RESUMEN

Glioblastoma multiforme (GBM) is the most common primary malignant brain tumor in adults with a poor prognosis. Despite significant progress in drug development, the blood-brain barrier (BBB) continues to limit the use of novel chemotherapeutics. Thus, our attention has been focused on the design, synthesis, and testing of small-molecule anticancer agents that are able to penetrate the BBB. One such compound is the D-glucose analog, 2-deoxy-D-glucose (2-DG), which inhibits glycolysis and induces GBM cell death. 2-DG has already been tested in clinical trials but was not approved as a drug, in part due to inadequate pharmacokinetics. To improve the pharmacokinetic properties of 2-DG, a series of novel derivatives was synthesized. Herein, we report the biological effects of WP1234, a 2-ethylbutyric acid 3,6-diester of 2-DG that can potentially release 2-ethylbutyrate and 2-DG inside the cells when metabolized. Using biochemical assays and examining cell viability, proliferation, protein synthesis, and apoptosis induction, we assessed the cytotoxic potential of WP1234. WP1234 significantly reduced the viability of GBM cells in a dose- and time-dependent manner. The lactate and ATP synthesis assays confirmed the inhibition of glycolysis elicited by released 2-DG. Furthermore, an evaluation of histone deacetylases (HDAC) activity revealed that the 2-ethylbutyrate action resulted in HDAC inhibition. Overall, these results demonstrated that WP1234 is a bifunctional molecule with promising anticancer potential. Further experiments in animal models and toxicology studies are needed to evaluate the efficacy and safety of this new 2-DG derivative.

RESUMEN

Interleukin (IL)-6, a known proinflammatory cytokine, is released in both visceral adipose tissue and contracting skeletal muscle. In this study, we used microRNA profiling as a screening method to identify miRNA species modified by IL-6 treatment in mouse 3T3-L1 adipocytes. miRNA microarray analysis and qRT-PCR revealed increased expression of miR-146b-3p in adipocytes exposed to IL-6 (1 ng/ml) during 8-day differentiation. On the basis of ontological analysis of potential targets, selected proteins associated with cytoskeleton and transport were examined in the context of adipocyte response to insulin, using immunofluorescence and confocal microscopy. We concluded that IL-6: (i) does not affect insulin action on actin cellular distribution; (ii) modulates the effect of insulin on myosin light chain kinase (Mylk) distribution by preventing its shift toward cytoplasm; (iii) mimics the effect of insulin on dynein distribution by increasing its near-nuclear accumulation; (iv) mimics the effect of insulin on glucose transporter Glut4 distribution, especially by increasing its near-nuclear accumulation; (v) supports insulin action on early endosome marker Rab4A near-nuclear accumulation. Moreover, as IL-6 did not disturb insulin-dependent glucose uptake, our results do not confirm the IL-6-induced impairment of insulin action observed in some in vitro studies, suggesting that the effect of IL-6 is dose dependent.

Asunto(s)

Interleucina-6 , MicroARNs , Células 3T3-L1 , Adipocitos/metabolismo , Animales , Proteínas del Citoesqueleto/metabolismo , Glucosa/metabolismo , Transportador de Glucosa de Tipo 4/metabolismo , Insulina/metabolismo , Insulina/farmacología , Interleucina-6/metabolismo , Ratones , MicroARNs/metabolismo

RESUMEN

Our concerns have been raised about whether prolonged exposure to ammunition-related chemicals could correlate with genomic instability predisposing to lung carcinogenesis. The group of professional soldiers engaged in routine ammunition analysis and its explosive properties testing. To assess the presence of an innate genetic profile, DNA isolated from swabs was analyzed with LungCarta and HS Lung Panels and MassARRAY Analyzer 4 mass spectrometry. The presence of new somatic single nucleotide polymorphisms (SNPs) was detected with liquid biopsy technique and circulating cell-free DNA (ccfDNA) isolation, which was further analyzed with UltraSeek Lung Panel. Both genetic profiles obtained for each individual were compared. Genetic analysis revealed that in professional soldiers with long-term exposure to ammunition-related toxic chemicals, new SNPs in genes related to lung carcinogenesis are present. The preliminary data indicate that using modern molecular techniques could be a valuable tool for monitoring the genome instability in context of occupational risk of harmful volatile toxic compounds and improving personnel safety. The analyzed group will be further enlarged, and follow-up studies will be continued.

