RESUMEN
Ewing sarcoma (EWS) is the second most common and aggressive type of metastatic bone tumor in adolescents and young adults. There is unmet medical need to develop and test novel pharmacological targets and novel therapies to treat EWS. Here, we found that EWS expresses high levels of a p53 isoform, delta133p53. We further determined that aberrant expression of delta133p53 induced HGF secretion resulting in tumor growth and metastasis. Thereafter, we evaluated targeting EWS tumors with HGF receptor neutralizing antibody (AMG102) in preclinical studies. Surprisingly, we found that targeting EWS tumors with HGF receptor neutralizing antibody (AMG102) in combination with GD2-specific, CAR-reengineered T-cell therapy synergistically inhibited primary tumor growth and establishment of metastatic disease in preclinical models. Furthermore, our data suggested that AMG102 treatment alone might increase leukocyte infiltration including efficient CAR-T access into tumor mass and thereby improves its antitumor activity. Together, our findings warrant the development of novel CAR-T-cell therapies that incorporate HGF receptor neutralizing antibody to improve therapeutic potency, not only in EWS but also in tumors with aberrant activation of the HGF/c-MET pathway.
Asunto(s)
Anticuerpos Monoclonales Humanizados/uso terapéutico , Antineoplásicos Inmunológicos/uso terapéutico , Neoplasias Óseas/tratamiento farmacológico , Receptores Quiméricos de Antígenos/inmunología , Sarcoma de Ewing/tratamiento farmacológico , Animales , Neoplasias Óseas/patología , Línea Celular Tumoral , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Factor de Crecimiento de Hepatocito/metabolismo , Humanos , Inmunoterapia Adoptiva/métodos , Ratones , Proteínas Proto-Oncogénicas c-met/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-met/inmunología , Sarcoma de Ewing/patología , Transducción de Señal/inmunología , Linfocitos T/inmunología , Microambiente Tumoral/inmunología , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
The mTOR complex-1 (mTORC1) coordinates cell growth and metabolism, acting as a restriction point under stress conditions such as low oxygen tension (hypoxia). Hypoxia suppresses mTORC1 signaling. However, the signals by which hypoxia suppresses mTORC1 are only partially understood, and a direct link between hypoxia-driven physiological stress and the regulation of mTORC1 signaling is unknown. Here we show that hypoxia results in ataxia telangiectasia mutated (ATM)-dependent phosphorylation of hypoxia-inducible factor 1-alpha (HIF-1α) on serine(696) and mediates downregulation of mTORC1 signaling. Deregulation of these pathways in pediatric solid tumor xenografts suggests a link between mTORC1 dysregulation and solid tumor development and points to an important role for hypoxic regulation of mTORC1 activity in tumor development.
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Proteínas de Ciclo Celular/metabolismo , Proteínas de Unión al ADN/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas/metabolismo , Transducción de Señal , Proteínas Supresoras de Tumor/metabolismo , Animales , Apoptosis , Proteínas de la Ataxia Telangiectasia Mutada , Hipoxia de la Célula , Daño del ADN , Activación Enzimática , Humanos , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Complejos Multiproteicos , Fosforilación , Fosfoserina/metabolismo , Serina-Treonina Quinasas TOR , Factores de Transcripción/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Under conditions of DNA damage, the mammalian target of rapamycin complex 1 (mTORC1) is inhibited, preventing cell cycle progression and conserving cellular energy by suppressing translation. We show that suppression of mTORC1 signaling to 4E-BP1 requires the coordinated activity of two tumor suppressors, p53 and p63. In contrast, suppression of S6K1 and ribosomal protein S6 phosphorylation by DNA damage is Akt-dependent. We find that loss of either p53, required for the induction of Sestrin 1/2, or p63, required for the induction of REDD1 and activation of the tuberous sclerosis complex, prevents the DNA damage-induced suppression of mTORC1 signaling. These data indicate that the negative regulation of cap-dependent translation by mTORC1 inhibition subsequent to DNA damage is abrogated in most human cancers.
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Daño del ADN , Complejos Multiproteicos/metabolismo , Neoplasias/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismo , Factores de Transcripción/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Proteínas de Ciclo Celular , Línea Celular Tumoral , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Ratones Noqueados , Complejos Multiproteicos/genética , Neoplasias/genética , Neoplasias/patología , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Fosforilación/genética , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Quinasas S6 Ribosómicas 70-kDa/genética , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , Serina-Treonina Quinasas TOR/genética , Factores de Transcripción/genética , Proteína p53 Supresora de Tumor/genética , Proteínas Supresoras de Tumor/genéticaRESUMEN
The p53 family comprises the tumor suppressor p53 and the structural homologs p63 and p73. How the three family members cooperate in tumor suppression remains unclear. Here, we report different but complementary functions of the individual members for regulating retinoblastoma protein (RB) function during myogenic differentiation. Whereas p53 transactivates the retinoblastoma gene, p63 and p73 induce the cyclin-dependent kinase inhibitor p57 to maintain RB in an active, hypophosphorylated state. DeltaNp73 inhibits these functions of the p53 family in differentiation control, prevents myogenic differentiation, and enables cooperating oncogenes to transform myoblasts to tumorigenicity. DeltaNp73 is frequently overexpressed in rhabdomyosarcoma and essential for tumor progression in vivo. These findings establish differentiation control as a key tumor suppressor activity of the p53 family.
