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A hexanuclear heterometallic cluster of composition [Dy2 Co4 (L)4 (NO3 )2 (OH)4 (C2 H5 OH)2 ] â 2â C2 H5 OH (1) was synthesized by employing a Schiff base 2-(((2-hydroxy-3-methoxybenzyl) imino)methyl)-4-methoxyphenol (H2 L) as ligand and utilizing Dy(NO3 )3 â 6H2 O and Co(NO3 )2 â 6H2 O as metal ion sources. X-ray single-crystal diffraction analysis indicated that complex 1 contains a defect tetracubane core and possesses central symmetric structure, with two DyIII ions being in the central body position of the molecule and four CoII ions being arranged at the outer sites. Magnetic studies reveal that complex 1 behaves as single-molecule magnet (SMM) with energy barrier of 27.50â K. To investigate the individual contribution of DyIII and CoII ions to the SMM behavior, another two complexes of formulae [Dy2 Zn4 (L)4 (NO3 )2 (OH)4 ] â 4CH3 OH (2) and [Y2 Co4 (L)4 (NO3 )2 (OH)4 (C2 H5 OH)2 ] â 2â C2 H5 OH (3) were prepared. Complexes 1 and 3 are isomorphous. The coordination geometries of DyIII ions in 1 and 2 are different. The DyIII ions are eight-coordinated in 2 and nine-coordinated in 1. Complex 2 exhibits SMM behavior with energy barrier of 69.67â K, but complex 3 does not display SMM property. These results reveal that the SMM behaviors of 1 and 2 are mainly originated from DyIII ions. It might be the higher symmetry of DyIII ions in 2 that results in the higher energy barrier.
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PURPOSE: SHP2 inhibitors offer an appealing and novel approach to inhibit receptor tyrosine kinase (RTK) signaling, which is the oncogenic driver in many tumors or is frequently feedback activated in response to targeted therapies including RTK inhibitors and MAPK inhibitors. We seek to evaluate the efficacy and synergistic mechanisms of combinations with a novel SHP2 inhibitor, TNO155, to inform their clinical development. EXPERIMENTAL DESIGN: The combinations of TNO155 with EGFR inhibitors (EGFRi), BRAFi, KRASG12Ci, CDK4/6i, and anti-programmed cell death-1 (PD-1) antibody were tested in appropriate cancer models in vitro and in vivo, and their effects on downstream signaling were examined. RESULTS: In EGFR-mutant lung cancer models, combination benefit of TNO155 and the EGFRi nazartinib was observed, coincident with sustained ERK inhibition. In BRAFV600E colorectal cancer models, TNO155 synergized with BRAF plus MEK inhibitors by blocking ERK feedback activation by different RTKs. In KRASG12C cancer cells, TNO155 effectively blocked the feedback activation of wild-type KRAS or other RAS isoforms induced by KRASG12Ci and greatly enhanced efficacy. In addition, TNO155 and the CDK4/6 inhibitor ribociclib showed combination benefit in a large panel of lung and colorectal cancer patient-derived xenografts, including those with KRAS mutations. Finally, TNO155 effectively inhibited RAS activation by colony-stimulating factor 1 receptor, which is critical for the maturation of immunosuppressive tumor-associated macrophages, and showed combination activity with anti-PD-1 antibody. CONCLUSIONS: Our findings suggest TNO155 is an effective agent for blocking both tumor-promoting and immune-suppressive RTK signaling in RTK- and MAPK-driven cancers and their tumor microenvironment. Our data provide the rationale for evaluating these combinations clinically.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Inhibidores de Puntos de Control Inmunológico/farmacología , Neoplasias/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Proteína Tirosina Fosfatasa no Receptora Tipo 11/antagonistas & inhibidores , Regulación Alostérica/efectos de los fármacos , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Línea Celular Tumoral , Quinasa 4 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 6 Dependiente de la Ciclina/antagonistas & inhibidores , Sinergismo Farmacológico , Receptores ErbB/antagonistas & inhibidores , Femenino , Humanos , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Ratones , Mutación , Neoplasias/genética , Neoplasias/inmunología , Neoplasias/patología , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas p21(ras)/antagonistas & inhibidores , Proteínas Proto-Oncogénicas p21(ras)/genética , Macrófagos Asociados a Tumores/efectos de los fármacos , Macrófagos Asociados a Tumores/inmunología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The reaction of Dy(NO3)3·6H2O and an asymmetric Schiff-base linker 5-chloro-2-(((2-hydroxy-3-methoxybenzyl)imino)methyl) phenol (H2L) afforded a dinuclear