RESUMEN
Protein arginine methyltransferases (PRMTs) regulate diverse biological processes and are increasingly being recognized for their potential as drug targets. Here we report the discovery of a potent, selective, and cell-active chemical probe for PRMT7. SGC3027 is a cell permeable prodrug, which in cells is converted to SGC8158, a potent, SAM-competitive PRMT7 inhibitor. Inhibition or knockout of cellular PRMT7 results in drastically reduced levels of arginine monomethylated HSP70 family stress-associated proteins. Structural and biochemical analyses reveal that PRMT7-driven in vitro methylation of HSP70 at R469 requires an ATP-bound, open conformation of HSP70. In cells, SGC3027 inhibits methylation of both constitutive and inducible forms of HSP70, and leads to decreased tolerance for perturbations of proteostasis including heat shock and proteasome inhibitors. These results demonstrate a role for PRMT7 and arginine methylation in stress response.
Asunto(s)
Arginina/metabolismo , Proteínas HSP70 de Choque Térmico/metabolismo , Proteína-Arginina N-Metiltransferasas/metabolismo , Estrés Fisiológico , Animales , Técnicas de Silenciamiento del Gen , Células HCT116 , Humanos , Metilación/efectos de los fármacos , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Proteína-Arginina N-Metiltransferasas/antagonistas & inhibidores , Proteína-Arginina N-Metiltransferasas/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Células Sf9RESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMEN
T-3775440 is an irreversible inhibitor of the chromatin demethylase LSD1, which exerts antiproliferative effects by disrupting the interaction between LSD1 and GFI1B, a SNAG domain transcription factor, inducing leukemia cell transdifferentiation. Here, we describe the anticancer effects and mechanism of action of T-3775440 in small-cell lung cancer (SCLC). T-3775440 inhibited proliferation of SCLC cells in vitro and retarded SCLC tumor growth in vivo T-3775440 disrupted the interaction between LSD1 and the transcriptional repressor INSM1, thereby inhibiting expression of neuroendocrine-associated genes, such as ASCL1 INSM1 silencing phenocopied the effects of T-3775440 on gene expression and cell proliferation, consistent with the likelihood T-3775440 mediated its effects in SCLC by inhibiting INSM1. T-3775440 also inhibited proliferation of an SCLC cell line that overexpressed GFI1B, rather than INSM1, by disrupting the interaction between LSD1 and GFI1B. Taken together, our results argue that LSD1 plays an important role in neuroendocrine-associated transcription and cell proliferation of SCLC via interactions with the SNAG domain proteins INSM1 and GFI1B. Targeting these critical interactions with LSD1 inhibitors offers a novel rational strategy to therapeutically manage SCLC. Cancer Res; 77(17); 4652-62. ©2017 AACR.
Asunto(s)
Anilidas/farmacología , Antineoplásicos/farmacología , Proliferación Celular/efectos de los fármacos , Ciclopropanos/farmacología , Histona Demetilasas/antagonistas & inhibidores , Dominios y Motivos de Interacción de Proteínas/efectos de los fármacos , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Represoras/antagonistas & inhibidores , Carcinoma Pulmonar de Células Pequeñas/tratamiento farmacológico , Animales , Femenino , Histona Demetilasas/metabolismo , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Ratones Endogámicos BALB C , Ratones Desnudos , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Carcinoma Pulmonar de Células Pequeñas/metabolismo , Carcinoma Pulmonar de Células Pequeñas/patología , Factores de Transcripción de la Familia Snail , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Dysregulation of lysine (K)-specific demethylase 1A (LSD1), also known as KDM1A, has been implicated in the development of various cancers, including leukemia. Here, we describe the antileukemic activity and mechanism of action of T-3775440, a novel irreversible LSD1 inhibitor. Cell growth analysis of leukemia cell lines revealed that acute erythroid leukemia (AEL) and acute megakaryoblastic leukemia cells (AMKL) were highly sensitive to this compound. T-3775440 treatment enforced transdifferentiation of erythroid/megakaryocytic lineages into granulomonocytic-like lineage cells. Mechanistically, T-3775440 disrupted the interaction between LSD1 and growth factor-independent 1B (GFI1B), a transcription factor critical for the differentiation processes of erythroid and megakaryocytic lineage cells. Knockdown of LSD1 and GFI1B recapitulated T-3775440-induced transdifferentiation and cell growth suppression, highlighting the significance of LSD1-GFI1B axis inhibition with regard to the anti-AML effects of T-3775440. Moreover, T-3775440 exhibited significant antitumor efficacy in AEL and AMKL xenograft models. Our findings provide a rationale for evaluating LSD1 inhibitors as potential treatments and indicate a novel mechanism of action against AML, particularly AEL and AMKL. Mol Cancer Ther; 16(2); 273-84. ©2016 AACR.