Asunto(s)

Inestabilidad Genómica , Humanos

RESUMEN

Over the last decade, we have seen tremendous progress in research on 2-deoxy-D-glucose (2-DG) and its analogs. Clinical trials of 2-DG have demonstrated the challenges of using 2-DG as a monotherapy, due to its poor drug-like characteristics, leading researchers to focus on improving its bioavailability to tissue and organs. Novel 2-DG analogs such as WP1122 and others have revived the old concept of glycolysis inhibition as an effective anticancer strategy. Combined with other potent cytotoxic agents, inhibitors of glycolysis could synergistically eliminate cancer cells. We focused our efforts on the development of new combinations of anticancer agents coupled with 2-DG and its derivatives, targeting glioblastoma, which is in desperate need of novel approaches and therapeutic options and is particularly suited to glycolysis inhibition, due to its reliance on aerobic glycolysis. Herein, we present evidence that a combined treatment of 2-DG analogs and modulation of histone deacetylases (HDAC) activity via HDAC inhibitors (sodium butyrate and sodium valproate) exerts synergistic cytotoxic effects in glioblastoma U-87 and U-251 cells and represents a promising therapeutic strategy.

RESUMEN

Muscle cachexia is one of the most critical unmet medical needs. Identifying the molecular background of cancer-induced muscle loss revealed a promising possibility of new therapeutic targets and new drug development. In this review, we will define the signal transducer and activator of transcription 3 (STAT3) protein's role in the tumor formation process and summarize the role of STAT3 in skeletal muscle cachexia. Finally, we will discuss a vast therapeutic potential for the STAT3-inhibiting single-agent treatment innovation that, as the desired outcome, could block tumor growth and generally prevent muscle cachexia.

Asunto(s)

Caquexia/tratamiento farmacológico , Janus Quinasa 2 , Terapia Molecular Dirigida/métodos , Atrofia Muscular/tratamiento farmacológico , Factor de Transcripción STAT3 , Animales , Caquexia/etiología , Caquexia/metabolismo , Humanos , Janus Quinasa 2/antagonistas & inhibidores , Janus Quinasa 2/metabolismo , Janus Quinasa 2/fisiología , Terapia Molecular Dirigida/tendencias , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Atrofia Muscular/etiología , Atrofia Muscular/metabolismo , Neoplasias/complicaciones , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Neoplasias/patología , Fosforilación/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Factor de Transcripción STAT3/antagonistas & inhibidores , Factor de Transcripción STAT3/metabolismo , Factor de Transcripción STAT3/fisiología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología

RESUMEN

Interleukin (IL)-6 is a proinflammatory cytokine released in injured and contracting skeletal muscles. In this study, we examined cellular expression of proteins associated with cytoskeleton organization and cell migration, chosen on the basis of microRNA profiling, in rat primary skeletal muscle cells (RSkMC) treated with IL-6 (1 ng/ml) for 11 days. MiRNA microarray analysis and qRT-PCR revealed increased expression of miR-154-3p and miR-338-3p in muscle cells treated with IL-6. Pacsin3 was downregulated post-transcriptionally by IL-6, but not by IGF-I. Ephrin4A protein was increased both in IL-6- and IGF-I-treated myocytes. IL-6, but not IGF-I, stimulated migratory ability of RSkMC, examined in wound healing assay. Alpha-actinin protein was slightly augmented in RSKMC treated with IL-6, similarly to IGF-I. IL-6, but not IGF-I, upregulated desmin in differentiating RSkMC. IL-6 supplementation caused accumulation of alpha-actinin and desmin in near-nuclear area of muscle cells, which was manifested by increased ratio: mean near-nuclear fluorescence/mean peripheral cytoplasm fluorescence of these proteins. We concluded that IL-6, a known proinflammatory cytokine and a physical activity-associated myokine, acting during differentiation of primary skeletal muscle cells, alters expression of nonmuscle-specific miRNAs. This cytokine causes differential effects on pacsin-3 and ephrinA4, through post-transcriptional inhibition and stimulation, respectively. IL-6-exerted modifications of cytoskeletal proteins in muscle cells include both transcriptional (desmin and dynein heavy chain 5) and post-transcriptional activation (alpha-actinin). Moreover, IL-6 augments near-nuclear distribution of cytoskeletal proteins, alpha-actinin and desmin and promotes migration of myocytes. Such effects suggest that IL-6 plays a role during skeletal muscle regeneration, acting through mechanisms independent of regulation of myogenic program.

