The FIREBIRD-II CubeSat mission: Focused investigations of relativistic electron burst intensity, range, and dynamics.

FIREBIRD-II is a National Science Foundation funded CubeSat mission designed to study the scale size and energy spectrum of relativistic electron microbursts. The mission consists of two identical 1.5 U CubeSats in a low earth polar orbit, each with two solid state detectors that differ only in the size of their geometric factors and fields of view. Having two spacecraft in close orbit allows the scale size of microbursts to be investigated through the intra-spacecraft separation when microbursts are observed simultaneously on each unit. Each detector returns high cadence (10 s of ms) measurements of the electron population from 200 keV to >1 MeV across six energy channels. The energy channels were selected to fill a gap in the observations of the Heavy Ion Large Telescope instrument on the Solar, Anomalous, and Magnetospheric Particle Explorer. FIREBIRD-II has been in orbit for 5 years and continues to return high quality data. After the first month in orbit, the spacecraft had separated beyond the expected scale size of microbursts, so the focus has shifted toward conjunctions with other magnetospheric missions. FIREBIRD-II has addressed all of its primary science objectives, and its long lifetime and focus on conjunctions has enabled additional science beyond the scope of the original mission. This paper presents a brief history of the FIREBIRD mission's science goals, followed by a description of the instrument and spacecraft. The data products are then discussed along with some caveats necessary for proper use of the data.

Novel, rapid, low-cost screen-printed (bio)sensors for the direct analysis of boar taint compounds androstenone and skatole in porcine adipose tissue: Comparison with a high-resolution gas chromatographic method.

This is the first report on the fabrication, characterisation and application of an electrochemical (bio)sensor system for the simultaneous measurement of skatole and androstenone. A biosensor for androstenone was fabricated using a Meldola's Blue modified SPCE (MB-SPCE) by depositing NADH and the enzyme 3α-hydroxysteroid dehydrogenase onto the MB-SPCE surface; samples of adipose tissue were analysed using the biosensors in conjunction with chronoamperometry. Cyclic voltammetry was used to investigate the electrochemical behaviour of skatole at a screen-printed carbon electrode (SPCE vs. Ag/AgCl). An oxidation peak was observed around +0.55 V (vs. Ag/AgCl) and differential pulse voltammetry was applied for quantification of skatole in adipose tissue (in-situ). Quantitative analysis was achieved using calibration plots obtained from fortified meat samples. The concentrations obtained by the electrochemical and gas chromatographic (GC) methods demonstrated a good positive correlation. The (bio)sensor system completed both measurements within 60 s, as compared to several hours for GC, and at a considerably reduced cost and complexity. Consequently, the novel (bio)sensor system should have applications for analysis of carcasses on the abattoir processing line.

A novel electroanalytical approach to the measurement of B vitamins in food supplements based on screen-printed carbon sensors.

This paper describes the development of a novel electrochemical assay for the measurement of water-soluble vitamins in food and pharmaceutical products. The optimum conditions for the determination of vitamin B1 (thiamine), B2 (riboflavin) and B6 (pyridoxine) in phosphate buffer were established using cyclic voltammetry in conjunction with screen printed carbon electrodes (SPCEs). The optimum current response for all three vitamins was achieved in 0.1M phosphate buffer pH 11 using an initial potential of -1.0V. Using square wave voltammetry, the linear ranges for thiamine, riboflavin, and pyridoxine were found to be: 15-110µg/ml, 0.1-20µg/ml, and 2-80µg/ml respectively. The application of the method to a commercial food product yielded a recovery of 95.78% for riboflavin, with a coefficient of variation (CV) of 3.38% (n = 5). The method was also applied to a multi-vitamin supplement for the simultaneous determination of thiamine, riboflavin and pyridoxine. In both cases only simple dilution with buffer followed by centrifugation was required prior to analysis. The resulting square wave voltammetric signals were completely resolved with Ep values of -0.7V, +0.2V, and +0.6V respectively. The recoveries determined for the vitamin B complex in a commercial supplement product were found to be 110%, 114%, and 112% respectively (CV = 7.14%, 6.28%. 5.66% respectively, n = 5).

