RESUMEN
Pediatric high-grade gliomas (pHGG) accounts for approximately 8-12% of primary brain tumors in children. Prognosis is poor, with a median survival of 9-15 months. Insulin-like growth factor 1-receptor (IGF-1R) gene amplifications have been identified in high-grade gliomas and may contribute to its highly aggressive phenotype, but the effect of IGF inhibitors on pHGG is yet to be determined. In the present study, we analyzed the response of patient-derived pediatric high-grade glioma cells to a novel IGF-1R inhibitor, the IGF-Trap. Using immunohistochemistry, we found that IGF-1R was localized to both the nucleus and cell membrane in different pHGG patient-derived xenograft (PDX) lines under basal conditions. In response to ligand binding, nuclear levels of the receptor increased, and this was associated with the transcriptional upregulation of both the receptor and cyclin D1, suggesting that IGF-1R could regulate its own expression and cell cycle progression in these cells. Insulin-like growth factor-1 (IGF-1) increased the proliferation of the pHGG cells DIPG13 and SGJ2, and this could be blocked by the addition of the IGF-Trap. The IGF-Trap reduced the colony formation of these cells in an optimal growth medium and impeded the ability of IGF-1 to rescue DIPG13 cells from starvation-induced apoptosis. Collectively, these results implicate the IGF-1 axis in the regulation of cell cycle progression, cellular proliferation, and cell survival in pHGG, and identify the IGF-axis as a target and the IGF-Trap as a potential inhibitor of this axis in pHGG.
RESUMEN
Triple negative breast cancer (TNBC) is associated with unfavorable prognosis and high relapse rates following chemotherapy. There is an urgent need to develop effective targeted therapy for this BC subtype. The type I insulin-like growth factor receptor (IGF-IR) was identified as a potential target for BC management. We previously reported on the production of the IGF-Trap, a soluble IGF-1R fusion protein that reduces the bioavailability of circulating IGF-1 and IGF-2 to the cognate receptor, impeding signaling. In nude mice xenotransplanted with the human TNBC MDA-MB-231 cells, we found variable responses to this inhibitor. We used this model to investigate potential resistance mechanisms to IGF-targeted therapy. We show here that prolonged exposure of MDA-MB-231 cells to the IGF-Trap in vitro selected a resistant subpopulation that proliferated unhindered in the presence of the IGF-Trap. We identified in these cells increased fibroblast growth factor receptor 1 (FGFR1) activation levels that sensitized them to the FGFR1-specific tyrosine kinase inhibitor PD166866. Treatment with this inhibitor caused cell cycle arrest in both the parental and resistant cells, markedly increasing cell death in the latter. When combined with the IGF-Trap, an increase in cell cycle arrest was observed in the resistant cells. Moreover, FGFR1 silencing increased the sensitivity of these cells to IGF-Trap treatment in vivo. Our data identify increased FGFR1 signaling as a resistance mechanism to targeted inhibition of the IGF-IR and suggest that dual IGF-1R/FGFR1 blockade may be required to overcome TNBC cell resistance to IGF-axis inhibitors.
Asunto(s)
Resistencia a Antineoplásicos , Receptor IGF Tipo 1/metabolismo , Neoplasias de la Mama Triple Negativas/patología , Animales , Apoptosis/efectos de los fármacos , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Humanos , Ligandos , Ratones , Ratones Desnudos , Inhibidores de Proteínas Quinasas/farmacología , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/antagonistas & inhibidores , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Receptor IGF Tipo 1/antagonistas & inhibidores , Transducción de Señal/efectos de los fármacos , Trasplante Heterólogo , Neoplasias de la Mama Triple Negativas/metabolismoRESUMEN
Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is a rare but often lethal cancer that is diagnosed at a median age of 24 years. Optimal management of patients is not well defined, and current treatment remains challenging, necessitating the discovery of novel therapeutic approaches. The identification of SMARCA4-inactivating mutations invariably characterizing this type of cancer provided insights facilitating diagnostic and therapeutic measures against this disease. We show here that the BET inhibitor OTX015 acts in synergy with the MEK inhibitor cobimetinib to repress the proliferation of SCCOHT in vivo Notably, this synergy is also observed in some SMARCA4-expressing ovarian adenocarcinoma models intrinsically resistant to BETi. Mass spectrometry, coupled with knockdown of newly found targets such as thymidylate synthase, revealed that the repression of a panel of proteins involved in nucleotide synthesis underlies this synergy both in vitro and in vivo, resulting in reduced pools of nucleotide metabolites and subsequent cell-cycle arrest. Overall, our data indicate that dual treatment with BETi and MEKi represents a rational combination therapy against SCCOHT and potentially additional ovarian cancer subtypes.
