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BACKGROUND AND OBJECTIVE: Human epidermal growth factor receptor 2 (HER2) protein overexpression is one of the most significant biomarkers for breast cancer diagnostics, treatment prediction, and prognostics. The high accessibility of HER2 inhibitors in routine clinical practice directly translates into the diagnostic need for precise and robust marker identification. Even though multigene next-generation sequencing methodologies have slowly taken over the field of single-biomarker molecular tests, the copy number alterations such as amplification of the HER2-coding ERBB2 gene are hard to validate on next-generation sequencing platforms as they are characterized by chromosomal structural heterogeneity, polysomy, and genomic context of ploidy. In our study, we tested the approach of using whole genome sequencing instead of next-generation sequencing panels to determine HER2 status in the clinical set-up. METHODS: We used a large dataset of 876 patients with breast cancer whole genomes with curated clinical data and an additional set of 551 patients' external genomic data. We used the decision-tree-based algorithm for optimization of the diagnostic tool for HER2 status assessment by whole genome sequencing. RESULTS: The most efficient approach to assess HER2 status in whole genome sequencing data was the ploidy-corrected copy number, utilizing ERBB2 copy number and mean tumor ploidy. The classifier achieved sensitivity of 91.18% and specificity of 98.69% on the internal validation dataset and 89.86% and 96.06% on the external data, which is similar to other next-generation sequencing methods, currently tested in the clinic. CONCLUSIONS: We provide evidence that the HER2 status may be reliably determined by whole genome sequencing and is applicable across different laboratory protocols and pipelines. We suggest using the ploidy-corrected copy number for diagnostic purposes.
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Neoplasias da Mama , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Variações do Número de Cópias de DNA , Feminino , Genes erbB-2 , Humanos , Ploidias , Receptor ErbB-2/genética , Sequenciamento Completo do GenomaRESUMO
Essential and non-essential elements deficiencies may lead to various birth complications. The aim of this paper was to determine calcium (Ca), copper (Cu), iron (Fe), potassium (K), magnesium (Mg), sodium (Na), phosphorus (P), lead (Pb), strontium (Sr), and zinc (Zn) concentrations in maternal blood and cord blood. Whole blood and cord blood samples collected from pregnant women (n = 136) were analyzed for the concentration of the elements by spectrophotometric atomic absorption in inductively coupled argon plasma (ICP-OES). The results showed that Ca, Pb, and Sr concentrations were similar in maternal and cord blood, while Fe and K levels were higher in cord blood than in maternal blood. The cord blood Cu, Na, and Zn concentrations were lower than those in maternal blood, suggesting transplacental transfer of these elements were limited. Moreover, checking the influence of studied elements on the anthropometric parameters of the newborns, we found that the highest number of associations was between Cu in cord blood. Due to the fact that the pregnant women were healthy, and the newborns were without any disorders, we suggest that the values obtained in our study are normal values of studied elements in whole blood and cord blood in patients from Poland.
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Sangue Fetal , Oligoelementos , Cobre , Feminino , Humanos , Recém-Nascido , Magnésio , Gravidez , ZincoRESUMO
Newborns, regardless of the method of termination of pregnancy, are exposed to the first exogenous stress factors during delivery. The purpose of the study was to evaluate the differences in newborns' thermal response to vaginal (VD) vs caesarean section (CS) delivery. The temperature was measured during the first minutes of life within 122 healthy full-term newborns, on the forehead, chest and upper-back by infrared camera (FLIR T1030sc HD). The lowest temperatures were recorded in the forehead of VD newborns (significantly difference with CS; p < 0.001), the warmest was the chest. A significant correlation was found between the duration of the second stage of natural childbirth and surface temperature and pO2 in the newborn blood. The temperatures of selected body surface areas correlate highly positively, regardless of the mode of delivery. In the case of healthy neonates, with normal birth weight and full-term, VD creates more favourable conditions stimulating the mechanisms of adaptation for a newborn than CS.