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Diferenciación Celular/genética , Genes p53 , Músculo Esquelético/citología , Músculo Esquelético/fisiología , Rabdomiosarcoma/genética , Animales , Línea Celular , Regulación Neoplásica de la Expresión Génica , Humanos , Inmunohistoquímica , Ratones , Ratones Desnudos , Análisis de Secuencia por Matrices de Oligonucleótidos , Rabdomiosarcoma/patología , Proteína p53 Supresora de Tumor/análisis , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Antibodies targeting insulin-like growth factor 1 receptor (IGF-1R) induce objective responses in only 5% to 15% of children with sarcoma. Understanding the mechanisms of resistance may identify combination therapies that optimize efficacy of IGF-1R-targeted antibodies. Sensitivity to the IGF-1R-targeting antibody TZ-1 was determined in rhabdomyosarcoma and Ewing sarcoma cell lines. Acquired resistance to TZ-1 was developed and characterized in sensitive Rh41 cells. The BRD4 inhibitor, JQ1, was evaluated as an agent to prevent acquired TZ-1 resistance in Rh41 cells. The phosphorylation status of receptor tyrosine kinases (RTK) was assessed. Sensitivity to TZ-1 in vivo was determined in Rh41 parental and TZ-1-resistant xenografts. Of 20 sarcoma cell lines, only Rh41 was sensitive to TZ-1. Cells intrinsically resistant to TZ-1 expressed multiple (>10) activated RTKs or a relatively less complex set of activated RTKs (â¼5). TZ-1 decreased the phosphorylation of IGF-1R but had little effect on other phosphorylated RTKs in all resistant lines. TZ-1 rapidly induced activation of RTKs in Rh41 that was partially abrogated by knockdown of SOX18 and JQ1. Rh41/TZ-1 cells selected for acquired resistance to TZ-1 constitutively expressed multiple activated RTKs. TZ-1 treatment caused complete regressions in Rh41 xenografts and was significantly less effective against the Rh41/TZ-1 xenograft. Intrinsic resistance is a consequence of redundant signaling in pediatric sarcoma cell lines. Acquired resistance in Rh41 cells is associated with rapid induction of multiple RTKs, indicating a dynamic response to IGF-1R blockade and rapid development of resistance. The TZ-1 antibody had greater antitumor activity against Rh41 xenografts compared with other IGF-1R-targeted antibodies tested against this model.
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Proteínas Nucleares , Sarcoma , Niño , Humanos , Factores de Transcripción , Receptor IGF Tipo 1 , Sarcoma/tratamiento farmacológico , Receptores de Somatomedina , Anticuerpos Monoclonales/farmacología , Línea Celular Tumoral , Proteínas de Ciclo Celular , Factores de Transcripción SOXFRESUMEN
Despite recent advances in cancer therapeutics, glioblastoma (GBM) remains one of the most difficult cancers to treat in both the primary and recurrent settings. GBM presents a unique therapeutic challenge given the immune-privileged environment of the brain and the aggressive nature of the disease. Furthermore, it can change phenotypes throughout the course of disease-switching between mesenchymal, neural, and classic gene signatures, each with specific markers and mechanisms of resistance. Recent advancements in the field of immunotherapy-which utilizes strategies to reenergize or alter the immune system to target cancer-have shown striking results in patients with many types of malignancy. Immune checkpoint inhibitors, adoptive cellular therapy, cellular and peptide vaccines, and other technologies provide clinicians with a vast array of tools to design highly individualized treatment and potential for combination strategies. There are currently over 80 active clinical trials evaluating immunotherapies for GBM, often in combination with standard secondary treatment options including re-resection and anti-angiogenic agents, such as bevacizumab. This review will provide a clinically focused overview of the immune environment present in GBM, which is frequently immunosuppressive and characterized by M2 macrophages, T cell exhaustion, enhanced transforming growth factor-ß signaling, and others. We will also outline existing immunotherapeutic strategies, with a special focus on immune checkpoint inhibitors, chimeric antigen receptor therapy, and dendritic cell vaccines. Finally, we will summarize key discoveries in the field and discuss currently active clinical trials, including combination strategies, burgeoning technology like nucleic acid and nanoparticle therapy, and novel anticancer vaccines. This review aims to provide the most updated summary of the field of immunotherapy for GBM and offer both historical perspective and future directions to help inform clinical practice.