compound [Dy2L2(HL)(NO3)(EtOH)]·0.5C2H5OH (1). Complex 1 features two inequivalent Dy(iii) sites, where three ligand sets (one HL- moiety and two L2- groups) are shared by two Dy(iii) ions. The strategic introduction of CH3COOH in the reaction system used for synthesizing 1 induces the replacement of the HL- ligand by the CH3COO- ion, consequently resulting in the generation of [Et3NH][Dy2L2(NO3)(CH3COO)2] (2). In complex 2, two Dy(iii) centers adopt NO7 (D2d geometry) and NO6 (C2v) coordination sphere, respectively. DC magnetic susceptibility studies for the two complexes indicate ferromagnetic interactions. Complexes 1 and 2 exhibit single-molecule magnet behavior with two-step slow relaxation processes due to the possession of inequivalent metal sites. The energy barriers of 69.19 and 45.73 K for 1, and 92.77 and 72.95 K for 2 are determined. Theoretical calculations reveal that the two-step relaxation processes in both 1 and 2 mainly originate from the single-ion magnetism of two distinct Dy(iii) ions with inequivalent coordination environments.
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Advanced ovarian cancers are a leading cause of cancer-related death in women and are currently treated with surgery and chemotherapy. This standard of care is often temporarily successful but exhibits a high rate of relapse, after which, treatment options are few. Here we investigate whether biomarker-guided use of multiple targeted therapies, including small molecules and antibody-drug conjugates, is a viable alternative. A panel of patient-derived ovarian cancer xenografts (PDX), similar in genetics and chemotherapy responsiveness to human tumors, was exposed to 21 monotherapies and combination therapies. Three monotherapies and one combination were found to be active in different subsets of PDX. Analysis of gene expression data identified biomarkers associated with responsiveness to each of the three targeted therapies, none of which directly inhibits an oncogenic driver. While no single treatment had as high a response rate as chemotherapy, nearly 90% of PDXs were eligible for and responded to at least one biomarker-guided treatment, including tumors resistant to standard chemotherapy. The distribution of biomarker positivity in The Cancer Genome Atlas data suggests the potential for a similar precision approach in human patients. SIGNIFICANCE: This study exploits a panel of patient-derived xenografts to demonstrate that most ovarian tumors can be matched to effective biomarker-guided treatments.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Biomarcadores de Tumor/genética , Neoplasias Ováricas/tratamiento farmacológico , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Antineoplásicos/farmacología , Carcinoma Epitelial de Ovario/tratamiento farmacológico , Carcinoma Epitelial de Ovario/genética , Carcinoma Epitelial de Ovario/mortalidad , Carcinoma Epitelial de Ovario/patología , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Estimación de Kaplan-Meier , Terapia Molecular Dirigida/métodos , Neoplasias Ováricas/genética , Neoplasias Ováricas/mortalidad , Neoplasias Ováricas/patología , Medicina de Precisión , Prueba de Estudio ConceptualRESUMEN
OBJECTIVE: TGF-beta plays a significant role in vascular injury-induced stenosis. This study evaluates the efficacy of a novel, small molecule inhibitor of ALK5/ALK4 kinase, in the rat carotid injury model of vascular fibrosis. METHODS AND RESULTS: The small molecule, SM16, was shown to bind with high affinity to ALK5 kinase ATP binding site using a competitive binding assay and biacore analysis. SM16 blocked TGF-beta and activin-induced Smad2/3 phosphorylation and TGF-beta-induced plasminogen activator inhibitor (PAI)-luciferase activity in cells. Good overall selectivity was demonstrated in a large panel of kinase assays, but SM16 also showed nanomolar inhibition of ALK4 and weak (micromolar) inhibition of Raf and p38. In the rat carotid injury model, SM16 dosed once daily orally at 15 or 30 mg/kg SM16 for 14 days caused significant inhibition of neointimal thickening and lumenal narrowing. SM16 also prevented induction of adventitial smooth muscle alpha-actin-positive myofibroblasts and the production of intimal collagen, but did not decrease the percentage of proliferative cells. CONCLUSIONS: These results are the first to demonstrate the efficacy of an orally active, small-molecule ALK5/ALK4 inhibitor in a vascular fibrosis model and suggest the potential therapeutic application of these inhibitors in vascular fibrosis.