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Temperatura Corporal , Cesárea , Parto Obstétrico , Adolescente , Adulto , Feminino , Humanos , Masculino , Consumo de Oxigênio , Parto , Gravidez , Adulto JovemRESUMO
<b>Introduction: </b>Pressure ulcers and their consequences can occur in any patient regardless of the underlying disease, so the knowledge of their prevention and treatment is extremely important. <br><b>The aim of the study:</b> The aim of the study was to analyze the state of students' knowledge on pressure ulcers.<br><b> Material and methods:</b> The research was carried out in the Pomeranian Medical University in Szczecin among students of nursing. The study involved 203 full-time students. Data was collected using a questionnaire consisting of 27 questions. The questions addressed the issues of the formation, prevention and treatment of pressure ulcers. The Kolmogorow-Smirnow, Fisher-Snedecor test, (ANOVA) with Spearman's correlation and Student's t-test were applied for statistical analysis. The statistical significance was assumed to be p ≤ 0.05. <br><b>Results:</b> Over half of the respondents (57.64%) had a sufficient level of knowledge on pressure ulcers prevention and treatment. The level increased with the duration of the study, the highest was in the last few years, both I and II cycle of studies. The better (the higher) subjects assessed their knowledge about prevention and treatment of pressure ulcers, they were characterized by a higher level of knowledge. <br><b>Conclusions:</b> The knowledge of nursing students on pressure ulcers increases with subsequent years of study. The selfassessment of students' knowledge about pressure ulcers, their prevention and treatment is accurate. Students of the consecutive years are better prepared to carry out anti-bedsore prevention. Students are aware of the need to have knowledge, as well as practice in the field of prevention and treatment of pressure ulcers.
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Bacharelado em Enfermagem/métodos , Avaliação Educacional/métodos , Conhecimentos, Atitudes e Prática em Saúde , Úlcera por Pressão/enfermagem , Higiene da Pele/enfermagem , Estudantes de Enfermagem/estatística & dados numéricos , Feminino , Humanos , Masculino , Guias de Prática Clínica como AssuntoRESUMO
Background: Preterm birth is the most frequent cause of neonatal death, but its aetiology remains unclear. It has been suggested that the imbalance of immunological mechanisms responsible for maintaining pregnancy is contributing to preterm birth pathogenesis. We aimed to investigate global gene expression and the levels of several complement system components in umbilical cord blood samples from preterm neonates and compare them to term newborns. We sought to examine how differentially expressed genes could affect various immune-related pathways that are believed to be crucial factors in preterm birth. Material and methods: We enrolled 27 preterm infants (<37 weeks GA) and 52 term infants (>37 weeks GA), from which umbilical cord blood samples were collected. From these samples, peripheral blood mononuclear cells were isolated and subsequent RNA isolation was performed. We used Affymetrix Human Gene 2.1 ST Array Strip for microarray experiment and DAVID resources for bioinformatics analysis of the obtained data. Concentrations of C2, C3a, C5/C5a, C9, FactorD, Properdin were measured in umbilical cord blood plasma samples using multiplex fluorescent bead-based immunoassays using Luminex technology. Results: The levels of C3a and C5/5a were significantly elevated in preterm neonates compared to term babies, whereas C9 concentration was evidently increased in term infants. The expression of 250 genes was upregulated at least 2-fold and 3781 genes were downregulated at least 2-fold in preterm neonates in comparison with term infants. Functional annotation analysis revealed that in preterm infants in comparison to term babies there was a significant downregulation of genes encoding several Toll-like receptors, interleukins and genes involved in major signalling pathways (e.g. NF-κB, MAPK, TNF, Notch, JAK) and vital cellular processes (e.g. intracellular signal transduction, protein ubiquitination, protein transport, RNA splicing, DNA-templated transcription). Conclusions: Preterm birth results in immediate and long-term complications. Our results indicate that infants born prematurely show significant differences in complement components concentration and a downregulation of over 3,000 genes, involved mainly in various immune-related pathways, including innate immune response, phagocytosis and TLR function, when compared to full-term babies. Further studies on larger cohorts are needed to elucidate the role of immunity in prematurity.