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Neoplasias Encefálicas , Glioblastoma , Médicos , Neoplasias Encefálicas/patología , Glioblastoma/patología , Humanos , Inhibidores de Puntos de Control Inmunológico , Factores Inmunológicos , Inmunoterapia/métodos , Linfocitos TRESUMEN
AIM: To evaluate the impact of polymorphisms in the cytochrome P450 (CYP) 2C9, 2C19 and 2C8 genes on the risk of mild hypoglycaemic attacks in patients treated with sulphonylureas. METHODS: One hundred and eight type 2 diabetic patients (50 men, 58 women), treated with oral antidiabetics, including at least one from the sulphonylurea group (glimepiride n = 50, gliclazide n = 46, or glipizide n = 12) for 3 months or longer, were included in the study. Symptoms of hypoglycaemia (sweating, tremor, anxiety and palpitations) during a 3 month period were recorded and confirmed by home glucose measurements. Gender, age, body mass index, creatinine clearance, HbA1c, oral antidiabetic dose and concomitant medication were assessed together with functional CYP2C9, CYP2C19 and CYP2C8 polymorphisms, analysed by real-time PCR methods. RESULTS: Fifteen patients (eight men, seven women) reported hypoglycaemia symptoms which were validated by their home glucose measurements (< 70 mg/dl). Heterozygosity and homozygosity for CYP2C9 variant alleles (*2 or *3) tended to be more frequent among patients who reported hypoglycaemic attacks (60 and 7%) than those who did not (39 and 3%). Similarly, the CYP2C8*1/*3 genotype tended to be more frequent in patients with (47%) than without (27%) hypoglycaemia, while no such trend was observed for CYP2C19 variants. However, only in the gliclazide group a significant association between CYP2C9 genotype and hypoglycaemic attacks was observed (P = 0.035). None of the other covariates showed any significant association with the risk of hypoglycaemic attacks. CONCLUSIONS: CYP2C9 polymorphisms leading to decreased enzyme activity show a modest impact on the risk of mild hypoglycaemia attacks during oral antidiabetic treatment, with a significant association in patients treated with gliclazide.
Asunto(s)
Hidrocarburo de Aril Hidroxilasas/genética , Hipoglucemia/inducido químicamente , Hipoglucemiantes/efectos adversos , Compuestos de Sulfonilurea/efectos adversos , Adulto , Anciano , Anciano de 80 o más Años , Citocromo P-450 CYP2C19 , Citocromo P-450 CYP2C8 , Citocromo P-450 CYP2C9 , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Polimorfismo GenéticoRESUMEN
BACKGROUND AND STUDY AIMS: The clinical significance of serum parameters of iron metabolism and hepcidin in liver disease remains unknown. Therefore, this study aimed to evaluate the association of serum hepcidin levels with fibrosis stage and serum iron parameters in patients with chronic hepatitis B (CHB). PATIENTS AND METHODS: This cross-sectional study included 126 treatment-naïve patients with CHB (median age, 39.0 years; 64.3% males) who were positive for hepatitis B surface antigen and 23 healthy controls (median age, 33.0 years; 52.2% males). Data on patient demographics, serum hepcidin levels, liver function tests and serum iron parameters and liver biopsy findings including fibrosis grade, histological activity index (HAI) and liver iron level were recorded. RESULTS: The median (minimum-maximum) serum hepcidin levels were significantly lower in the CHB group than in the control group [71.2 (13.3-672.7) vs. 657.5 (201.7-2714.2) pg/mL, p < 0.001]. Higher fibrosis stage was associated with higher transferrin saturation (p = 0.029), serum ferritin level (p < 0.001) and viral load (p < 0.001). Fibrosis stage and HAI were positively correlated with ferritin (r = 0.407, p < 0.001 and r = 0.415, p < 0.001, respectively) and transferrin saturation (r = 0.219, p = 0.026 and r = 0.290, p = 0.003, respectively) levels, whereas hepcidin level was negatively correlated with fibrosis stage (r = -0.175, p = 0.051), viral load (r = -0.209, p = 0.020) and ferritin level (r = -0.244, p = 0.006) level. There were no significant differences in serum iron level, total iron binding capacity and liver iron level among patients with different stages of fibrosis. CONCLUSION: Reduced hepcidin levels and elevated transferrin saturation and ferritin levels are linked to fibrosis severity and HAI in patients with CHB.