BET protein proteolysis targeting chimera (PROTAC) exerts potent lethal activity against mantle cell lymphoma cells.

Bromodomain extraterminal protein (BETP) inhibitors transcriptionally repress oncoproteins and nuclear factor-κB (NF-κB) target genes that undermines the growth and survival of mantle cell lymphoma (MCL) cells. However, BET bromodomain inhibitor (BETi) treatment causes accumulation of BETPs, associated with reversible binding and incomplete inhibition of BRD4 that potentially compromises the activity of BETi in MCL cells. Unlike BETi, BET-PROTACs (proteolysis-targeting chimera) ARV-825 and ARV-771 (Arvinas, Inc.) recruit and utilize an E3-ubiquitin ligase to effectively degrade BETPs in MCL cells. BET-PROTACs induce more apoptosis than BETi of MCL cells, including those resistant to ibrutinib. BET-PROTAC treatment induced more perturbations in the mRNA and protein expressions than BETi, with depletion of c-Myc, CDK4, cyclin D1 and the NF-κB transcriptional targets Bcl-xL, XIAP and BTK, while inducing the levels of HEXIM1, NOXA and CDKN1A/p21. Treatment with ARV-771, which possesses superior pharmacological properties compared with ARV-825, inhibited the in vivo growth and induced greater survival improvement than the BETi OTX015 of immune-depleted mice engrafted with MCL cells. Cotreatment of ARV-771 with ibrutinib or the BCL2 antagonist venetoclax or CDK4/6 inhibitor palbociclib synergistically induced apoptosis of MCL cells. These studies highlight promising and superior preclinical activity of BET-PROTAC than BETi, requiring further in vivo evaluation of BET-PROTAC as a therapy for ibrutinib-sensitive or -resistant MCL.

Novel BET protein proteolysis-targeting chimera exerts superior lethal activity than bromodomain inhibitor (BETi) against post-myeloproliferative neoplasm secondary (s) AML cells.

The PROTAC (proteolysis-targeting chimera) ARV-825 recruits bromodomain and extraterminal (BET) proteins to the E3 ubiquitin ligase cereblon, leading to degradation of BET proteins, including BRD4. Although the BET-protein inhibitor (BETi) OTX015 caused accumulation of BRD4, treatment with equimolar concentrations of ARV-825 caused sustained and profound depletion (>90%) of BRD4 and induced significantly more apoptosis in cultured and patient-derived (PD) CD34+ post-MPN sAML cells, while relatively sparing the CD34+ normal hematopoietic progenitor cells. RNA-Seq, Reverse Phase Protein Array and mass cytometry 'CyTOF' analyses demonstrated that ARV-825 caused greater perturbations in messenger RNA (mRNA) and protein expressions than OTX015 in sAML cells. Specifically, compared with OTX015, ARV-825 treatment caused more robust and sustained depletion of c-Myc, CDK4/6, JAK2, p-STAT3/5, PIM1 and Bcl-xL, while increasing the levels of p21 and p27. Compared with OTX015, PROTAC ARV-771 treatment caused greater reduction in leukemia burden and further improved survival of NSG mice engrafted with luciferase-expressing HEL92.1.7 cells. Co-treatment with ARV-825 and JAK inhibitor ruxolitinib was synergistically lethal against established and PD CD34+ sAML cells. Notably, ARV-825 induced high levels of apoptosis in the in vitro generated ruxolitinib-persister or ruxolitinib-resistant sAML cells. These findings strongly support the in vivo testing of the BRD4-PROTAC based combinations against post-MPN sAML.

A screen-printed, amperometric biosensor array incorporated into a novel automated system for the simultaneous determination of organophosphate pesticides.

