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Rheumatoid arthritis (RA) is globally treated with several commercially available anti-inflammatory and analgesic drugs, which pose adverse side effects in many cases. Due to increasing population affected by autoimmune disorder of joints inflammation, it is crucial to use natural therapies, which are less toxic at metabolic level and promote gut health. In this study, we investigated the potential role of a locally developed traditional Chinese medicine (TCM), namely Duzheng tablet (DZGP) in controlling the RA. For this purpose, we introduced RA in male mice and divided them into 5 different groups. High throughput transcriptome analysis of synovial cells after DZGP treatment in arthritic mice revealed a significant alteration of gene expression. The correlation analysis of transcriptome with metabolites revealed that DZGP specifically targeted the B cells mediated immunity pathways. Treatment with DZGP inhibited the cytokines production, while reducing the production of inflammatory TNF-α, which led to the alleviation of inflammatory response in arthritic mice. Additionally, we applied integrated approach using 16S rDNA sequencing to understand the microbial population in relation to metabolites accumulation. The results showed that DZGP promoted the healthy gut microbiota by maintaining the ratio of Firmicutes and Bacteroidota and introduction of two additional phyla namely, Verrucomicrobiota and Cyanobacteria. Therefore, it is concluded that DZGP offers an advantage over commercial drug by changing the metabolic profile, gut microbiota while exhibiting lower cellular toxicity.
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Sweet potatoes are rich in flavonoids and phenolic acids, showing incomparable nutritional and health value. In this investigation, we comprehensively analyzed the secondary metabolite profiles in the flesh of different-colored sweet potato flesh. We determined the metabolomic profiles of white sweet potato flesh (BS), orange sweet potato flesh (CS), and purple sweet potato flesh (ZS) using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The CS vs. BS, ZS vs. BS, and ZS vs. CS comparisons identified a total of 4447 secondary metabolites, including 1540, 1949, and 1931 differentially accumulated metabolites. Among them, there were significant differences in flavonoids and phenolic acids. There were 20 flavonoids and 13 phenolic acids that were common differential metabolites among the three comparison groups. The accumulation of paeoniflorin-like and delphinidin-like compounds may be responsible for the purple coloration of sweet potato flesh. These findings provide new rationale and insights for the development of functional foods for sweet potatoes. List of compounds: Kaempferol (PubChem CID: 5280863); Peonidin 3-(6"-p-coumarylglucoside) (PubChem CID: 44256849); Swerchirin (PubChem CID: 5281660); Trilobatin (PubChem CID: 6451798); 3-Geranyl-4-hydroxybenzoate (PubChem CID: 54730540); Eupatorin (PubChem CID: 97214); Icaritin (PubChem CID: 5318980); Isorhamnetin (PubChem CID: 5281654); Glucoliquiritin apioside (PubChem CID: 74819335); Brazilin (PubChem CID: 73384).
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Topological electronic transition is the very promising strategy for achieving high band degeneracy (NV) and for optimizing thermoelectric performance. Herein, this work verifies in p-type Mg3Sb2- xBix that topological electronic transition could be the key mechanism responsible for elevating the NV of valence band edge from 1 to 6, leading to much improved thermoelectric performance. Through comprehensive spectroscopy characterizations and theoretical calculations of electronic structures, the topological electronic transition from trivial semiconductor is unambiguously demonstrated to topological semimetal of Mg3Sb2- xBix with increasing the Bi content, due to the strong spin-orbit coupling of Bi and the band inversion. The distinct evolution of Fermi surface configuration and the multivalley valence band edge with NV of 6 are discovered in the Bi-rich compositions, while a peculiar two-step band inversion is revealed for the first time in the end compound Mg3Bi2. As a result, the optimal p-type Mg3Sb0.5Bi1.5 simultaneously obtains a positive bandgap and high NV of 6, and thus acquires the largest thermoelectric power factor of 3.54 and 6.93 µW cm-1 K-2 at 300 and 575 K, respectively, outperforming the values in other compositions. This work provides important guidance on improving thermoelectric performance of p-type Mg3Sb2- xBix utilizing the topological electronic transition.