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Compuestos de Azabiciclo/farmacología , Traumatismos de las Arterias Carótidas/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Receptores de Factores de Crecimiento Transformadores beta/antagonistas & inhibidores , Receptores de Activinas Tipo I/antagonistas & inhibidores , Adenosina Trifosfato/metabolismo , Administración Oral , Animales , Compuestos de Azabiciclo/administración & dosificación , Compuestos de Azabiciclo/metabolismo , Sitios de Unión , Traumatismos de las Arterias Carótidas/patología , Traumatismos de las Arterias Carótidas/fisiopatología , Línea Celular , Fibroblastos/efectos de los fármacos , Fibroblastos/patología , Fibrosis , Humanos , Masculino , Mioblastos del Músculo Liso/efectos de los fármacos , Mioblastos del Músculo Liso/patología , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/metabolismo , Ratas , Ratas Sprague-Dawley , Receptor Tipo I de Factor de Crecimiento Transformador beta , Factor de Crecimiento Transformador beta/fisiologíaRESUMEN
BACKGROUND: Transforming growth factor beta (TGF-beta) plays a complex role in breast carcinogenesis. Initially functioning as a tumor suppressor, this cytokine later contributes to the progression of malignant cells by enhancing their invasive and metastatic potential as well as suppressing antitumor immunity. The purpose of this study was to investigate the efficacy of SM16, a novel small molecule ALK5 kinase inhibitor, to treat a highly metastatic, TGF-beta-producing murine mammary carcinoma (4T1). MATERIALS AND METHODS: Mice bearing established 4T1 tumors were treated with SM16 intraperitoneally (i.p.) or orally, and primary and metastatic tumor growth was assessed. RESULTS: SM16 inhibited Smad2 phosphorylation in cultured 4T1 tumor cells as well as primary and metastatic 4T1 tumor tissue. Blockade of TGF-beta signal transduction in 4T1 tumor cells by SM16 prevented TGF-beta-induced morphological changes and inhibited TGF-beta-induced invasion in vitro. When delivered via daily i.p. injection or orally through mouse chow, SM16 inhibited the growth of primary and metastatic 4T1 tumors. Splenocytes isolated from mice on the SM16 diet displayed enhanced IFN-gamma production and antitumor CTL activity. Furthermore, SM16 failed to inhibit the growth and metastasis of established 4T1 tumors in immunodeficient SCID mice. CONCLUSION: Taken together, the data indicate that the antitumor efficacy of SM16 is dependent on an immune-mediated mechanism and that SM16 may represent a safe and effective treatment for metastatic breast cancer.
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Compuestos de Azabiciclo/administración & dosificación , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/secundario , Neoplasias Mamarias Experimentales/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/administración & dosificación , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Receptores de Factores de Crecimiento Transformadores beta/antagonistas & inhibidores , Administración Oral , Animales , Apoptosis/efectos de los fármacos , Western Blotting , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Técnica del Anticuerpo Fluorescente , Neoplasias Mamarias Experimentales/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Desnudos , Proteínas Serina-Treonina Quinasas/metabolismo , Receptor Tipo I de Factor de Crecimiento Transformador beta , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Malignant mesothelioma is an aggressive and lethal pleural cancer that overexpresses transforming growth factor beta (TGFbeta). We investigated the efficacy of a novel small-molecule TGFbeta type I receptor (ALK5) kinase inhibitor, SM16, in the AB12 syngeneic model of malignant mesothelioma. SM16 inhibited TGFbeta signaling seen as decreased phosphorylated Smad2/3 levels in cultured AB12 cells (IC(50), approximately 200 nmol/L). SM16 penetrated tumor cells in vivo, suppressing tumor phosphorylated Smad2/3 levels for at least 3 h following treatment of tumor-bearing mice with a single i.p. bolus of 20 mg/kg SM16. The growth of established AB12 tumors was significantly inhibited by 5 mg/kg/d SM16 (P < 0.001) delivered via s.c. miniosmotic pumps over 28 days. The efficacy of SM16 was a result of a CD8+ antitumor response because (a) the antitumor effects were markedly diminished in severe combined immunodeficient mice and (b) CD8+ T cells isolated from spleens of mice treated with SM16 showed strong antitumor cytolytic effects whereas CD8+ T cells isolated from spleens of tumor-bearing mice treated with control vehicle showed minimal activity. Treatment of mice bearing large tumors with 5 mg/kg/d SM16 after debulking surgery reduced the extent of tumor recurrence from 80% to <20% (P < 0.05). SM16 was also highly effective in blocking and regressing tumors when given p.o. at doses of 0.45 or 0.65 g/kg in mouse chow. Thus, SM16 shows potent activity against established AB12 malignant mesothelioma tumors using an immune-mediated mechanism and can significantly prevent tumor recurrence after resection of bulky AB12 malignant mesothelioma tumors. These data suggest that ALK5 inhibitors, such as SM16, offer significant potential for the treatment of malignant mesothelioma and possibly other cancers.
