timp3). The involvement of the SIRT1/TIMP3/TACE axis was confirmed by the release of TNF-α, which upregulated AATF, a key molecular driver of MASH-HCC. Interestingly, TACE inhibition by Marimastat reduced liver injury, dyslipidemia, AATF expression, and oncogenic signaling, effectively preventing hepatocarcinogenesis. Furthermore, Marimastat inhibited the activation of JNK, ERK1/2, and AKT, which are key regulators of tumorigenesis in WD/CCl4 mice and in AATF control cells, but had no effect on AATF knockdown cells. This study shows that TACE inhibition prevents AATF-mediated inflammation, fibrosis, and oncogenesis in MASH-HCC, offering a potential target for therapeutic intervention.
Asunto(s)
Proteína ADAM17 , Carcinoma Hepatocelular , Neoplasias Hepáticas , Ratones Endogámicos C57BL , Animales , Humanos , Masculino , Ratones , Proteína ADAM17/metabolismo , Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/genética , Carcinogénesis/patología , Carcinogénesis/efectos de los fármacos , Carcinogénesis/metabolismo , Carcinogénesis/genética , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/genética , Hígado Graso/patología , Hígado Graso/metabolismo , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/tratamiento farmacológico , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Fortunately, ample efforts are being made to find the best strategy to improve the anti-leukemia capacity of NK cells for treating different types of cancer. Despite the favorable ADCC capacity of functional CD16â¯+â¯NK cells for immunotherapy, when NK cells face leukemia cells, the CD16 receptor is cleaved during the process mediated by a disintegrin and metalloproteinase-17(ADAM17). Reduced CD16 expression on NK cells weakens their cytotoxicity against leukemia cells. In addition, the expression of the CD47 receptor is high in acute lymphoblastic leukemia (ALL) compared to normal cells and can be correlated with poor prognosis. In the present study, ADAM17 was inhibited in cord blood-derived CD16â¯+â¯NK cells, and their activity against ALL cell lines was evaluated following blockage with anti-CD47 antibody. As the results showed, the CD16 expression was reduced in the NK cells co-cultured with ALL cell lines. However, the ADAM17 inhibition increased the CD16 expression on the NK cells. This enhanced the cytotoxicity of those cells as well as cytokine production was evaluated by measuring expression of CD107-a expression, and IFN-γ production. Moreover, the presence of the ADAM17 inhibitor increased the apoptosis effect of the generated NK cells in response to ALL cells. Therefore, the inhibition of ADAM17 is useful for the activity of CD16â¯+â¯NK cells against cancer cells.
Asunto(s)
Proteína ADAM17 , Sangre Fetal , Células Asesinas Naturales , Leucemia-Linfoma Linfoblástico de Células Precursoras , Receptores de IgG , Humanos , Células Asesinas Naturales/inmunología , Leucemia-Linfoma Linfoblástico de Células Precursoras/inmunología , Proteína ADAM17/metabolismo , Proteína ADAM17/antagonistas & inhibidores , Receptores de IgG/metabolismo , Sangre Fetal/citología , Línea Celular Tumoral , Citotoxicidad Inmunológica , Proteínas Ligadas a GPI/metabolismo , Técnicas de Cocultivo , Apoptosis , Citotoxicidad Celular Dependiente de Anticuerpos , Interferón gamma/metabolismo , Antígeno CD47RESUMEN
ZLDI-8 is an A disintegrin and metalloproteinase domain 17 (ADAM17) inhibitor that suppresses the shedding of Notch1 to the Notch1 intracellular domain (NICD). In previous studies, we found that ZLDI-8 was able to sensitize HCC to sorafenib, but the mechanism of action remains unclear. The sensitizing effects of ZLDI-8 were tested both in vitro and in vivo. EMT-related factors, sorafenib sensitivity-related proteins and ECM-related gene expression were assessed using immunohistochemistry, RTPCR and Western blotting. Knockdown assays were conducted to determine the relationship between the Notch and Integrin pathways. CoIP assays, nuclear and cytoplasmic fractionation and immunofluorescence colocalization were applied to explore the interaction between the Notch and Integrin pathways. Appropriate statistical analysis methods were used to assess the significance of the experimental results and to ensure the scientific validity and reliability of the experimental design. We found that ECM- and EMT-related proteins were downregulated after ZLDI-8 treatment (P<0.05). ZLDI-8 significantly downregulated Integrinß1 and Integrinß3 in HCC in vitro and in vivo (P<0.05), possibly through Foxc2-dependent regulation. Mechanistically, interfering with the expression of both Integrin-linked kinase (ILK) and the NICD may downregulate the expression of proteins targeted by sorafenib, thereby sensitizing cells to sorafenib. The retroregulation of Integrinß by ILK may occur through the interaction between the NICD and ILK and may be the result of the translocation of the complexus. Our study indicates that blocking the Notch pathway may affect Integrinß through crosstalk between the Notch1 and Integrinß/ILK signaling pathways, thus providing a potential therapeutic strategy for HCC.
