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Circulating tumor DNA in urine and its use as a potential biomarker of colorectal cancer
Surkovová, Veronika ; Ptáčková, Renata (advisor) ; Dračínská, Helena (referee)
Colorectal cancer (CRC) is a serious disease whose incidence has been steadily increasing. There is a high chance of cure and a good quality of life, if the treatment is started in the early stages of this disease. The so-called liquid biopsy, in which biomarkers from various body fluids are obtained and analyzed, could help with early detection of the disease, monitoring of the patient's condition and appropriate determination of the diagnosis. The aim of this study is to isolate and analyse circulating tumor DNA (ctDNA) from the urine of patients with advanced CRC. In these patients the ctDNA was detected in plasma. The method used for analysis is based on the principle of PCR with the formation of heteroduplexes, followed by the separation of "wild type" and mutated DNA fragments by denaturing capillary electrophoresis. Unlike plasma samples, the presence of KRAS mutations was not confirmed in urine samples. Neither the established procedure for plasma analysis, the attempt to enrich ctDNA by so-called purification, nor other tested isolation kits using the selected detection method allowed detection of the KRAS mutation in urine via ctDNA. Although urine ctDNA testing in general appears to be a promising method for improving the treatment level of (colorectal) cancer, and although its use could...
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Long-term monitoring of ctDNA levels in patients with metastatic colorectal cancer for early detection of progression or recurrence of the disease
Kopalová, Dominika ; Benešová, Lucie (advisor) ; Kološtová, Katarína (referee)
Circulating tumor DNA (ctDNA) in peripheral blood of patients with metastatic colorectal cancer appears to be a promising molecular marker that provides various applications. ctDNA levels vary depending on the presence, alternatively on the volume of tumor mass within patient's body, which can be used primarily for early detection of disease progression or recurrence and moreover for evaluating radicality of surgical treatment, all within long-term postoperative follow-up of the patient. Due to minimal invasivity of ctDNA analysis from peripheral blood (so-called liquid biopsy), it is possible to perform it repeatedly at relatively short time intervals. On account of very low fraction of ctDNA in total cell-free DNA (cfDNA) ranging between units and hundreds of percent, the key factor is optimal methodology covering all steps from the isolation process to a sufficiently sensitive detection technology. In this thesis I focus on an optimization of isolation process and analysis of ctDNA obtained from tumor tissue and plasma of selected patients with metastatic colorectal cancer in connection with surgical radicality and correlation with a clinical status of the patients.
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Circulating cell-free DNA and its potential in oncology
Karasová, Dominika ; Žižková, Hana (advisor) ; Novák, Josef (referee)
The circulating free DNA ("Cell-free DNA",cfDNA) is a type of extracellular DNA present in blood, with clinical use in prenatal diagnosis. The cfDNA is given great emphasis in connection with tumor diseases, where there is a great potential for its use in oncology, where free circulating tumor DNA is the most commonly used type. A suitable alternative to classical biopsy, which is safer and less painful for the patient and reflects the heterogeneity of the tumor is called Liquid biopsy, which can be used to obtain free circulating DNA from the blood. Most studies show that cfDNA provides the same information as tissue DNA, regarding both genetic and epigenetic changes detected in tumor cells and in some cases it enabled more sensitive or earlier detection of relapse in comparison with routinely used examinations. The high specificity and sensitivity of this non-invasive approach can be a great benefit to patients. However, to confirm its diagnostic and prognostic significance, it is necessary to expand the group of patients with individual diagnoses and, not only for the purpose of data comparison, standardize methods for the isolation and detection of cfDNA, The aim of this thesis is to gain a general overview of free circulating DNA and its potential use in oncology. Keywords: circulating free...
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