Asunto(s)
Hepatitis B Crónica , Hepcidinas/sangre , Cirrosis Hepática , Adulto , Biomarcadores/sangre , Correlación de Datos , Estudios Transversales , Femenino , Ferritinas/sangre , Hepatitis B Crónica/sangre , Hepatitis B Crónica/complicaciones , Hepatitis B Crónica/diagnóstico , Hepatitis B Crónica/epidemiología , Humanos , Hierro/metabolismo , Hígado/metabolismo , Hígado/patología , Cirrosis Hepática/diagnóstico , Cirrosis Hepática/etiología , Cirrosis Hepática/metabolismo , Pruebas de Función Hepática/métodos , Masculino , Persona de Mediana Edad , Gravedad del Paciente , Transferrina/metabolismo , Turquía/epidemiologíaRESUMEN
The pre-metastatic niche (PMN) represents an abnormal microenvironment devoid of cancer cells, but favoring tumor growth. Little is known about the mechanisms that generate the PMN or their effects on host cells within metastasis-prone organs. Here, we investigated by using spontaneous metastatic models whether lung epithelial cells are essential for primary tumor induced neutrophil recruitment in lung and subsequently initiating PMN formation in osteosarcoma. We found that serum levels of ANGPTL2 in osteosarcoma patients are significantly higher compared to those in healthy controls and that ANGPTL2 secretion by tumor cells plays an essential role in osteosarcoma metastasis. We determined that tumor-derived ANGPTL2 stimulates lung epithelial cells, which is essential for primary tumor-induced neutrophil recruitment in lung and subsequent pre-metastatic niche formation. Lastly, we identified that a p63 isoform, ΔNp63, drives high level of ANGPTL2 secretion and pharmaceutical inhibition of ANGPTL2 signaling by a non-RGD-based integrin binding peptide (ATN-161) diminished metastatic load in lungs likely due to reduction of the lung pre-metastatic niche formation.
RESUMEN
BACKGROUND: Glioblastoma (GBM) remains one of the least successfully treated cancers. It is essential to understand the basic biology of this lethal disease and investigate novel pharmacological targets to treat GBM. The aims of this study were to determine the biological consequences of elevated expression of ΔNp73, an N-terminal truncated isoform of TP73, and to evaluate targeting of its downstream mediators, the angiopoietin 1 (ANGPT1)/tunica interna endothelial cell kinase 2 (Tie2) axis, by using a highly potent, orally available small-molecule inhibitor (rebastinib) in GBM. METHODS: ΔNp73 expression was assessed in glioma sphere cultures, xenograft glioblastoma tumors, and glioblastoma patients by western blot, quantitative reverse transcription PCR, and immunohistochemistry. Immunoprecipitation, chromatin immunoprecipitation (ChiP) and sequential ChIP were performed to determine the interaction between ΔNp73 and E26 transformation-specific (ETS) proto-oncogene 2 (ETS2) proteins. The oncogenic consequences of ΔNp73 expression in glioblastomas were examined by in vitro and in vivo experiments, including orthotopic zebrafish and mouse intracranial-injection models. Effects of rebastinib on growth of established tumors and survival were examined in an intracranial-injection mouse model. RESULTS: ΔNp73 upregulates both ANGPT1 and Tie2 transcriptionally through ETS conserved binding sites on the promoters by interacting with ETS2. Elevated expression of ΔNp73 promotes tumor progression by mediating angiogenesis and survival. Therapeutic targeting of downstream ΔNp73 signaling pathways by rebastinib inhibits growth of established tumors and extends survival in preclinical models of glioblastoma. CONCLUSION: Aberrant expression of ΔNp73 in GBM promotes tumor progression through autocrine and paracrine signaling dependent on Tie2 activation by ANGPT1. Disruption of this signaling by rebastinib improves tumor response to treatment in glioblastoma.
Asunto(s)
Antineoplásicos/administración & dosificación , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Glioblastoma/metabolismo , Glioblastoma/patología , Proteína Proto-Oncogénica c-ets-2/metabolismo , Pirazoles/administración & dosificación , Piridinas/administración & dosificación , Quinolinas/administración & dosificación , Proteína Tumoral p73/metabolismo , Animales , Neoplasias Encefálicas/tratamiento farmacológico , Línea Celular Tumoral/efectos de los fármacos , Modelos Animales de Enfermedad , Glioblastoma/tratamiento farmacológico , Humanos , Ratones Transgénicos , Neovascularización Patológica/metabolismo , Proto-Oncogenes Mas , Análisis de Supervivencia , Pez CebraRESUMEN
Osteosarcoma (OS), a malignant tumor of bone, kills through aggressive metastatic spread almost exclusively to the lung. Mechanisms driving this tropism for lung tissue remain unknown, though likely invoke specific interactions between tumor cells and other cells within the lung metastatic niche. Aberrant overexpression of ΔNp63 in OS cells directly drives production of IL-6 and CXCL8. All these factors were expressed at higher levels in OS lung metastases than in matched primary tumors from the same patients. Expression in cell lines correlated strongly with lung colonization efficiency in murine xenograft models. Lentivirus-mediated expression endowed poorly metastatic OS cells with increased metastatic capacity. Disruption of IL-6 and CXCL8 signaling using genetic or pharmaceutical inhibitors had minimal effects on tumor cell proliferation in vitro or in vivo, but combination treatment inhibited metastasis across multiple models of metastatic OS. Strong interactions occurred between OS cells and both primary bronchial epithelial cells and bronchial smooth muscle cells that drove feed-forward amplification of IL-6 and CXCL8 production. These results identify IL-6 and CXCL8 as primary mediators of OS lung tropism and suggest pleiotropic, redundant mechanisms by which they might effect metastasis. Combination therapy studies demonstrate proof of concept for targeting these tumor-lung interactions to affect metastatic disease.