Asunto(s)
Proteína ADAM17 , Antineoplásicos , Carcinoma Hepatocelular , Neoplasias Hepáticas , Receptor Notch1 , Sorafenib , Sorafenib/farmacología , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Humanos , Animales , Receptor Notch1/metabolismo , Receptor Notch1/genética , Antineoplásicos/farmacología , Línea Celular Tumoral , Proteína ADAM17/metabolismo , Proteína ADAM17/antagonistas & inhibidores , Ratones Desnudos , Masculino , Cadenas beta de Integrinas/metabolismo , Cadenas beta de Integrinas/genética , Ratones Endogámicos BALB C , Transducción de Señal/efectos de los fármacos , Transición Epitelial-Mesenquimal/efectos de los fármacos , RatonesRESUMEN
Melatonin, as a dietary supplement, has a potent neuroprotective effect and exerts a certain antidepressant effect. This study explored the molecular mechanisms and targets of melatonin on chronic stress-induced hippocampal damage from the perspective of inhibiting inflammatory cytokines release. Our results indicated that melatonin alleviated chronic restraint stress (CRS)-induced inflammatory response and apoptosis, thus improving hippocampal structural damage and subsequent depression-like behaviors in rats. The radar map displayed that the change of TNF-α content was the most significant. Meanwhile, correlation analysis showed that TNF-α content was highly positively correlated with apoptosis. Molecular autodocking studies suggested that TNF-α converting enzyme ADAM17 as a potential target has a priority in docking with melatonin. Molecular mechanism studies indicated that melatonin inhibited CRS-induced activation of the ADAM17/TNF-α axis and its downstream proteins p38 and p53 phosphorylation in the hippocampus. Analogously, Both ADAM17 inhibitor TMI-1 and TNF-α inhibitor thalidomide relieved the effects of CRS on ADAM17/TNF-α axis and its downstream proteins phosphorylation, hippocampal apoptosis, hippocampal inflammatory response, and depression-like behaviors in rats. Altogether, these findings reveal that melatonin relieves CRS-induced inflammatory response and apoptosis, and subsequent depression-like behaviors by inhibiting ADAM17/TNF-α axis.
Asunto(s)
Proteína ADAM17 , Apoptosis , Hipocampo , Melatonina , Enfermedades Neuroinflamatorias , Fármacos Neuroprotectores , Estrés Psicológico , Animales , Ratas , Proteína ADAM17/antagonistas & inhibidores , Citocinas/metabolismo , Hipocampo/efectos de los fármacos , Hipocampo/patología , Melatonina/farmacología , Melatonina/uso terapéutico , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Talidomida/farmacología , Factor de Necrosis Tumoral alfa/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Enfermedades Neuroinflamatorias/tratamiento farmacológico , Enfermedades Neuroinflamatorias/etnología , Estrés Psicológico/complicaciones , Enfermedad CrónicaRESUMEN
Angiotensin Converting Enzyme 2 (ACE2) is the primary cell entry receptor for SARS-CoV and SARS-CoV-2 viruses. A disintegrin and metalloproteinase 17 (ADAM17) is a protease that cleaves ectodomains of transmembrane proteins, including that of ACE2 and the proinflammatory cytokine TNF-α, from cell surfaces upon cellular activation. We hypothesized that blockade of ADAM17 activity would alter COVID-19 pathogenesis. To assess this pathway, we blocked the function of ADAM17 using the monoclonal antibody MEDI3622 in the K18-hACE2 transgenic mouse model of COVID-19. Antibody-treated mice were healthier, less moribund, and had significantly lower lung pathology than saline-treated mice. However, the viral burden in the lungs of MEDI3622-treated mice was significantly increased. Thus, ADAM17 appears to have a critical anti-viral role, but also may promote inflammatory damage. Since the inflammatory cascade is ultimately the reason for adverse outcomes in COVID-19 patients, there may be a therapeutic application for the MEDI3622 antibody.