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Receptores de Activinas Tipo I/antagonistas & inhibidores , Mesotelioma/tratamiento farmacológico , Mesotelioma/cirugía , Recurrencia Local de Neoplasia/prevención & control , Inhibidores de Proteínas Quinasas/uso terapéutico , Receptores de Factores de Crecimiento Transformadores beta/antagonistas & inhibidores , Animales , Proliferación Celular/efectos de los fármacos , Supervivencia sin Enfermedad , Evaluación Preclínica de Medicamentos , Femenino , Humanos , Inmunidad Celular/efectos de los fármacos , Mesotelioma/patología , Ratones , Ratones Endogámicos BALB C , Ratones SCID , Trasplante de Neoplasias , Proteínas Serina-Treonina Quinasas , Receptor Tipo I de Factor de Crecimiento Transformador beta , Células Tumorales CultivadasRESUMEN
Inhibitors targeting BRAF and its downstream kinase MEK produce robust response in patients with advanced BRAF V600-mutant melanoma. However, the duration and depth of response vary significantly between patients; therefore, predicting response a priori remains a significant challenge. Here, we utilized the Novartis collection of patient-derived xenografts to characterize transcriptional alterations elicited by BRAF and MEK inhibitors in vivo, in an effort to identify mechanisms governing differential response to MAPK inhibition. We show that the expression of an MITF-high, "epithelial-like" transcriptional program is associated with reduced sensitivity and adaptive response to BRAF and MEK inhibitor treatment. On the other hand, xenograft models that express an MAPK-driven "mesenchymal-like" transcriptional program are preferentially sensitive to MAPK inhibition. These gene-expression programs are somewhat similar to the MITF-high and -low phenotypes described in cancer cell lines, but demonstrate an inverse relationship with drug response. This suggests a discrepancy between in vitro and in vivo experimental systems that warrants future investigations. Finally, BRAF V600-mutant melanoma relies on either MAPK or alternative pathways for survival under BRAF and MEK inhibition in vivo, which in turn predicts their response to further pathway suppression using a combination of BRAF, MEK, and ERK inhibitors. Our findings highlight the intertumor heterogeneity in BRAF V600-mutant melanoma, and the need for precision medicine strategies to target this aggressive cancer.
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MAP Quinasa Quinasa 2/antagonistas & inhibidores , Sistema de Señalización de MAP Quinasas/genética , Proteínas Proto-Oncogénicas B-raf/genética , Animales , Línea Celular Tumoral , Modelos Animales de Enfermedad , Humanos , RatonesRESUMEN
Trichoderma reesei is well known as an industrial workhorse fungus in cellulase production. The low dissolved oxygen supply in the highly viscous medium of T. reesei remains a major bottleneck that hampers growth and cellulase production in submerged fermentation. Vitreoscilla hemoglobin (VHb) has been demonstrated to improve metabolism and protein production in different heterologous hosts under hypoxic conditions, but the use of VHb in T. reesei remains uninvestigated. This study examines the effect of VHb in improving T. reesei performance in submerged fermentation. The VHb gene (vgb)-expressing cassette was successfully transformed into the TU-6 strain, integrated into the genome of T. reesei, and functionally expressed with biological activity, which was confirmed by carbon monoxide difference analysis. Compared to the parent strain, the expression of VHb increased the glucose consumption rate of the transformant. Moreover, in cellulase-inducing medium total protein secretion of the VHb expressing strain was 2.2-fold of the parental strain and the filter paper cellulase activity was increased by 58% under oxygen-limiting conditions. In summary, our results demonstrate that VHb has beneficial effects on improving total protein secretion and cellulase activity of T. reesei in submerged fermentation.