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Neoplasias Óseas/patología , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Neoplasias Pulmonares/patología , Adolescente , Adulto , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias Óseas/tratamiento farmacológico , Huesos/patología , Línea Celular Tumoral , Proliferación Celular , Niño , Receptor gp130 de Citocinas/antagonistas & inhibidores , Receptor gp130 de Citocinas/metabolismo , Evaluación Preclínica de Medicamentos , Estudios de Seguimiento , Humanos , Hidrazinas/farmacología , Hidrazinas/uso terapéutico , Pulmón/patología , Neoplasias Pulmonares/prevención & control , Neoplasias Pulmonares/secundario , Masculino , Ratones , Osteosarcoma/tratamiento farmacológico , Osteosarcoma/prevención & control , Osteosarcoma/secundario , Cultivo Primario de Células , Quinoxalinas/farmacología , Quinoxalinas/uso terapéutico , Receptores de Interleucina-8A/antagonistas & inhibidores , Sulfonamidas/farmacología , Sulfonamidas/uso terapéutico , Ensayos Antitumor por Modelo de Xenoinjerto , Adulto JovenRESUMEN
Endemic water borne fluorosis is a public health problem in Isparta, a city located in southern Turkey. In order to investigate the association between osteosclerosis and fluorosis, we retrospectively screened the results of lumbar spine and femur neck bone mineral density (BMD) of 1500 patients who were examined before, for any reason in between 2001-2003. Sixty nine patients (67 females and 2 males, mean age 52.6+/-10.2) with vertebra T-scores>or=+2 were found only except a patient with osteoid osteoma in the femur neck (femur T-score+6.64). Thirty-four of the patients could be reexamined with lateral vertebra BMD and investigated for fluorosis and the other etiologic causes of osteosclerosis. Of 34 patients, 14 had either mottled tooth enamel or urine fluoride level greater than 1.5 mg/l. Other etiologic causes were hypothyroidism (2), hypoparathyroidism (1), history of lumbar fracture (1), use of retinoids (1), vitamin D (7), oral calcium preparations (9), and bisphosphanates (3). Lateral lumbar vertebral T-score was greater than+2 in 12 patients (35.3%). Femur T-score was greater than+2 in 7 patients (20.6%). Fourteen patients (41.2%) had lateral vertebral or femur T-score>or=+2. Five (35.7%) of these patients had signs of fluorosis, as discussed before. Mean body mass index of individuals with fluorosis was 36.4+/-7.9 and this result was significantly higher than other osteosclerotic subjects (31.6+/-4.4). In conclusion we believe that approximately one third of the osteosclerosis in our region was due to endemic skeletal fluorosis and obesity may enhance this osteosclerotic type bone changes in endemic fluorosis.
Asunto(s)
Fluoruros/toxicidad , Osteosclerosis/etiología , Adulto , Anciano , Femenino , Cuello Femoral/diagnóstico por imagen , Fluoruros/orina , Fluorosis Dental/etiología , Humanos , Vértebras Lumbares/diagnóstico por imagen , Masculino , Persona de Mediana Edad , Osteosclerosis/diagnóstico por imagen , Osteosclerosis/epidemiología , Osteosclerosis/orina , Radiografía , Turquía/epidemiologíaRESUMEN
BACKGROUND: STAT3 is a transcription factor involved in cytokine and receptor kinase signal transduction that is aberrantly activated in a variety of sarcomas, promoting metastasis and chemotherapy resistance. The purpose of this work was to develop and test a novel putative STAT3 inhibitor, LY5. METHODS AND FINDINGS: An in silico fragment-based drug design strategy was used to create LY5, a small molecule inhibitor that blocks the STAT3 SH2 domain phosphotyrosine binding site, inhibiting homodimerization. LY5 was evaluated in vitro demonstrating good biologic activity against rhabdomyosarcoma, osteosarcoma and Ewing's sarcoma cell lines at high nanomolar/low micromolar concentrations, as well as specific inhibition of STAT3 phosphorylation without effects on other STAT3 family members. LY5 exhibited excellent oral bioavailability in both mice and healthy dogs, and drug absorption was enhanced in the fasted state with tolerable dosing in mice at 40 mg/kg BID. However, RNAi-mediated knockdown of STAT3 did not phenocopy the biologic effects of LY5 in sarcoma cell lines. Moreover, concentrations needed to inhibit ex vivo metastasis growth using the PuMA assay were significantly higher than those needed to inhibit STAT3 phosphorylation in vitro. Lastly, LY5 treatment did not inhibit the growth of sarcoma xenografts or prevent pulmonary metastasis in mice. CONCLUSIONS: LY5 is a novel small molecule inhibitor that effectively inhibits STAT3 phosphorylation and cell proliferation at nanomolar concentrations. LY5 demonstrates good oral bioavailability in mice and dogs. However LY5 did not decrease tumor growth in xenograft mouse models and STAT3 knockdown did not induce concordant biologic effects. These data suggest that the anti-cancer effects of LY5 identified in vitro were not mediated through STAT3 inhibition.