Organophosphate pesticides present serious risks to human and environmental health. A rapid reliable, economical and portable analytical system will be of great benefit in the detection and prevention of contamination. A biosensor array based on six acetylcholinesterase enzymes for use in a novel automated instrument incorporating a neural network program is described. Electrochemical analysis was carried out using chronoamperometry and the measurement was taken 10s after applying a potential of 0 V vs. Ag/AgCl. The total analysis time for the complete assay was less than 6 min. The array was used to produce calibration data with six organophosphate pesticides (OPs) in the concentration range of 10(-5) M to 10(-9) M to train a neural network. The output of the neural network was subsequently evaluated using different sample matrices. There were no detrimental matrix effects observed from water, phosphate buffer, food or vegetable extracts. Furthermore, the sensor system was not detrimentally affected by the contents of water samples taken from each stage of the water treatment process. The biosensor system successfully identified and quantified all samples where an OP was present in water, food and vegetable extracts containing different OPs. There were no false positives or false negatives observed during the evaluation of the analytical system. The biosensor arrays and automated instrument were evaluated in situ in field experiments where the instrument was successfully applied to the analysis of a range of environmental samples. It is envisaged that the analytical system could provide a rapid detection system for the early warning of contamination in water and food.

Development of an anodic stripping voltammetric assay, using a disposable mercury-free screen-printed carbon electrode, for the determination of zinc in human sweat.

This paper reports on the development of a novel electrochemical assay for Zn(2+) in human sweat, which involves the use of disposable screen-printed carbon electrodes (SPCEs). Initially, SPCEs were used in conjunction with cyclic voltammetry to study the redox characteristics of Zn(2+) in a selection of supporting electrolytes. The best defined cathodic and anodic peaks were obtained with 0.1 M NaCl/0.1 M acetate buffer pH 6.0. The anodic peak was sharp and symmetrical which is typical for the oxidation of a thin metal film on the electrode surface. This behaviour was exploited in the development of a differential pulse anodic stripping voltammetric (DPASV) assay for zinc. It was shown that a deposition potential of -1.6 V versus Ag/AgCl and deposition time of 60 s with stirring (10 s equilibration) produced a well-defined stripping peak with E(pa) = -1.2 V versus Ag/AgCl. Using these conditions, the calibration plot was linear over the range 1x10(-8) to 5x10(-6) M Zn(2+). The precision was examined by carrying out six replicate measurements at a concentration of 2x10(-6) M; the coefficient of variation was calculated to be 5.6%. The method was applied to the determination of the analyte in sweat from 10 human volunteers. The concentrations were between 0.39 and 1.56 microg/mL, which agrees well with previously reported values. This simple, low-cost sensitive assay should have application in biomedical studies and for stress and fatigue in sports studies.

Improved glucose homeostasis and enhanced insulin signalling in Grb14-deficient mice.

Gene targeting was used to characterize the physiological role of growth factor receptor-bound (Grb)14, an adapter-type signalling protein that associates with the insulin receptor (IR). Adult male Grb14(-/-) mice displayed improved glucose tolerance, lower circulating insulin levels, and increased incorporation of glucose into glycogen in the liver and skeletal muscle. In ex vivo studies, insulin-induced 2-deoxyglucose uptake was enhanced in soleus muscle, but not in epididymal adipose tissue. These metabolic effects correlated with tissue-specific alterations in insulin signalling. In the liver, despite lower IR autophosphorylation, enhanced insulin-induced tyrosine phosphorylation of insulin receptor substrate (IRS)-1 and activation of protein kinase B (PKB) was observed. In skeletal muscle, IR tyrosine phosphorylation was normal, but signalling via IRS-1 and PKB was increased. Finally, no effect of Grb14 ablation was observed on insulin signalling in white adipose tissue. These findings demonstrate that Grb14 functions in vivo as a tissue-specific modulator of insulin action, most likely via repression of IR-mediated IRS-1 tyrosine phosphorylation, and highlight this protein as a potential target for therapeutic intervention.

Up-regulation of the protein tyrosine phosphatase SHP-1 in human breast cancer and correlation with GRB2 expression.

