RESUMEN
The pathogenesis of acute kidney injury (AKI) is associated with the activation of multiple signaling pathways, including Wnt/ß-catenin signaling. However, the mechanism of Wnt/ß-catenin pathway activation in renal interstitial fibroblasts during AKI is unclear. S100 calcium-binding protein A16 (S100A16), a new member of calcium-binding protein S100 family, is a multi-functional signaling factor involved in various pathogenies, including tumors, glycolipid metabolism disorder, and chronic kidney disease (CKD). We investigated the potential participation of S100A16 in Wnt/ß-catenin pathway activation during AKI by subjecting wild-type (WT) and S100A16 knockout (S100A16+/-) mice to the ischemia-reperfusion injury (IRI), and revealed S100A16 upregulation in this model, in which knockout of S100A16 impeded the Wnt/ß-catenin signaling pathway activation and recovered the expression of downstream hepatocyte growth factor (HGF). We also found that S100A16 was highly expressed in Platelet-derived growth factor receptor beta (PDGFRß) positive renal fibroblasts in vivo. Consistently, in rat renal interstitial fibroblasts (NRK-49F cells), both hypoxia/reoxygenation and S100A16 overexpression exacerbated fibroblasts apoptosis and inhibited HGF secretion; whereas S100A16 knockdown or Wnt/ß-catenin pathway inhibitor ICG-001 reversed these changes. Mechanistically, we showed that S100A16 promoted Wnt/ß-catenin signaling activation via the ubiquitylation and degradation of ß-catenin complex members, glycogen synthase kinase 3ß (GSK3ß) and casein kinase 1α (CK1α), mediated by E3 ubiquitin ligase, the HMG-CoA reductase degradation protein 1 (HRD1). Our study identified the S100A16 as a key regulator in the activation of Wnt/ß-catenin signaling pathway in AKI.
Asunto(s)
Lesión Renal Aguda/patología , Caseína Quinasa Ialfa/metabolismo , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Proteínas S100/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Lesión Renal Aguda/metabolismo , Animales , Modelos Animales de Enfermedad , Fibroblastos/citología , Fibroblastos/metabolismo , Riñón/metabolismo , Riñón/patología , Masculino , Ratones , Ratones Noqueados , Unión Proteica , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Ratas , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Proteínas S100/antagonistas & inhibidores , Proteínas S100/deficiencia , Proteínas S100/genética , Ubiquitinación , Vía de Señalización Wnt/efectos de los fármacos , Vía de Señalización Wnt/genética , Proteína X Asociada a bcl-2/metabolismoRESUMEN
Allopregnanolone (allo) is a physiological regulator of neuronal activity that treats multiple neurological disorders. Allo penetrates the blood-brain barrier with very high efficiency, implying that allo can treat CNS-related diseases, including glioblastoma (GBM), which always recurs after standard therapy. Hence, this study aimed to determine whether allo has a therapeutic effect on GBM. We found that allo enhanced temozolomide (TMZ)-suppressed cell survival and proliferation of TMZ-resistant cells. In particular, allo enhanced TMZ-inhibited cell migration and TMZ-induced apoptosis. Additionally, allo strongly induced DNA damage characterized by γH2Ax. Furthermore, quantitative proteomic analysis, iTRAQ, showed that allo significantly decreased the levels of DPYSL3, S100A11, and S100A4, reflecting the poor prognosis of patients with GBM confirmed by differential gene expression and survival analysis. Moreover, single-cell RNA-Seq revealed that S100A11, expressed in malignant cells, oligodendrocytes, and macrophages, was significantly associated with immune cell infiltration. Furthermore, overexpression of DPYSL3 or S100A11 prevented allo-induced cell death. In conclusion, allo suppresses GBM cell survival by decreasing DPYSL3/S100A11 expression and inducing DNA damage.