Asunto(s)
Epigénesis Genética , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Nucleótidos/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Animales , Azetidinas/farmacología , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Sinergismo Farmacológico , Epigénesis Genética/efectos de los fármacos , Femenino , Técnicas de Silenciamiento del Gen , Células HEK293 , Humanos , Ratones Endogámicos NOD , Ratones SCID , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Piperidinas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Fase S/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Plasticity of neoplasia, whereby cancer cells attain stem-cell-like properties, is required for disease progression and represents a major therapeutic challenge. We report that in breast cancer cells NANOG, SNAIL and NODAL transcripts manifest multiple isoforms characterized by different 5' Untranslated Regions (5'UTRs), whereby translation of a subset of these isoforms is stimulated under hypoxia. The accumulation of the corresponding proteins induces plasticity and "fate-switching" toward stem cell-like phenotypes. Mechanistically, we observe that mTOR inhibitors and chemotherapeutics induce translational activation of a subset of NANOG, SNAIL and NODAL mRNA isoforms akin to hypoxia, engendering stem-cell-like phenotypes. These effects are overcome with drugs that antagonize translational reprogramming caused by eIF2α phosphorylation (e.g. ISRIB), suggesting that the Integrated Stress Response drives breast cancer plasticity. Collectively, our findings reveal a mechanism of induction of plasticity of breast cancer cells and provide a molecular basis for therapeutic strategies aimed at overcoming drug resistance and abrogating metastasis.
Asunto(s)
Regiones no Traducidas 5'/genética , Neoplasias de la Mama/genética , Regulación Neoplásica de la Expresión Génica/genética , Biosíntesis de Proteínas/genética , Isoformas de ARN/genética , Antineoplásicos/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Hipoxia de la Célula , Línea Celular Tumoral , Factor 2 Eucariótico de Iniciación/genética , Factor 2 Eucariótico de Iniciación/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Células MCF-7 , Proteína Homeótica Nanog/genética , Proteína Nodal/genética , Fosforilación/efectos de los fármacos , Factores de Transcripción de la Familia Snail/genéticaRESUMEN
Estrogen receptor alpha (ERα) activity is associated with increased cancer cell proliferation. Studies aiming to understand the impact of ERα on cancer-associated phenotypes have largely been limited to its transcriptional activity. Herein, we demonstrate that ERα coordinates its transcriptional output with selective modulation of mRNA translation. Importantly, translational perturbations caused by depletion of ERα largely manifest as "translational offsetting" of the transcriptome, whereby amounts of translated mRNAs and corresponding protein levels are maintained constant despite changes in mRNA abundance. Transcripts whose levels, but not polysome association, are reduced following ERα depletion lack features which limit translation efficiency including structured 5'UTRs and miRNA target sites. In contrast, mRNAs induced upon ERα depletion whose polysome association remains unaltered are enriched in codons requiring U34-modified tRNAs for efficient decoding. Consistently, ERα regulates levels of U34-modifying enzymes and thereby controls levels of U34-modified tRNAs. These findings unravel a hitherto unprecedented mechanism of ERα-dependent orchestration of transcriptional and translational programs that may be a pervasive mechanism of proteome maintenance in hormone-dependent cancers.