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Because of its tumor-suppressive effect, interferon-based therapy has been used for the treatment of melanoma. However, limited data are available regarding the antitumor effects of pegylated interferons, either alone or in combination with approved anticancer drugs. We report that treatment of human WM-266-4 melanoma cells with peginterferon beta-1a induced apoptotic markers. Additionally, peginterferon beta-1a significantly inhibited the growth of human SK-MEL-1, A-375, and WM-266-4 melanoma xenografts established in immunocompromised mice. Peginterferon beta-1a regressed large, established WM-266-4 xenografts in nude mice. Treatment of SK-MEL-1 tumor-bearing mice with a combination of peginterferon beta-1a and the MEK inhibitor PD325901 ((R)-N-(2,3-dihydroxypropoxy)-3,4-difluoro-2-(2-fluoro-4-iodophenylamino)benzamide) significantly improved tumor growth inhibition compared with either agent alone. Examination of the antitumor activity of peginterferon beta-1a in combination with approved anticancer drugs in breast and renal carcinomas revealed improved antitumor activity in these preclinical xenograft models, as did the combination of peginterferon beta-1a and bevacizumab in a colon carcinoma xenograft model.
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Antineoplásicos/farmacología , Interferón beta/farmacología , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Polietilenglicoles/farmacología , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/farmacocinética , Antineoplásicos Inmunológicos/farmacología , Apoptosis/efectos de los fármacos , Biomarcadores de Tumor , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Línea Celular Tumoral , Supervivencia Celular , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/patología , Modelos Animales de Enfermedad , Quimioterapia Combinada , Femenino , Humanos , Interferón beta/administración & dosificación , Interferón beta/farmacocinética , Neoplasias Renales/tratamiento farmacológico , Neoplasias Renales/patología , Masculino , Melanoma/tratamiento farmacológico , Melanoma/patología , Ratones , Ratones Noqueados , Mutación , Neoplasias/genética , Polietilenglicoles/administración & dosificación , Polietilenglicoles/farmacocinética , Inhibidores de Proteínas Quinasas/farmacología , Resultado del Tratamiento , Carga Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Despite an improving therapeutic landscape, significant challenges remain in treating the majority of patients with advanced ovarian or renal cancer. We identified the cell-cell adhesion molecule cadherin-6 (CDH6) as a lineage gene having significant differential expression in ovarian and kidney cancers. HKT288 is an optimized CDH6-targeting DM4-based antibody-drug conjugate (ADC) developed for the treatment of these diseases. Our study provides mechanistic evidence supporting the importance of linker choice for optimal antitumor activity and highlights CDH6 as an antigen for biotherapeutic development. To more robustly predict patient benefit of targeting CDH6, we incorporate a population-based patient-derived xenograft (PDX) clinical trial (PCT) to capture the heterogeneity of response across an unselected cohort of 30 models-a novel preclinical approach in ADC development. HKT288 induces durable tumor regressions of ovarian and renal cancer models in vivo, including 40% of models on the PCT, and features a preclinical safety profile supportive of progression toward clinical evaluation.Significance: We identify CDH6 as a target for biotherapeutics development and demonstrate how an integrated pharmacology strategy that incorporates mechanistic pharmacodynamics and toxicology studies provides a rich dataset for optimizing the therapeutic format. We highlight how a population-based PDX clinical trial and retrospective biomarker analysis can provide correlates of activity and response to guide initial patient selection for first-in-human trials of HKT288. Cancer Discov; 7(9); 1030-45. ©2017 AACR.This article is highlighted in the In This Issue feature, p. 920.
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Antineoplásicos/uso terapéutico , Cadherinas/antagonistas & inhibidores , Neoplasias Renales/tratamiento farmacológico , Neoplasias Ováricas/tratamiento farmacológico , Animales , Antineoplásicos/farmacología , Cadherinas/genética , Cadherinas/metabolismo , Femenino , Humanos , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Macaca fascicularis , Ratones Desnudos , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Ratas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Oncogenic mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) occur in several types of cancer, but the metabolic consequences of these genetic changes are not fully understood. In this study, we performed (13)C metabolic flux analysis on a panel of isogenic cell lines containing heterozygous IDH1/2 mutations. We observed that under hypoxic conditions, IDH1-mutant cells exhibited increased oxidative tricarboxylic acid metabolism along with decreased reductive glutamine metabolism, but not IDH2-mutant cells. However, selective inhibition of mutant IDH1 enzyme function could not reverse the defect in reductive carboxylation activity. Furthermore, this metabolic reprogramming increased the sensitivity of IDH1-mutant cells to hypoxia or electron transport chain inhibition in vitro. Lastly, IDH1-mutant cells also grew poorly as subcutaneous xenografts within a hypoxic in vivo microenvironment. Together, our results suggest therapeutic opportunities to exploit the metabolic vulnerabilities specific to IDH1 mutation.