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Aminopiridinas/farmacocinética , Aminopiridinas/uso terapéutico , Osteosarcoma/tratamiento farmacológico , Rabdomiosarcoma/tratamiento farmacológico , Factor de Transcripción STAT3/antagonistas & inhibidores , Sarcoma de Ewing/tratamiento farmacológico , Sulfonamidas/farmacocinética , Sulfonamidas/uso terapéutico , Aminopiridinas/farmacología , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Citocinas/farmacología , Perros , Femenino , Humanos , Neoplasias Pulmonares/secundario , Ratones , Osteosarcoma/patología , Fosforilación/efectos de los fármacos , Rabdomiosarcoma/patología , Factor de Transcripción STAT3/metabolismo , Sarcoma de Ewing/patología , Sulfonamidas/farmacología , Resultado del Tratamiento , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND/AIMS: This study is designed to determine which drug forms provide ideal pharyngeal anesthesia when used during upper gastrointestinal system endoscopy. MATERIALS AND METHODS: A total of 180 patients were included in the study. Using the random number table, these patients were divided into three groups. Group 1, lidocaine gel+isotonic spray; Group 2, base lubricant gel+lidocaine spray; and Group 3: lidocaine gel+lidocaine spray. Data were collected from the patient identification form, compliance to operation form, and State Anxiety Inventory. RESULTS: Anesthetization and compliance to procedure scores were higher and anxiety scores were lower in Group 3 than in other groups (p<0.05). It was observed that as the compliance score increased, the anesthetization and satisfaction scores also increased; however, coughing during the procedure, duration of the procedure, and anxiety scores decreased (p<0.05). It was determined that as anesthetization scores increased, discomfort in the throat caused by the device, coughing during the procedure, and anxiety scores decreased (p<0.05). CONCLUSION: Lidocaine gel and spray combination is the most ideal pharyngeal anesthesia to ensure the adaptation of the patient to the procedure and to decrease anxiety and discomfort during the procedure.
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Anestésicos Combinados/administración & dosificación , Anestésicos Locales/administración & dosificación , Endoscopía Gastrointestinal/métodos , Soluciones Isotónicas/administración & dosificación , Lidocaína/administración & dosificación , Adulto , Ansiedad/epidemiología , Ansiedad/etiología , Tos/epidemiología , Tos/etiología , Método Doble Ciego , Endoscopía Gastrointestinal/psicología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Tempo Operativo , Satisfacción del Paciente , Faringe/cirugíaRESUMEN
p63 is a structural homolog within the 53 family encoding two isoforms, ΔNp63 and TAp63. The oncogenic activity of ΔNp63 has been demonstrated in multiple cancers, however the underlying mechanisms that contribute to tumorigenesis are poorly characterized. Osteosarcoma (OSA) is the most common primary bone tumor in dogs, exhibiting clinical behavior and molecular biology essentially identical to its human counterpart. The purpose of this study was to evaluate the potential contribution of ΔNp63 to the biology of canine OSA. As demonstrated by qRT-PCR, nearly all canine OSA cell lines and tissues overexpressed ΔNp63 relative to normal control osteoblasts. Inhibition of ΔNp63 by RNAi selectively induced apoptosis in the OSA cell lines overexpressing ΔNp63. Knockdown of ΔNp63 upregulated expression of the proapoptotic Bcl-2 family members Puma and Noxa independent of p53. However the effects of ΔNp63 required transactivating isoforms of p73, suggesting that ΔNp63 promotes survival in OSA by repressing p73-dependent apoptosis. In addition, ΔNp63 modulated angiogenesis and invasion through its effects on VEGF-A and IL-8 expression, and STAT3 phosphorylation. Lastly, the capacity of canine OSA cell lines to form pulmonary metastasis was directly related to expression levels of ΔNp63 in a murine model of metastatic OSA. Together, these data demonstrate that ΔNp63 inhibits apoptosis and promotes metastasis, supporting continued evaluation of this oncogene as a therapeutic target in both human and canine OSA.