Asunto(s)
Neoplasias Encefálicas , Glioblastoma , Proteínas Musculares , Pregnanolona , Proteínas S100 , Antineoplásicos Alquilantes , Apoptosis/efectos de los fármacos , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Resistencia a Antineoplásicos , Glioblastoma/tratamiento farmacológico , Glioblastoma/metabolismo , Glioblastoma/patología , Humanos , Proteínas Musculares/antagonistas & inhibidores , Proteínas Musculares/biosíntesis , Recurrencia Local de Neoplasia , Pregnanolona/farmacología , Proteómica , Proteínas S100/antagonistas & inhibidores , Proteínas S100/biosíntesis , Temozolomida/farmacologíaRESUMEN
In silico approaches identified 1, N-(6-((4-bromo- benzyl)amino)hexyl)-3,5-bis(trifluoromethyl)benzene sulfonamide, as a potential inhibitor of the S100A2-p53 protein-protein interaction, a validated pancreatic cancer drug target. Subsequent cytotoxicity screening revealed it to be a 2.97â µM cell growth inhibitor of the MiaPaCa-2 pancreatic cell line. This is in keeping with our hypothesis that inhibiting this interaction would have an anti-pancreatic cancer effect with S100A2, the validated PC drug target. A combination of focused library synthesis (three libraries, 24 compounds total) and cytotoxicity screening identified a propyl alkyl diamine spacer as optimal; the nature of the terminal phenyl substituent had limited impact on observed cytotoxicity, whereas N-methylation was detrimental to activity. In total 15 human cancer cell lines were examined, with most analogues showing broad-spectrum activity. Near uniform activity was observed against a panel of six pancreatic cancer cell lines: MiaPaCa-2, BxPC-3, AsPC-1, Capan-2, HPAC and PANC-1. In all cases there was good to excellent correlation between the predicted docking pose in the S100A2-p53 binding groove and the observed cytotoxicity, especially in the pancreatic cancer cell line with high endogenous S100A2 expression. This supports S100A2 as a pancreatic cancer drug target.
Asunto(s)
Antineoplásicos/farmacología , Factores Quimiotácticos/antagonistas & inhibidores , Proteínas S100/antagonistas & inhibidores , Proteína p53 Supresora de Tumor/antagonistas & inhibidores , Antineoplásicos/síntesis química , Antineoplásicos/química , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Factores Quimiotácticos/metabolismo , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Simulación del Acoplamiento Molecular , Estructura Molecular , Proteínas S100/metabolismo , Relación Estructura-Actividad , Células Tumorales Cultivadas , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
In silico screening predicted 1 (N-(4-((4-(3-(4-(3-methoxyphenyl)-1H-1,2,3-triazol-1-yl)propyl)piperazin-1-yl) sulfonyl)-phenyl)acetamide) as an inhibitor of the S100A2-p53 protein-protein interaction. S100A2 is a validated pancreatic cancer drug target. In the MiaPaCa-2 pancreatic cell line, 1 was a â¼50â µM growth inhibitor. Synthesis of five focused compound libraries and cytotoxicity screening revealed increased activity from the presence of electron withdrawing moieties on the sulfonamide aromatic ring, with the 3,5-bis-CF3 Library 3 analogues the most active, with GI50 values of 0.91 (3-ClPh; 13 i; BxPC-3, Pancreas) to 9.0â µM (4-CH3 ; 13 d; PANC-1, Pancreas). Activity was retained against an expanded pancreatic cancer cell line panel (MiaPaCa-2, BxPC-3, AsPC-1, Capan-2, PANC-1 and HPAC) and the normal cell line MCF10A (breast). Bulky 4-disposed substituents on the terminal phenyl ring enhanced broad spectrum activity with growth inhibition values spanning 1.1 to 3.1â µM (4-C(CH3 )3 ; 13 e; BxPC-3 and AsPC-1 (pancreas), respectively). Central alkyl spacer contraction from propyl to ethyl proved detrimental to activity with Library 4 and 5.5- to 10-fold less cytotoxic than the propyl linked Library 2 and Library 3. The data herein was consistent with the predicted binding poses of the compounds evaluated. The highest levels of cytotoxicity were observed with those analogues best capable of adopting a near identical pose to the p53-peptide in the S100A2-p53 binding groove.
Asunto(s)
Antineoplásicos/farmacología , Factores Quimiotácticos/antagonistas & inhibidores , Proteínas S100/antagonistas & inhibidores , Triazoles/farmacología , Proteína p53 Supresora de Tumor/antagonistas & inhibidores , Antineoplásicos/síntesis química , Antineoplásicos/química , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Factores Quimiotácticos/metabolismo , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Simulación del Acoplamiento Molecular , Estructura Molecular , Proteínas S100/metabolismo , Relación Estructura-Actividad , Triazoles/síntesis química , Triazoles/química , Células Tumorales Cultivadas , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
S100B, a biomarker of malignant melanoma, interacts with the p53 protein and diminishes its tumor suppressor function, which makes this S100 family member a promising therapeutic target for treating malignant melanoma. However, it is a challenge to design inhibitors that are specific for S100B in melanoma versus other S100-family members that are important for normal cellular activities. For example, S100A1 is most similar in sequence and structure to S100B, and this S100 protein is important for normal skeletal and cardiac muscle function. Therefore, a combination of NMR and computer aided drug design (CADD) was used to initiate the design of specific S100B inhibitors. Fragment-based screening by NMR, also termed "SAR by NMR," is a well-established method, and was used to examine spectral perturbations in 2D [1H, 15N]-HSQC spectra of Ca2+-bound S100B and Ca2+-bound S100A1, side-by-side, and under identical conditions for comparison. Of the 1000 compounds screened, two were found to be specific for binding Ca2+-bound S100A1 and four were found to be specific for Ca2+-bound S100B, respectively. The NMR spectral perturbations observed in these six data sets were then used to model how each of these small molecule fragments showed specificity for one S100 versus the other using a CADD approach termed Site Identification by Ligand Competitive Saturation (SILCS). In summary, the combination of NMR and computational approaches provided insight into how S100A1 versus S100B bind small molecules specifically, which will enable improved drug design efforts to inhibit elevated S100B in melanoma. Such a fragment-based approach can be used generally to initiate the design of specific inhibitors for other highly homologous drug targets.
