Recombinant Interleukin-24 Lacks Apoptosis-Inducing Properties in Melanoma Cells

IL-24, also known as melanoma differentiation antigen 7 (mda-7), is a member of the IL-10 family of cytokines and is mainly produced by Th2 cells as well as by activated monocytes. Binding of IL-24 to either of its two possible heterodimeric receptors IL-20R1/IL-20R2 and IL-22R/IL-20R2 activates STAT3 and/or STAT1 in target tissues such as lung, testis, ovary, keratinocytes and skin. To date, the physiological properties of IL-24 are still not well understood but available data suggest that IL-24 affects epidermal functions by increasing proliferation of dermal cells. In stark contrast to its “normal” and physiological behaviour, IL-24 has been reported to selectively and efficiently kill a vast variety of cancer cells, especially melanoma cells, independent of receptor expression and Jak-STAT signalling. These intriguing properties have led to the development of adenovirally-expressed IL-24, which is currently being evaluated in clinical trials. Using three different methods, we have analysed a large panel of melanoma cell lines with respect to IL-24 and IL-24 receptor expression and found that none of the investigated cell lines expressed sufficient amounts of functional receptor pairs and therefore did not react to IL-24 stimulation with Jak/STAT activation. Results for three cell lines contrasted with previous studies, which reported presence of IL-24 receptors and activation of STAT3 following IL-24 stimulation. Furthermore, evaluating four different sources and modes of IL-24 administration (commercial recombinant IL-24, bacterially expressed GST-IL-24 fusion protein, IL-24 produced from transfected Hek cells, transiently over-expressed IL-24) no induction or increase in cell death was detected when compared to appropriate control treatments. Thus, we conclude that the cytokine IL-24 itself has no cancer-specific apoptosis-inducing properties in melanoma cells

CCT196969 effectively inhibits growth and survival of melanoma brain metastasis cells

Melanomas frequently metastasize to the brain. Despite recent progress in the treatment of melanoma brain metastasis, therapy resistance and relapse of disease remain unsolved challenges. CCT196969 is a SRC family kinase (SFK) and Raf proto-oncogene, serine/threonine kinase (RAF) inhibitor with documented effects in primary melanoma cell lines in vitro and in vivo. Using in vitro cell line assays, we studied the effects of CCT196969 in multiple melanoma brain metastasis cell lines. The drug effectively inhibited proliferation, migration, and survival in all examined cell lines, with viability IC50 doses in the range of 0.18–2.6 μM. Western blot analysis showed decreased expression of p-ERK, p-MEK, p-STAT3 and STAT3 upon CCT196969 treatment. Furthermore, CCT196969 inhibited viability in two B-Raf Proto-Oncogene (BRAF) inhibitor resistant metastatic melanoma cell lines. Further in vivo studies should be performed to determine the treatment potential of CCT196969 in patients with treatment-naïve and resistant melanoma brain metastasis.publishedVersio

Oncostatin M-induced and constitutive activation of the JAK2/STAT5/CIS pathway suppresses CCL1, but not CCL7 and CCL8, chemokine expression

The recruitment of leukocytes to injured tissue is crucial for the initiation of inflammatory responses as well as for immune surveillance to fight tumor progression. In this study, we show that oncostatin M, a member of the IL-6-type cytokine family and potent proinflammatory cytokine stimulates the expression of the chemokines CCL1, CCL7, and CCL8 in primary human dermal fibroblasts at a faster kinetic than IL-1beta or TNF-alpha. The production of CCL1 and CCL8 is important for migration of monocytes, while specific Abs against CCL1 additionally inhibit the migration of T lymphocytes. We identify the mitogen-activated protein kinases ERK1/2 and p38 as crucial factors for the enhanced expression of CCL1 and CCL8. Depletion of the ERK1/2 target genes c-Jun or c-Fos strongly decrease CCL1 and CCL8 expression, while p38 MAPK prolongs the half-life of CCL1, CCL7, and CCL8 mRNA through inhibition of tristetraprolin. None of the STAT transcription factors STAT1, STAT3, or STAT5 stimulate transcription of CCL1 or CCL8. However, we identify a negative regulatory function of activated STAT5 for the gene expression of CCL1. Importantly, not STAT5 itself, but its target gene cytokine inducible SH2-domain containing protein is required for the STAT5 inhibitory effect on CCL1 expression. Finally, we show that constitutive activation of STAT5 through a mutated form of JAK2 (JAK2 V617F) occurring in patients with myeloproliferative disorders similarly suppresses CCL1 expression. Taken together, we identify novel important inflammatory target genes of OSM which are independent of STAT signaling per se, but depend on MAPK activation and are partly repressed through STAT5-dependent expression of cytokine inducible SH2-domain containing protein