Asunto(s)

Proteínas Adaptadoras Transductoras de Señales/biosíntesis , Efrina-A4/biosíntesis , Interleucina-6/farmacología , Mioblastos Esqueléticos/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Diferenciación Celular/efectos de los fármacos , Proteínas del Citoesqueleto/biosíntesis , Proteínas del Citoesqueleto/genética , Modelos Animales de Enfermedad , Efrina-A4/genética , Factor I del Crecimiento Similar a la Insulina/farmacología , MicroARNs/biosíntesis , MicroARNs/genética , MicroARNs/metabolismo , Mioblastos Esqueléticos/citología , Mioblastos Esqueléticos/efectos de los fármacos , Procesamiento Postranscripcional del ARN , Ratas , Proteínas Recombinantes/farmacología , Transcripción Genética

RESUMEN

Interleukin (IL)-8 is released both in visceral adipose tissue and in contracting skeletal muscles. In this study, we examined cellular pathways associated with muscle hypertrophy, chosen on the basis of microRNA profiling, in differentiating rat primary skeletal muscle cells (RSkMC) treated with IL-8 (1 ng/ml) for 11 days. IL-8 increased myocilin expression, Akt phosphorylation, FoxO3 dispersion throughout the cytoplasm, and reduced FoxO3 level. IL-8 decreased the expression of atrogin and MuRF1 and increased myotube length and diameter. We concluded that IL-8 present in extracellular environment of myoblasts induced to differentiation stimulates expression of myocilin, a protein important for skeletal muscle hypertrophy. This phenomenon was associated with: (a) activation of myogenic transcription, (b) increased phosphorylation and activation of PKB/Akt, leading to (c) cytoplasm distribution and degradation of a transcription factor FoxO3, (d) decreased expression of gene markers of proteolysis, atrogin and Murf1, and (e) increased myotube length and diameter. In this regard, IL-8 affects skeletal muscle cells similarly to IGF-I and can be considered as a potent anticatabolic factor for skeletal muscle.

Asunto(s)

Proteínas del Citoesqueleto/genética , Proteínas del Ojo/genética , Proteína Forkhead Box O3/genética , Glicoproteínas/genética , Interleucina-8/genética , Desarrollo de Músculos/genética , Músculo Esquelético/metabolismo , Animales , Diferenciación Celular/genética , Factor I del Crecimiento Similar a la Insulina/genética , Fibras Musculares Esqueléticas/metabolismo , Proteínas Musculares/genética , Mioblastos/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Ratas , Transducción de Señal , Proteínas de Motivos Tripartitos/genética , Ubiquitina-Proteína Ligasas/genética

RESUMEN

Microarray-based transcriptomic profiling enables simultaneous measurement of expression of multiple genes from one biological sample. Here we describe a detailed protocol, which serves to examine global gene expression using whole genome oligonucleotide microarrays. We also provide examples of bioinformatics tools, which are helpful in analyses and interpretation of microarray data, and propose further biological assays, to warrant conclusions drawn from transcriptomic signature.

Asunto(s)

Perfilación de la Expresión Génica , Desarrollo de Músculos/genética , Transcriptoma , Biología Computacional/métodos , Perfilación de la Expresión Génica/métodos , Ontología de Genes , Redes Reguladoras de Genes , Humanos , Anotación de Secuencia Molecular , Análisis de Secuencia por Matrices de Oligonucleótidos , Reproducibilidad de los Resultados , Programas Informáticos , Flujo de Trabajo