Asunto(s)
Resonancia Magnética Nuclear Biomolecular , Subunidad beta de la Proteína de Unión al Calcio S100/antagonistas & inhibidores , Proteínas S100/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/farmacología , Propuestas de Licitación , Humanos , Ligandos , Subunidad beta de la Proteína de Unión al Calcio S100/química , Proteínas S100/química , Bibliotecas de Moléculas Pequeñas/químicaRESUMEN
S100P, a calcium-binding protein, is known to advance tumor progression and metastasis in pancreatic and several other cancers. Herein is described the in silico identification of a putative binding pocket of S100P to identify, synthesize and evaluate novel small molecules with the potential to selectively bind S100P and inhibit its activation of cell survival and metastatic pathways. The virtual screening of a drug-like database against the S100P model led to the identification of over 100 clusters of diverse scaffolds. A representative test set identified a number of structurally unrelated hits that inhibit S100P-RAGE interaction, measured by ELISA, and reduce in vitro cell invasion selectively in S100P-expressing pancreatic cancer cells at 10 µM. This study establishes a proof of concept in the potential for rational design of small molecule S100P inhibitors for drug candidate development.
Asunto(s)
Antineoplásicos/farmacología , Diseño de Fármacos , Neoplasias Pancreáticas/patología , Proteínas S100/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/farmacología , Antineoplásicos/química , Línea Celular Tumoral , Relación Dosis-Respuesta a Droga , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Invasividad Neoplásica , Bibliotecas de Moléculas Pequeñas/químicaRESUMEN
Interactions of the receptor for advanced glycation end product (RAGE) and its ligands in the context of their role in diabetes mellitus, inflammation, and carcinogenesis have been extensively investigated. This review focuses on the role of RAGE-ligands and anti-RAGE drugs capable of controlling cancer progression. Different studies have demonstrated interaction of RAGE with a diverse range of acidic (negatively charged) ligands such as advanced glycation end products (AGEs), high-mobility group box1 (HMGB1), and S100s, and their importance to cancer progression. Some RAGE-ligands displayed effects on anti- and pro-apoptotic proteins through upregulation of the phosphatidylinositide 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR), mitogen-activated protein kinases (MAPKs), matrix metalloproteinases (MMPs), vascular endothelial growth factor (VEGF), and nuclear factor kappa B (NF-κB) pathways, while downregulating p53 in cancer progression. In addition, RAGE may undergo ligand-driven multimodal dimerization or oligomerization mediated through self-association of some of its subunits. We conclude our review by proposing possible future lines of study that could result in control of cancer progression through RAGE inhibition.
Asunto(s)
Productos Finales de Glicación Avanzada/metabolismo , Neoplasias/metabolismo , Animales , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Productos Finales de Glicación Avanzada/antagonistas & inhibidores , Proteína HMGB1/antagonistas & inhibidores , Proteína HMGB1/metabolismo , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/patología , Proteínas S100/antagonistas & inhibidores , Proteínas S100/metabolismo , Transducción de SeñalRESUMEN
The protective effect of methylene blue (MB) was investigated on the model of focal one-sided traumatic brain injury (TBI) of the sensorimotor cortex region from 1 to 7 days after the injury. TBI caused a reliable disruption of the functions of the limbs contralateral to injury focus, an increase in the expression of S100 protein and blood-brain barrier (BBB) permeability in the ipsilateral hemisphere. The single intravenous injection of MB (1 mg/kg body weight) 30 min after TBI significantly reduced the limb function impairment as well as a TBI-induced increase in the expression of inflammatory marker S100 protein, and BBB permeability. When modeling inflammation in vitro, MB was found to protect cultured neurons from the toxic effects of lipopolysaccharide. In conclusion, the preservation of blood-brain barrier and a decrease in the expression of S100 protein may be an important mechanism by means of which MB improves neurological outcome. Our data demonstrate that MB can be a very promising pharmacological compound with neuroprotective properties for TBI treatment.