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Ageing is a phenomenon in which cells, tissues and organs undergo systemic pathological changes as individuals age, leading to the occurrence of ageing-related diseases and the end of life. It is associated with many phenotypes known as ageing characteristics, such as genomic instability, nutritional imbalance, mitochondrial dysfunction, cell senescence, stem cell depletion, and an altered microenvironment. The sirtuin family (SIRT), known as longevity proteins, is thought to delay ageing and prolong life, and mammals, including humans, have seven family members (SIRT1-7). SIRT4 has been studied less among the sirtuin family thus far, but it has been reported that it has important physiological functions in organisms, such as promoting DNA damage repair, participating in the energy metabolism of three substances, inhibiting inflammatory reactions and apoptosis, and regulating mitochondrial function. Recently, some studies have demonstrated the involvement of SIRT4 in age-related processes, but knowledge in this field is still scarce. Therefore, this review aims to analyse the relationship between SIRT4 and ageing characteristics as well as some age-related diseases (e.g., cardiovascular diseases, metabolic diseases, neurodegenerative diseases and cancer).
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Neoplasias , Sirtuinas , Animales , Humanos , Envejecimiento/metabolismo , Senescencia Celular , Longevidad , Neoplasias/genética , Sirtuinas/metabolismo , Proteínas Mitocondriales/metabolismo , Mamíferos/metabolismo , Microambiente TumoralRESUMEN
PURPOSE: PAX-fusion negative rhabdomyosarcoma (FN RMS) is driven by alterations in the RAS/MAP kinase pathway and is partially responsive to MEK inhibition. Overexpression of IGF1R and its ligands is also observed in FN RMS. Preclinical and clinical studies have suggested that IGF1R is itself an important target in FN RMS. Our previous studies revealed preclinical efficacy of the MEK1/2 inhibitor, trametinib, and an IGF1R inhibitor, BMS-754807, but this combination was not pursued clinically due to intolerability in preclinical murine models. Here, we sought to identify a combination of an MEK1/2 inhibitor and IGF1R inhibitor, which would be tolerated in murine models and effective in both cell line and patient-derived xenograft models of RAS-mutant FN RMS. EXPERIMENTAL DESIGN: Using proliferation and apoptosis assays, we studied the factorial effects of trametinib and ganitumab (AMG 479), a mAb with specificity for human and murine IGF1R, in a panel of RAS-mutant FN RMS cell lines. The molecular mechanism of the observed synergy was determined using conventional and capillary immunoassays. The efficacy and tolerability of trametinib/ganitumab was assessed using a panel of RAS-mutated cell-line and patient-derived RMS xenograft models. RESULTS: Treatment with trametinib and ganitumab resulted in synergistic cellular growth inhibition in all cell lines tested and inhibition of tumor growth in four of six models of RAS-mutant RMS. The combination had little effect on body weight and did not produce thrombocytopenia, neutropenia, or hyperinsulinemia in tumor-bearing SCID beige mice. Mechanistically, ganitumab treatment prevented the phosphorylation of AKT induced by MEK inhibition alone. Therapeutic response to the combination was observed in models without a mutation in the PI3K/PTEN axis. CONCLUSIONS: We demonstrate that combined trametinib and ganitumab is effective in a genomically diverse panel of RAS-mutated FN RMS preclinical models. Our data also show that the trametinib/ganitumab combination likely has a favorable tolerability profile. These data support testing this combination in a phase I/II clinical trial for pediatric patients with relapsed or refractory RAS-mutated FN RMS.
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Rabdomiosarcoma , Humanos , Animales , Ratones , Niño , Línea Celular Tumoral , Ratones SCID , Rabdomiosarcoma/tratamiento farmacológico , Rabdomiosarcoma/genética , Rabdomiosarcoma/patología , Inhibidores de Proteínas Quinasas/farmacología , Quinasas de Proteína Quinasa Activadas por MitógenosRESUMEN
Activating RAS mutations are found in a subset of fusion-negative rhabdomyosarcoma (RMS), and therapeutic strategies to directly target RAS in these tumors have been investigated, without clinical success to date. A potential strategy to inhibit oncogenic RAS activity is the disruption of RAS prenylation, an obligate step for RAS membrane localization and effector pathway signaling, through inhibition of farnesyltransferase (FTase). Of the major RAS family members, HRAS is uniquely dependent on FTase for prenylation, whereas NRAS and KRAS can utilize geranylgeranyl transferase as a bypass prenylation mechanism. Tumors driven by oncogenic HRAS may therefore be uniquely sensitive to FTase inhibition. To investigate the mutation-specific effects of FTase inhibition in RMS we utilized tipifarnib, a potent and selective FTase inhibitor, in in vitro and in vivo models of RMS genomically characterized for RAS mutation status. Tipifarnib reduced HRAS processing, and plasma membrane localization leading to decreased GTP-bound HRAS and decreased signaling through RAS effector pathways. In HRAS-mutant cell lines, tipifarnib reduced two-dimensional and three-dimensional cell growth, and in vivo treatment with tipifarnib resulted in tumor growth inhibition exclusively in HRAS-mutant RMS xenografts. Our data suggest that small molecule inhibition of FTase is active in HRAS-driven RMS and may represent an effective therapeutic strategy for a genomically-defined subset of patients with RMS.
