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AV-101 (imatinib) powder for inhalation, an investigational dry powder inhaled formulation of imatinib designed to target the underlying pathobiology of pulmonary arterial hypertension, was generally well tolerated in healthy adults in a phase 1 single and multiple ascending dose study. Inhaled Imatinib Pulmonary Arterial Hypertension Clinical Trial (IMPAHCT; NCT05036135) is a phase 2b/3, randomized, double-blind, placebo-controlled, dose-ranging, and confirmatory study. IMPAHCT is designed to identify an optimal AV-101 dose (phase 2b primary endpoint: pulmonary vascular resistance) and assess the efficacy (phase 3 primary endpoint: 6-min walk distance), safety, and tolerability of AV-101 dose levels in subjects with pulmonary arterial hypertension using background therapies. The study has an operationally seamless, adaptive design allowing for continuous recruitment. It includes three parts; subjects enrolled in Part 1 (phase 2b dose-response portion) or Part 2 (phase 3 intermediate portion) will be randomized 1:1:1:1 to 10, 35, 70 mg AV-101, or placebo (twice daily), respectively. Subjects enrolled in Part 3 (phase 3 optimal dose portion) will be randomized 1:1 to the optimal dose of AV-101 and placebo (twice daily), respectively. All study parts include a screening period, a 24-week treatment period, and a 30-day safety follow-up period; the total duration is â¼32 weeks. Participation is possible in only one study part. IMPAHCT has the potential to advance therapies for patients with pulmonary arterial hypertension by assessing the efficacy and safety of a novel investigational drug-device combination (AV-101) using an improved study design that has the potential to save 6-12 months of development time. ClinicalTrials.gov Identifier: NCT05036135.
RESUMEN
Background: Oral imatinib has been shown to be effective, but poorly tolerated, in patients with advanced pulmonary arterial hypertension (PAH). To maintain efficacy while improving tolerability, AV-101, a dry powder inhaled formulation of imatinib, was developed to deliver imatinib directly to the lungs. Methods: This phase 1, placebo-controlled, randomised single ascending dose (SAD) and multiple ascending dose (MAD) study evaluated the safety/tolerability and pharmacokinetics of AV-101 in healthy adults. The SAD study included five AV-101 cohorts (1 mg, 3 mg, 10 mg, 30 mg, 90â mg) and placebo, and a single-dose oral imatinib 400-mg cohort. The MAD study included three AV-101 cohorts (10 mg, 30â mg, 90â mg) and placebo; dosing occurred twice daily for 7â days. Results: 82 participants (SAD n=48, MAD n=34) were enrolled. For the SAD study, peak plasma concentrations of imatinib occurred within 3 h of dosing with lower systemic exposure compared to oral imatinib (p<0.001). For the MAD study, systemic exposure of imatinib was higher after multiple doses of AV-101 compared to a single dose, but steady-state plasma concentrations were lower for the highest AV-101 cohort (90â mg) compared to simulated steady-state oral imatinib at day 7 (p=0.0002). Across AV-101 MAD dose cohorts, the most common treatment-emergent adverse events were cough (n=7, 27%) and headache (n=4, 15%). Conclusions: AV-101 was well tolerated in healthy adults, and targeted doses of AV-101 significantly reduced the systemic exposure of imatinib compared with oral imatinib. An ongoing phase 2b/phase 3 study (IMPAHCT; clinicaltrials.gov identifier NCT05036135) will evaluate the safety/tolerability and clinical benefit of AV-101 for PAH.
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Luminal breast cancers are typically estrogen receptor-positive and generally have the best prognosis. However, a subset of luminal tumors, namely luminal B cancers, frequently metastasize and recur. Unfortunately, the causal events that drive their progression are unknown, and therefore it is difficult to identify individuals who are likely to relapse and should receive escalated treatment. Here, we identify a bifunctional RasGAP tumor suppressor whose expression is lost in almost 50% of luminal B tumors. Moreover, we show that two RasGAP genes are concomitantly suppressed in the most aggressive luminal malignancies. Importantly, these genes cooperatively regulate two major oncogenic pathways, RAS and NF-κB, through distinct domains, and when inactivated drive the metastasis of luminal tumors in vivo Finally, although the cooperative effects on RAS drive invasion, NF-κB activation triggers epithelial-to-mesenchymal transition and is required for metastasis. Collectively, these studies reveal important mechanistic insight into the pathogenesis of luminal B tumors and provide functionally relevant prognostic biomarkers that may guide treatment decisions. SIGNIFICANCE: The lack of insight into mechanisms that underlie the aggressive behavior of luminal B breast cancers impairs treatment decisions and therapeutic advances. Here, we show that two RasGAP tumor suppressors are concomitantly suppressed in aggressive luminal B tumors and demonstrate that they drive metastasis by activating RAS and NF-κB. Cancer Discov; 7(2); 202-17. ©2016 AACR.See related commentary by Sears and Gray, p. 131This article is highlighted in the In This Issue feature, p. 115.