The protein tyrosine phosphatase SHP-1 is predominantly expressed in hemopoietic cell lineages, where its function is relatively well defined. However, its expression profile also extends to certain epithelial cell types. Furthermore, the negative regulatory role of this enzyme in hemopoietic cell signaling may not apply to other systems, where positive effects on particular tyrosine kinase signaling pathways have been described. Expression of SHP-1 was therefore investigated in human breast cancer cell lines and primary breast cancers. Differential expression of SHP-1 mRNA was observed among the 19 breast cancer cell lines examined, and in an analysis of 72 primary breast cancers, SHP-1 mRNA expression was increased 2- to 12-fold relative to normal breast epithelial cells in 58% of the samples. Interestingly, a subset of the cancers also over-expressed GRB2 mRNA by 2- to 7-fold, and a significant (p < 0.01) positive correlation was observed between SHP-1 and GRB2 mRNA expression. Since these proteins can bind to each other and regulate MEK/MAP kinase activation, their co-ordinate up-regulation may amplify tyrosine kinase signaling in breast cancer cells.

Estrogen regulation of transforming growth factor-alpha in ovarian cancer.

Transforming growth factor alpha (TGFalpha) may be induced by estrogen in estrogen responsive systems and can contribute to the growth-modulatory effects of this hormone. To test whether TGFalpha contributes to estrogen-regulated growth in ovarian cancers, we have compared the effects of 17beta-estradiol (E2) and TGFalpha in a range of ovarian carcinoma cell lines. Addition of E2 to the estrogen receptor (ER)-positive cell lines (PE01, PE04 and PE01CDDP) produced a 2-4 fold increase in TGFalpha protein concentrations in media conditioned by the cells. Both E2 and TGFalpha stimulated the growth of the PE01 and PE04 lines and inhibited the growth of the PE01CDDP line. Furthermore, the E2-mediated growth effects could be reversed by an epidermal growth factor (EGF) receptor-targeted antibody. E2 also down-regulated EGF receptor expression in ER-positive cell lines. In a series of primary ovarian tumors, higher concentrations of ER were associated with an increased percentage of tumors expressing TGFalpha mRNA and a decreased percentage expressing EGF receptor protein. All these data are consistent with E2 increasing production of TGFalpha in ER-positive ovarian cancer and this in turn acting through the EGF receptor to modulate growth in an autocrine manner.

The t(X;1)(p11.2;q21.2) translocation in papillary renal cell carcinoma fuses a novel gene PRCC to the TFE3 transcription factor gene.

The specific chromosomal translocation t(X;1)(p11.2;q21.2) has been observed in human papillary renal cell carcinomas. In this study we demonstrated that this translocation results in the fusion of a novel gene designated PRCC at 1q21.2 to the TFE3 gene at Xp11.2. TFE3 encodes a member of the basic helix-loop-helix (bHLH) family of transcription factors originally identified by its ability to bind to microE3 elements in the immunoglobin heavy chain intronic enhancer. The translocation is predicted to result in the fusion of the N-terminal region of the PRCC protein, which includes a proline-rich domain, to the entire TFE3 protein. Notably the generation of the chimaeric PRCC-TFE3 gene appears to be accompanied by complete loss of normal TFE3 transcripts. This work establishes that the disruption of transcriptional control by chromosomal translocation is important in the development of kidney carcinoma in addition to its previously established role in the aetiology of sarcomas and leukaemias.

Fusion of SYT to two genes, SSX1 and SSX2, encoding proteins with homology to the Kruppel-associated box in human synovial sarcoma.

We demonstrate that the cytogenetically defined translocation t(X;18)(p11.2;q11.2) found in human synovial sarcoma results in the fusion of the chromosome 18 SYT gene to either of two distinct genes, SSX1 or SSX2, at Xp11.2. The SSX1 and SSX2 genes encode closely related proteins (81% identity) of 188 amino acids that are rich in charged amino acids. The N-terminal portion of each SSX protein exhibits homology to the Kruppel-associated box (KRAB), a transcriptional repressor domain previously found only in Kruppel-type zinc finger proteins. PCR analysis demonstrates the presence of SYT-SSX1 or SYT-SSX2 fusion transcripts in 29 of 32 of the synovial sarcomas examined, indicating that the detection of these hybrid transcripts by PCR may represent a very useful diagnostic method. Sequence analysis has demonstrated heterogeneity in the fusion transcripts with the formation of two distinct SYT-SSX1 fusion junctions and two distinct SYT-SSX2 fusion junctions.