Asunto(s)
Proteína ADAM17 , Anticuerpos Neutralizantes , COVID-19 , SARS-CoV-2 , Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/inmunología , Enzima Convertidora de Angiotensina 2/metabolismo , Animales , Anticuerpos Neutralizantes/inmunología , Anticuerpos Neutralizantes/farmacología , COVID-19/inmunología , COVID-19/terapia , COVID-19/virología , Modelos Animales de Enfermedad , Humanos , Ratones , Ratones Transgénicos , Peptidil-Dipeptidasa A/metabolismo , SARS-CoV-2/inmunología , Carga ViralRESUMEN
Understanding the mechanisms of modulators' action on enzymes is crucial for optimizing and designing pharmaceutical substances. The acute inflammatory response, in particular, is regulated mainly by a disintegrin and metalloproteinase (ADAM) 17. ADAM17 processes several disease mediators such as TNFα and APP, releasing their soluble ectodomains (shedding). A malfunction of this process leads to a disturbed inflammatory response. Chemical protease inhibitors such as TAPI-1 were used in the past to inhibit ADAM17 proteolytic activity. However, due to ADAM17's broad expression and activity profile, the development of active-site-directed ADAM17 inhibitor was discontinued. New 'exosite' (secondary substrate binding site) inhibitors with substrate selectivity raised the hope of a substrate-selective modulation as a promising approach for inflammatory disease therapy. This work aimed to develop a high-throughput screen for potential ADAM17 modulators as therapeutic drugs. By combining experimental and in silico methods (structural modeling and docking), we modeled the kinetics of ADAM17 inhibitor. The results explain ADAM17 inhibition mechanisms and give a methodology for studying selective inhibition towards the design of pharmaceutical substances with higher selectivity.
Asunto(s)
Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/efectos de los fármacos , Proteína ADAM17/metabolismo , Proteínas ADAM/metabolismo , Sitios de Unión/efectos de los fármacos , Dominio Catalítico/efectos de los fármacos , Simulación por Computador , Evaluación Preclínica de Medicamentos/métodos , Células HEK293 , Ensayos Analíticos de Alto Rendimiento/métodos , Humanos , Cinética , Inhibidores de Proteasas/farmacología , Especificidad por Sustrato/efectos de los fármacosRESUMEN
Metastasis is the major cause of death in cancer patients. Circulating tumor cells need to migrate through the endothelial layer of blood vessels to escape the hostile circulation and establish metastases at distant organ sites. Here, we identified the membrane-bound metalloprotease ADAM17 on endothelial cells as a key driver of metastasis. We show that TNFR1-dependent tumor cell-induced endothelial cell death, tumor cell extravasation, and subsequent metastatic seeding is dependent on the activity of endothelial ADAM17. Moreover, we reveal that ADAM17-mediated TNFR1 ectodomain shedding and subsequent processing by the γ-secretase complex is required for the induction of TNF-induced necroptosis. Consequently, genetic ablation of ADAM17 in endothelial cells as well as short-term pharmacological inhibition of ADAM17 prevents long-term metastases formation in the lung. Thus, our data identified ADAM17 as a novel essential regulator of necroptosis and as a new promising target for antimetastatic and advanced-stage cancer therapies.
Asunto(s)
Proteína ADAM17/antagonistas & inhibidores , Células Endoteliales/metabolismo , Necroptosis , Neoplasias/etiología , Neoplasias/patología , Animales , Antineoplásicos/farmacología , Biomarcadores , Biomarcadores de Tumor , Comunicación Celular , Muerte Celular , Susceptibilidad a Enfermedades/inmunología , Humanos , Necroptosis/genética , Invasividad Neoplásica , Metástasis de la Neoplasia , Siembra Neoplásica , Neoplasias/metabolismo , Neoplasias/terapia , Proteolisis , Receptores Tipo I de Factores de Necrosis Tumoral/metabolismo , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/genética , Microambiente Tumoral/inmunología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
It was realized in the 1990s that some membrane proteins such as TNFα, both TNF receptors, ligands of the EGF-R and the Interleukin-6 receptor are proteolytically cleaved and are shed from the cell membrane as soluble proteins. The major responsible protease is a metalloprotease named ADAM17. So far, close to 100 substrates, including cytokines, cytokine receptors, chemokines and adhesion molecules of ADAM17 are known. Therefore, ADAM17 orchestrates many different signaling pathways and is a central signaling hub in inflammation and carcinogenesis. ADAM17 plays an important role in the biology of Interleukin-6 (IL-6) since the generation of the soluble Interleukin-6 receptor (sIL-6R) is needed for trans-signaling, which has been identified as the pro-inflammatory activity of this cytokine. In contrast, Interleukin-6 signaling via the membrane-bound Interleukin-6 receptor is mostly regenerative and protective. Probably due to its broad substrate spectrum, ADAM17 is essential for life and most of the few human individuals identified with ADAM17 gene defects died at young age. Although the potential of ADAM17 as a therapeutic target has been recognized, specific blockade of ADAM17 is not trivial since the metalloprotease domain of ADAM17 shares high structural homology with other proteases, in particular matrix metalloproteases. Here, the critical functions of ADAM17 in IL-6, TNFα and EGF-R pathways and strategies of therapeutic interventions are discussed.
