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Preparation of probes for the cancer imaging based on single photon emission computer tomography (SPECT) based on peptidomimetic inhibitors of FAP protease
Trajhan, Filip ; Konvalinka, Jan (advisor) ; Baszczyňski, Ondřej (referee)
Fibroblast activation protein (FAP) is a membrane serine protease that is mainly expressed in epithelial tumor cells. The specific occurrence in the tumor microenvironment and its absence in most healthy tissues makes this protein an important target for the development of diagnostic probes or targeted endoradiotherapy. The goal of this work is the preparation of a new series of potential probes for SPECT imaging of tumours using low molecular weight FAP inhibitors. The structure and synthetic strategy for four new probes were proposed. At the same time two already published probes serving as references for biodistribution experiments were prepared. Two probes were based on the compound IOCB22-AP446, the most potent FAP inhibitor published so far, which was developed in the laboratory of prof. Konvalinka at the IOCB, Prague. The other four prepared probes used the inhibitor UAMC1110, which is utilized as a targeting molecule in a number of compounds in the clinical testing phase. The degree of FAP inhibition in vitro was determined for all prepared compounds. Furthermore, the ability of the probes to visualise tumour tissue in vivo was tested on two animal models of human tumours. Comparison of the results with published data led to the validation of one of the models for use in further...
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Structural biochemistry of protein NKp80, an activation receptor of human lymphocytes
Pazderová, Kristýna ; Vaněk, Ondřej (advisor) ; Konvalinka, Jan (referee)
Natural Killer (NK) cells can spontaneously destroy target cells, such as infected, damaged, or malignant cells. NK cell cytotoxicity is mediated by activating receptors on their surface, such as NKp80 (Natural Killer protein 80 kDa). The human receptor NKp80 stimulates cytotoxicity through its interaction with its ligand AICL (Activation Induced C-type Lectin), which is constitutively expressed by all myeloid cells. In pathological conditions, such as cancerous or damaged cells, AICL is often upregulated, resulting in the lysis of these cells by NK cells expressing NKp80. This interaction is thus a promising immunotherapeutic target for treating myeloid leukaemia. However, the structures of both proteins have remained elusive. Therefore, we have focused on the successful production of the extracellular domain of these proteins. To improve the production yield of NKp80, we introduced a series of mutations in the stalk region to study their effect on production, stability, and homodimer formation. The introduced mutations significantly increased the production yield allowing for a large-scale production with subsequent crystallisation of the protein. The crystallisation resulted in the elucidation of the hitherto unknown structure of NKp80 homodimer at a resolution of 2.9 Å. However, the large-scale...
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Methods of measurement of bone density
Konvalinka, Jan ; Valášek, Jiří (referee) ; Marcián, Petr (advisor)
This bachelor's thesis focuses on methods for measuring bone density and consequences of changes in bone density on biomechanical properties of bone tissue. In first part are described methods of bone densitometry, especially radiological methods, which are currently most widely used. Second part focuses on measuring bone density from CT images (hog femur bone, healthy and damaged human femur bone and lumbar vertebra) using software ROI Analysis. Third part discusses influence of changes in bone density on strain-stress quantities of trabecular bone. For the assessment was created 2D computational model of hip using FEM.
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Stress-strain analysis of tumorous total endoprosthesis of hip joint
Vystrk, Tomáš ; Konvalinka, Jan (referee) ; Marcián, Petr (advisor)
Tumorous total endoprosthesis of hip joint is used for reconstruction of the limb affected by bone tumor and to regain its functnion. Longterm load conditions must be considered by the reconstruction and the joint mobility must be enabled. Suitability of the endoprosthesis depends on its construction considering shape, material and with this related mechanical properties. This thesis deals with creation of computational model and stress-strain analysis of tumorous total endoprosthesis of hip joint. There are two different types of endoprosthesis modeled in this thesis, each with three different length of resected bone replacement. Based on stress-strain analysis an assessment of possible limit states is made.
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Relationship of force and deformation characteristics to the energy of deformed system
Kupka, Oldřich ; Konvalinka, Jan (referee) ; Horníková, Jana (advisor)
The bachelor´s thesis deals with the description of energy in deformed systems and their use to determine the force acting on the system and displacements caused by forces. In the first part of the thesis there are described the basic types of stresses and how it is possible to determine their strain energy and displacements. After that, strain and potential energy including their complementary components are described. Next part is explanation of the principle of virtual work, Betti's theorem and Maxwell's reciprocity theorem. In the second part of thesis, the first and the second Castigliano´s theorems are derived. Castigliano´s theorems are applied to a statically determinate and indeterminate problems.