Оценка выбросов в атмосферу отходящих дымовых газов ТЭЦ и технологий их переработки

Сравнительная оценка выбросов в атмосферу отходящих дымовых газов, при сжигании органических топлив и способы их переработки.Comparative assessment of emissions of flue gases into the atmosphere, during combustion of organic fuels and methods of their processing

A novel nuclear antigen

The human oestrogen receptor (hER) mediates some effects of the steroid hormone oestrogen and functions as a ligand-dependent transcription factor in the nuclei of oestrogen-sensitive cells. The measurement of hER levels in breast cancer biopsies provides useful clinical information regarding therapy and prognosis. The current study describes a monoclonal antibody raised against hER aa 497-507 which recognises a novel nuclear antigen. Monoclonal antibodies were raised by immunising mice with a synthetic fragment of the hER (aa 497-507) conjugated to keyhole lymphocyte haemocyanin. Thirty antibody secreting hybridomas were identified. Hybridoma supernatants were characterised by enzyme-linked immunosorbent assay (ELISA), immunological staining using MCF-7 cells, binding studies, and SDS-PAGE Western blotting. One supernatant (15F6) displayed nuclear staining in fixed MCF -7 cells. Staining could be abolished by pre-incubation of the supernatant with the aa 497-507 peptide and peptide conjugates, but not with an unrelated hER peptide (aa 256-275). This antibody also stained the nuclei of hER negative breast cell lines MDA-MB-231 and MDA-MB-330, the breast cell line T47D, and liver cell line HepG2. Immunological staining of human tissue sections reveal the antigen to be present in the nuclei of keratocytes in skin and tubule and luminal endothelial cells of the kidney. The antibody identified a 120 kD band on Western blots with cytosols prepared from human breast cell lines and in solubilised cells. The antibody does not precipitate 16a-iodooestradiol-labelled ER from MCF-7 cells. Expression-linked screening of the MCF-7 cDNA library with antibody 15F6 identified nine positive clones. Antibody staining could be blocked by pre-incubating the antibody with hER aa 497-507-BSA conjugate, but not with an unrelated hER peptide conjugate. The (260 bp) clones were found to be identical. Submission of sequence to BLAST protein and nucleotide databases revealed a lack of homology to known proteins and genes. Sequence was matched to expressed sequence tags (ESTs) from brain, liver/spleen, uterus, ovary, colon, heart, and placenta. To further define the epitope of antibody 15F6, the sequence was translated and three peptides containing potential epitopes, comparable to the hER aa 497-507 region, were synthesised and tested by ELISA. The putative epitope was shown to be contained within one of these peptides

The PD-L1- and IL6-mediated dampening of the IL27/STAT1 anticancer responses are prevented by a-PD-L1 or a-IL6 antibodies

Interleukin-27 (IL27) is a type-I cytokine of the IL6/IL12 family and is predominantly secreted by activated macrophages and dendritic cells.We show that IL27 induces STAT factor phosphorylation in cancerous cell lines of different tissue origin. IL27 leads to STAT1 phosphorylation and recapitulates an IFN- -like response in the microarray analyses, with up-regulation of genes involved in antiviral defense, antigen presentation, and immune suppression. Like IFN- , IL27 leads to an up-regulation of TAP2 and MHC-I proteins, which mediate increased tumor immune clearance. However, both cytokines also upregulate proteins such as PD-L1 (CD274) and IDO-1, which are associatedwith immune escape of cancer. Interestingly, differential expression of these geneswas observed within the different cell lines and when comparing IL27 to IFN- . In coculture experiments of hepatocellular carcinoma (HCC) cells with peripheral blood mononuclear cells, pre-treatment of the HCC cells with IL27 resulted in lowered IL2 production by anti-CD3/-CD28 activated T-lymphocytes. Addition of anti-PD-L1 antibody, however, restored IL2 secretion. The levels of other TH1 cytokines were also enhanced or restored upon administration of anti-PD-L1. In addition, we show that the suppression of IL27 signaling by IL6-type cytokine prestimulation— mimicking a situation occurring, for example, in IL6-secreting tumors or in tumor inflammation–induced cachexia—can be antagonized by antibodies against IL6-type cytokines or their receptors. Therapeutically, the antitumor effects of IL27 (mediated, e.g., by increased antigen presentation) might thus be increased by combining IL27with blocking antibodies against PD-L1 or/and IL6-type cytokines