Fusion of the EWS gene to a DNA segment from 9q22-31 in a human myxoid chondrosarcoma.

Southern blot analyses revealed a rearrangement of the EWS gene in a skeletal human myxoid chondrosarcoma. Interphase fluorescence in situ hybridization (FISH) studies, using cosmid clones F7 and G9 that flank the EWS locus on 22q12, confirmed the presence of this EWS gene abnormality. Cloning the rearranged EWS DNA fragment and mapping by FISH demonstrated that the EWS gene is joined to DNA sequences localised in 9q22-31. These findings are consistent with previous cytogenetic reports of a recurrent t(9;22)(q22-31;q11-12) in the myxoid variant of chondrosarcoma and reveal involvement of the EWS gene in a fourth type of human sarcoma.

Identification of novel genes, SYT and SSX, involved in the t(X;18)(p11.2;q11.2) translocation found in human synovial sarcoma.

Human synovial sarcomas contain a recurrent and specific chromosomal translocation t(X;18)(p11.2;q11.2). By screening a synovial sarcoma cDNA library with a yeast artificial chromosome spanning the X chromosome breakpoint, we have identified a hybrid transcript that contains 5' sequences (designated SYT) mapping to chromosome 18 and 3' sequences (designated SSX) mapping to chromosome X. An SYT probe detected genomic rearrangements in 10/13 synovial sarcomas. Sequencing of cDNA clones shows that the normal SYT gene encodes a protein rich in glutamine, proline and glycine, and indicates that in synovial sarcoma rearrangement of the SYT gene results in the formation of an SYT-SSX fusion protein. Both SYT and SSX failed to exhibit significant homology to known gene sequences.

The t(X;18)(p11.2;q11.2) translocation found in human synovial sarcomas involves two distinct loci on the X chromosome.

A high proportion of synovial sarcomas contain the reciprocal translocation t(X;18)(p11.2;q11.2). We have previously localized the breakpoint on the X chromosome between the X chromosome marker DXS255 and an ornithine aminotransferase (OAT) pseudogene region designated OATL2. Subsequently by fluorescence in situ hybridization (FISH) we provided evidence that YACs corresponding to the OATL2 locus spanned the break-point. In order to confirm the position of this breakpoint cosmids corresponding to the OATL2 region were isolated. Most of these cosmids mapped to four cosmid contigs designated C1-C4. Analysis of two contigs, C1- and C4, using FISH established that in four of six synovial sarcomas examined the breakpoint occurs between these two contigs: C1 lies distal to the break-point while C4 is proximal. In contrast we provide evidence that the breakpoint in the remaining two tumours mapped to a second pseudogene region called OATL1 that is telomeric to the OATL2 locus. This heterogeneity of the breakpoint position on the X chromosome explains why in previous mapping studies there have been discrepancies between the results obtained by different laboratories.

Growth inhibition of oestrogen receptor-positive human ovarian carcinoma by anti-oestrogens in vitro and in a xenograft model.

This paper presents results of the in vitro and in vivo effects of anti-oestrogens on the growth of human ovarian cancer cells. Tamoxifen and the "pure" anti-oestrogens, ICI 164,384 and ICI 182,780, inhibited the oestrogen-stimulated growth of the oestrogen receptor (ER)-positive PE04 and PE01 cell lines grown in culture, the latter two compounds being more potent than tamoxifen. In the absence of 17 beta-oestradiol (E2), tamoxifen, but not the pure anti-oestrogens, produced a small degree of growth stimulation in the PE01 and PE04 lines at concentrations between 10((7) and 10(-9) M. In contrast, growth of the ER-negative PE014 line was unaffected by E2 and all three anti-oestrogens. The effects of tamoxifen and ICI 182,780 on PE04 cells grown as xenografts in nude mice were also studied. Both anti-oestrogens produce significant growth inhibitory effects. These results indicate that ovarian carcinoma cells may be sensitive to anti-oestrogens in vitro and in vivo, and support the view that anti-oestrogens merit further clinical studies in patients with ER-positive tumours.