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Sangue Fetal/metabolismo , Imunidade Inata/genética , Nascimento Prematuro/genética , Nascimento a Termo/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Idade Gestacional , Humanos , Lactente , Recém-Nascido , Recém-Nascido Prematuro/crescimento & desenvolvimento , Recém-Nascido Prematuro/metabolismo , Leucócitos Mononucleares/metabolismo , Masculino , NF-kappa B/genética , Gravidez , Nascimento Prematuro/patologia , Transdução de Sinais/genéticaRESUMO
Triptycene derivatives are widely utilized in different fields of chemistry and materials sciences. Their physicochemical properties, often of pivotal importance for the rational design of triptycene-based functional materials, are influenced by noncovalent interactions between substituents mounted on the triptycene skeleton. Herein, a unique interaction between electron-rich substituents in the peri position and the silyl group located on the bridgehead sp3 -carbon is discussed on the example of 1,4-dichloro-9-(p-methoxyphenyl)-silyltriptycene (TRPCl) which exists in solution in the form of two rotamers differing by dispositions, syn or anti, of the Si-CPh (the CPh atom is from the p-methoxyphenyl group) bond against the peri-Cl atom. For the first time, substantial differences between the Si-CPh bonds in these two dispositions are identified, based on indirect experimental and direct theoretical evidence. For these two orientations, the experimental 1 J(Si,CPh ) values differ by as much as 10 percent. The differences are explained in terms of effective electron density transfer from the peri-Cl atom to the antibonding σ* orbitals of the Si-X bonds (X=H, CPh ) oriented anti to that atom. The electronic effects are revealed by an NBO analysis. Connections of these observations with the notion of blue-shifting hydrogen bonds are discussed.
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The aim of the study was to investigate relationships between the concentrations of macroelements (Ca), microelements (Cr, Cu, Fe, Mn, Mo, Ni, Sn, Sr, V, Zn) and heavy metals (Ag, Cd, Pb) in the placenta, fetal membrane and umbilical cord. âªFurthermore, we examined relationships between the concentrations of these metals in the studied afterbirths and maternal age, gestational age, placenta parameters (breadth, length, weight) and newborn parameters (length, weight and Apgar score). This study confirms previously reported Zn-Cd, Pb-Cd and Ni-Pb interactions in the placenta. New types of interactions in the placenta, fetal membrane and umbilical cord were also noted. Analysis of the correlations between metal elements in the afterbirths (placenta, fetal membrane and umbilical cord) and biological parameters showed the following relationships: maternal age and Mn (in the fetal membrane); gestational age and Cr, Fe, Zn (in the fetal membrane), Ag and Cu (in the umbilical cord); newborn's length and Sr (in the placenta), Ag (in the umbilical cord); newborn's weight and Sr (in the placenta), Cu (in the fetal membrane), Ag (in the umbilical cord); Apgar score and Ca, Cr and Ni (in the umbilical cord); placenta's length and Cr and Sn (in the fetal membrane), Cu (in the umbilical cord); placenta's width and Mo, Pb (in the placenta) and placenta weight and Sr (in the placenta), Ag, Fe, Mn (in the fetal membrane). The results show the influence of metals on the placenta, mother and newborn parameters, and the same point indicates the essential trace elements during the course of pregnancy.
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Membranas Extraembrionárias/metabolismo , Metais Pesados/metabolismo , Placenta/metabolismo , Cordão Umbilical/metabolismo , Feminino , Idade Gestacional , Humanos , Recém-Nascido , Idade Materna , GravidezRESUMO
The aim of the study was to perform an evaluation of chosen body surface temperatures in neonates immediately after birth, and to seek a relationship between those temperatures and the factors related both to the mother and newborn. The study included 74 healthy newborns. Maternal age, body weight, body mass index before pregnancy and on delivery day, birth and pregnancy order, newborn sex, birth weight, body length, pregnancy week on delivery, as well as newborn gasometric test results were collected. The highest temperature values were observed in the chest of the newborn. Significant relationships between the temperature of the evaluated areas were found. The parameters that correlated positively with the temperature of the back region were maternal body weight (both before pregnancy and on delivery day) as well as weight gain during pregnancy. The core and surface temperatures of the body are one of the most important elements of neonatal homeostasis and any changes constitute a risk to the newborn's health. It seemed that according to the surface temperature, the most important area that must be evaluated is the neonate's back, as it is most affected by appropriate weight gain during pregnancy.