Asunto(s)
Proteína ADAM17/metabolismo , Receptores ErbB/metabolismo , Interleucina-17/metabolismo , Neoplasias/metabolismo , Transducción de Señal , Factor de Necrosis Tumoral alfa/metabolismo , Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/genética , Animales , Antiinflamatorios/farmacología , Receptores ErbB/genética , Humanos , Interleucina-17/genética , Inhibidores de Proteasas/farmacología , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
BACKGROUND: Peritumoral fibroblasts are key components of the tumor microenvironment. Through remodeling of the extracellular matrix (ECM) and secretion of pro-tumorigenic cytokines, peritumoral fibroblasts foster an immunosuppressive milieu conducive to tumor cell proliferation. In this study, we investigated if peritumoral fibroblasts could be therapeutically engineered to elicit an anti-cancer response by abolishing the proteolytic activities of membrane-bound metalloproteinases involved in ECM modulation. METHODS: A high affinity, glycosylphosphatidylinositol (GPI)-anchored Tissue Inhibitor of Metalloproteinase (TIMP) named "T1PrαTACE" was created for dual inhibition of MT1-MMP and TACE. T1PrαTACE was expressed in fibroblasts and its effects on cancer cell proliferation investigated in 3D co-culture models. RESULTS: T1PrαTACE abrogated the activities of MT1-MMP and TACE in host fibroblasts. As a GPI protein, T1PrαTACE could spontaneously detach from the plasma membrane of the fibroblast to co-localize with MT1-MMP and TACE on neighboring cancer cells. In a 3D co-culture model, T1PrαTACE promoted adherence between the cancer cells and surrounding fibroblasts, which led to an attenuation in tumor development. CONCLUSION: Peritumoral fibroblasts can be modulated with the TIMP for the elimination of cancer cells. As a novel anti-tumor strategy, our approach could potentially be used in combination with conventional chemo- and immunotherapies for a more effective cancer therapy.
Asunto(s)
Fibroblastos/efectos de los fármacos , Neoplasias/patología , Inhibidores Tisulares de Metaloproteinasas/farmacología , Proteínas ADAM/antagonistas & inhibidores , Proteína ADAM17/antagonistas & inhibidores , Línea Celular Tumoral , Células Endoteliales de la Vena Umbilical Humana , HumanosRESUMEN
A disintegrin and metalloproteinase 17 (ADAM17) is a zinc-dependent enzyme that catalyzes the cleavage of the extracellular domains of various transmembrane proteins. ADAM17 is regarded as a promising drug target for the suppression of various diseases, including cancer metastasis. We synthesized a new ADAM17 inhibitor, SN-4, composed of a zinc-binding dithiol moiety and an appendage that specifically binds to a pocket of ADAM17. We show that SN-4 inhibits the ability of ADAM17 to cleave tumor necrosis factor α (TNF-α) in vitro. This activity was reduced by the addition of zinc, indicating the importance of the zinc chelating dithiol moiety. Inhibition of TNF-α cleavage by SN-4 in cells was also observed, and with an IC50 of 3.22 µM, SN-4 showed slightly higher activity than the well-studied ADAM17 inhibitor marimastat. Furthermore, SN-4 was shown to inhibit cleavage of CD44 by ADAM17, but not by ADAM10, and to suppress cell invasion. Molecular docking showed good fitting of the specificity pocket-binding group and one SH of SN-4 and hinted at possible means of structural optimization. This study provides clues for the development of potent and selective ADAM17 inhibitors.