Asunto(s)
Antineoplásicos/farmacología , Transdiferenciación Celular/efectos de los fármacos , Histona Demetilasas/antagonistas & inhibidores , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Complejos Multiproteicos/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Animales , Antineoplásicos/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Análisis por Conglomerados , Biología Computacional/métodos , Modelos Animales de Enfermedad , Resistencia a Antineoplásicos , Femenino , Perfilación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Hematopoyesis/genética , Histona Demetilasas/genética , Histona Demetilasas/metabolismo , Humanos , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Ratones , Terapia Molecular Dirigida , Unión Proteica , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Recurrent mutations in histone-modifying enzymes imply key roles in tumorigenesis, yet their functional relevance is largely unknown. Here, we show that JARID1B, encoding a histone H3 lysine 4 (H3K4) demethylase, is frequently amplified and overexpressed in luminal breast tumors and a somatic mutation in a basal-like breast cancer results in the gain of unique chromatin binding and luminal expression and splicing patterns. Downregulation of JARID1B in luminal cells induces basal genes expression and growth arrest, which is rescued by TGFß pathway inhibitors. Integrated JARID1B chromatin binding, H3K4 methylation, and expression profiles suggest a key function for JARID1B in luminal cell-specific expression programs. High luminal JARID1B activity is associated with poor outcome in patients with hormone receptor-positive breast tumors.
Asunto(s)
Neoplasias de la Mama/genética , Histona Demetilasas con Dominio de Jumonji/genética , Proteínas Nucleares/genética , Oncogenes , Proteínas Represoras/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Factor de Unión a CCCTC , Procesos de Crecimiento Celular/genética , Línea Celular Tumoral , Linaje de la Célula , Femenino , Amplificación de Genes , Regulación Neoplásica de la Expresión Génica , Histonas/genética , Histonas/metabolismo , Humanos , Histona Demetilasas con Dominio de Jumonji/metabolismo , Células MCF-7 , Mutación , Proteínas Nucleares/metabolismo , Regiones Promotoras Genéticas , Pirazoles/farmacología , Pirroles/farmacología , ARN Interferente Pequeño/administración & dosificación , ARN Interferente Pequeño/genética , Proteínas Represoras/metabolismo , Transfección , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
The human epidermal growth factor receptor (HER) family plays a major role in cancer cell proliferation. Overexpression of these receptors occurs in various cancers, including breast cancer, and correlates with shorter time to relapse and lower overall survival. We recently reported that TAK-285, an orally bioavailable small molecule inhibitor of HER kinases, is not a p-glycoprotein substrate and penetrates the blood-brain barrier, suggesting favorable activity for the treatment of brain metastases. To identify the determinants of sensitivity to TAK-285, we examined the relationship between the IC50 values of TAK-285 for cell growth inhibition and the expression of candidate genes that are involved in the HER family signaling pathway and trastuzumab resistance in a panel of human breast cancer cell lines, other types of cancer cells, and non-transformed cells in vitro. These analyses showed an inverse correlation between sensitivity to TAK-285 (IC50 values) and HER2 or HER3 expression. HER3 was highly phosphorylated in TAK-285-sensitive cells, where TAK-285 treatment reduced HER3 phosphorylation level. Because HER3 does not possess kinase activity and a selective inhibitor of HER2 but not of an epidermal growth factor receptor reduced the phospho-HER3 level, HER3 was suggested to be trans-phosphorylated by HER2. HER3 knockdown using small interfering RNA (siRNA) inhibited cancer cell growth in TAK-285-sensitive cells but not in TAK-285-insensitive cells. These results suggest that HER2 and HER3 mainly regulate cancer cell growth in TAK-285-sensitive cells and that phospho-HER3 could be used as a potential molecular marker to select patients most likely to respond to TAK-285.
