National Repository of Grey Literature 27 records found  1 - 10nextend  jump to record: Search took 0.00 seconds. 
Synthesis of biologically active compounds with quinazoline scaffold
Štěpánek, Ondřej
This thesis concludes my contribution to research of HIV-1 capsid assembly inhibitors. It has been shown that 2,4-disubstituted quinazoline derivatives are able to inhibit this process both, in competitive biochemical assay based on the AlphaScreen technology as well as in tissue cultures. The main objective of the work was to prepare the aforementioned quinazolines, to design and prepare new candidates with higher activity based on results of biochemical tests, and also to try to increase the solubility of otherwise poorly soluble compounds. Disubstituted quinazolines are relatively easily accessible from the commercially available anthranilic acid derivatives. These are converted to the corresponding quinazolin- 4(3H)-ones by the condensation reactions. In this work, two methods were used for the preparation of quinazolin-4(3H)-ones: reaction of acyl chlorides with aromatic anthranilamides provided 2-arylamidobenzamides whose subsequent cyclisation under basic conditions led to derivatives of 2-arylquinazolin-4(3H)-one; reaction of anthranilic acid esters with aromatic nitriles, which afforded desired quinazolin-4(3H)-one in one reaction step. Chlorination of 2-arylquinazolin-4(3H)-ones using POCl3 then led to 2-aryl-4- chloroderivatives as key intermediates. Nucleophilic substitution of...
The Role of Lck Kinase in T-cell Antigen Receptor Signaling
Němec, Dušan ; Štěpánek, Ondřej (advisor) ; Rösel, Daniel (referee)
LCK activity is crucial for the triggering of the entire T cell activation process. The primary function of LCK is to convert the signal of TCR:pMHC ligation into the intracellular environment. The outcome of the LCK-triggered pathway is T cell activation, cytokine production, differentiation, and clonal expansion. This thesis provides a summary of recent knowledge about the unique position of LCK in the T cell signaling machinery as well as an overview of molecules and interacting partners that regulate LCK activity. It describes the importance of the LCK-coreceptor association for optimal TCR signaling and physiological thymocyte development and mentions discussed adaptor role of LCK in the T cells. Keywords: LCK, T-cell, antigen, kinase, enzyme
The Role of B-cell Receptor Signaling in Lymphoid Malignancies
Kochmannová, Kateřina ; Havránek, Ondřej (advisor) ; Štěpánek, Ondřej (referee)
The aim of this thesis is to review current knowledge about tumor-specific B cell receptor (BCR) signaling and related novel therapy options in B-cell malignancies with the main focus on non-Hodgkin lymphomas (NHL). To a certain degree, the pathogenic BCR signaling mirrors normal forms of BCR signaling, antigen-induced and tonic. Differences between antigen- dependent and antigen-independent forms of BCR signaling are well characterized in two major subtypes of diffuse large B-cell lymphoma, the most common type of NHL. In addition to the conventional chemotherapy, several BCR inhibitors targeting BTK and PI3K have been approved for the treatment of lymphoid malignancies. However, improvements in the tumor specificity, toxicity profiles and patients selection are needed. A better understanding of BCR signaling deregulation and overall tumor pathogenesis is believed to further improve NHL treatment outcomes. Keywords: B-cell malignancies, non-Hodgkin lymphoma, B-cell receptor, tumor signaling, targeted therapy, inhibitors
The role of S100A11 protein (calgizzarin) in rheumatoid arthritis pathogenesis
Navrátilová, Adéla ; Andrés Cerezo, Lucie (advisor) ; Štěpánek, Ondřej (referee)
Rheumatoid arthritis (RA) is a systemic connective tissue autoimmune disease which is characterized by persistent inflammation of the synovial tissue, joint destruction and visceral organs and vessels damage. The RA joint environment contains pro-inflammatory molecules and immune cells. Small calcium-binding proteins of the S100 family are also known to be involved in the inflammatory process of RA, and some of them have been studied in our laboratory. One member of this family, S100A11 (calgizzarin), is well described in oncological diseases but its role in autoimmune diseases has not yet been described. Neutrophil extracellular traps (NETs), which are a potential source of autoantigens and immunoactive molecules, also contribute to the chronic inflammatory process in RA. The aim of our study was to investigate S100A11 with the focus on its role in the processes of inflammation and tissue destruction and to assess its possible association with NETs. Using in vitro experiments and molecular methods (reverse transcription, quantitative polymerase chain reaction (qPCR) and multiplex assay) we have shown that S100A11 exerts its pro-inflammatory effect on mononuclear cells via toll-like receptor (TLR)-4, however there was not any significant effect of S100A11 on the production of pro-inflammatory...
Characterization of T-cell clones from naïve and virtual memory compartment
Přibíková, Michaela ; Štěpánek, Ondřej (advisor) ; Drbal, Karel (referee)
Virtual memory (VM) CD8+ T cells represent a population of antigen-inexperienced T cells with an apparent memory phenotype. In lymphoreplete germ-free mice VM CD8+ T cells represent 10-20% of all peripheral CD8+ T cells. Their origin correlates with the levels of self-reactivity where the main factor that determinates the T-cell fate decision is the strength of homeostatic signals. In the first part of this thesis, we demonstrated that VM CD8+ T cells and naïve CD8+ T cells had distinct TCR repertoire and T-cell subsets contained different clonotypes. Moreover, 'VM clones' were enriched among VM T cells and were also present in naïve T cells. In contrast, 'naïve clones' were almost exclusively detected in naïve T cells. Next, we characterized the signaling of particular OVA-reactive TCRs from both naïve and VM subsets. We confirmed that 6 out of 8 tested TCRs were responsive to Kb-OVA. In the last part of the thesis, we developed and optimized a qPCR-based method for the relative quantification of specific T-cell clonotypes prior to and during the immune response. This method will serve as a tool for studying the biology of particular VM and naïve T-cell subsets and their role during the immune response. Keywords: T-cell receptor, homeostatic signaling, self-reactivity, virtual memory cells, T cells
Snižují regulační T lymfocyty riziko autoimmunity indukované CD8+ T lymfocyty?