Asunto(s)
Barrera Hematoencefálica/efectos de los fármacos , Lesiones Traumáticas del Encéfalo/tratamiento farmacológico , Azul de Metileno/administración & dosificación , Enfermedades del Sistema Nervioso/tratamiento farmacológico , Fármacos Neuroprotectores/administración & dosificación , Proteínas S100/antagonistas & inhibidores , Administración Intravenosa , Animales , Barrera Hematoencefálica/metabolismo , Lesiones Traumáticas del Encéfalo/metabolismo , Células Cultivadas , Inhibidores Enzimáticos/administración & dosificación , Expresión Génica , Fuerza de la Mano/fisiología , Masculino , Enfermedades del Sistema Nervioso/metabolismo , Ratas , Ratas Wistar , Proteínas S100/biosíntesis , Proteínas S100/genéticaRESUMEN
High fat diet is a risk factor for the development of atherosclerosis. Hence, research studies are important to understand the cellular and molecular mechanisms of atherosclerosis pathogenesis. The current study was conducted to evaluate the role of vitamin D in modulation of aortic histopathological, immunohistochemical alterations and biochemical changes induced by high fat diet in male albino rats. Forty adult rats were divided into three major groups; group I (control), group II (High fat diet) and group III (High fat diet with vitamin D). At the end of the experiment, blood cholesterol and triglycerides were determined. Aortic arches specimens were collected for histopathological study and immunohistochemical staining. Aorta of high fat diet group showed intimal thickening with vacuolated endothelial cells. The tunica media showed areas of fibrosis and irregular vacuolated smooth muscle cells. Many inflammatory cells were detected in the tunica adventitia. Significant reduction in area percentage of smooth muscle actin-α (SMA-α) immunoreactivity and increase in number of S100 positive dendritic cells (DCs) with significant increase in serum cholesterol and triglycerides were also detected. Concomitant vitamin D supplementation, with high fat diet, showed amelioration in histopathological aortic changes with significant increase in SMA-α immunoreactivity and decrease in S100 positive (DCs). However, serum cholesterol and triglyceride showed non-significant decrease after vitamin D supplementation. In conclusion, vitamin D administration ameliorates aortic wall histoopathological changes induced by high fat diet most probably through local modulation of S100 and SMA-α immunoreactivity. Hence, vitamin D could be suggested as a protective agent against aortic atherosclerotic changes.
Asunto(s)
Actinas/genética , Aterosclerosis/dietoterapia , Suplementos Dietéticos , Proteínas S100/genética , Vitamina D/administración & dosificación , Actinas/agonistas , Actinas/metabolismo , Animales , Aorta/metabolismo , Aorta/patología , Aterosclerosis/etiología , Aterosclerosis/genética , Aterosclerosis/metabolismo , Grosor Intima-Media Carotídeo , Colesterol/sangre , Células Dendríticas/metabolismo , Células Dendríticas/patología , Dieta Alta en Grasa , Células Endoteliales/metabolismo , Células Endoteliales/patología , Inmunohistoquímica , Masculino , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/patología , Ratas , Proteínas S100/antagonistas & inhibidores , Proteínas S100/metabolismo , Triglicéridos/sangre , Vitamina D/sangreRESUMEN
Antibodies to 5100 proteins (anti-5100) in release-active form (RA anti-5100) are an active component of some domestic drugs(tenoten, tenoten for children, divaza, brizantin, kolofort and proproten-100). The authors present the results of preclinical and clinical trials (with detailed consideration of experimental data) which demonstrated a wide spectrum of specific pharmacological activity and safety as well as mechanisms of anti-5100 action.
Asunto(s)
Anticuerpos/farmacología , Proteínas S100/antagonistas & inhibidores , Proteínas S100/inmunología , Estrés Psicológico/terapia , Anticuerpos/efectos adversos , Anticuerpos/uso terapéutico , Estudios Clínicos como Asunto , Evaluación Preclínica de Medicamentos , Humanos , Estrés Psicológico/inmunologíaRESUMEN
Structure-based drug discovery is under way to identify and develop small-molecule S100B inhibitors (SBiXs). Such inhibitors have therapeutic potential for treating malignant melanoma, since high levels of S100B downregulate wild-type p53 tumor suppressor function in this cancer. Computational and X-ray crystallographic studies of two S100B-SBiX complexes are described, and both compounds (apomorphine hydrochloride and ethidium bromide) occupy an area of the S100B hydrophobic cleft which is termed site 3. These data also reveal novel protein-inhibitor interactions which can be used in future drug-design studies to improve SBiX affinity and specificity. Of particular interest, apomorphine hydrochloride showed S100B-dependent killing in melanoma cell assays, although the efficacy exceeds its affinity for S100B and implicates possible off-target contributions. Because there are no structural data available for compounds occupying site 3 alone, these studies contribute towards the structure-based approach to targeting S100B by including interactions with residues in site 3 of S100B.
Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Proteínas S100/antagonistas & inhibidores , Proteínas S100/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Línea Celular Tumoral , Cristalografía por Rayos X , Diseño de Fármacos , Humanos , Melanoma/tratamiento farmacológico , Simulación del Acoplamiento Molecular , Proteínas S100/químicaRESUMEN
Using mouse gene expression microarray analysis, we obtained dynamic expression profiles of the whole genome in a depilation-induced hair growth mouse model. S100A3 expression increased during the anagen phase and returned to normal during the telogen phase. The effects of S100A3 blockade on the hair growth cycle were examined in mice after subcutaneous injection of an anti-mouse S100A3 antibody. Protein localization of S100A3 was confined to the hair shafts during the anagen phase and the sebaceous glands during the telogen phase. S100A3 blockade delayed hair follicle entry into the anagen phase, decreased hair elongation, and reduced the number of hair follicles in the subcutis, which correlated with the downregulated expression of hair growth induction-related genes in vivo. The present study demonstrates that anti-S100A3 antibody inhibits mouse hair growth, suggesting that S100A3 can be used as a target for hair loss treatment.
Asunto(s)
Cabello/crecimiento & desarrollo , Cabello/metabolismo , Proteínas S100/metabolismo , Animales , Anticuerpos/farmacología , Cabello/efectos de los fármacos , Folículo Piloso/efectos de los fármacos , Folículo Piloso/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas S100/antagonistas & inhibidores , Proteínas S100/inmunología , Glándulas Sebáceas/efectos de los fármacos , Glándulas Sebáceas/metabolismoRESUMEN
OBJECTIVE: To explore the role of S100A4 in the epithelial-mesenchymal transition (EMT) in esophageal squamous cell carcinoma and its possible molecular mechanism. METHODS: Three chemically synthesized S100A4 siRNA sequences were transiently transfected into esophageal carcinoma EC9706 cells. EC9706 cells transfected with negative siRNA, lipofectamine 2000, and vacant EC9706 cells were used as control. Fluorescence quantitative RT-PCR and Western blot were used to detect the inhibition rate of S100A4 siRNA. S100A4 siRNA2 with the best inhibition rate was chosen to transiently transfect into EC9706 cells under the same conditions. The EC9706 cells transfected with negative siRNA, lipofectamine 2000 and vacant EC9706 cells were also used as control. Fluorescence quantitative RT-PCR and Western blot were used to detect the mRNA and protein expressions of E-cadherin, vimentin and snail. The morphology of EC9706 cells was observed under an inverted microscope. Boyden chamber and scratch test were used to detect the invasion and migration ability of EC9706 cells, and CCK8 assay was used to detect the proliferation ability of EC9706 cells. EC9706 cells transfected with S100A4 siRNA2 were further transfected with snail eukaryotic expression vector. The EC9706 cells transfected with S100A4 siRNA, EC9706 cells transfected with snail eukaryotic expression vector and vacant EC9706 cells were used as control. The above indexes of all the groups were observed, too. RESULTS: The S100A4 mRNA and protein expression levels of the S100A4 siRNA2 group were 0.417 ± 0.041 and 0.337 ± 0.039, the transmembrane cell number was 61.608 ± 8.937, the scratch healing distance was (0.216 ± 0.064) mm, the A value was 0.623 ± 0.084, the E-cadherin mRNA and protein levels were 0.619 ± 0.032 and 0.495 ± 0.034, the vimentin mRNA and protein levels were 0.514 ± 0.032 and 0.427 ± 0.028, the snail mRNA and protein levels were 0.573 ± 0.029 and 0.429 ± 0.041. These data were significantly different with the liposome group, the negative control group and the blank group (P < 0.05 for all). After the S100A4 siRNA2 treatment for 24 h, the appearance of EC9706 cells changed to epithelial cell morphology. The transmembrane cell number and the scratch healing distance of the S100A4 siRNA2+snail eukaryotic expression vector group were (69.382 ± 9.666) cells and (0.274 ± 0.029) mm, the A value was 0.823 ± 0.101, the snail mRNA and protein levels were 0.704 ± 0.037 and 0.625 ± 0.031, the vimentin mRNA and protein levels were 0.712 ± 0.046 and 0.609 ± 0.038, and these data were significantly higher than those of the Sl00A4 siRNA2 group (P < 0.05 for all). The E-cadherin mRNA and protein levels of the S100A4 siRNA2+eukaryotic expression vector group were 0.437 ± 0.038 and 0.381 ± 0.031, significantly lower than those of the S100A4 siRNA2 group (P < 0.05 for all). However, snail had no effect on the morphology of EC9706 cells. CONCLUSIONS: S100A4 may be involved in the EMT process of esophageal squamous-cell carcinoma by regulating the expression of snail and then plays a role in the invasion and metastasis of esophageal carcinoma.