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Ciclo del Ácido Cítrico , Isocitrato Deshidrogenasa/genética , Mitocondrias/metabolismo , Mutación Missense , Animales , Antineoplásicos/farmacología , Hipoxia de la Célula , Inhibidores Enzimáticos/farmacología , Glutamina/metabolismo , Células HCT116 , Humanos , Isocitrato Deshidrogenasa/antagonistas & inhibidores , Isocitrato Deshidrogenasa/metabolismo , Ratones , Oxidación-Reducción , Estrés Fisiológico , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Agonistic antibodies targeting Fn14, the receptor for TWEAK, have demonstrated anti-tumor activity in xenograft models. Herein, we further explore the therapeutic potential of the humanized anti-Fn14 agonistic antibody, BIIB036, as a single agent and in combination with standard of care cancer therapeutics. Pharmacokinetic studies of BIIB036 in tumor-bearing mice revealed a half-life of approximately three days suggesting twice a week dosing would be necessary to maintain efficacy. However, in multiple xenograft models, BIIB036 treatment resulted in extended tumor growth inhibition up to 40-50 d following cessation of dosing, suggesting that frequent administration of BIIB036 may not be necessary to maintain prolonged anti-tumor activity. Subsequent xenograft studies revealed that maximal efficacy was achieved with BIIB036 dosing once every two weeks, by either intraperitoneal or subcutaneous administration. Xenograft tumors that were initially treated with BIBI036 and then re-grew up to 1000 mm³ following cessation of the first cycle of treatment remained sensitive to a second cycle of treatment. BIIB036 was also evaluated in patient derived primary colon tumor models, where efficacy compared favorably with a standard of care agent. Lastly, BIIB036 enhanced the efficacy of several standard of care chemotherapeutics, including paclitaxel in MDA-MBA-231 breast tumor xenografts, paclitaxel or carboplatin in HOP62 non-small cell lung xenografts, and 5-FU in NCI-N87 gastric xenografts, with no overlapping toxicities. These studies thus establish BIIB036 as a promising therapeutic agent with durable anti-tumor activity in human xenografts as well as patient derived primary tumor models, and enhanced activity and tolerability in combination with standard of care chemotherapeutics. Taken together, the data presented herein suggest that BIIB036 warrants evaluation in the clinic.
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Anticuerpos Monoclonales Humanizados/administración & dosificación , Antineoplásicos/administración & dosificación , Neoplasias del Colon/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Receptores del Factor de Necrosis Tumoral/inmunología , Neoplasias Gástricas/tratamiento farmacológico , Animales , Anticuerpos Monoclonales Humanizados/farmacocinética , Antineoplásicos/farmacocinética , Carboplatino/administración & dosificación , Carboplatino/farmacología , Línea Celular Tumoral , Neoplasias del Colon/patología , Sinergismo Farmacológico , Femenino , Fluorouracilo/administración & dosificación , Fluorouracilo/farmacología , Semivida , Humanos , Neoplasias Pulmonares/patología , Ratones , Ratones Desnudos , Ratones SCID , Paclitaxel/administración & dosificación , Paclitaxel/farmacología , Neoplasias Gástricas/patología , Receptor de TWEAK , Carga Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Locally produced transforming growth factor-beta (TGF-beta) promotes tumor-induced immunosuppression and contributes to resistance to immunotherapy. This article explores the potential for increased efficacy when combining immunotherapies with TGF-beta suppression using the TGF-beta type I receptor kinase inhibitor SM16. Adenovirus expressing IFN-beta (Ad.IFN-beta) was injected intratumorally once in established s.c. AB12 (mesothelioma) and LKR (lung cancer) tumors or intratracheally in a Kras orthotopic lung tumor model. Mice bearing TC1 (lung cancer) tumors were vaccinated with two injections of adenovirus expressing human papillomavirus-E7 (HPV-E7; Ad.E7). SM16 was administered orally in formulated chow. Tumor growth was assessed and cytokine expression and cell populations were measured in tumors and spleens by real-time PCR and flow cytometry. SM16 potentiated the efficacy of both immunotherapies in each of the models and caused changes in the tumor microenvironment. The combination of SM16 and Ad.IFN-beta increased the number of intratumoral leukocytes (including macrophages, natural killer cells, and CD8(+) cells) and increased the percentage of T cells expressing the activation marker CD25. SM16 also augmented the antitumor effects of Ad.E7 in the TC1 flank tumor model. The combination did not increase HPV-E7 tetramer-positive CD8(+) T cells in the spleens but did induce a marked increase in the tumors. Tumors from SM16-treated mice showed increased mRNA and protein for immunostimulatory cytokines and chemokines, as well as endothelial adhesion molecules, suggesting a mechanism for the increased intratumoral leukocyte trafficking. Blockade of the TGF-beta signaling pathway augments the antitumor effects of Ad.IFN-beta immune-activating or Ad.E7 vaccination therapy. The addition of TGF-beta blocking agents in clinical trials of immunotherapies may increase efficacy.