Contrasting effects of 17 beta-estradiol on the growth of human ovarian carcinoma cells in vitro and in vivo.

A human ovarian adenocarcinoma cell line (PE04) has been established as a xenograft in nude mice. In vitro, this cell line is estrogen receptor (ER)-positive and its growth is stimulated by 17 beta-estradiol at concentrations between 10(-12) and 10(-6) M. When xenografted, PE04 cells remain ER-positive and also possess progesterone receptors (PR); treatment with 17 beta-estradiol reduces the concentration of ER and increases levels of PR. Growth of the xenograft is reduced in ovariectomized animals while implantation of estrogen pellets also results in growth inhibition. Similar treatment with estrogen does not inhibit the ER-negative HOX 60 ovarian xenograft, and stimulates growth of the ER-positive ZR-75-I breast carcinoma xenograft. Serum measurements of 17 beta-estradiol confirm that ovariectomy reduces the level of 17 beta-estradiol while implantation of estrogen pellets results in raised levels of the hormone. Tamoxifen inhibits growth of the PE04 xenograft but not that of the HOX 60 xenograft, consistent with ER status. These results indicate that ER-positive PE04 ovarian cancer cells are sensitive to 17 beta-estradiol in vivo but that the response may be of a different type from the in vitro response. This lends further support to the concept that ovarian cancer may be hormone-sensitive and potentially responsive to endocrine therapy.

Growth control by epidermal growth factor and transforming growth factor-alpha in human lung squamous carcinoma cells.

Although EGF receptor expression is generally elevated in human lung squamous carcinoma, the biological significance of this phenomenon and the role of EGF and TGF-alpha in this disease are poorly understood. We have investigated three human lung squamous carcinoma cell lines (NX002, CX140 and CX143) and have shown, using an antibody (EGFR1) directed against the EGF receptor, that the majority of cells in all three lines express the EGF receptor. Using a ligand binding assay, Scatchard analysis indicated high concentrations (1,300-2,700 fmol mg-1 protein) of a single low affinity binding site (Kd = 3-5 nM) within these lines. Addition of EGF or TGF-alpha at concentrations greater than 0.1 nM resulted in growth inhibition of all three lines and this was associated with an accumulation of cells in the G2/M phase of the cell cycle. Growth inhibitory effects were not explained by an enhancement of cellular differentiation as monitored by involucrin expression and the ability to form cornified envelopes. While the presence of EGF could not be detected in medium conditioned by the NX002 cell line, mRNA for TGF-alpha was detected in all three lines suggesting the possibility of an autocrine loop. These results together with reports of growth inhibition by EGF and TGF-alpha in other systems suggest that EGF and similar molecules might have a growth regulatory role in lung cancer cells and modulation of such may have therapeutic potential.

Mitogenic effects of epidermal growth factor and transforming growth factor-alpha on EGF-receptor positive human ovarian carcinoma cell lines.

The role of epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha) in the growth modulation of three human ovarian adenocarcinoma cell lines, PEO1, PEO4 and PEO14, has been examined by measuring responses of the cells growing in monolayer culture to exogenous addition of the growth factors. The presence of EGF receptors in the cell lines has been confirmed by ligand binding and immunocytochemical staining using a monoclonal antibody directed against the EGF receptor. The growth of all three cell lines was stimulated by both EGF and TGF-alpha. Dose-response effects were noted with the greatest growth stimulation occurring at concentrations between 0.1 and 10 nmol/l. The stimulatory effects of EGF and TGF-alpha were accompanied by changes in the cell cycle distribution as detected by flow cytometric analysis. It is concluded that EGF and TGF-alpha are important growth regulators in these EGF-receptor positive ovarian cancer cells.