|
|
Inhibition of P-glycoprotein-mediated multidrug resistance and STAT3 signaling pathway through polymeric conjugates bearing protease inhibitor derivatives
Starenko, Daniil ; Kovář, Marek (advisor) ; Truksa, Jaroslav (referee)
Tumor cells expressing high levels of some ABC transporters (mainly P-glycoprotein) can become resistant to many structurally and functionally different drugs. Such multidrug resistance can be a significant barrier for a successful chemotherapy of malignant diseases. There is a considerable amount of small-molecular-weight compounds capable of potent inhibition of P-glycoprotein, but none of them are approved for the clinical use. STAT3 is a transcription factor important for many physiological processes, but its constitutive activation may lead to the malignant transformation and chemotherapy resistance in tumor cells. This molecule is thus potential target for anticancer drugs. The inhibition of STAT3 signaling should lead to lower cancer cell proliferation and their increased susceptibility to induction of apoptosis. Considerable attention is given to increase the effectiveness and to lower the adverse effects of conventional cytostatic agents via using nanomaterials and drug delivery systems in the research of new cancer therapy approaches. Polymeric carriers based on N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers are promising candidates in this field. The main aim of this diploma thesis was to evaluate the effectiveness of several HIV protease inhibitor (ritonavir, lopinavir, indinavir,...
|
|
|
Antitumor activity of the polymeric conjugates bearing derivatives of protease inhibitors and conventional cytostatics for the treatment of head and neck tumors
Běhalová, Kateřina ; Kovář, Marek (advisor) ; Vodička, Pavel (referee)
Head and neck cancers account for about 4,6 % of all malignancies worldwide and their incidence is increasing. However, the development of chemotherapeutics in this field is rather stagnating. One promising approach seems to be the repurposing of drugs originally developed and clinically used as HIV protease inhibitors, which have also been described to have anticancer activity. Esterification of the OH group of these drugs with 5-methyl-4-oxohexanoic acid allows their attachment via pH-sensitive hydrazone linkage to a hydrophilic and biocompatible HPMA copolymer carrier. This binding provides an improvement in the pharmacokinetics of the drug, prolongs its circulation time in the bloodstream, lowers its side effects and it also allows passive accumulation of the drug in the tumor tissue due to the EPR effect. Six protease inhibitors in total (ritonavir, lopinavir, saquinavir, indinavir, nelfinavir and atazanavir) were derivatized as described above. Ritonavir and its derivative had been tested in a previous project and were used as reference substances. After initial determination of in vitro cytostatic and cytotoxic activity in FaDu (human head and neck carcinoma), SCC7 (murine squamous cell carcinoma), 4T1 (murine breast carcinoma) and CT26 (murine colon carcinoma) cell lines, lopinavir derivative was...
|
|
|
Parasite cystatins as inhibitors of cysteine proteases: structural aspects of functional specificity and their evolution
Buša, Michal ; Mareš, Michael (advisor) ; Hudeček, Jiří (referee) ; Kukačka, Zdeněk (referee)
Members of the cystatin family are important inhibitors of cathepsin-type cysteine proteases and are involved in a number of pathologies. Parasite cystatins are attractive target molecules for parasite control, but our knowledge about them is still limited. This work is focused on cystatins of two blood-feeding parasites: the common tick (Ixodes ricinus) as the main vector of Lyme disease and tick-borne encephalitis, and the liver fluke (Fasciola hepatica), the causative agent of fasciolosis. Four novel cystatins were functionally and structurally characterized to determine the structural determinants of their inhibitory specificity and describe them in the context of evolution and physiological role of cystatins. The cystatin FhCyLS-2 from F. hepatica has broad inhibitory specificity and is suggested to play a dual role in the regulation of proteolytic systems in host tissue and the parasite gut. FhCyLS-2 combines the characteristics of two cystatin subfamilies in a unique way and is a model representative of a novel evolutionary group of cystatins identified in several orders of parasitic flukes. Ricistatin and iristatin are salivary cystatins of I. ricinus with immunomodulatory effects on the host caused by an exceptionally narrow inhibitory specificity. It was explained by structural modifications of...
|
|
|
Functional analysis of tick salivary serine and cysteine protease inhibitors
KOTÁL, Jan
The proposed thesis focuses on the characterization of two protease inhibitors present in tick saliva. More specifically, the thesis presents immunomodulatory properties, biochemical specificity and structure of a cysteine protease inhibitor named Iristatin. Another characterized protein, IRS-8, comes from a serpin family (serine protease inhibitors) and inhibits blood coagulation and complement system in the host. Furthermore, the thesis provides a literature overview and discussion of tick salivary molecules in the context of tick-host-pathogen interaction, vaccination potential and medicine potential. Two review manuscripts, which are part of this thesis summarize the effects of tick saliva and protease inhibitors on host immune mechanisms.
|
|
|
IrAM4: Partial characterisation of a molecule similar to \recke{alpha}\dindex{2}-macroglobulin from a tick \kur{Ixodes ricinus}
ABSOLONOVÁ, Markéta
Ixodes ricinus is a hard tick that can transmit several diseases that are capable of affecting humans. Among those are tick-borne encephalitis and Lyme disease. The focus of this work is on IrAM4, a member of tick ?2-macroglobulin family (?2M-F) of proteins which belong to the evolutionarily oldest constituents of the innate immune system. ?2-macroglobulins are protease inhibitors and act primarily in inactivation of proteases secreted by invading pathogens within their infection cycle. The aim of this study was to identify ?2-M of Ixodes ricinus (IrAM4) from corresponding ortholog of protein ?2-M named IsAM4 present in the genome of closely related Ixodes scapularis. The partial sequence was determined by amplification of cDNA and subsequent sequencing of PCR products. RT-PCR tissue profiling revealed that IrAM4 is present in ovaries and salivary glands but not in the tick gut. The recombinant fragment of IrAM4 was afterwards prepared for immunization of a rabbit and the obtained polyclonal antibodies were used for Western blot analysis. The results showed that IrAM4 is mainly present in the hemolymph and probably in salivary glands and ovaries but it is not expressed in the gut. The native IrAM4 seems to be composed of two disulfide bound subunits. However, the exact structure of the molecule was not analyzed in this work.
|
guest ::