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Compuestos de Azabiciclo/farmacología , Vacunas contra el Cáncer/farmacología , Inmunogenética/métodos , Neoplasias Pulmonares/terapia , Mesotelioma/terapia , Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Adenoviridae/genética , Animales , Linfocitos T CD8-positivos/inmunología , Vacunas contra el Cáncer/genética , Vacunas contra el Cáncer/inmunología , Terapia Combinada , Citocinas/biosíntesis , Citocinas/genética , Citocinas/inmunología , Epítopos de Linfocito T/inmunología , Femenino , Inmunoterapia/métodos , Molécula 1 de Adhesión Intercelular/biosíntesis , Molécula 1 de Adhesión Intercelular/genética , Molécula 1 de Adhesión Intercelular/inmunología , Interferón beta/genética , Interferón beta/inmunología , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/inmunología , Masculino , Mesotelioma/genética , Mesotelioma/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Proteínas Oncogénicas Virales/genética , Proteínas Oncogénicas Virales/inmunología , ARN Mensajero/biosíntesis , ARN Mensajero/genéticaRESUMEN
Moxifloxacin has enhanced potency against Staphylococcus aureus, lower propensity to select for resistant mutants, and higher bactericidal activity against highly resistant strains than ciprofloxacin. Despite similar activity against purified S. aureus topoisomerase IV and DNA gyrase, it selects for topoisomerase IV mutants, making topoisomerase IV the preferred target in vivo.
Asunto(s)
Antiinfecciosos/farmacología , Compuestos Aza , Fluoroquinolonas , Quinolinas , Staphylococcus aureus/efectos de los fármacos , Ciprofloxacina/farmacología , Girasa de ADN/genética , Topoisomerasa de ADN IV/antagonistas & inhibidores , Topoisomerasa de ADN IV/genética , Farmacorresistencia Bacteriana , Moxifloxacino , Mutación , Staphylococcus aureus/genética , Inhibidores de Topoisomerasa IIRESUMEN
We characterized a Staphylococcus aureus norA gene expression regulator, NorR, initially identified from its binding to the norA promoter. The norR gene was 444 bp in length, located approximately 7 kb upstream from the norA gene, and encoded a predicted 17.6-kDa protein. Overexpression of norR in wild-type S. aureus strain ISP794 led to a fourfold decrease in sensitivity to quinolones and ethidium bromide and an increase in the level of norA transcripts, suggesting that NorR acts as a positive regulator of norA expression. Overexpression of norR in sarA and agr mutants did not alter quinolone sensitivity or levels of norA transcription, indicating that the presence of these two global regulatory systems is necessary for NorR to affect the expression of norA. Insertion and disruption of norR in ISP794 increased resistance to quinolones by 4- to 16-fold but had no effect on norA transcription, suggesting that NorR acts as a repressor for another unidentified efflux pump or pumps. These mutants also exhibited an exaggerated clumping phenotype in liquid media, which was complemented fully by a plasmid-encoded norR gene. Collectively, these results indicate that NorR is a multifunctional regulator, affecting cell surface properties as well as the expression of NorA and likely other multidrug resistance efflux pumps.