Asunto(s)
Neoplasias Óseas/patología , Neoplasias Pulmonares/patología , Osteosarcoma/patología , Factor de Transcripción STAT3/metabolismo , Sarcoma Experimental/patología , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Animales , Apoptosis , Neoplasias Óseas/veterinaria , Línea Celular Tumoral , Supervivencia Celular , Perros , Femenino , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Interleucina-8/metabolismo , Neoplasias Pulmonares/secundario , Ratones , Ratones SCID , Invasividad Neoplásica/patología , Neovascularización Patológica/patología , Osteoblastos , Osteosarcoma/veterinaria , Fosforilación , Isoformas de Proteínas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Sarcoma Experimental/secundario , Factores de Transcripción/genética , Proteína Tumoral p73/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Proteínas Supresoras de Tumor/genética , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Efficient expression of genes transferred by retroviral vectors is a prerequisite for gene therapy, especially when the biological effect depends on the amount of transgene product. High-level gene expression is desirable for several gene therapy approaches involving T lymphocytes. We evaluated standard retroviral vectors with cis-regulatory control elements of the Moloney murine leukemia virus (Mo-MLV) with or without the human T cell-specific CD2 enhancer. For comparison, vectors containing the long terminal repeat (LTR) of myeloproliferative sarcoma virus (MPSV) and an improved 5' untranslated region were used (MP71 vectors), with or without the woodchuck hepatitis virus posttranscriptional regulatory element (PRE). All vectors expressed the enhanced green fluorescent protein (GFP) to measure transgene expression. In mouse T cells MP71 vectors with and without the PRE yielded an up to 10-fold higher expression level compared with the Mo-MLV-based vectors currently used for gene transfer into T lymphocytes. A high multiplicity of infection (MOI) of standard Mo-MLV vectors could not reach expression levels obtained with a low MOI of MP71 vector. Ex vivo-transduced mouse T lymphocytes maintained the vector-dependent differences in level of transgene expression in Rag-1-deficient mice when adoptively transferred. In four human T cell lines and human primary T lymphocytes MP71 vectors yielded an up to 75-fold higher GFP expression level in comparison with the standard Mo-MLV vector. In contrast to mouse T cells, the integration of the PRE into MP71 vectors induced in human T cells a further significant increase in transgene expression level. Southern blot analysis of CEM T cells revealed that the superior performance of MP71 vectors was not due to a higher rate of viral integration. In summary, MP71 vectors are useful tools for stable, high-level gene expression in T lymphocytes, for example, in the expression of T cell receptor genes.
Asunto(s)
Vectores Genéticos , Retroviridae/genética , Linfocitos T/inmunología , Transgenes , Células 3T3 , Animales , Antígenos CD2/genética , Línea Celular , Supervivencia Celular , Células Cultivadas , Elementos de Facilitación Genéticos/genética , Expresión Génica , Terapia Genética , Proteínas Fluorescentes Verdes , Humanos , Proteínas Luminiscentes/genética , Ratones , Virus de la Leucemia Murina de Moloney/genética , Secuencias Reguladoras de Ácidos Nucleicos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Transducción GenéticaRESUMEN
The tumor suppressor gene p53 and its family members p63/p73 are critical determinants of tumorigenesis. ΔNp63 is a splice variant of p63, which lacks the N-terminal transactivation domain. It is thought to antagonize p53-, p63-, and p73-dependent translation, thus blocking their tumor suppressor activity. In our studies of the pediatric solid tumors neuroblastoma and osteosarcoma, we find overexpression of ΔNp63; however, there is no correlation of ΔNp63 expression with p53 mutation status. Our data suggest that ΔNp63 itself endows cells with a gain-of-function that leads to malignant transformation, a function independent of any p53 antagonism. Here, we demonstrate that ΔNp63 overexpression, independent of p53, increases secretion of interleukin (IL)-6 and IL-8, leading to elevated phosphorylation of STAT3 (Tyr-705). We show that elevated phosphorylation of STAT3 leads to stabilization of hypoxia-inducible factor 1α (HIF-1α) protein, resulting in VEGF secretion. We also show human clinical data, which suggest a mechanistic role for ΔNp63 in osteosarcoma metastasis. In summary, our studies reveal the mechanism by which ΔNp63, as a master transcription factor, modulates tumor angiogenesis.