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Peso ao Nascer , Temperatura Corporal/fisiologia , Recém-Nascido , Estatura , Índice de Massa Corporal , Feminino , Idade Gestacional , Humanos , Masculino , Idade Materna , Projetos Piloto , Gravidez , Aumento de PesoRESUMO
The understanding of the molecular and biochemical characteristics of the human leukocyte antigen-G (HLA-G) is important because of the diverse influence of this antigen's polymorphisms on the course of a pregnancy. The aim of our study was to assess how the variation of the HLA-G allele and the HLA-G 14-bp ins/del polymorphism influence predisposition to a complicated pregnancy. The clinical material consisted of parental pairs with complicated pregnancies (210 women; 190 men). The control group included parental pairs without complications during pregnancy (89 women; 86 men). The study involved isolation of genome DNA from peripheral blood leukocytes, sequencing, and analysis of the 14-bp ins/del polymorphism in the 3'-untranslated region (3'-UTR) of the HLA-G gene based on polymerase chain reaction (PCR). The most common HLA-G allele in the group of women with complicated pregnancies was the HLA-G 10101 allele. There were no statistically significant differences in the frequencies of the 14-bp ins/del polymorphism in the 3'UTR of the HLA-G gene between the groups. Our results suggest that the risk of complications in pregnancy is influenced by the HLA-G 10101, HLA-G 10108, and HLA-G 10106 alleles and is not influenced by the 14-bp ins/del polymorphism in the 3'UTR of the HLA-G gene.
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Antígenos HLA-G/genética , Complicações na Gravidez/genética , Gravidez de Alto Risco/genética , Regiões 3' não Traduzidas , Adulto , Alelos , Estudos de Casos e Controles , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Masculino , Polimorfismo Genético , GravidezRESUMO
Use of chimeric antigen receptors (CARs) as the basis of targeted adoptive T cell therapies has enabled dramatic efficacy against multiple hematopoietic malignancies, but potency against bulky and solid tumors has lagged, potentially due to insufficient CAR-T cell expansion and persistence. To improve CAR-T cell efficacy, we utilized a potent activation switch based on rimiducid-inducible MyD88 and CD40 (iMC)-signaling elements. To offset potential toxicity risks by this enhanced CAR, an orthogonally regulated, rapamycin-induced, caspase-9-based safety switch (iRC9) was developed to allow in vivo elimination of CAR-T cells. iMC costimulation induced by systemic rimiducid administration enhanced CAR-T cell proliferation, cytokine secretion, and antitumor efficacy in both in vitro assays and xenograft tumor models. Conversely, rapamycin-mediated iRC9 dimerization rapidly induced apoptosis in a dose-dependent fashion as an approach to mitigate therapy-related toxicity. This novel, regulatable dual-switch system may promote greater CAR-T cell expansion and prolonged persistence in a drug-dependent manner while providing a safety switch to mitigate toxicity concerns.
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The aim of the study was to evaluate Hg and Se concentrations and Se:Hg molar ratios in the placenta, umbilical cord and fetal membranes, and to examine the relationship between the concentrations of the elements and selected factors. The study material consisted of the placenta, umbilical cord and fetal membranes obtained from 91 healthy women from northwestern and central Poland. In our study mean Hg and Se concentrations in afterbirth were ~ 0.01 mg/kg dry weight (dw) and ≤ 0.5 mg/kg dw, respectively. Correlation analysis showed negative relationships between placenta weight and Se concentration in the placenta and umbilical cord, as well as between placenta length and Se levels in the umbilical cord. We found negative correlations between THg concentration in the placenta and birth weight and between Se concentration in the placenta and umbilical cord and the morphological parameters of the placenta. Furthermore, we noted new types of interactions in specific parts of the afterbirth. In our study, Se:THg molar ratios ranged from 5 to 626; these values indicate protection against Hg toxicity.