Chadimová, Tereza ; Štěpánek, Ondřej (advisor) ; Šenolt, Ladislav (referee)
5 Regulatory T cells (Tregs) are essential for the maintenance of peripheral self-tolerance and prevention of autoimmunity by suppressing the response of self-reactive CD8+ and CD4+ T cells. However, while interactions of Tregs with CD4+ T cells have been extensively studied, their effect on the self-tolerance of CD8+ T cells has not been explored in detail. The main aim of this diploma project was to provide evidence whether and how Tregs prevent autoimmunity induced by CD8+ T cells. We used an experimental mouse model of autoimmune diabetes allowing us to acutely deplete Tregs and titrate the number of self-reactive T cells, self- antigen affinity, and self-antigen doses. We found out that Tregs play an important role in the prevention of CD8+ T-cell mediated autoimmunity. Moreover, we revealed that Tregs suppress both high-affinity T cells that escape negative selection and relatively weakly self-reactive, but numerous, positively selected T cells. Tregs do so by increasing requirement for the number of self-reactive CD8+ T cells required for the autoimmunity induction. Intriguingly, presence of Tregs does not impact threshold for self-antigen. Moreover, for the first time, we showed that Tregs can suppress CD8+ T-cell-mediated autoimmunity in the absence of conventional CD4+ T cells. This means that...
Pathophysiological development and differentiation of cells during hematopoiesis
Moudrá, Alena ; Štěpánek, Ondřej (advisor) ; Černý, Jan (referee) ; Kalina, Tomáš (referee)
In recent years, a great effort has been deployed towards a better understanding of the molecular changes in cells and in the bone marrow (BM) environment that contribute to the development and progression of myelodysplastic syndrome (MDS) to acute myeloid leukemia (AML). Among others, the aberrant hematopoietic stem cells in MDS often display increase in DNA double strand breaks, genomic instability with common loss or rearrangement of chromosomes and an ineffective response to DNA damage, a phenomenon that has been linked to the onset of cellular senescence. Additionally, the BM microenvironment can become more pro-inflammatory. In our effort to better understand the contribution of the BM microenvironment on MDS progression, we analyzed the expression profiles of cytokines in the BM microenvironment in all stages of MDS/AML and found several proinflammatory cytokines that increase with disease progression. Also, by repeated sampling of patients over the course of 5-azacytidine therapy, we were able to assess the changes in the proinflammatory cytokine milieu with the progression of the disease. Additionally, we aimed to identify the candidate markers for the improvement of MDS prognosis. We focused on naturally occurring germline polymorphism of NAD(P)H dehydrogenase (quinone 1) gene (NQO1*2)...
Modern mathematical methods in the research of immune reconstitutions and immunodeficiencies
Stuchlý, Jan ; Kalina, Tomáš (advisor) ; Šinkora, Jiří (referee) ; Štěpánek, Ondřej (referee)
In this thesis we present new analytical and integrative approaches for broad spectrum of applications of flow cytometry. CVID Large (88 individuals) cohort of CVID patients was immunopheno- typed by flow cytometry. The "probability binning" algorithm was used to au- tomatically assess the immunophenotype which presented unprecedented stabi- lity. We were able to define sub-group of CVID patients with highly activated immunophenotype showing hallmarks of immunosenescence and common clinical characteristics (thrombocytopenia, lung fibrosis and bronchiectasis). The severity of clinical complications correlated quantitatively with the immunophenotype. Analysis of cellular proteome We have designed technics which allow for highly multiplexed (>1000 of antibodies) analysis of human proteome using af- finity proteomics. We analyzed the changes of proteome of human cell lines and characterized the proteome of acute leukemias. The yet undescribed stability of proteome with respect the sub-cellular localization is shown and theoretical and practical background for antibody standardization and validation for the use in a nity proteomics is presented. Topological analysis of cytometry data Multiparametric flow cytometry allows for description of complex topological relationships of the cells in di erent stages of...
Role of Bardet-Biedl syndrome (BBS) protein complex in T cells
Niederlová, Veronika ; Štěpánek, Ondřej (advisor) ; Černý, Jan (referee)
BBSome is a protein complex crucial for trafficking of specific cargoes to the primary cilium. Although primary cilia are typically not present in cells of haematopoietic origin, such as T cells, recent research has revealed striking parallels between the primary cilium and the immunological synapse. Amongst other similarities, both structures are supposed to use the same transport machinery involving Rab8 and IFT20, the close interaction partners of BBSome. The first goal of this thesis was to investigate the role of BBSome in the biology of T cells. Using RT-qPCR, we have shown that BBSome subunits are expressed in lymphoid tissues and T cells. Studies of localization of BBSome subunits in Jurkat cell line and primary OT-I T cells revealed that the subunits have distinct localization patterns with BBS4 localizing to the centrosome and BBS1, BBS5, and BBip10 having dispersed localization. After the contact with an antigen presenting cell, BBS4 re-localizes to the immunological synapse. Mutations in BBSome encoding genes cause Bardet-Biedl syndrome (BBS), a rare ciliopathy presenting with multiorganic symptoms. The second goal of this thesis was to examine the associations between BBS and the immune system. Examination of medical records of more than 450 BBS patients revealed that autoimmune...

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