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Neoplasias de la Mama/patología , Proteínas Portadoras/genética , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Proteínas Activadoras de ras GTPasa/genética , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Proteínas Portadoras/metabolismo , Línea Celular Tumoral , Progresión de la Enfermedad , Transición Epitelial-Mesenquimal , Femenino , Proteínas Activadoras de GTPasa , Humanos , Células MCF-7 , Ratones , Mutación , Metástasis de la Neoplasia , Trasplante de Neoplasias , Transducción de Señal , Proteínas Activadoras de ras GTPasa/metabolismoRESUMEN
The JmjC domain histone H3K36me2/me1 demethylase NDY1/KDM2B is overexpressed in various types of cancer. Here we show that knocking down NDY1 in a set of 10 cell lines derived from a broad range of human tumors inhibited their anchorage-dependent and anchorage-independent growth by inducing senescence and/or apoptosis in some and by inhibiting G1 progression in all. We further show that the knockdown of NDY1 in mammary adenocarcinoma cell lines decreased the number, size, and replating efficiency of mammospheres and downregulated the stem cell markers ALDH and CD44, while upregulating CD24. Together, these findings suggest that NDY1 is required for the self-renewal of cancer stem cells and are in agreement with additional findings showing that tumor cells in which NDY1 was knocked down undergo differentiation and a higher number of them is required to induce mammary adenocarcinomas, upon orthotopic injection in animals. Mechanistically, NDY1 functions as a master regulator of a set of miRNAs that target several members of the polycomb complexes PRC1 and PRC2, and its knockdown results in the de-repression of these miRNAs and the downregulation of their polycomb targets. Consistent with these observations, NDY1/KDM2B is expressed at higher levels in basal-like triple-negative breast cancers, and its overexpression is associated with higher rates of relapse after treatment. In addition, NDY1-regulated miRNAs are downregulated in both normal and cancer mammary stem cells. Finally, in primary human breast cancer, NDY1/KDM2B expression correlates negatively with the expression of the NDY1-regulated miRNAs and positively with the expression of their PRC targets.
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Neoplasias de la Mama/metabolismo , Proteínas F-Box/metabolismo , Histona Demetilasas con Dominio de Jumonji/metabolismo , Células Madre Neoplásicas/metabolismo , Proteínas del Grupo Polycomb/metabolismo , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Proliferación Celular , Proteína Potenciadora del Homólogo Zeste 2 , Proteínas F-Box/genética , Femenino , Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Inmunofenotipificación , Histona Demetilasas con Dominio de Jumonji/genética , Fenotipo , Complejo Represivo Polycomb 2/genética , Complejo Represivo Polycomb 2/metabolismo , Proteínas del Grupo Polycomb/química , Subunidades de Proteína/metabolismo , Interferencia de ARNRESUMEN
BACKGROUND: Ascending aortic dissection and rupture remain a life-threatening complication in patients with Marfan syndrome. The extracellular matrix provides strength and elastic recoil to the aortic wall, thereby preventing radial expansion. We have previously shown that ascending aortic aneurysm formation in Marfan mice (Fbn1(C1039G/+)) is associated with decreased aortic wall elastogenesis and increased elastin breakdown. In this study, we test the feasibility of quantifying aortic wall elastin content using MRI with a gadolinium-based elastin-specific magnetic resonance contrast agent in Fbn1(C1039G/+) mice. METHODS AND RESULTS: Ascending aorta elastin content was measured in 32-week-old Fbn1(C1039G/+) mice and wild-type (n=9 and n=10, respectively) using 7-T MRI with a T1 mapping sequence. Significantly lower enhancement (ie, lower R1 values, where R1=1/T1) was detected post-elastin-specific magnetic resonance contrast agent in Fbn1(C1039G/+) compared with wild-type ascending aortas (1.15±0.07 versus 1.36±0.05; P<0.05). Post-elastin-specific magnetic resonance contrast agent R1 values correlated with ascending aortic wall gadolinium content directly measured by inductively coupled mass spectroscopy (P=0.006). CONCLUSIONS: Herein, we demonstrate that MRI with elastin-specific magnetic resonance contrast agent accurately measures elastin bound gadolinium within the aortic wall and detects a decrease in aortic wall elastin in Marfan mice compared with wild-type controls. This approach has translational potential for noninvasively assessing aneurysm tissue changes and risk, as well as monitoring elastin content in response to therapeutic interventions.