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Membranas Extraembrionárias/metabolismo , Mercúrio/metabolismo , Placenta/metabolismo , Selênio/metabolismo , Cordão Umbilical/metabolismo , Adulto , Feminino , Humanos , GravidezRESUMO
According to the damped quantum rotation (DQR) theory, hindered rotation of methyl groups, evidenced in nuclear magnetic resonance (NMR) line shapes, is a nonclassical process. It comprises a number of quantum-rate processes measured by two different quantum-rate constants. The classical jump model employing only one rate constant is reproduced if these quantum constants happen to be equal. The values of their ratio, or the nonclassicallity coefficient, determined hitherto from NMR spectra of single crystals and solutions range from about 1.20 to 1.30 in the latter case to above 5.0 in the former, with the value of 1 corresponding to the jump model. Presently, first systematic investigations of the DQR effects in wide-line NMR spectra of a powder sample are reported. For 1,1,1-triphenylethane deuterated in the aromatic positions, the relevant line-shape effects were monitored in the range 99-121 K. The values of the nonclassicality coefficient dropping from 2.7 to 1.7 were evaluated in line shape fits to the experimental powder spectra from the range 99-108 K. At these temperatures, the fits with the conventional line-shape model are visibly inferior to the DQR fits. Using a theoretical model reported earlier, a semiquantitative interpretation of the DQR parameters evaluated from the spectra is given. It is shown that the DQR effects as such can be detected in wide-line NMR spectra of powdered samples, which are relatively facile to measure. However, a fully quantitative picture of these effects can only be obtained from the much more demanding experiments on single crystals.
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Hindered rotation of the -NH3+ group in a sterically crowded molecular environment was frozen on the NMR time scale. These effects were observed for solutions of 1,2,3,4-tetrabromotriptycyl-9-ammonium tetrafluoroborate in tetrahydrofuran with and without excess of HBF4. In the absence of acid, the hindered rotation is accompanied by a number of other effects originating in part from practically nonremovable traces of water in the solvent. Because of these additional effects, an effective rate constant of the hindered rotation could be evaluated at only one temperature for the neutral solution of the salt. For one of the acidified samples, which was prepared with special precautions against moisture, these complications are practically nonexistent at low and intermediate temperatures. For the latter sample, the Arrhenius parameters of the hindered rotation could be determined from the rotation rates evaluated in the range 194-226 K.
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Extending our earlier studies on cyclophanes, we here report the structure, chemical shifts, spin-spin coupling constants, absorption and emission properties of [m.n]paracyclophanes, m, n = 2-4, obtained using a combination of experimental and computational techniques. Accurate values of proton chemical shifts as well as of JHH for the bridges are determined. The experimental chemical shifts, coupling constants, absorption and emission wavelengths are satisfactorily reproduced using density functional theory calculations, using both the B3LYP and ωB97X-D functionals. The geometries predicted using a functional that includes dispersion corrections (ωB97X-D) are in a better agreement with available experimental values than those obtained using the B3LYP method. Up to 8 UV-vis absorption/emission bands have been observed (or anticipated in the region below 200 nm) and assigned on the basis of quantum-chemical calculations. Optimized excited-state geometries showed that the distances between the aromatic bridgehead carbon atoms of all the [m.n]paracyclophanes in the excited state decrease compared to the ground-state geometries by ca. 0.2-0.9 Å, the largest being for [4.4]paracyclophane, though the rather large differences in the calculated emission wavelength compared to experiment cast some doubts on the accuracy of the excited-state geometries.