Asunto(s)
Carcinoma de Células Escamosas/fisiopatología , Transición Epitelial-Mesenquimal , Neoplasias Esofágicas/fisiopatología , ARN Interferente Pequeño/fisiología , Proteínas S100/fisiología , Cadherinas/análisis , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Células Epiteliales , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/patología , Carcinoma de Células Escamosas de Esófago , Humanos , Indicadores y Reactivos , Lípidos , ARN Mensajero/análisis , ARN Interferente Pequeño/análisis , Proteína de Unión al Calcio S100A4 , Proteínas S100/antagonistas & inhibidores , Proteínas S100/genética , Factores de Transcripción de la Familia Snail , Factores de Transcripción/análisis , Factores de Transcripción/genética , Transfección , Vimentina/análisis , Vimentina/genéticaRESUMEN
BACKGROUND: The tumor microenvironment plays a determinative role in stimulating tumor progression and metastasis. Notably, tumor-stroma signals affect the pattern of infiltrated immune cells and the profile of tumor-released cytokines. Among the known molecules that are engaged in stimulating the metastatic spread of tumor cells is the S100A4 protein. S100A4 is known as an inducer of inflammatory processes and has been shown to attract T-cells to the primary tumor and to the pre-metastatic niche. The present study aims to examine the immunomodulatory role of S100A4 in vivo and in vitro and assess the mode of action of 6B12, a S100A4 neutralizing antibody. METHODS: The therapeutic effect of the 6B12 antibody was evaluated in two different mouse models. First, in a model of spontaneous breast cancer we assessed the dynamics of tumor growth and metastasis. Second, in a model of metastatic niche formation we determined the expression of metastatic niche markers. The levels of cytokine expression were assessed using antibody as well as PCR arrays and the results confirmed by qRT-PCR and ELISA. T-cell phenotyping and in vitro differentiation analyses were performed by flow cytometry. RESULTS: We show that the S100A4 protein alters the expression of transcription factor and signal transduction pathway genes involved in the T-cell lineage differentiation. T-cells challenged with S100A4 demonstrated reduced proportion of Th1-polarized cells shifting the Th1/Th2 balance towards the Th2 pro-tumorigenic phenotype. The 6B12 antibody restored the Th1/Th2 balance. Furthermore, we provide evidence that the 6B12 antibody deploys its anti-metastatic effect, by suppressing the attraction of T-cells to the site of primary tumor and pre-metastatic niche. This was associated with delayed primary tumor growth, decreased vessel density and inhibition of metastases. CONCLUSION: The S100A4 blocking antibody (6B12) reduces tumor growth and metastasis in a model of spontaneous breast cancer. The 6B12 antibody treatment inhibits T cell accumulation at the primary and pre-metastatic tumor sites. The 6B12 antibody acts as an immunomodulatory agent and thus supports the view that the 6B12 antibody is a promising therapeutic candidate to fight cancer.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Anticuerpos Neutralizantes/farmacología , Neoplasias/inmunología , Neoplasias/metabolismo , Proteínas S100/antagonistas & inhibidores , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/inmunología , Animales , Diferenciación Celular , Línea Celular Tumoral , Citocinas/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Activación de Linfocitos/genética , Activación de Linfocitos/inmunología , Recuento de Linfocitos , Ratones , Ratones Noqueados , Metástasis de la Neoplasia , Neoplasias/genética , Neoplasias/patología , Proteína de Unión al Calcio S100A4 , Proteínas S100/genética , Proteínas S100/metabolismo , Transducción de Señal , Bazo/inmunología , Subgrupos de Linfocitos T/efectos de los fármacos , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Linfocitos T/metabolismo , Microambiente Tumoral/genéticaRESUMEN
S100A4, a member of a calcium-regulated protein family, is involved in various cellular signaling pathways. From many studies over the last decade or so, it has become clear that it is involved in tumor metastasis, probably playing a determinative role. However, except the phenothiazine group of drugs, no significant inhibitor of S100A4 has been reported. Even the phenothiazines are very weak inhibitors of S100A4 action. In this study, we report design and development of a conformationally constrained helical peptide modeled on the non-muscle myosin peptide that binds to S100A4. This conformationally constrained peptide binds to S100A4 with a dissociation constant in the nanomolar range. We also synthesized a peptide for experimental control that bears several alanine mutations in the peptide-protein interface. We demonstrate that the former peptide specifically inhibits motility of H1299 and MCF-7 cells in a wound-healing assay. Structures of several S100A4-ligand complexes suggest that it may be possible to develop a smaller peptide-small molecule conjugate having high affinity for S100A4. Peptide-drug conjugates of this kind may play an important role in developing drug leads against this antimetastasis target.