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Rabdomiosarcoma Embrionario , Rabdomiosarcoma , Farnesiltransferasa/genética , Genes ras , Humanos , Prenilación , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Rabdomiosarcoma/tratamiento farmacológico , Rabdomiosarcoma/genéticaRESUMEN
Relapsed pediatric rhabdomyosarcomas (RMS) and neuroblastomas (NBs) have a poor prognosis despite multimodality therapy. In addition, the current standard of care for these cancers includes vinca alkaloids that have severe toxicity profiles, further underscoring the need for novel therapies for these malignancies. Here, we show that the small-molecule rigosertib inhibits the growth of RMS and NB cell lines by arresting cells in mitosis, which leads to cell death. Our data indicate that rigosertib, like the vinca alkaloids, exerts its effects mainly by interfering with mitotic spindle assembly. Although rigosertib has the ability to inhibit oncogenic RAS signaling, we provide evidence that rigosertib does not induce cell death through inhibition of the RAS pathway in RAS-mutated RMS and NB cells. However, the combination of rigosertib and the MEK inhibitor trametinib, which has efficacy in RAS-mutated tumors, synergistically inhibits the growth of an RMS cell line, suggesting a new avenue for combination therapy. Importantly, rigosertib treatment delays tumor growth and prolongs survival in a xenograft model of RMS. In conclusion, rigosertib, through its impact on the mitotic spindle, represents a potential therapeutic for RMS.
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Glicina/análogos & derivados , Neuroblastoma/tratamiento farmacológico , Rabdomiosarcoma/tratamiento farmacológico , Huso Acromático/metabolismo , Sulfonas/uso terapéutico , Apoptosis , Glicina/farmacología , Glicina/uso terapéutico , Humanos , Sulfonas/farmacologíaRESUMEN
Inflammation aggravates the lethal consequences of intracerebral hemorrhage. Recently, many studies have found that nuclear factor-κB (NF-κB) is a crucial transcription factor that initiates inflammation in the perihematomal region of ICH. NF-κB essential modulator (NEMO)-binding domain (NBD) peptide, a cell-permeable peptide spanning the NBD of IKKα or IKKß, functions as a highly specific inhibitor of NF-κB. This peptide can negatively regulate the NF-κB pathway. The present study aimed to explore the effects and underlying pathomechanisms of NBD peptides after ICH. Striatum infusion of whole blood or saline was performed on C57BL/6 mice (nâ¯=â¯198). Experimental animals were administered NBD or control (mutated) peptides 2â¯h before or after ICH by intracerebroventricular injection (icv.). NBD peptides significantly inhibited edema formation, ameliorated the neurological deficits, markedly reduced IκBα and p65 phosphorylation, blocked nuclear translocation of p65, and upregulated IκBα expression by NF-κB after ICH induction. Using an in vitro hemin toxicity model, we investigated the effects of NBD peptides on microglial inflammation. We found that NBD peptides suppressed microglia inflammation and lowered the expression of TNF-α and IL-1ß in both in vivo and in vitro experiments. Further experiments were performed in mice and cultured microglia, which treated with NBD peptides in the presence of p65 siRNA confirmed that the specificity of NBD peptides inhibit ICH-induced NF-κB activation. This study demonstrated that NBD peptides exert a neuroprotective role after ICH and might be a potential candidate for a novel therapeutic strategy for ICH.