Asunto(s)
Proteína ADAM17/antagonistas & inhibidores , Inhibidores de Proteasas/síntesis química , Sulfonamidas/síntesis química , Tolueno/análogos & derivados , Proteína ADAM10/metabolismo , Humanos , Receptores de Hialuranos/metabolismo , Ácidos Hidroxámicos/química , Ácidos Hidroxámicos/farmacología , Simulación del Acoplamiento Molecular , Inhibidores de Proteasas/metabolismo , Inhibidores de Proteasas/farmacología , Unión Proteica , Conformación Proteica , Relación Estructura-Actividad , Sulfonamidas/metabolismo , Sulfonamidas/farmacología , Tolueno/química , Factor de Necrosis Tumoral alfa/metabolismo , Zinc , BencenosulfonamidasRESUMEN
The current pandemic caused by SARS-CoV-2 virus infection is known as Covid-19 (coronavirus disease 2019). This disease can be asymptomatic or can affect multiple organ systems. Damage induced by the virus is related to dysfunctional activity of the immune system, but the activity of molecules such as C-reactive protein (CRP) as a factor capable of inducing an inflammatory status that may be involved in the severe evolution of the disease, has not been extensively evaluated. A systematic review was performed using the NCBI-PubMed database to find articles related to Covid-19 immunity, inflammatory response, and CRP published from December 2019 to December 2020. High levels of CRP were found in patients with severe evolution of Covid-19 in which several organ systems were affected and in patients who died. CRP activates complement, induces the production of pro-inflammatory cytokines and induces apoptosis which, together with the inflammatory status during the disease, can lead to a severe outcome. Several drugs can decrease the level or block the effect of CRP and might be useful in the treatment of Covid-19. From this review it is reasonable to conclude that CRP is a factor that can contribute to severe evolution of Covid-19 and that the use of drugs able to lower CRP levels or block its activity should be evaluated in randomized controlled clinical trials.
Asunto(s)
Antiinflamatorios/uso terapéutico , Proteína C-Reactiva/antagonistas & inhibidores , Tratamiento Farmacológico de COVID-19 , Proteínas del Sistema Complemento/inmunología , Síndrome de Liberación de Citoquinas/tratamiento farmacológico , SARS-CoV-2/patogenicidad , Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/genética , Proteína ADAM17/inmunología , Enzima Convertidora de Angiotensina 2/antagonistas & inhibidores , Enzima Convertidora de Angiotensina 2/genética , Enzima Convertidora de Angiotensina 2/inmunología , Biomarcadores/sangre , Proteína C-Reactiva/genética , Proteína C-Reactiva/inmunología , COVID-19/inmunología , COVID-19/patología , COVID-19/virología , Celecoxib/uso terapéutico , Proteínas del Sistema Complemento/genética , Síndrome de Liberación de Citoquinas/inmunología , Síndrome de Liberación de Citoquinas/patología , Síndrome de Liberación de Citoquinas/virología , Citocinas/antagonistas & inhibidores , Citocinas/genética , Citocinas/inmunología , Progresión de la Enfermedad , Doxiciclina/uso terapéutico , Regulación de la Expresión Génica , Humanos , Ensayos Clínicos Controlados Aleatorios como Asunto , Índice de Severidad de la Enfermedad , Análisis de SupervivenciaRESUMEN
Signaling via death receptor family members such as TNF-R1 mediates pleiotropic biological outcomes ranging from inflammation and proliferation to cell death. Pro-survival signaling is mediated via TNF-R1 complex I at the cellular plasma membrane. Cell death induction requires complex IIa/b or necrosome formation, which occurs in the cytoplasm. In many cell types, full apoptotic or necroptotic cell death induction requires the internalization of TNF-R1 and receptosome formation to properly relay the signal inside the cell. We interrogated the role of the enzyme A disintegrin and metalloprotease 17 (ADAM17)/TACE (TNF-α converting enzyme) in death receptor signaling in human hematopoietic cells, using pharmacological inhibition and genetic ablation. We show that in U937 and Jurkat cells the absence of ADAM17 does not abrogate, but rather increases TNF mediated cell death. Likewise, cell death triggered via DR3 is enhanced in U937 cells lacking ADAM17. We identified ADAM17 as the key molecule that fine-tunes death receptor signaling. A better understanding of cell fate decisions made via the receptors of the TNF-R1 superfamily may enable us, in the future, to more efficiently treat infectious and inflammatory diseases or cancer.