RESUMEN
Breast cancer therapy has improved following the development of drugs with specific molecular targets, exemplified by inhibitors of human epidermal growth factor receptor-2 (HER2) or epidermal growth factor receptor (EGFR) such as trastuzumab and lapatinib. However, these drugs have little effect on brain metastasis due to the combined effects of poor penetration of the blood-brain barrier and their removal from the central nervous system (CNS) by the p-glycoprotein (Pgp) drug efflux pump. We investigated the effects of TAK-285, a novel, investigational, dual EGFR/HER2 inhibitor that has been shown to penetrate the CNS and has comparable inhibitory efficacy to lapatinib which is a known Pgp substrate. Tested against a panel of 96 kinases, TAK-285 showed specificity for inhibition of HER family kinases. Unlike lapatinib, TAK-285 is not a substrate for Pgp efflux. In mouse and rat xenograft tumor models, TAK-285 showed antitumor activity against cancers that expressed HER2 or EGFR. TAK-285 was as effective as lapatinib in antitumor activity in a mouse subcutaneous BT-474 breast cancer xenograft model. TAK-285 was examined in a model of breast cancer brain metastasis using direct intracranial injection of BT-474-derived luciferase-expressing cells and showed greater inhibition of brain tumor growth compared to animals treated with lapatinib. Our studies suggest that investigational drugs such as TAK-285 that have strong antitumor activity and are not Pgp substrates may be useful in the development of agents with the potential to treat brain metastases.
RESUMEN
The epidermal growth factor receptor (EGFR) family plays a critical role in vital cellular processes and in various cancers. Known EGFR inhibitors exhibit distinct antitumor responses against the various EGFR mutants associated with nonsmall-cell lung cancer. The L858R mutation enhances clinical sensitivity to gefitinib and erlotinib as compared with wild type and reduces the relative sensitivity to lapatinib. In contrast, the T790M mutation confers drug resistance to gefitinib and erlotinib. We determined crystal structures of the wild-type and T790M/L858R double mutant EGFR kinases with reversible and irreversible pyrrolo[3,2-d]pyrimidine inhibitors based on analogues of TAK-285 and neratinib. In these structures, M790 adopts distinct conformations to accommodate different inhibitors, whereas R858 allows conformational variations of the activation loop. These results provide structural insights for understanding the structure-activity relationships that should contribute to the development of potent inhibitors against drug-sensitive or -resistant EGFR mutations.
Asunto(s)
Brotes de Enfermedades , Contaminación de Alimentos , Intoxicación Alimentaria por Salmonella/epidemiología , Intoxicación Alimentaria por Salmonella/microbiología , Salmonella enteritidis/clasificación , Electroforesis en Gel de Campo Pulsado , Gastroenteritis/epidemiología , Gastroenteritis/microbiología , Humanos , Japón/epidemiología , Salmonella enteritidis/genética , Salmonella enteritidis/aislamiento & purificaciónRESUMEN
The aim of this study was to evaluate the usefulness of targeted intra-arterial carboplatin chemoradiotherapy in allowing less invasive surgery for patients with oral and oropharyngeal squamous cell carcinoma. Twenty patients with previously untreated squamous cell carcinoma of the oral cavity and oropharynx (T4; 8, T2; 12 patients) were treated with targeted transfemoral intra-arterial carboplatin infusion with concurrent hyperfractionated radiotherapy and the administration of tegafur/uracil (UFT). Of 20 patients, 15 underwent surgery after completion of one course of targeted chemoradiotherapy, and five were given another course or radiotherapy only. Eighteen (90%) of 20 patients had a clinically complete response at the primary site and two (10%) had a partial response. Of the 15 patients who underwent tumor resection, 11 (73%) showed histopathological disappearance of cancer cells at the primary site. Sixteen (80 %) of 20 tumors were controlled at the primary site within a mean follow-up of 30 months. Adverse effects were relatively mild. Targeted intra-arterial chemoradiotherapy caused a down-staging of tumors and facilitated the use of less invasive surgery in patients with squamous cell carcinoma of the oral cavity and oropharynx as a result of its favorable anti-tumor effect.