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Edema Encefálico/tratamiento farmacológico , Hemorragia Cerebral/tratamiento farmacológico , Fármacos Neuroprotectores/uso terapéutico , Péptidos/uso terapéutico , Regulación hacia Arriba/efectos de los fármacos , Animales , Edema Encefálico/metabolismo , Hemorragia Cerebral/metabolismo , Quinasa I-kappa B/metabolismo , Interleucina-1beta/metabolismo , Ratones , Microglía/efectos de los fármacos , Microglía/metabolismo , FN-kappa B/metabolismo , Fármacos Neuroprotectores/farmacología , Péptidos/farmacología , Fosforilación/efectos de los fármacos , ARN Interferente Pequeño , Transducción de Señal/efectos de los fármacos , Factor de Transcripción ReIA/metabolismo , Resultado del Tratamiento , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
To determine what alternative pathways may act as mechanisms of bypass resistance to type 1 insulin-like growth factor receptor (IGF-1R) blockade in rhabdomyosarcoma (RMS), we compared expression of receptor tyrosine kinase activity in a number of IGF-1R antibody-resistant and -sensitive RMS cell lines. We found that platelet-derived growth factor receptor ß (PDGFR-ß) activity was upregulated in three xenograft-derived IGF-1R antibody-resistant cell lines that arose from a highly sensitive fusion-positive RMS cell line (Rh41). Furthermore, we identified four additional fusion-negative RMS cell lines that similarly upregulated PDGFR-ß activity when selected for IGF-1R antibody resistance in vitro. In the seven cell lines described, we observed enhanced growth inhibition when cells were treated with dual IGF-1R and PDGFR-ß inhibition in vitro. In vivo studies have confirmed the enhanced effect of targeting IGF-1R and PDGFR-ß in several mouse xenograft models of fusion-negative RMS. These findings suggest that PDGFR-ß acts as a bypass resistance pathway to IGF-1R inhibition in a subset of RMS. Therapy co-targeting these receptors may be a promising new strategy in RMS care.
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The insulin-like growth factor 1 receptor (IGF-1R) has surfaced as a significant target in multiple solid cancers due to its fundamental roles in pro-survival and anti-apoptotic signaling. However, development of resistance to IGF-1R blockade represents a significant hindrance and limits treatment efficacy in the clinic. In this study, we identified acquired resistance to IGF-1R blockade with R1507, an antibody against IGF-1R, and with BMS-754807, a small molecular inhibitor of IGF-1R/insulin receptor (IR). We showed that treatment with an IGF-IR antibody, R1507, or an IR/IGF-IR kinase inhibitor, BMS-754807, was associated with increased activation of YES/SRC family tyrosine kinase (SFK) in rhabdomyosarcoma (RMS). Combining anti-IGF-1R agents with SFK inhibitors resulted in blockade of IGF-1R inhibition-induced activation of YES/SFK and displayed advantageous antitumor activity in vitro and in vivo. Our data provide evidence that IGF-1R blockade results in activation of the YES/SRC family kinase bypass resistance pathway in vitro and in vivo. This may be of particular clinical relevance since both Yes and IGF components are overexpressed in RMS. Increased YES/SFK activation might serve as a clinical biomarker for predicting tumor resistance to IGF-1R inhibition. Dual inhibition of IGF-1R and SFK may have a broader and enhanced clinical benefit for patients with RMS.
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Resistencia a Antineoplásicos/efectos de los fármacos , Proteínas Proto-Oncogénicas c-yes/metabolismo , Receptor IGF Tipo 1/antagonistas & inhibidores , Receptor IGF Tipo 1/metabolismo , Rabdomiosarcoma/tratamiento farmacológico , Rabdomiosarcoma/metabolismo , Animales , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales Humanizados , Antineoplásicos/farmacología , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Femenino , Humanos , Ratones , Inhibidores de Proteínas Quinasas/farmacología , Pirazoles/farmacología , Receptor de Insulina/antagonistas & inhibidores , Receptor de Insulina/metabolismo , Triazinas/farmacología , Ensayos Antitumor por Modelo de Xenoinjerto , Familia-src Quinasas/metabolismoRESUMEN
We identified Bub1b as an essential element for the growth and survival of rhabdomyosarcoma (RMS) cells using a bar-coded, tetracycline-inducible short hairpin RNA (shRNA) library screen. Knockdown of Bub1b resulted in suppression of tumor growth in vivo, including the regression of established tumors. The mechanism by which this occurs is via postmitotic endoreduplication checkpoint and mitotic catastrophe. Furthermore, using a chromatin immunoprecipitation assay, we found that Bub1b is a direct transcriptional target of Forkhead Box M1 (FoxM1). Suppression of FoxM1 either by shRNA or the inhibitor siomycin A resulted in reduction of Bub1b expression and inhibition of cell growth and survival. These results show the important role of the Bub1b/FoxM1 pathway in RMS and provide potential therapeutic targets.