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Analysis of impact of mechanical properties of components of pathological arterial wall on its state of stress
Bartoňová, Petra ; Konvalinka, Jan (referee) ; Burša, Jiří (advisor)
The thesis deals with measurements of the atherosclerothic carotid plaques mechanical properties with wiew towards the improvements of their constitutive models and then towards the impact of application of those constitutive models in fibrous cap. The introductory part provides a brief summary of medical knowledge needed to understand atheroma formation and the risks involved. Next, there's a survey of plaque mechanical testing methods and the obtained results on tissue properties. The previously used constitutive models of atherosclerothic tissues are listed as well. Main body of the thesis describes the preparation of testing samples from obtained atheroma, methodology and course of experiments execution and subsequent processing of the experimental data to evaluate the behavior of the atheromatous tissue individual parts. Then the discussion and evaluation of our results and findings is given. Along with the evaluated mechanical properties, experimental data fitted to models are presented with respect to their predictive abilities. The final part is dedvoted to the finite element analysis of an idealized 2D model with emphasis on the effect of necrotic core and fibrous tissue properties on von Mises equivalent stress in the fibrous cap.
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Strain and stress analysis of the femur with distraction intramedullary nail
Konvalinka, Jan ; Marcián, Petr (referee) ; Florian, Zdeněk (advisor)
This master's thesis is focused on determination and analysis of stress and strain in femur with distraction intramedullary nail for treating leg length discrepancy with the method of distraction osteogenesis. Thesis is mainly focused on states after distraction when the callus consolidates. Problem of determining stress and strain is solved by computational modeling using FEM. Detailed description of modeling is included in this thesis, complicated 3D geometry of bone was acquired from segmentation of CT images. Computational model is solve with 4 different types of callus geometry and also material properties of callus are varied. The influence on stress and strain when the middle distal screw is not applied is also analyzed.
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Studies on structure and biological functions of NKR-P1 receptors
Rozbeský, Daniel ; Novák, Petr (advisor) ; Konvalinka, Jan (referee) ; Drbal, Karel (referee)
Natural killer (NK) cells play a significant role in the detection and destruction of virally infected and tumor cells. The NKR-P1 receptors regulate NK cell function by an alternative missing-self recognition system. Although the NKR-P1 receptors were among the first surface NK receptors identified on rodent NK cells more than 20 years ago, there is still very little known about their biological function and their physiological ligands. Furthermore, no three-dimensional structure of any of the NKR-P1 family receptors has been published so far. To understand the functional architecture of mouse NKR-P1 receptors, we developed a simple and efficient protocol providing large amounts of pure soluble NKR-P1 proteins. The crystal structure of mouse NKR-P1A, determined at 1.7 A resolution, is the first structure of a representative of the NKR-P1 family. Crystal structure is formed by a compact C-type lectin-like domain and an extended loop that participates in domain swapping. A potential role of the swapped loop has been suggested in natural ligand binding by in silico studies. However, chemical cross-linking and H/D exchange in combination with high resolution mass spectrometry revealed this loop in close proximity to the compact core in solution. The discrepancy between the crystal and solution structure...
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Production and characterisation of human C1 inhibitor and Plasmodium falciparum PfMSP3.1 recombinant proteins for structural studies
Čápová, Kateřina ; Konvalinka, Jan (advisor) ; Heidingsfeld, Olga (referee)
PfMSP3.1 is one of the surface proteins of the intracellular parasite Plasmodium falciparum, which causes malaria. As one of the evasion strategies of the immunity system of the human host this protein interacts with one regulator of the complement system - C1 inhibitor. Determining the exact binding site and its structural assessment would help to better understand the interaction between the parasite and the host, which is necessary for the disease progression and thus for the development of a potential therapy. In the theoretical part of the thesis, the life cycle of Plasmodium falciparum, the role of the parasite stage called merozoite, the role of its surface proteins, including merozoite surface protein 3, in the attack of red blood cells by the parasite, are described in more detail. It also briefly describes the complement system, its activation pathways and the regulation of these pathways. The experimental part includes the cloning of plasmids to produce C1 inhibitor and various forms of merozoite surface protein PfMSP3.1, transfection of S2 insect cells with these plasmids, subsequent protein expression in S2 cells and their purification. In the second half of the experimental part, we tried to create complexes of C1 inhibitor with individual PfMSP3.1 forms and an attempt to crystallize...
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