Asunto(s)
Antineoplásicos/administración & dosificación , Carboplatino/administración & dosificación , Carcinoma de Células Escamosas/terapia , Neoplasias de la Boca/terapia , Neoplasias Orofaríngeas/terapia , Adulto , Anciano , Anciano de 80 o más Años , Antimetabolitos Antineoplásicos/administración & dosificación , Terapia Combinada , Sistemas de Liberación de Medicamentos , Femenino , Humanos , Infusiones Intraarteriales , Masculino , Persona de Mediana Edad , Tegafur/administración & dosificación , Resultado del Tratamiento , Uracilo/administración & dosificaciónRESUMEN
This study was performed to analyze treatment of fractures of the edentulous mandible and to discuss this method in relation to the mandibular height at the fracture site. Fifteen fracture sites in 11 patients with an edentulous mandible were retrospectively examined. These fractures were located: nine fractures in the mandibular body, three in the paramedian region, and three in the mandibular angle. Fractures in a mandible measuring more than 10 mm in the vertical height were treated with one miniplate. Fractures in an extremely atrophic mandible with 10 mm or less were treated using one or two miniplates, also using a modified Champy plate with 1.3 mm in thickness. A mandibular fracture with a height of 5 mm was treated with a combination of a microplate on the buccal side and a miniplate on the inferior border of the mandible with additional direct circumferential wiring. Oblique or splitting fractures were treated with direct circumferential wiring or a Herbert screw, at one fracture site each, respectively. Complications, including infection, fibrous union, nonunion and trismus, were not seen. In one patient, hypesthesia of the lower lip was, however, persistent 1 month after surgery. Miniplate osteosynthesis is the less invasive treatment, and it is suitable for fractures of the atrophic edentulous mandible, except for comminuted or defect fractures. To obtain stable fixation in severely atrophic mandibles, we need to consider the use of two miniplates or a combination with microplates.
Asunto(s)
Placas Óseas , Fijación Interna de Fracturas/instrumentación , Arcada Edéntula/cirugía , Fracturas Mandibulares/cirugía , Anciano , Anciano de 80 o más Años , Atrofia , Tornillos Óseos , Hilos Ortopédicos , Femenino , Estudios de Seguimiento , Fijación Interna de Fracturas/métodos , Humanos , Hipoestesia/etiología , Arcada Edéntula/patología , Enfermedades de los Labios/etiología , Masculino , Mandíbula/patología , Fracturas Mandibulares/clasificación , Fracturas Mandibulares/patología , Persona de Mediana Edad , Procedimientos Quirúrgicos Mínimamente Invasivos , Complicaciones Posoperatorias , Estudios RetrospectivosRESUMEN
Wnt/beta-catenin signaling plays important roles in tumorigenesis in certain tumors as well as during development. However, the mechanism of tumorigenesis mediated by this signaling remains to be elucidated. We investigated the response of rodent fibroblasts to activation of Wnt/beta-catenin signaling by treatment with conditioned medium containing soluble Wnt-3a protein (W3a-CM) and by expression of a constitutive active beta-catenin gene harbored by an adenovirus vector. W3a-CM induced transcriptional activation of a beta-catenin/T-cell factor (Tcf)-responsive promoter in rodent fibroblasts such as NIH3T3, Rat-1, Swiss3T3 and Balb3T3 cells. In these cells, an increase in saturation density and an inhibition of apoptosis and/or promotion of growth in low-serum medium were induced by treatment with W3a-CM. In Rat-1 cells, morphologic changes were also induced. All these alterations were reversible. Moreover, the inhibition of apoptosis of NIH3T3 cells in low-serum medium and the morphologic changes in Rat-1 cells, but not the increase in saturation density, were also induced by ectopic expression of a constitutive active beta-catenin gene. These results suggested that activation of Wnt/beta-catenin signaling induces inhibition of apoptosis and morphologic changes in these cells.