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The theory of nuclear spin-lattice relaxation in methyl groups in solids has been a recurring problem in nuclear magnetic resonance (NMR) spectroscopy. The current view is that, except for extreme cases of low torsional barriers where special quantum effects are at stake, the relaxation behaviour of the nuclear spins in methyl groups is controlled by thermally activated classical jumps of the methyl group between its three orientations. The temperature effects on the relaxation rates can be modelled by Arrhenius behaviour of the correlation time of the jump process. The entire variety of relaxation effects in protonated methyl groups have recently been given a consistent quantum mechanical explanation not invoking the jump model regardless of the temperature range. It exploits the damped quantum rotation (DQR) theory originally developed to describe NMR line shape effects for hindered methyl groups. In the DQR model, the incoherent dynamics of the methyl group include two quantum rate (i.e., coherence-damping) processes. For proton relaxation only one of these processes is relevant. In this paper, temperature-dependent proton spin-lattice relaxation data for the methyl groups in polycrystalline methyltriphenyl silane and methyltriphenyl germanium, both deuterated in aromatic positions, are reported and interpreted in terms of the DQR model. A comparison with the conventional approach exploiting the phenomenological Arrhenius equation is made. The present observations provide further indications that incoherent motions of molecular moieties in the condensed phase can retain quantum character over much broader temperature range than is commonly thought.
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Blastocystis hominis (B. hominis) is a cosmopolitan pro- tozoa which parasitizes the human large intestine. This parasite had been considered to be commensal of the large intestine for a long time, because even an intense invasion may be asymptomatic. However, this species is now being regarded as a parasitic organism. In this paper the latest data concerning the epidemiology, diagnostics and treatment of B. hominis invasion have been cited and discussed.
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Infecções por Blastocystis/diagnóstico , Infecções por Blastocystis/parasitologia , Blastocystis hominis/fisiologia , Interações Hospedeiro-Parasita/fisiologia , Intestino Grosso/parasitologia , Animais , Infecções por Blastocystis/terapia , Infecções por Blastocystis/veterinária , Blastocystis hominis/classificação , Blastocystis hominis/citologia , Humanos , Especificidade da EspécieRESUMO
Tris(pentafluorophenyl)corrole and its (15)N-enriched isotopomer were studied in [D8]toluene solution by 1D and 2D variable-temperature NMR techniques to establish the mechanisms of tautomerization of the NH protons inside the interior of the corrole macrocycle. Three such rate processes could be identified of which two modulate the spectral line shapes at temperatures above 205â K and the third is NMR-inaccessible as it is very fast. The latter involves the proton engaged in an unsymmetrical proton sponge unit formed by two pyrrole nitrogen atoms. Temperature and concentration dependences of the two remaining processes were determined. One of them is purely intramolecular and the other is intermolecular at low temperatures, with growing contribution of an intramolecular mechanism at elevated temperatures. The proposed microscopic mechanisms of all these processes are semi-quantitatively confirmed by quantum chemical calculations using density functional theory.
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Espectroscopia de Ressonância Magnética , Porfirinas/química , Isótopos de Carbono/análise , Flúor/análise , Hidrogênio/análise , Ligação de Hidrogênio , Isomerismo , Isótopos de Nitrogênio/análise , TemperaturaRESUMO
The RIO kinases are essential protein factors required for the synthesis of new ribosomes in eukaryotes. Conserved in archaeal organisms as well, RIO kinases are among the most ancient of protein kinases. Their exact molecular mechanisms are under investigation and progress of this research would be significantly improved with the availability of suitable molecular probes that selectively block RIO kinases. RIO kinases contain a canonical eukaryotic protein kinase fold, but also display several unusual structural features that potentially create opportunity for the design of selective inhibitors. In an attempt to identify structural leads to target the RIO kinases, a series of pyridine caffeic acid benzyl amides (CABA) were tested for their ability to inhibit the autophosphorylation activity of Archeaoglobus fulgidus Rio1 (AfRio1). Screening of a small library of CABA molecules resulted in the identification of four compounds that measurably inhibited AfRio1 activity. Additional biochemical characterization of binding and inhibition activity of these compounds demonstrated an ATP competitive inhibition mode, and allowed identification of the functional groups that result in the highest binding affinity. In addition, docking of the compound to the structure of Rio1 and determination of the X-ray crystal structure of a model compound (WP1086) containing the desired functional groups allowed detailed analysis of the interactions between these compounds and the enzyme. Furthermore, the X-ray crystal structure demonstrated that these compounds stabilize an inactive form of the enzyme. Taken together, these results provide an important step in identification of a scaffold for the design of selective molecular probes to study molecular mechanisms of Rio1 kinases in vitro and in vivo. In addition, it provides a rationale for the future design of potent inhibitors with drug-like properties targeting an inactive form of the enzyme. This article is part of a Special Issue entitled: Inhibitors of Protein Kinases (2012).