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Aorta Torácica/química , Aneurisma de la Aorta Torácica/diagnóstico , Disección Aórtica/diagnóstico , Medios de Contraste , Elastina/deficiencia , Imagen por Resonancia Magnética/métodos , Síndrome de Marfan/complicaciones , Disección Aórtica/etiología , Disección Aórtica/metabolismo , Animales , Aorta Torácica/patología , Aneurisma de la Aorta Torácica/etiología , Aneurisma de la Aorta Torácica/metabolismo , Quelantes , Modelos Animales de Enfermedad , Elastina/análisis , Estudios de Factibilidad , Compuestos Heterocíclicos con 1 Anillo , Masculino , Síndrome de Marfan/diagnóstico , Síndrome de Marfan/metabolismo , Ratones , Ratones Endogámicos C57BL , Reproducibilidad de los ResultadosRESUMEN
RAS genes are commonly mutated in cancer; however, RAS mutations are rare in breast cancer, despite frequent hyperactivation of Ras and ERK. Here, we report that the RasGAP gene, RASAL2, functions as a tumor and metastasis suppressor. RASAL2 is mutated or suppressed in human breast cancer, and RASAL2 ablation promotes tumor growth, progression, and metastasis in mouse models. In human breast cancer, RASAL2 loss is associated with metastatic disease; low RASAL2 levels correlate with recurrence of luminal B tumors; and RASAL2 ablation promotes metastasis of luminal mouse tumors. Additional data reveal a broader role for RASAL2 inactivation in other tumor types. These studies highlight the expanding role of RasGAPs and reveal an alternative mechanism of activating Ras in cancer.
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Proteínas Portadoras/genética , Neoplasias/genética , Neoplasias/patología , Proteínas Supresoras de Tumor/genética , Proteínas Activadoras de ras GTPasa/genética , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/mortalidad , Neoplasias de la Mama/patología , Proteínas Portadoras/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Transformación Celular Neoplásica/genética , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Proteínas Activadoras de GTPasa , Genes p53 , Humanos , Ratones , Ratones Noqueados , Mutación , Invasividad Neoplásica , Metástasis de la Neoplasia , Neoplasias/metabolismo , Neoplasias/mortalidad , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Trasplante Heterólogo , Proteínas Supresoras de Tumor/metabolismo , Proteínas Activadoras de ras GTPasa/metabolismoRESUMEN
Zinc-finger nucleases (ZFNs) have been used for genome engineering in a wide variety of organisms; however, it remains challenging to design effective ZFNs for many genomic sequences using publicly available zinc-finger modules. This limitation is in part because of potential finger-finger incompatibility generated on assembly of modules into zinc-finger arrays (ZFAs). Herein, we describe the validation of a new set of two-finger modules that can be used for building ZFAs via conventional assembly methods or a new strategy-finger stitching-that increases the diversity of genomic sequences targetable by ZFNs. Instead of assembling ZFAs based on units of the zinc-finger structural domain, our finger stitching method uses units that span the finger-finger interface to ensure compatibility of neighbouring recognition helices. We tested this approach by generating and characterizing eight ZFAs, and we found their DNA-binding specificities reflected the specificities of the component modules used in their construction. Four pairs of ZFNs incorporating these ZFAs generated targeted lesions in vivo, demonstrating that stitching yields ZFAs with robust recognition properties.