Asunto(s)
Neoplasias de la Mama/genética , Receptor alfa de Estrógeno/genética , Regulación Neoplásica de la Expresión Génica , Polirribosomas/genética , Biosíntesis de Proteínas , ARN Mensajero/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Proliferación Celular , Receptor alfa de Estrógeno/metabolismo , Femenino , Humanos , Células MCF-7 , Polirribosomas/metabolismo , ARN Mensajero/metabolismo , Transducción de Señal , Activación TranscripcionalRESUMEN
Dysregulation of histone modifications promotes carcinogenesis by altering transcription. Breast cancers frequently overexpress the histone methyltransferase EZH2, the catalytic subunit of Polycomb Repressor Complex 2 (PRC2). However, the role of EZH2 in this setting is unclear due to the context-dependent functions of PRC2 and the heterogeneity of breast cancer. Moreover, the mechanisms underlying PRC2 overexpression in cancer are obscure. Here, using multiple models of breast cancer driven by the oncogene ErbB2, we show that the tyrosine kinase c-Src links energy sufficiency with PRC2 overexpression via control of mRNA translation. By stimulating mitochondrial ATP production, c-Src suppresses energy stress, permitting sustained activation of the mammalian/mechanistic target of rapamycin complex 1 (mTORC1), which increases the translation of mRNAs encoding the PRC2 subunits Ezh2 and Suz12. We show that Ezh2 overexpression and activity are pivotal in ErbB2-mediated mammary tumourigenesis. These results reveal the hitherto unknown c-Src/mTORC1/PRC2 axis, which is essential for ErbB2-driven carcinogenesis.
Asunto(s)
Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Epigénesis Genética , Complejo Represivo Polycomb 2/genética , Receptor ErbB-2/metabolismo , Familia-src Quinasas/metabolismo , Adenosina Trifosfato/metabolismo , Adulto , Animales , Neoplasias de la Mama/patología , Proteína Tirosina Quinasa CSK , Carcinogénesis , Línea Celular Tumoral , Proteína Potenciadora del Homólogo Zeste 2/genética , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Femenino , Humanos , Glándulas Mamarias Humanas/metabolismo , Glándulas Mamarias Humanas/patología , Diana Mecanicista del Complejo 1 de la Rapamicina/genética , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Ratones , Ratones Endogámicos NOD , Ratones Transgénicos , Persona de Mediana Edad , Mitocondrias/genética , Mitocondrias/metabolismo , Complejo Represivo Polycomb 2/metabolismo , Biosíntesis de Proteínas , Receptor ErbB-2/genética , Familia-src Quinasas/genéticaRESUMEN
There is increasing interest in therapeutically exploiting metabolic differences between normal and cancer cells. We show that kinase inhibitors (KIs) and biguanides synergistically and selectively target a variety of cancer cells. Synthesis of non-essential amino acids (NEAAs) aspartate, asparagine, and serine, as well as glutamine metabolism, are major determinants of the efficacy of KI/biguanide combinations. The mTORC1/4E-BP axis regulates aspartate, asparagine, and serine synthesis by modulating mRNA translation, while ablation of 4E-BP1/2 substantially decreases sensitivity of breast cancer and melanoma cells to KI/biguanide combinations. Efficacy of the KI/biguanide combinations is also determined by HIF-1α-dependent perturbations in glutamine metabolism, which were observed in VHL-deficient renal cancer cells. This suggests that cancer cells display metabolic plasticity by engaging non-redundant adaptive mechanisms, which allows them to survive therapeutic insults that target cancer metabolism.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Resistencia a Antineoplásicos , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Neoplasias , Transducción de Señal/efectos de los fármacos , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Aminoácidos/metabolismo , Animales , Biguanidas/farmacología , Proteínas de Ciclo Celular , Factores Eucarióticos de Iniciación/metabolismo , Regulación Neoplásica de la Expresión Génica , Células HCT116 , Humanos , Células K562 , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Ratones , Ratones Desnudos , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Fosfoproteínas/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , ARN Mensajero/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Dysregulated mRNA translation and aberrant energy metabolism are frequent in cancer. Considering that mRNA translation is an energy demanding process, cancer cells must produce sufficient ATP to meet energy demand of hyperactive translational machinery. In recent years, the mammalian/mechanistic target of rapamycin (mTOR) emerged as a central regulatory node which coordinates energy consumption by the translation apparatus and ATP production in mitochondria. Aberrant mTOR signaling underpins the vast majority of cancers whereby increased mTOR activity is thought to be a major determinant of both malignant translatomes and metabolomes. Nonetheless, the role of mTOR and other related signaling nodes (e.g. AMPK) in orchestrating protein synthesis and cancer energetics is only recently being unraveled. In this review, we discuss recent findings that provide insights into the molecular underpinnings of coordination of translational and metabolic programs of cancer cells, and potential strategies to translate these findings into clinical treatments.