Asunto(s)
Proteína ADAM17/metabolismo , Receptores Tipo I de Factores de Necrosis Tumoral/metabolismo , Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/deficiencia , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Muerte Celular , Supervivencia Celular , Endocitosis , Humanos , Células Jurkat , Células MCF-7 , Modelos Biológicos , FN-kappa B/metabolismo , Miembro 25 de Receptores de Factores de Necrosis Tumoral/metabolismo , Transducción de Señal , Factor de Necrosis Tumoral alfa/farmacología , Células U937RESUMEN
Very low-density lipoprotein receptor (VLDLR) is a multifunctional transmembrane protein. Beyond the function of the full-length VLDLR in lipid transport, the soluble ectodomain of VLDLR (sVLDLR) confers anti-inflammatory and antiangiogenic roles in ocular tissues through inhibition of canonical Wnt signaling. However, it remains unknown how sVLDLR is shed into the extracellular space. In this study, we present the first evidence that a disintegrin and metalloprotease 17 (ADAM17) is responsible for sVLDLR shedding in human retinal pigment epithelium cells using pharmacological and genetic approaches. Among selected proteinase inhibitors, an ADAM17 inhibitor demonstrated the most potent inhibitory effect on sVLDLR shedding. siRNA-mediated knockdown or CRISPR/Cas9-mediated KO of ADAM17 diminished, whereas plasmid-mediated overexpression of ADAM17 promoted sVLDLR shedding. The amount of shed sVLDLR correlated with an inhibitory effect on the Wnt signaling pathway. Consistent with these in vitro findings, intravitreal injection of an ADAM17 inhibitor reduced sVLDLR levels in the extracellular matrix in the mouse retina. In addition, our results demonstrated that ADAM17 cleaved VLDLR only in cells coexpressing these proteins, suggesting that shedding occurs in a cis manner. Moreover, our study demonstrated that aberrant activation of Wnt signaling was associated with decreased sVLDLR levels, along with downregulation of ADAM17 in ocular tissues of an age-related macular degeneration model. Taken together, our observations reveal the mechanism underlying VLDLR cleavage and identify a potential therapeutic target for the treatment of disorders associated with dysregulation of Wnt signaling.
Asunto(s)
Proteína ADAM17/metabolismo , Degeneración Macular/metabolismo , Receptores de LDL/metabolismo , Epitelio Pigmentado de la Retina/metabolismo , Vía de Señalización Wnt , Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/genética , Animales , Modelos Animales de Enfermedad , Humanos , Degeneración Macular/genética , Ratones , Ratones Noqueados , Dominios Proteicos , Receptores de LDL/genéticaRESUMEN
The article describes the possible pathophysiological origin of COVID-19 and the crucial role of renin-angiotensin system (RAS), providing several "converging" evidence in support of this hypothesis. SARS-CoV-2 has been shown to initially upregulate ACE2 systemic activity (early phase), which can subsequently induce compensatory responses leading to upregulation of both arms of the RAS (late phase) and consequently to critical, advanced and untreatable stages of COVID-19 disease. The main and initial actors of the process are ACE2 and ADAM17 zinc-metalloproteases, which, initially triggered by SARS-CoV-2 spike proteins, work together in increasing circulating Ang 1-7 and Ang 1-9 peptides and downstream (Mas and Angiotensin type 2 receptors) pathways with anti-inflammatory, hypotensive and antithrombotic activities. During the late phase of severe COVID-19, compensatory secretion of renin and ACE enzymes are subsequently upregulated, leading to inflammation, hypertension and thrombosis, which further sustain ACE2 and ADAM17 upregulation. Based on this hypothesis, COVID-19-phase-specific inhibition of different RAS enzymes is proposed as a pharmacological strategy against COVID-19 and vaccine-induced adverse effects. The aim is to prevent the establishment of positive feedback-loops, which can sustain hyperactivity of both arms of the RAS independently of viral trigger and, in some cases, may lead to Long-COVID syndrome.
Asunto(s)
Proteína ADAM17/biosíntesis , Enzima Convertidora de Angiotensina 2/biosíntesis , COVID-19/metabolismo , Sistema Renina-Angiotensina , SARS-CoV-2/metabolismo , Glicoproteína de la Espiga del Coronavirus/metabolismo , Proteína ADAM17/antagonistas & inhibidores , Angiotensina I/metabolismo , Enzima Convertidora de Angiotensina 2/antagonistas & inhibidores , Regulación Enzimológica de la Expresión Génica , Humanos , Fragmentos de Péptidos/metabolismo , Glicoproteína de la Espiga del Coronavirus/antagonistas & inhibidores , Regulación hacia Arriba , Tratamiento Farmacológico de COVID-19RESUMEN
Natural killer (NK) cells are innate cytotoxic lymphocytes that can recognize assorted determinants on tumor cells and rapidly kill these cells. Due to their anti-tumor effector functions and potential for allogeneic use, various NK cell platforms are being examined for adoptive cell therapies. However, their limited in vivo persistence is a current challenge. Cytokine-mediated activation of these cells is under extensive investigation and interleukin-15 (IL-15) is a particular focus since it drives their activation and proliferation. IL-15 efficacy though is limited in part by its induction of regulatory checkpoints. A disintegrin and metalloproteinase-17 (ADAM17) is broadly expressed by leukocytes, including NK cells, and it plays a central role in cleaving cell surface receptors, a process that regulates cell activation and cell-cell interactions. We report that ADAM17 blockade with a monoclonal antibody markedly increased human NK cell proliferation by IL-15 both in vitro and in a xenograft mouse model. Blocking ADAM17 resulted in a significant increase in surface levels of the homing receptor CD62L on proliferating NK cells. We show that NK cell proliferation in vivo by IL-15 and the augmentation of this process upon blocking ADAM17 are dependent on CD62L. Hence, our findings reveal for the first time that ADAM17 activation in NK cells by IL-15 limits their proliferation, presumably functioning as a feedback system, and that its substrate CD62L has a key role in this process in vivo. ADAM17 blockade in combination with IL-15 may provide a new approach to improve NK cell persistence and function in cancer patients.