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Desoxirribonucleasas de Localización Especificada Tipo II/metabolismo , Dedos de Zinc , Animales , Sitios de Unión , ADN/química , ADN/metabolismo , Desoxirribonucleasas/química , Desoxirribonucleasas de Localización Especificada Tipo II/genética , Células HEK293 , Humanos , Nucleótidos/química , Ingeniería de Proteínas , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/metabolismo , Pez CebraRESUMEN
RATIONALE: Marfan syndrome (MFS) is a systemic connective tissue disorder notable for the development of aortic root aneurysms and the subsequent life-threatening complications of aortic dissection and rupture. Underlying fibrillin-1 gene mutations cause increased transforming growth factor-ß (TGF-ß) signaling. Although TGF-ß blockade prevents aneurysms in MFS mouse models, the mechanisms through which excessive TGF-ß causes aneurysms remain ill-defined. OBJECTIVE: We investigated the role of microRNA-29b (miR-29b) in aneurysm formation in MFS. METHODS AND RESULTS: Using quantitative polymerase chain reaction, we discovered that miR-29b, a microRNA regulating apoptosis and extracellular matrix synthesis/deposition genes, is increased in the ascending aorta of Marfan (Fbn1(C1039G/+)) mice. Increased apoptosis, assessed by increased cleaved caspase-3 and caspase-9, enhanced caspase-3 activity, and decreased levels of the antiapoptotic proteins, Mcl-1 and Bcl-2, were found in the Fbn1(C1039G/+) aorta. Histological evidence of decreased and fragmented elastin was observed exclusively in the Fbn1(C1039G/+) ascending aorta in association with repressed elastin mRNA and increased matrix metalloproteinase-2 expression and activity, both targets of miR-29b. Evidence of decreased activation of nuclear factor κB, a repressor of miR-29b, and a factor suppressed by TGF-ß, was also observed in Fbn1(C1039G/+) aorta. Furthermore, administration of a nuclear factor κB inhibitor increased miR-29b levels, whereas TGF-ß blockade or losartan effectively decreased miR-29b levels in Fbn1(C1039G/+) mice. Finally, miR-29b blockade by locked nucleic acid antisense oligonucleotides prevented early aneurysm development, aortic wall apoptosis, and extracellular matrix deficiencies. CONCLUSIONS: We identify increased miR-29b expression as key to the pathogenesis of early aneurysm development in MFS by regulating aortic wall apoptosis and extracellular matrix abnormalities.
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Aorta/metabolismo , Aneurisma de la Aorta/metabolismo , Síndrome de Marfan/metabolismo , MicroARNs/metabolismo , Factores de Edad , Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacología , Animales , Aorta/patología , Aneurisma de la Aorta/genética , Aneurisma de la Aorta/patología , Aneurisma de la Aorta/prevención & control , Apoptosis , Proteínas Reguladoras de la Apoptosis/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Elastina/genética , Elastina/metabolismo , Femenino , Fibrilina-1 , Fibrilinas , Terapia Genética/métodos , Losartán/farmacología , Masculino , Síndrome de Marfan/complicaciones , Síndrome de Marfan/genética , Síndrome de Marfan/patología , Síndrome de Marfan/terapia , Metaloproteinasa 2 de la Matriz/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , MicroARNs/genética , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , FN-kappa B/metabolismo , Oligonucleótidos Antisentido/administración & dosificación , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor de Crecimiento Transformador beta/metabolismo , Regulación hacia ArribaRESUMEN
BACKGROUND: Type 1 regulatory T (Tr1) cells, characterized by the secretion of high levels of the anti-inflammatory cytokine interleukin-10 (IL-10), play an important role in the regulation of autoimmune diseases and transplantation. However, effective strategies that specifically induce Tr1 cells in vivo are limited. Furthermore, the pathways controlling the induction of these cells in vivo are not well understood. METHODOLOGY/PRINCIPAL FINDINGS: Here we report that nasal administration of anti-CD3 antibody induces suppressive Tr1 cells in mice. The in vivo induction of Tr1 cells by nasal anti-CD3 is dependent on IL-27 produced by upper airway resident dendritic cells (DCs), and is controlled by the transcription factors aryl hydrocarbon receptor (AHR) and c-Maf. Subsequently, IL-21 acts in an autocrine fashion to expand and maintain the Tr1 cells induced in vivo by nasally administered anti-CD3. CONCLUSIONS/SIGNIFICANCE: Our findings identify a unique approach to generate Tr1 cells in vivo and provide insights into the mechanisms by which these cells are induced.