Asunto(s)
Metabolismo Energético , Proteínas de Neoplasias/biosíntesis , Neoplasias/metabolismo , Biosíntesis de Proteínas , ARN Mensajero/metabolismo , ARN Neoplásico/metabolismo , Serina-Treonina Quinasas TOR/biosíntesis , Animales , Humanos , Proteínas de Neoplasias/genética , Neoplasias/genética , Neoplasias/patología , ARN Mensajero/genética , ARN Neoplásico/genética , Serina-Treonina Quinasas TOR/genéticaRESUMEN
We report that the temperature-sensitive (ts) phenotype in Saccharomyces cerevisiae associated with a variant tRNA nucleotidyltransferase containing an amino acid substitution at position 189 results from a reduced ability to incorporate AMP and CMP into tRNAs. We show that this defect can be compensated for by a second-site suppressor converting residue arginine 64 to tryptophan. The R64W substitution does not alter the structure or thermal stability of the enzyme dramatically but restores catalytic activity in vitro and suppresses the ts phenotype in vivo. R64 is found in motif A known to be involved in catalysis and nucleotide triphosphate binding while E189 lies within motif C previously thought only to connect the head and neck domains of the protein. Although mutagenesis experiments indicate that residues R64 and E189 do not interact directly, our data suggest a critical role for residue E189 in enzyme structure and function. Both R64 and E189 may contribute to the organization of the catalytic domain of the enzyme. These results, along with overexpression and deletion analyses, show that the ts phenotype of cca1-E189F does not arise from thermal instability of the variant tRNA nucleotidyltransferase but instead from the inability of a partially active enzyme to support growth only at higher temperatures.
Asunto(s)
Arginina/química , Ácido Aspártico/química , ARN Nucleotidiltransferasas/química , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/química , Triptófano/química , Adenosina Monofosfato/química , Secuencias de Aminoácidos , Sustitución de Aminoácidos , Arginina/genética , Ácido Aspártico/genética , Dominio Catalítico , Citidina Monofosfato/química , Calor , Simulación de Dinámica Molecular , Datos de Secuencia Molecular , Fenotipo , Estructura Secundaria de Proteína , ARN Nucleotidiltransferasas/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Alineación de Secuencia , Triptófano/genéticaRESUMEN
tRNA-NTs (tRNA nucleotidyltransferases) are required for the maturation or repair of tRNAs by ensuring that they have an intact cytidine-cytidine-adenosine sequence at their 3'-termini. Therefore this enzymatic activity is found in all cellular compartments, namely the nucleus, cytoplasm, plastids and mitochondria, in which tRNA synthesis or translation occurs. A single gene codes for tRNA-NT in plants, suggesting a complex targeting mechanism. Consistent with this, distinct signals have been proposed for plastidic, mitochondrial and nuclear targeting. Our previous research has shown that in addition to N-terminal targeting information, the mature domain of the protein itself modifies targeting to mitochondria and plastids. This suggests the existence of an as yet unknown determinate for the distribution of dual-targeted proteins between these two organelles. In the present study, we explore the enzymatic and physicochemical properties of tRNA-NT variants to correlate the properties of the enzyme with the intracellular distribution of the protein. We show that alteration of tRNA-NT stability influences its intracellular distribution due to variations in organelle import capacities. Hence the fate of the protein is determined not only by the transit peptide sequence, but also by the physicochemical properties of the mature protein.