Asunto(s)
Proteína ADAM17/metabolismo , Interleucina-15/farmacología , Células Asesinas Naturales/citología , Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/inmunología , Traslado Adoptivo , Animales , División Celular , Activación Enzimática , Femenino , Xenoinjertos , Humanos , Interleucina-15/metabolismo , Células Asesinas Naturales/enzimología , Selectina L/metabolismo , Ratones , Ratones Endogámicos NOD , Ratones SCID , Receptores de Superficie Celular/metabolismo , Proteínas Recombinantes/farmacologíaRESUMEN
For decades, disintegrin and metalloproteinase 17 (ADAM17) has been the object of deep investigation. Since its discovery as the tumor necrosis factor convertase, it has been considered a major drug target, especially in the context of inflammatory diseases and cancer. Nevertheless, the development of drugs targeting ADAM17 has been harder than expected. This has generally been due to its multifunctionality, with over 80 different transmembrane proteins other than tumor necrosis factor α (TNF) being released by ADAM17, and its structural similarity to other metalloproteinases. This review provides an overview of the different roles of ADAM17 in disease and the effects of its ablation in a number of in vivo models of pathological conditions. Furthermore, here, we comprehensively encompass the approaches that have been developed to accomplish ADAM17 selective inhibition, from the newest non-zinc-binding ADAM17 synthetic inhibitors to the exploitation of iRhom2 to specifically target ADAM17 in immune cells.
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Proteína ADAM17/antagonistas & inhibidores , Antiinflamatorios/farmacología , Antineoplásicos/farmacología , Inflamación/tratamiento farmacológico , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neoplasias/tratamiento farmacológico , Proteína ADAM17/metabolismo , Animales , Humanos , Inflamación/metabolismo , Inflamación/patología , Neoplasias/metabolismo , Neoplasias/patologíaRESUMEN
ADAM17 is a transmembrane protease expressed by most cells in humans and mice that cleaves cell surface substrates primarily in a cis manner, a process referred to as ectodomain shedding. ADAM17 has numerous substrates and plays a broad role in various physiological processes, including as a key regulator of inflammation. At this time, little is known about ADAM17 expression and function in dogs. A well-established ADAM17 substrate is the leukocyte adhesion protein CD62L (L-selectin). We show that a selective inhibitor of ADAM17, but not an inhibitor of its most closely related family member ADAM10, blocks CD62L shedding upon canine neutrophil activation. We also tested several anti-human ADAM17 monoclonal antibodies (mAbs) for staining canine neutrophils. Although most did not recognize canine neutrophils, the mAbs MEDI3622 and D1(A12) did. They also blocked the downregulation of CD62L upon neutrophil activation. MEDI3622 is a human IgG antibody and we found that a canine chimeric version of this mAb also blocked CD62L shedding by canine leukocytes. Taken together, our findings provide the first direct evidence of ADAM17 expression and sheddase activity in dogs, establishing a potential therapeutic target for various inflammatory disorders.
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Proteína ADAM17/metabolismo , Perros/sangre , Neutrófilos/metabolismo , Proteína ADAM17/antagonistas & inhibidores , Proteína ADAM17/inmunología , Proteína ADAM17/fisiología , Animales , Anticuerpos Monoclonales/inmunología , Regulación hacia Abajo , Selectina L/metabolismoRESUMEN
Sepsis is the culmination of hyperinflammation and immune suppression in response to severe infection. Neutrophils are critical early responders to bacterial infection but can become highly dysfunctional during sepsis and other inflammatory disorders. The transmembrane protease ADAM17 (a disintegrin and metalloproteinase 17) is expressed by leukocytes and most other cells and has many substrates that regulate inflammation. We have reported that conditional knockout mice lacking ADAM17 in all leukocytes had a survival advantage during sepsis, which was associated with improved neutrophil effector functions. These and other findings indicate aberrant ADAM17 activity during sepsis. For this study, we evaluated for the first time the effects of an ADAM17 function blocking monoclonal antibody (mAb) on the pathogenesis of polymicrobial sepsis. Mice treated with the ADAM17 mAb MEDI3622 prior to sepsis induction exhibited significantly decreased mortality. When the ADAM17 mAb was combined with antibiotic administration, sepsis survival was markedly enhanced compared to either intervention alone, which was associated with a significant reduction in plasma levels of various inflammation-related factors. MEDI3622 and antibiotic administration after sepsis induction also significantly improved survival. Our results indicate that the combination of blocking ADAM17 as an immune modulator and appropriate antibiotics may provide a new therapeutic avenue for sepsis treatment.