Asunto(s)
Neoplasias Óseas/irrigación sanguínea , Proteínas de la Membrana/metabolismo , Neuroblastoma/irrigación sanguínea , Osteosarcoma/irrigación sanguínea , Adolescente , Animales , Neoplasias Óseas/genética , Neoplasias Óseas/metabolismo , Línea Celular Tumoral , Niño , Modelos Animales de Enfermedad , Femenino , Xenoinjertos , Humanos , Proteínas de la Membrana/genética , Ratones , Ratones SCID , Neovascularización Patológica/genética , Neovascularización Patológica/metabolismo , Neuroblastoma/genética , Neuroblastoma/metabolismo , Osteosarcoma/genética , Osteosarcoma/metabolismo , Factores de Transcripción , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
The Interferon (IFN) which is the standard treatment for Hepatitis C, may cause a lot of side effects including dermatological anomalies. This paper presents a psoriasis case which occurred in relation with the treatment of acute hepatitis C (AHC) with peginterferon alfa (peg-IFN-α). A 60-year-old male patient came to the hospital with symptoms of high liver enzymes. The patient with history of a recent operation showed anti-HCV(+), HCVRNA 3.5 million IU/mL and HCV genotype 1b in the tests. Without any other etiological factors found in the patient, we started a treatment of peg-IFNα-2b with the diagnosis of AHC. After 3 weeks, psoriatic plaques were observed in various parts of the body. Antiviral treatment of the patient was concluded within 6 months. His psoriasis treatment initially commenced with local agents followed by phototherapy. Permanent viral response was seen in the patient and his lesions recovered rapidly after the antipsoriatic and antiviral treatment. Psoriasis and other autoimmune diseases should be considered even though they are encountered rarely,and the patients should be informed of the possible risks before planning treatment with peg-IFN-α.
Asunto(s)
Erupciones por Medicamentos/diagnóstico , Erupciones por Medicamentos/etiología , Hepatitis C/tratamiento farmacológico , Interferón-alfa/efectos adversos , Interferón-alfa/uso terapéutico , Polietilenglicoles/efectos adversos , Polietilenglicoles/uso terapéutico , Psoriasis/inducido químicamente , Psoriasis/diagnóstico , Diagnóstico Diferencial , Erupciones por Medicamentos/prevención & control , Hepatitis C/complicaciones , Humanos , Masculino , Persona de Mediana Edad , Psoriasis/prevención & control , Proteínas Recombinantes/efectos adversos , Proteínas Recombinantes/uso terapéutico , Resultado del TratamientoRESUMEN
Eisenmenger syndrome is associated with irreversible increase in pulmonary vascular resistance causing reduced survival. Bosentan, a non-selective endothelin receptor antagonist is the commonly used specific pulmonary arterial hypertension drug in Eisenmenger syndrome. In this paper, we present a case of recurrent gastrointestinal bleeding in a 23-year-old female with Eisenmenger syndrome who was only under bosentan treatment, which has not been reported previously. Most common adverse effect of bosentan is elevation in the liver enzymes however, bleeding complication is very rare. On the contrary, it was proposed that bosentan might be a potential protector against hyperacidity and mucosal erosion that occur as a consequence of stress. Although the mechanistic relationship of bleeding tendency and role of Eisenmenger syndrome concomitant with bosentan treatment is far from conclusive statement for now, this association warrants and should draw attention of clinicians and researches in this field.
Asunto(s)
Antihipertensivos/efectos adversos , Complejo de Eisenmenger/complicaciones , Hemorragia Gastrointestinal/etiología , Sulfonamidas/efectos adversos , Antihipertensivos/farmacología , Bosentán , Complejo de Eisenmenger/tratamiento farmacológico , Hipertensión Pulmonar Primaria Familiar , Femenino , Humanos , Hipertensión Pulmonar/tratamiento farmacológico , Hipertensión Pulmonar/etiología , Recurrencia , Sulfonamidas/farmacología , Sulfonamidas/uso terapéutico , Adulto JovenRESUMEN
Deregulation of the mTOR pathway is closely associated with tumorigenesis. Accordingly, mTOR inhibitors such as rapamycin and mTOR-selective kinase inhibitors have been tested as cancer therapeutic agents. Inhibition of mTOR results in sensitization to DNA-damaging agents; however, the molecular mechanism is not well understood. We found that an mTOR-selective kinase inhibitor, AZD8055, significantly enhanced sensitivity of a pediatric rhabdomyosarcoma xenograft to radiotherapy and sensitized rhabdomyosarcoma cells to the DNA interstrand cross-linker (ICL) melphalan. Sensitization correlated with drug-induced downregulation of a key component of the Fanconi anemia pathway, FANCD2 through mTOR regulation of FANCD2 gene transcripts via mTORC1-S6K1. Importantly, we show that FANCD2 is required for the proper activation of ATM-Chk2 checkpoint in response to ICL and that mTOR signaling promotes ICL-induced ATM-Chk2 checkpoint activation by sustaining FANCD2. In FANCD2-deficient lymphoblasts, FANCD2 is essential to suppress endogenous and induced DNA damage, and FANCD2-deficient cells showed impaired ATM-Chk2 and ATR-Chk1 activation, which was rescued by reintroduction of wild-type FANCD2. Pharmacologic inhibition of PI3K-mTOR-AKT pathway in Rh30 rhabdomyosarcoma cells attenuated ICL-induced activation of ATM, accompanied with the decrease of FANCD2. These data suggest that the mTOR pathway may promote the repair of DNA double-strand breaks by sustaining FANCD2 and provide a novel mechanism of how the Fanconi anemia pathway modulates DNA damage response and repair.