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Factores de Transcripción Forkhead/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Rabdomiosarcoma/metabolismo , Animales , Proteínas de Ciclo Celular , Procesos de Crecimiento Celular/fisiología , Línea Celular Tumoral , Supervivencia Celular/fisiología , Código de Barras del ADN Taxonómico/métodos , Femenino , Proteína Forkhead Box M1 , Técnicas de Silenciamiento del Gen , Humanos , Células MCF-7 , Ratones , Ratones Desnudos , Ratones SCID , Regiones Promotoras Genéticas , Proteínas Serina-Treonina Quinasas/genética , ARN Interferente Pequeño/genética , Sistema del Grupo Sanguíneo Rh-Hr/biosíntesis , Rabdomiosarcoma/genética , Rabdomiosarcoma/patología , Transducción de Señal , Trasplante HeterólogoRESUMEN
The use of multimodality therapy has resulted in markedly improved cure rates for patients with sarcomas in the past 25 years. However, for virtually all patients with metastatic or recurrent disease, survival remains dismal. The important role that members of the insulin-like growth factor (IGF) family play in tumorigenesis has been known for decades. But it has only been in the last five years, that humanized and fully human antibodies targeting insulin-like growth factor receptor 1 (IGF-1R) have been developed. The use of these agents in clinical trials has been accompanied by several dramatic responses in patients with recurrent and refractory sarcomas. In this review, we will focus on preclinical highlights in the past, current clinic trials and discuss some exciting research opportunities to foster advances in the future.
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Anticuerpos Monoclonales/uso terapéutico , Receptor IGF Tipo 1/antagonistas & inhibidores , Sarcoma/terapia , Animales , Antineoplásicos/uso terapéutico , Resistencia a Antineoplásicos , Predicción , Humanos , Ratones , Sarcoma/metabolismo , Somatomedinas/metabolismoRESUMEN
Insulin-like growth factor I receptor (IGF-IR) and its ligands are overexpressed by tumors, mediating proliferation and protecting against stress-induced apoptosis. Accordingly, there has been a considerable amount of interest in developing therapeutic agents against IGF-IR. IGF-IR is believed to be ubiquitously expressed without detectable mutation or amplification in cancer. We explored the determinants of cellular response to a humanized anti-IGF-IR antibody. Our results showed a large variation in IGF-IR levels in rhabdomyosarcoma tumor specimens that were comparable with those in rhabdomyosarcoma cell lines. In vitro analysis revealed a direct and very significant correlation between elevated IGF-IR levels and antiproliferative effects of the antibody and defined a receptor number that would predict sensitivity. Our data further suggested a strong dependence on IGF-IR for AKT signaling in cells with elevated IGF-IR. The sensitivity of the high IGF-IR-expressing cells was blocked with a constitutively active AKT. The extracellular signal-regulated kinase pathway was not affected by the antibody. In vivo studies showed that anti-IGF-IR had single-agent antitumor activity; furthermore, predictions of responses based on IGF-IR levels were accurate. In vivo biomarker analysis suggested that h7C10 down-regulated both IGF-IR and p-AKT initially, concordant with antitumor activity. Subsequent progression of tumors was associated with reactivation of p-AKT despite sustained suppression of IGF-IR. These results identified the first predictive biomarker for anti-IGF-IR therapies in cancer.