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Proteína ADAM17/antagonistas & inhibidores , Anticuerpos Monoclonales/farmacología , Sepsis/tratamiento farmacológico , Animales , Antibacterianos/farmacología , Modelos Animales de Enfermedad , Inflamación/tratamiento farmacológico , Leucocitos/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neutrófilos/efectos de los fármacosRESUMEN
AIMS: We previously reported that fenugreek-derived 4-hydroxyisoleucine ameliorates insulin resistance via regulation of TNF-α converting enzyme (TACE) expression. In the present study, we further investigate the effects and mechanisms of fenugreek on obesity-induced inflammation and insulin signaling in the high-fat diet (HFD)-challenged obese mice. MAIN METHODS: After 12 weeks of HFD intervention, mice were treated with the low or high dosages of fenugreek. Serum levels of glucose, insulin, lipid profile, inflammation cytokines, and adipokines were detected. Macrophage infiltration and adipose tissue morphology were observed. Western blot was conducted to investigate the expressions of inactive rhomboid 2 (iRhom2) and TACE as well as other signaling pathways in subcutaneous adipose tissue. KEY FINDINGS: We showed that fenugreek significantly suppressed body weight gain and fat accumulation in HFD-challenged obese mice. Meanwhile, fasting glucose, insulin, and HOMA-IR in fenugreek-treated mice were remarkably decreased, which were properly explained by fenugreek-induced activation of the insulin receptor signaling pathway. Moreover, the anti-inflammatory properties of fenugreek were shown by the decrease of systemic and local expressions of pro-inflammatory cytokines as well as reduced macrophage infiltration into adipose tissue. Additionally, fenugreek markedly deactivated NF-κB and JNK pathways. Finally, we demonstrated that fenugreek strikingly repressed the transcriptions and expressions of iRhom2 and TACE. SIGNIFICANCE: Fenugreek shows an encouraging and promising property in ameliorating insulin resistance and suppressing inflammation in obesity, which might be realized by fenugreek-mediated inhibition of iRhom2/TACE axis-facilitated TNF-α release from adipocytes.
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Proteína ADAM17/antagonistas & inhibidores , Proteínas Portadoras/antagonistas & inhibidores , Mediadores de Inflamación/antagonistas & inhibidores , Resistencia a la Insulina/fisiología , Obesidad/tratamiento farmacológico , Trigonella , Proteína ADAM17/sangre , Animales , Proteínas Portadoras/sangre , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/fisiología , Inflamación/sangre , Inflamación/tratamiento farmacológico , Mediadores de Inflamación/sangre , Masculino , Ratones , Ratones Endogámicos C57BL , Obesidad/sangre , Extractos Vegetales/aislamiento & purificación , Extractos Vegetales/farmacología , Extractos Vegetales/uso terapéutico , SemillasRESUMEN
A key feature of osteoarthritis is the gradual loss of articular cartilage and bone deformation, resulting in the impairment of joint function. The primary cause of cartilage destruction is considered to be the presence of elevated proteases, such as matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTSs). However, clinically tested global MMP inhibitors have low efficacy that may be due to their lack of selectivity. We previously demonstrated in vitro that a variant of tissue inhibitor of metalloproteinase-3 ([-1A]TIMP3) inhibits ADAMTSs but not MMPs. In this study, we tested whether the selectivity of [-1A]TIMP3 is beneficial compared with that of the wild-type TIMP3 in preventing or delaying the onset of the degenerative effects in a mouse model of osteoarthritis. We generated transgenic mice that overexpressed TIMP3 or [-1A]TIMP3 driven by a chondrocyte-specific type II collagen promoter. TIMP3 transgenic mice showed compromised bone integrity as opposed to [-1A]TIMP3 mice. After surgically induced joint instability, TIMP3 overexpression proved to be less protective in cartilage destruction than [-1A]TIMP3 at late stages of OA. The selective inhibition of ADAMTSs provides the possibility of modifying TIMP3 to specifically target a class of cartilage-degrading proteinases and to minimize adverse effects on bone and possibly other tissues.