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Antineoplásicos/química , Antineoplásicos/farmacología , Movimiento Celular/efectos de los fármacos , Péptidos/química , Péptidos/farmacología , Proteínas S100/antagonistas & inhibidores , Secuencia de Aminoácidos , Línea Celular Tumoral , Humanos , Células MCF-7 , Modelos Moleculares , Datos de Secuencia Molecular , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Neoplasias/patología , Miosina Tipo IIA no Muscular/química , Miosina Tipo IIA no Muscular/farmacología , Miosina Tipo IIB no Muscular/química , Miosina Tipo IIB no Muscular/farmacología , Estructura Secundaria de Proteína , Proteína de Unión al Calcio S100A4 , Proteínas S100/metabolismoRESUMEN
S100A4 expression is associated with poor clinical outcomes of patients with pancreatic cancer. The effects of loss or gain of S100A4 were examined in pancreatic cancer cell lines. S100A4 downregulation remarkably reduces cell migration and invasion, inhibits proliferation, and induces apoptosis in pancreatic tumor cells. S100A4 downregulation results in significant cell growth inhibition and apoptosis in response to TGF-ß1, supporting a non-canonical role of S100A4 in pancreatic cancer. The role of S100A4 in tumor progression was studied by using an orthotopic human pancreatic cancer xenograft mouse model. Tumor mass is remarkably decreased in animals injected with S100A4-deficient pancreatic tumor cells. P27(Kip1) expression and cleaved caspase-3 are increased, while cyclin E expression is decreased, in S100A4-deficient pancreatic tumors in vivo. S100A4-deficient tumors have lower expression of vascular endothelial growth factor, suggesting reduced angiogenesis. Biochemical assays revealed that S100A4 activates Src and focal adhesion kinase (FAK) signaling events, and inhibition of both kinases is required to maximally block the tumorigenic potential of pancreatic cancer cells. These findings support that S100A4 plays an important role in pancreatic cancer progression in vivo and S100A4 promotes tumorigenic phenotypes of pancreatic cancer cells through the Src-FAK mediated dual signaling pathway.
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Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Proteínas S100/metabolismo , Familia-src Quinasas/metabolismo , Animales , Caspasa 3/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Ciclina E/metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Regulación hacia Abajo , Femenino , Humanos , Ratones , Ratones SCID , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Proteína de Unión al Calcio S100A4 , Proteínas S100/antagonistas & inhibidores , Proteínas S100/genética , Transducción de Señal , Factor de Crecimiento Transformador beta1/farmacología , Trasplante Heterólogo , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
In humans, the S100 protein family is composed of 21 members that exhibit a high degree of structural similarity, but are not functionally interchangeable. This family of proteins modulates cellular responses by functioning both as intracellular Ca(2+) sensors and as extracellular factors. Dysregulated expression of multiple members of the S100 family is a common feature of human cancers, with each type of cancer showing a unique S100 protein profile or signature. Emerging in vivo evidence indicates that the biology of most S100 proteins is complex and multifactorial, and that these proteins actively contribute to tumorigenic processes such as cell proliferation, metastasis, angiogenesis and immune evasion. Drug discovery efforts have identified leads for inhibiting several S100 family members, and two of the identified inhibitors have progressed to clinical trials in patients with cancer. This Review highlights new findings regarding the role of S100 family members in cancer diagnosis and treatment, the contribution of S100 signalling to tumour biology, and the discovery and development of S100 inhibitors for treating cancer.
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Diseño de Fármacos , Neoplasias/diagnóstico , Neoplasias/tratamiento farmacológico , Proteínas S100/metabolismo , Animales , Calcio/metabolismo , Femenino , Humanos , Ratones , Unión Proteica , Proteínas S100/antagonistas & inhibidores , Proteínas S100/genética , Transducción de Señal/genéticaRESUMEN
OBJECTIVE: To investigate the role of S100A3 and the effect of S100A3 inhibition on human castration-resistant prostate cancer (CRPC) cells by using in vitro and in vivo functional assays. MATERIALS AND METHODS: Using human CRPC cells (PC3 and DU145), S100A3 expression levels were assessed by reverse transcription-polymerase chain reaction and Western blot analysis. After S100A3-specific small interfering ribonucleic acid (RNA) treatment, cell viability was determined by Cell Counting Kit-8 assay, and apoptotic cell fractions were evaluated by flow cytometry. The invasive properties of these cells and the expression pattern of matrix metalloproteinases (MMPs) were assessed using transwell migration assays, reverse transcription-polymerase chain reaction, and gelatin zymography. Finally, the in vivo efficacy of S100A3 inhibition on human CRPC cells was investigated using human tumor xenograft models in nude mice. RESULTS: Human CRPC cells showed overexpression of S100A3, and its suppression reduced cell viability owing to apoptotic cell death. Additionally, S100A3 inhibition decreased the invasiveness of human CRPC cells. Moreover, MMP-2 and MMP-9 were downregulated in PC3, whereas only MMP-9 was downregulated in D145 after S100A3 inhibition. In human CRPC xenograft models, we noted a marked reduction in tumor growth in mice injected with S100A3 short hairpin RNA-transfected PC3 and DU145 cells. CONCLUSION: Human CRPC cells showed upregulation of S100A3 expression, and S100A3 inhibition reduced tumor cell viability. S100A3 inhibition reduced invasion capability with downregulation of MMP expression. More importantly, S100A3 inhibition resulted in tumor growth suppression in human CRPC xenograft models, suggesting S100A3 could serve as a novel therapeutic target for the treatment of human CRPC.