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Anticuerpos/uso terapéutico , Factor I del Crecimiento Similar a la Insulina/genética , Neoplasias Hormono-Dependientes/terapia , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor IGF Tipo 1/antagonistas & inhibidores , Receptor IGF Tipo 1/genética , Rabdomiosarcoma/terapia , Animales , Anticuerpos/farmacología , Proliferación Celular/efectos de los fármacos , Sistemas de Liberación de Medicamentos , Resistencia a Antineoplásicos/genética , Resistencia a Antineoplásicos/fisiología , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Inmunoterapia , Factor I del Crecimiento Similar a la Insulina/metabolismo , Factor I del Crecimiento Similar a la Insulina/fisiología , Ratones , Ratones SCID , Neoplasias Hormono-Dependientes/genética , Neoplasias Hormono-Dependientes/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Receptor IGF Tipo 1/inmunología , Receptor IGF Tipo 1/metabolismo , Rabdomiosarcoma/genética , Rabdomiosarcoma/metabolismo , Transducción de Señal/genética , Transducción de Señal/fisiología , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Dysregulation of the mammalian target of rapamycin (mTOR) pathway has been found in many human tumors and implicated in the promotion of cancer cell growth and survival. Hence, the mTOR pathway is considered an important target for anticancer drug development. Currently, the mTOR inhibitor rapamycin and its derivatives CCI-779, RAD001, and AP23573 are being evaluated in cancer clinical trials. To date, clinical results have shown good tolerability of treatment with mTOR inhibitors in most reports and varying effectiveness of mTOR inhibitors in a variety of tumors in a subset of patients. For the targeted treatment of sarcomas, AP23573 has shown promising clinical efficacy and low toxicity profiles in patients. Further studies should define the optimal dose/schedule, patient selection, and combination strategies with other biological agents, especially those targeting signaling pathways crucial for cell survival. Disclosure of potential conflicts of interest is found at the end of this article.
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Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Quinasas/efectos de los fármacos , Sarcoma/tratamiento farmacológico , Sarcoma/enzimología , Sirolimus/análogos & derivados , Ensayos Clínicos como Asunto , Everolimus , Humanos , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Sirolimus/farmacología , Sirolimus/uso terapéutico , Serina-Treonina Quinasas TORRESUMEN
Mapping of protein signaling networks within tumors can identify new targets for therapy and provide a means to stratify patients for individualized therapy. Despite advances in combination chemotherapy, the overall survival for childhood rhabdomyosarcoma remains approximately 60%. A critical goal is to identify functionally important protein signaling defects associated with treatment failure for the 40% nonresponder cohort. Here, we show, by phosphoproteomic network analysis of microdissected tumor cells, that interlinked components of the Akt/mammalian target of rapamycin (mTOR) pathway exhibited increased levels of phosphorylation for tumors of patients with short-term survival. Specimens (n = 59) were obtained from the Children's Oncology Group Intergroup Rhabdomyosarcoma Study (IRS) IV, D9502 and D9803, with 12-year follow-up. High phosphorylation levels were associated with poor overall and poor disease-free survival: Akt Ser(473) (overall survival P < 0.001, recurrence-free survival P < 0.0009), 4EBP1 Thr(37/46) (overall survival P < 0.0110, recurrence-free survival P < 0.0106), eIF4G Ser(1108) (overall survival P < 0.0017, recurrence-free survival P < 0.0072), and p70S6 Thr(389) (overall survival P < 0.0085, recurrence-free survival P < 0.0296). Moreover, the findings support an altered interrelationship between the insulin receptor substrate (IRS-1) and Akt/mTOR pathway proteins (P < 0.0027) for tumors from patients with poor survival. The functional significance of this pathway was tested using CCI-779 in a mouse xenograft model. CCI-779 suppressed phosphorylation of mTOR downstream proteins and greatly reduced the growth of two different rhabdomyosarcoma (RD embryonal P = 0.00008; Rh30 alveolar P = 0.0002) cell lines compared with controls. These results suggest that phosphoprotein mapping of the Akt/mTOR pathway should be studied further as a means to select patients to receive mTOR/IRS pathway inhibitors before administration of chemotherapy.
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Proteínas Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Rabdomiosarcoma/metabolismo , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Niño , Preescolar , Supervivencia sin Enfermedad , Femenino , Humanos , Lactante , Proteínas Sustrato del Receptor de Insulina , Masculino , Ratones , Ratones SCID , Fosfoproteínas/metabolismo , Fosforilación , Rabdomiosarcoma/tratamiento farmacológico , Transducción de Señal , Tasa de Supervivencia , Serina-Treonina Quinasas TOR , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Angiogenesis is one of the critical steps in tumor growth and metastasis. The goal of this study was to evaluate whether the antitumor activity of CCI-779 is related to antiangiogenic effects in vivo in tumors of mice bearing human rhabdomyosarcoma (RMS) xenografts. We now demonstrate that CCI-779 rapidly inhibits mTOR activity, as indicated by S6 reduction and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) phosphorylation in two xenograft models of RMS within 24 hours of treatment. Treatment with a single 20-mg/kg dose of CCI-779 suppressed S6 phosphorylation for more than 72 hours and 4E-BP1 phosphorylation for more than 96 hours. Based on these data, an intermittent treatment schedule (every 3 days for 30 days) was chosen and displayed a significant suppression of both tumor growth and mTOR signaling. Western blot analysis and immunohistochemical studies demonstrated that the antitumor activity of CCI-779 was associated with antiangiogenesis, as indicated by impaired levels of hypoxia-inducible factor-1alpha (Hif-1alpha) and vascular endothelial growth factor (VEGF) protein expression and by decreased microvessel density in Rh30 and RD xenografts. Together, these data suggest that CCI-779 inhibits human RMS xenograft growth by an antiangiogenic mechanism associated with the targeting of mTOR/Hif-1alpha/VEGF signaling.
