Ratio of proliferation markers and HSP90 gene expression as a predictor of pathological complete response in breast cancer neoadjuvant chemotherapy

Introduction. Prediction of response to preoperative breast cancer chemotherapy may offer a substantial optimization of medical management of this disease. The most efficient prediction would be done a priori, before the start of chemotherapy and based on the biological features of patient and tumor. Numerous markers have been proposed but none of them has been applied as a routine. The role of MKI67 and HSP90 expression has been recently suggested to predict treatment sensitivity in HER2-positive breast cancer. The aim of this study was to validate the utility of proliferation based markers (MKI67 and CDK1) and heat shock proteins (namely HSP90) to predict response to chemotherapy in cohort of breast cancer patients treated preoperatively. Material and methods. Ninety-three patients with breast cancer, all females, mean age 42.2 years, among them 32% T1-T2 patients, 49% T3 patients and 13% with T4 tumor stage, 27% N0, 42% N1, 16% N2, 15% N3 were subjected to initial chemotherapy. The majority of patients (86%) received anthracycline and taxane chemotherapy. Among the patients there were 9 individuals with metastatic disease (M1) at initial presentation, and 11 patients were not treated surgically after initial chemotherapy (no sufficient disease response). From 82 patients operated on, 20 patients (24%) showed pathological complete response (pCR), while in 62 patients there was no pCR. 42% of patients were hormone-sensitive HER2-negative, 20% hormone-sensitive HER2-positive, 9% only HER-positive and 29% with triple negative breast cancer. Four gene transcripts (MKI67, cyclin-dependent kinase 1 [CDK1], heat shock proteins HSP90AA1 and HSP- 90AB1) were analyzed in total RNA isolated from single core obtained during preoperative core needle biopsy by quantitative real-time PCR with fluorescent probes (Universal Probe Library, Roche). Results were normalized to the panel of reference genes. Results. There were no statistically significant differences in MKI67 and CDK1 expression between pCR and no pCR groups (p = 0.099 and 0.35, respectively), although the median expression of both genes was slightly higher in pCR group. In contrast, both HSP90AA1 and HSP90AB1 transcripts showed decreased expression in pCR group (medians 0.77 and 0.55) when compared to no p CR group (median 0.86 and 0.73), statistically significant for HSP90AA1 (p = 0.031) and of borderline significance for HSP90AB1 (p = 0.054). The most significant predictor of pCR was the ratio of CDK1 transcript to HSP90AA transcript. This ratio was significantly higher in CR group (median 0.99) than in no CR group (median 0.68, p = 0.0023), and showed a potential diagnostic utility (area under receiver operating characteristic [ROC] curve 0.72). Conclusions. HSP90AA1 and AB1 genes exhibit low expression in breast cancers highly sensitive to chemotherapy and may indicate the patients with higher probability of pathological complete response. The ratio of HSP90AA1 to proliferation-related markers (CDK1 or MKI67) may be even better predictor of pCR chance, with higher expression of proliferation genes and lower stress response in patients sensitive to chemotherapy

LOVTRAP: an optogenetic system for photoinduced protein dissociation

Here we introduce LOVTRAP, an optogenetic approach for reversible, light-induced protein dissociation. LOVTRAP is based on protein A fragments that bind to the LOV domain only in the dark, with tunable kinetics and a >150-fold change in Kd. By reversibly sequestering proteins at mitochondria, we precisely modulated the proteins’ access to the cell edge, demonstrating a naturally occurring 3 mHz cell edge oscillation driven by interactions of Vav2, Rac1 and PI3K

Magnetic signature reproduction of ferromagnetic ships at arbitrary geographical position, direction and depth using a multi-dipole model

Abstract The reproduction of magnetic signatures is an important issue concerning the safety of ship traffic, as well as the identification and classification of vessels. Moreover, military applications of magnetic signatures and their reproduction refer to the activation or protection against activation of magnetic naval mines. Previous works on this subject focused on recording and replicating the signatures under the same conditions as those under which they were measured, e.g., on the same ship courses. In this article, much greater capabilities of the multi-dipole model are presented, including simultaneous identification of permanent and induced magnetism. Determining the dipole values using the data from cardinal directions gives the possibility of determining the magnetic field density at any trajectory (position), direction, or depth, with further reconstruction of the entire magnetic field on the basis of residual measurements. For the purpose of this article, a numerical test model of a corvette-type ship has been modelled in Opera simulation software for different geographical positions. The synthetic data from the simulator served as the data source for determining the parameters of the multi-dipole model and the reference data for the verification of the signatures reconstructed for other positions, directions, and depths than those used to determine the model parameters. To determine all permanent magnetization components, data sets were used for two different values of the external magnetic field vertical component. Finally, as a culmination of the demonstration of model universality, the entire magnetic field around the ship was reproduced for different control points on Earth, and for different courses and depths. Investigating the possibility of reconstructing the magnetic signature at a different geographic location than the place where the measurement was made for model synthesis is the main original issue considered in this paper