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Archaea/enzimologia , Proteínas Arqueais/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Trifosfato de Adenosina/química , Trifosfato de Adenosina/metabolismo , Trifosfato de Adenosina/farmacologia , Proteínas Arqueais/química , Proteínas Arqueais/metabolismo , Sítios de Ligação , Ligação Competitiva , Domínio Catalítico , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Cinética , Modelos Moleculares , Conformação Molecular , Estrutura Molecular , Fosforilação/efeitos dos fármacos , Ligação Proteica , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/metabolismo , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/metabolismo , Estrutura Terciária de Proteína , Piridinas/química , Piridinas/metabolismo , Piridinas/farmacologia , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
JAK (Janus kinase)/STAT (signal transducers and activators of transcription) signaling is involved in the regulation of cell growth, differentiation and apoptosis. Constitutive activation of STATs, in particular STAT3, is observed in a large number of human tumors, including gliomas and may contribute to oncogenesis by stimulating cell proliferation and preventing apoptosis, thus it emerges as a promising target for anti-cancer therapy. To investigate the therapeutic potential of blocking STAT3 in glioma cells a set of small synthetic molecules - caffeic acid derivatives, structurally related to AG490 was screened for its ability to inhibit STAT3. Inhibitor 2 (E)-2-cyano-N-[(S)-1-phenylethyl]-3-(pyridin-2-yl)acrylamide was the most effective in inhibition of JAK/STAT3 signaling and at doses ≥ 25 µM significantly reduced the level of phosphorylated JAK1, JAK2 and STAT3 (at Tyr705) and downregulated the expression of known STAT3 targets. In treated cells we observed rapid detachment and rounding of cells associated with reduction of focal adhesion kinase phosphorylation and activity, followed by upregulation of phosphorylated p38, JNK and ERK1/2 levels. Accumulation of cells with fragmented DNA, increases of the cleaved caspase 3 and fragmented PARP levels were detected 24 h after the treatment suggesting ongoing apoptotic cell death. Three human malignant glioblastoma cell lines defective in tumor suppressors TP53 and/or PTEN were susceptible to inhibitor 2 that induced the programmed cell death. Global gene expression profiling revealed modulation of numerous genes in cells treated with inhibitor 2 revealing novel, potential JAK/STAT targets. Our study demonstrates that suitably modified caffeic acid molecules exhibit significant cytotoxic potential toward glioma cells.
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Antineoplásicos/farmacologia , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Glioma/metabolismo , Janus Quinases/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tirfostinas/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Análise por Conglomerados , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioma/genética , Humanos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosforilação/efeitos dos fármacos , Ratos , Tirfostinas/química , Tirfostinas/toxicidadeRESUMO
The calculated structures of several known and hypothetical cyclophanes with ethylene bridges (cyclophenes) are reported together with experimental and calculated values of their NMR parameters. Of the exchange-correlation functionals and basis sets used in this work, only the ωB97X-D/6-311++G(2d,2p) and ωB97X-D/cc-pVQZ yielded values of the C(sp3)-C(sp3) bond length close to the experimental data, although significant differences still remain. As far as the NMR parameters are concerned, except for close-lying signals, chemical shifts and coupling constants calculated at the ωB97X-D/cc-pVQZ level reproduce in most cases the experimental trends. Contrary to the calculations of geometries, an agreement between the values of the NMR parameters obtained at ωB97X-D/cc-pVQZ level and the experimental ones is the poorest compared with that of the ωB97X-D/6-311++G(2d,2p) one. Taking into account that the results of the different calculations show the same qualitative trends in most cases, we believe that they correctly describe the structure and properties of the hypothetical molecules studied here.