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Inhibidores de la Angiogénesis/farmacología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Neovascularización Patológica , Proteínas Quinasas/metabolismo , Rabdomiosarcoma/tratamiento farmacológico , Sirolimus/análogos & derivados , Factor A de Crecimiento Endotelial Vascular/metabolismo , Animales , Antineoplásicos/farmacología , Línea Celular Tumoral , Femenino , Humanos , Ratones , Ratones SCID , Trasplante de Neoplasias , Fosforilación , Transducción de Señal , Sirolimus/farmacología , Serina-Treonina Quinasas TOR , Factores de TiempoRESUMEN
Osteosarcoma is the most frequent primary malignant tumor of bone with a high propensity for metastasis. We have previously showed that ezrin expression is necessary for metastatic behavior in a murine model of osteosarcoma (K7M2). In this study, we found that a mechanism of ezrin-related metastatic behavior is linked to an Akt-dependent mammalian target of rapamycin (mTOR)/p70 ribosomal protein S6 kinase (S6K1)/eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) pathway. Suppression of ezrin expression either by antisense transfection or by small interfering RNAs or disruption of ezrin function by transfection of a dominant-negative ezrin-T567A mutant led to decreased expression and decreased phosphorylation of both S6K1 and 4E-BP1. Proteosomal inhibition by MG132 reversed antisense-mediated decrease of S6K1 and 4E-BP1 protein expression, but failed to affect the effect of ezrin on phosphorylation of S6K1 and 4E-BP1. Blockade of the mTOR pathway with rapamycin or its analog, cell cycle inhibitor-779 led to significant inhibition of experimental lung metastasis in vivo. These results suggest that blocking the mTOR/S6K1/4E-BP1 pathway may be an appropriate target for strategies to reduce tumor cell metastasis.
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Neoplasias Óseas/patología , Osteosarcoma/patología , Fosfoproteínas/antagonistas & inhibidores , Sirolimus/análogos & derivados , Sirolimus/farmacología , Proteínas Adaptadoras Transductoras de Señales , Animales , Neoplasias Óseas/genética , Proteínas Portadoras/antagonistas & inhibidores , Proteínas Portadoras/metabolismo , Proteínas de Ciclo Celular , Proteínas del Citoesqueleto , Factores Eucarióticos de Iniciación , Femenino , Ratones , Ratones SCID , Osteosarcoma/genética , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt , ARN sin Sentido/genética , ARN Interferente Pequeño/genética , Proteínas Quinasas S6 Ribosómicas 70-kDa/antagonistas & inhibidores , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , TransfecciónRESUMEN
Metastatic cancers, once established, are the primary cause of mortality associated with cancer. Previously, we used a genomic approach to identify metastasis-associated genes in cancer. From this genomic data, we selected ezrin for further study based on its role in physically and functionally connecting the actin cytoskeleton to the cell membrane. In a mouse model of osteosarcoma, a highly metastatic pediatric cancer, we found ezrin to be necessary for metastasis. By imaging metastatic cells in the lungs of mice, we showed that ezrin expression provided an early survival advantage for cancer cells that reached the lung. AKT and MAPK phosphorylation and activity were reduced when ezrin protein was suppressed. Ezrin-mediated early metastatic survival was partially dependent on activation of MAPK, but not AKT. To define the relevance of ezrin in the biology of metastasis, beyond the founding mouse model, we examined ezrin expression in dogs that naturally developed osteosarcoma. High ezrin expression in dog tumors was associated with early development of metastases. Consistent with this data, we found a significant association between high ezrin expression and poor outcome in pediatric osteosarcoma patients.