Asunto(s)
Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Regulación Bacteriana de la Expresión Génica , Staphylococcus aureus/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteínas Bacterianas/aislamiento & purificación , Secuencia de Bases , Sitios de Unión , Membrana Celular/genética , Farmacorresistencia Bacteriana Múltiple/genética , Genes Reguladores , Datos de Secuencia Molecular , Proteínas Asociadas a Resistencia a Múltiples Medicamentos , Mutación , Regiones Promotoras Genéticas , Quinolonas/farmacología , ARN Mensajero/metabolismo , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/metabolismo , Factores de Transcripción/aislamiento & purificación , Transcripción GenéticaRESUMEN
Gemifloxacin, a novel quinolone with potent activity against Staphylococcus aureus, was 8- to 16-fold more active against wild-type S. aureus than ciprofloxacin. The two- to fourfold increase in the MIC of gemifloxacin in genetically defined grlBA mutants and the twofold increase in a single gyrA mutant, supported by the low frequency of selection of resistant mutants at twice the MIC (7.4 x 10(-11) to 1.1 x 10(-10)), suggested similar targeting of the two enzymes by gemifloxacin. Dual mutations in both gyrase and topoisomerase IV caused a 64- to 128-fold increase in the MIC of gemifloxacin, similar to that seen with ciprofloxacin. Gemifloxacin also had similar activity in vitro against topoisomerase IV and gyrase purified from S. aureus (50% inhibitory concentrations of 0.25 and 0.31 micro g/ml, respectively). This activity was 10- to 20-fold higher than that of ciprofloxacin for topoisomerase IV and 33-fold higher than that for gyrase. In contrast to the in vitro findings, only topoisomerase IV mutants were selected in first-step mutants. Overexpression of the NorA efflux pump had a minimal effect on resistance to gemifloxacin, and a mutation in the promoter region of the gene for NorA was selected only in the sixth step of serial selection of mutants. Our data show that although gemifloxacin targets purified topoisomerase IV and gyrase similarly in vitro, topoisomerase IV is the preferred target in the bacteria. Selection of novel resistance mutations in grlA requires further expansion of quinolone-resistance-determining regions, and their study may provide increased insight into enzyme-quinolone interactions.
Asunto(s)
Antiinfecciosos/farmacología , ADN-Topoisomerasas/efectos de los fármacos , Farmacorresistencia Bacteriana/genética , Fluoroquinolonas , Pruebas de Sensibilidad Microbiana , Naftiridinas/farmacología , Staphylococcus aureus/enzimología , ADN-Topoisomerasas/clasificación , ADN-Topoisomerasas/genética , Gemifloxacina , Mutación , Plásmidos/genética , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/genéticaRESUMEN
We determined the target enzyme interactions of garenoxacin (BMS-284756, T-3811ME), a novel desfluoroquinolone, in Staphylococcus aureus by genetic and biochemical studies. We found garenoxacin to be four- to eightfold more active than ciprofloxacin against wild-type S. aureus. A single topoisomerase IV or gyrase mutation caused only a 2- to 4-fold increase in the MIC of garenoxacin, whereas a combination of mutations in both loci caused a substantial increase (128-fold). Overexpression of the NorA efflux pump had minimal effect on resistance to garenoxacin. With garenoxacin at twice the MIC, selection of resistant mutants (<7.4 x 10(-12) to 4.0 x 10(-11)) was 5 to 6 log units less than that with ciprofloxacin. Mutations inside or outside the quinolone resistance-determining regions (QRDR) of either topoisomerase IV, or gyrase, or both were selected in single-step mutants, suggesting dual targeting of topoisomerase IV and gyrase. Three of the novel mutations were shown by genetic experiments to be responsible for resistance. Studies with purified topoisomerase IV and gyrase from S. aureus also showed that garenoxacin had similar activity against topoisomerase IV and gyrase (50% inhibitory concentration, 1.25 to 2.5 and 1.25 micro g/ml, respectively), and although its activity against topoisomerase IV was 2-fold greater than that of ciprofloxacin, its activity against gyrase was 10-fold greater. This study provides the first genetic and biochemical data supporting the dual targeting of topoisomerase IV and gyrase in S. aureus by a quinolone as well as providing genetic proof for the expansion of the QRDRs to include the 5' terminus of grlB and the 3' terminus of gyrA.