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Development of nanochemical tools targeting receptors in the tumor microenvironment
Blažková, Kristýna ; Konvalinka, Jan (advisor) ; Abramson, Jakub (referee) ; Bušek, Petr (referee)
Development of nanochemical tools targeting receptors in the tumor microenvironment Targeting the receptors in the tumor microenvironment is crucial for the future development of targeted therapies, precision medicine and immunotherapy of cancer. The options available now are, however, limited by the availability of specific ligands. The advances in the field strongly rely on the use of antibodies and genetic modifications of immune cells. Availability of small molecules targeting the receptors of interest would allow further development of alternative strategies as well as deepen our understanding of the underlying mechanisms of cancer development, progression and clearance. In the search for new small-molecule ligands and their use for receptor targeting, the prostate-specific membrane antigen (PSMA) and the immune receptors CD3 and CD64 were selected as model targets. The selected method - the phage display of bicyclic peptides - utilizes chemical modification of the displayed three-cysteine peptides to achieve their cyclization and formation of bicycles. The panning of a peptide library displayed on the phages and probed with PSMA revealed a reproducibly-selected amino acid sequence. Interestingly, the phage clone carrying this sequence was a specific binder of PSMA, but the synthesized peptide alone...
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Selection approaches in directed evolution of binding proteins
Huličiak, Maroš ; Malý, Petr (advisor) ; Hlouchová, Klára (referee)
Artificial binding proteins derived from small protein domains attract attention as a promising alternative to monoclonal antibodies and can be used in many kinds of applications. They are useful in diagnosis of human diseases, seem to be a clue for more efficient vaccine development preventing from global diseases such as AIDS, can exhibit a therapeutic potential or improve purification techniques. For the selection of protein variants with desired properties such as high specificity and binding affinity, more than 10 different selection techniques have been developed. So called display techniques such as phage display, yeast display, retroviral display or baculovirus display are based on protein expression from different vectors. Contrary that, ribosome display, mRNA display and CIS display are cell-free systems based on in vitro translation. Development of different selection approaches allows production of post- translationally glycosylated, phosphorylated and acetylated proteins, increased yield of the produced binders and improved their binding properties. The submitted work provides an overview of current selection techniques, compare their parametres regarding to combinatorial libraries, describes their advantageous properties and limitations, and focus on a practical utilization of...
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