Systematic transcriptional profiling of responses to STAT1- and STAT3- activating cytokines in different cancer types

Cytokines orchestrate responses to pathogens and in inflammatory processes but they also play an important role in cancer by shaping the expression levels of cytokine response genes. Here, we conducted a large profiling study comparing miRNome and mRNA transcriptome data generated following different cytokine stimulations. Transcriptomic responses to STAT1- (IFN, IL-27) and STAT3-activating cytokines (IL6, OSM) were systematically compared in nine cancerous and nonneoplastic cell lines of different tissue origins (skin, liver and colon). The largest variation in our datasets was seen between cell lines of the three different tissues rather than stimuli. Notably, the variability in miRNome datasets was a lot more pronounced than in mRNA data. Our data also revealed that cells of skin, liver and colon tissues respond very differently to cytokines and that the cell signaling networks activated or silenced in response to STAT1- or STAT3- activating cytokines are specific to the tissue and the type of cytokine. However, globally, STAT1-activating cytokines had stronger effects than STAT3-inducing cytokines with most significant responses in liver cells, showing more genes up-regulated and with higher fold change. A more detailed analysis of gene regulations upon cytokine stimulation in these cells provided insights into STAT1- versus STAT3-driven processes in hepatocarcinogenesis. Finally, independent component analysis revealed interconnected transcriptional networks distinct between cancer cells and their healthy counterparts

CCT196969 effectively inhibits growth and survival of melanoma brain metastasis cells

Melanomas frequently metastasize to the brain. Despite recent progress in the treatment of melanoma brain metastasis, therapy resistance and relapse of disease remain unsolved challenges. CCT196969 is a SRC family kinase (SFK) and Raf proto-oncogene, serine/thre onine kinase (RAF) inhibitor with documented effects in primary melanoma cell lines in vitro and in vivo. Using in vitro cell line assays, we studied the effects of CCT196969 in multiple melanoma brain metastasis cell lines. The drug effectively inhibited proliferation, migration, and survival in all examined cell lines, with viability IC50 doses in the range of 0.18–2.6 μM. Western blot analysis showed decreased expression of p-ERK, p-MEK, p-STAT3 and STAT3 upon CCT196969 treatment. Furthermore, CCT196969 inhibited viability in two B Raf Proto-Oncogene (BRAF) inhibitor resistant metastatic melanoma cell lines. Further in vivo studies should be performed to determine the treatment potential of CCT196969 in patients with treatment-naïve and resistant melanoma brain metastasis

Совершенствование системы вознаграждения персонала бюджетного учреждения на основе комплексной оценки его деятельности

Объектом исследования является МБОУ 슫СОШ № 18슻 Цель работы - является оценка системы вознаграждения труда персонала бюджетного учреждения, ее оптимизации на основе комплексной оценке его деятельности В процессе исследования проводились способы группировки и сравнения, графические и аналитические методы. В результате исследования были выявлены недостатки и предложен комплекс мер по их устранению. Степень внедрения: сделать систему оплаты труда взаимосвязанной с конкретным результатом каждого работника и учреждения в целом. Область применения: бюджетные учреждения. Экономическая эффективность работы заключается в объективной оценке оплаты труда в бюджетном учреждении и в разработке практических рекомендаций по совершенствованию системы оплаты труда в анализируемом учреждении.The object of study is MBOU "SOSH № 18" Purpose - is to assess the remuneration system of the personnel budget of the institution, its optimization on the basis of a comprehensive assessment of its activities In the process of investigation the methods of grouping and comparison of graphical and analytical methods. The study revealed shortcomings and proposed the complex of measures on their elimination. Level of implementation: to make the wage system is interconnected with the concrete result of each employee and the institution as a whole. Scope: budget companies. Economic efficiency is an objective assessment of remuneration in budgetary institutions and in the development of practical recommendations for improving the system of remuneration in the analyzed institution