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Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Neoplasias de la Próstata Resistentes a la Castración/patología , Proteínas S100/antagonistas & inhibidores , Animales , Humanos , Técnicas In Vitro , Masculino , Ratones Desnudos , Invasividad Neoplásica , Células Tumorales CultivadasRESUMEN
AIMS: The hedgehog signaling pathway plays an important role in EMT of pancreatic cancer cells, but the precise mechanisms remain elusive. Because S100A4 as a key EMT moleculer marker was found to be upregulated upon Gli1 in pancreatic cancer cells, we focused on the relationship between Shh-Gli1 signals and S100 genes family. METHODS: On the base of cDNA microarray data, we investigated regulating mechanism of Gli1 to some members of S100A genes family in pancreatic cancer cell lines firstly. Then, the regulation of Gli1 to S100A4 gene was studied by molecular biology assays and the pro-metastasis effection of Gli1-dependent S100A4 was investigated in vitro. Finally, the expressions of Shh, Gli1, S100A4 and E-cadherin in pancreatic cancer tissues were studied by using immunohistochemistry assays. RESULTS: Five members of the S100 genes family, S100A2, S100A4, S100A6, S100A11, and S100A14 were found to be downregulated significantly upon Gli1 knockdown. Gli1 enhancer prediction combining with in vitro data demonstrated that Gli1 primarily regulates S100A family members via cis-acting elements. Indeed, the data indicate S100A4 and vimentin genes were upregulated significantly by Shh/Gli1-expression increasing and E-cadherin was significantly reduced at the same time. Migration of PC cells was increased significantly in a dose-dependent manner of Gli1 expression (P<0.05) and siS100A4 significantly reversed the response of PC cells induced by L-Shh transduction (P<0.01). CONCLUSION: Our data establish a novel connection between Shh-Gli1 signaling and S100A4 regulation, which imply that S100A4 might be one of the key factors in EMT mediated by Shh-Gli1 signaling in pancreatic cancer.
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Proteínas Hedgehog/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , Proteínas S100/genética , Proteínas S100/metabolismo , Factores de Transcripción/metabolismo , Adulto , Anciano , Secuencia de Bases , Sitios de Unión , Cadherinas/metabolismo , Línea Celular Tumoral , Movimiento Celular , Regulación hacia Abajo , Transición Epitelial-Mesenquimal , Femenino , Células HT29 , Proteínas Hedgehog/genética , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Pancreáticas/metabolismo , Regiones Promotoras Genéticas , Unión Proteica , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Proteína de Unión al Calcio S100A4 , Proteínas S100/antagonistas & inhibidores , Transducción de Señal , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/genética , Regulación hacia Arriba , Vimentina/metabolismo , Proteína con Dedos de Zinc GLI1RESUMEN
S100A4, also known as the mts1 gene, has been reported as an invasive and metastatic marker for many types of cancers. S100A4 interacts with various target genes that affect tumor cell metastasis; however, little is known about cellular signaling pathways elicited by S100A4. In the current study, we demonstrate an inhibitory effect of S100A4 on ß-catenin signaling in breast cancer cells. By overexpressing S100A4 in MCF-7, MDA-MB-231 and MDA-MB-453 breast cancer cells, we observed the down-regulation of ß-catenin expression and ß-catenin-dependent TCF/LEF transcriptional activities. The activity of GSK3ß, which phosphorylates ß-catenin and induces proteasomal degradation of ß-catenin, was increased in S100A4-overexpressing cell lines. Blocking Glycogen Synthase Kinase (GSK3ß) activity by lithium chloride or Dvl gene overexpression restored ß-catenin expression. We also found that increased GSK3ß activity was due to decrease in Akt activity resulting from Egr-1-induced phosphatase and tensin homolog (PTEN) expression. S100A4 induced Egr-1 nuclear localization by increasing the association between Egr-1 and importin-7 and this effect was reduced in S100A4 mutants that harbored a defect in nuclear localization signals. Collectively, we verify herein that S100A4 may act as a tumor suppressor in breast cancers by down-regulating the central signaling axis for tumor cell survival.