Heterologous expression and initial characterization of recombinant RbcX protein from Thermosynechococcus elongatus BP-1 and the role of RbcX in RuBisCO assembly

In the cyanobacterial RuBisCO operon from Thermosynechococcus elongatus the rbcX gene is juxtaposed and cotranscribed with the rbcL and rbcS genes which encode large and small RuBisCO subunits, respectively. It has been suggested that the rbcX position is not random and that the RbcX protein could be a chaperone for RuBisCO. In this study, the RbcX protein from T. elongatus was overexpressed, purified and preliminary functional studies were conducted. The recombinant protein purified from Escherichia coli extracts was predominantly present in a soluble fraction in a dimeric form. Coexpression experiments have demonstrated that RbcX can mediate RbcL dimer (L2) formation, and that it is essential for the L8 core complex assembly. This is the first characterization of the RbcX protein from a thermophilic organism

Descripción del desempeño reproductivo de cerdas puras y cruzadas en la Escuela Agrícola Panamericana

32 P.La Granja Porcina Educativa de la Escuela Agrícola Panamericana maneja tres razas puras y cruces, se realizó un análisis creando grupos contemporáneos por años desde el 2000 hasta el 2012, analizando nueve variables reproductivas, raza por raza y también por paridad. Además se realizaron correlaciones entre las variables: Lechones nacidos totales – peso promedio del lechón al nacimiento, lechones destetados – peso promedio al destete, longitud de lactancia – peso promedio del lechón al destete, largo de longitud – periodo abierto. Todos los datos obtenidos fueron generados por el programa Pigchamp®. Para cada variable reproductiva se crearon dos cuadros, uno por genética y otro por paridad. Las correlaciones se hicieron tomando las medias anuales de cada variable. El principal problema observado en la granja porcina es la alta mortalidad predestete (14%). Además la tasa de concepción no es la adecuada (72%), encontrándose muy por debajo de lo recomendado (84%). Las variables tuvieron mejor eficiencia en las cerdas cruzadas, lo cual nos indica que una correcta recombinación genética, potencializa el rendimiento del cerdo.1. Índice de cuadro, figuras y anexos 2. Introducción 3. Materiales y métodos 4. Resultados y discusión 5. Conclusiones 6. Recomendaciones 7. Literatura citada 8. Anexos

Engineering extrinsic disorder to control protein activity in living cells

Optogenetic and chemogenetic control of proteins has revealed otherwise inaccessible facets of signaling dynamics. Here, we use light- or ligand-sensitive domains to modulate the structural disorder of diverse proteins, thereby generating robust allosteric switches. Sensory domains were inserted into nonconserved, surface-exposed loops that were tight and identified computationally as allosterically coupled to active sites. Allosteric switches introduced into motility signaling proteins (kinases, guanosine triphosphatases, and guanine exchange factors) controlled conversion between conformations closely resembling natural active and inactive states, as well as modulated the morphodynamics of living cells. Our results illustrate a broadly applicable approach to design physiological protein switches

Chemogenetic Control of Nanobodies

Engineered nanobody allows reversible control of activity in cells through the binding of small molecules.We introduce an engineered nanobody whose affinity to green fluorescent protein (GFP) can be switched on and off with small molecules. By controlling the cellular localization of GFP fusion proteins, the engineered nanobody allows interrogation of their roles in basic biological processes, an approach that should be applicable to numerous previously described GFP fusions. We also outline how the binding affinities of other nanobodies can be controlled by small molecules

Kinetic and Structural Characterization of the Self-Labeling Protein Tags HaloTag7, SNAP-tag, and CLIP-tag

The self-labeling protein tags (SLPs) HaloTag7, SNAP-tag, and CLIP-tag allow the covalent labeling of fusion proteins with synthetic molecules for applications in bioimaging and biotechnology. To guide the selection of an SLP-substrate pair and provide guidelines for the design of substrates, we report a systematic and comparative study of the labeling kinetics and substrate specificities of HaloTag7, SNAP-tag, and CLIP-tag. HaloTag7 reaches almost diffusion-limited labeling rate constants with certain rhodamine substrates, which are more than 2 orders of magnitude higher than those of SNAP-tag for the corresponding substrates. SNAP-tag labeling rate constants, however, are less affected by the structure of the label than those of HaloTag7, which vary over 6 orders of magnitude for commonly employed substrates. Determining the crystal structures of HaloTag7 and SNAP-tag labeled with fluorescent substrates allowed us to rationalize their substrate preferences. We also demonstrate how these insights can be exploited to design substrates with improved labeling kinetics.ISSN:0006-2960ISSN:1520-499