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Molecular Biomarkers of Anticancer Drugs Efficacy in Experimental Models and in Patients with Solid Tumors
Šeborová, Karolína ; Václavíková, Radka (advisor) ; Mohelníková Duchoňová, Beatrice (referee) ; Indra, Radek (referee)
Ovarian carcinoma is a heterogeneous group of diseases usually of epithelial origin. The vast majority of diagnosed cases are high-grade serous carcinomas, which account for over 80% of all cases. Ovarian carcinoma is one of the most common causes of death in women from gynecological malignancies. The main cause of high mortality is diagnosis of the disease in late stages due to the long-term asymptomatic course. The second important cause is the development of multidrug resistance to the administered treatment, which is in the case of ovarian carcinoma patients still mainly based on a combination of paclitaxel and platinum derivatives - cisplatin or carboplatin. These are some of the reasons that motivate scientists to search for new biomarkers, which could help detect the disease in early stages, or monitor the success of treatment, and predict the development of resistance. Molecular characterization of cancer in general may also reveal new therapeutic targets. The development of resistance to already existing drugs stimulates the direction of scientists to prepare new drugs that would be especially effective against already resistant tumors. The main aim of the presented theses was to expand information on the molecular profile of ovarian carcinoma, to reveal prognostic and predictive molecular...
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Analysis of hereditary genetic variants predisposing to the development of familial forms of ovarian cancer.
Lhotová, Klára ; Soukupová, Jana (advisor) ; Mohelníková Duchoňová, Beatrice (referee) ; Weinberger, Vít (referee)
Ovarian cancer (OC) is the deadliest gynecologic malignancy with a substantial proportion of hereditary cases and a frequent association with breast cancer (BC). Genetic testing facilitates preventive management for carriers of mutations in OC-susceptibility genes. However, the prevalence of germline mutations varies among populations and many rarely mutated OC predisposition genes remain to be identified. We analyzed 219 genes in 1333 Czech OC patients and 2278 population-matched controls (PMC) using next-generation sequencing. Altogether, 427/1333 (32%) patients and 58 /2278 (2,5%) PMC carried pathogenic mutations in 18 known/anticipated OC predisposition genes. Mutations in BRCA1, BRCA2, RAD51C, RAD51D, BARD1 and mismatch repair genes conferred a high OC risk (with OR>5). Mutations in BRIP1 and NBN were associated with moderate risk (both OR ≥2 - <5). BRCA1/2 mutations dominated in almost all clinicopathological subgroups including sporadic borderline tumors of ovary (BTO). Analysis of remaining 201 genes revealed somatic mosaics in PPM1D and germline mutations in SHPRH and NAT1 associating with a high/moderate OC risk significantly; however, further studies are warranted to delineate their contribution to OC development in other populations. Results of this study demonstrate the high proportion...
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Extracorporeal Elimination of Circulating Pegylated Liposomal Doxorubicin to Enhance the Benefit of Cytostatic Therapy in Ovarian Cancer
Kubeček, Ondřej ; Filip, Stanislav (advisor) ; Mohelníková Duchoňová, Beatrice (referee) ; Demlová, Regina (referee)
Extracorporeal elimination of circulating pegylated liposomal doxorubicin to enhance the benefit of cytostatic therapy in ovarian cancer Background: Encapsulation of cytotoxic drugs into liposomes changes their pharmacokinetic and toxicologic properties in order to enhance the benefit of chemotherapy. Pegylated liposomal doxo- rubicin (PLD) is one of the most prominent drugs in this group. Despite its reduced cardio- and hematologic toxicity, it confers a higher risk of mucocutaneous toxicity in comparison to its parent drug (i.e. doxorubicin, DOX). The specific pharmacokinetic properties of PLD enable its partial elimination from plasma after reaching maximum concentration within the tumor tissue in order to reduce its toxicity. Aim: This doctoral thesis evaluates the efficacy and safety of plasmafiltration (PF) as a me- thod of extracorporeal PLD elimination, seeking to enhance its therapeutical benefit. The reduc- tion of PLD-related toxicity thanks to the application of PF was determined. Further, PLD pharmacokinetics and the factors affecting it, as well as the effect of PF on the outcome of cancer therapy, were also analyzed. Material and methods: Sixteen patients with platinum-resistant epithelial ovarian cancer were enrolled in the study. PLD was administered as a one-hour i.v. infusion of...
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Morphological and Genomic Profiling of Circulating Tumor Cells in Metastatic Colorectal Cancer
Thiele, Jana-Aletta ; Pitule, Pavel (advisor) ; Mohelníková Duchoňová, Beatrice (referee) ; Kasimir-Bauer, Sabine (referee)
Colorectal cancer (CRC) is the third most common cancer worldwide; it is responsible for nearly 10% of all newly diagnosed cancers and is the second most cause of cancer related death in Europe. Biomarkers for therapy guidance, targeted therapy and survival prognosis are still limited. As CRC is a heterogeneous disease, different parts of the tumor might have varying molecular characteristics which may change during therapy or disease progression. Through solid biopsies and screenings, these local or temporal differences are impossible to monitor. To facilitate detection of these possible temporal changes, a regularly and non-invasively accessible biomarker is required for disease monitoring. Circulating tumor cells (CTCs) might represent such a biomarker as they have been shown to be fluid surrogates of the solid tumor. EpCAM positive CTCs have shown to be prognostic in CRC for survival, but their full potential has not yet been evaluated further. By using the High Definition Single Cell Analysis (HD-SCA) workflow, we were able to analyze the entire spectrum of CTCs and categorize them as the regular CTCs (HD-CTC), CTCs with a smaller nuclear area (CTC-Small), CTCs with low expression of epithelial marker cytokeratin (CTC-LowCK) and CTCs undergoing apoptosis and therefore releasing cell free DNA...
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The Changes of the Tumor Vascular Endothelial Growth Factor Expression and the Density CD8+ Tumor Infiltrating Lymphocytes after Neoadjuvant Chemoradiation in Patients with Rectal Adenocarcinoma
Buka, David ; Petera, Jiří (advisor) ; Vrána, David (referee) ; Mohelníková Duchoňová, Beatrice (referee)
Rectum adenocarcinoma is one of the most frequent tumour diseases in the Czech Republic. Therefore any progress in therapy or prevention of this disease can have a significant impact on mortality and morbidity of major amount of oncological patients. Within therapy the neoadjuvant concomitant radiochemotherapy or the sole radiotherapy is used before surgical resection in one third to half of all newly diagnosed cases. The goal is to achieve tumor downstaging and local relaps probability decrease. There is a large amount of processes on molecular and cellular level in organism reaction to neoplasia genesis, whose understanding can have a substantial impact on therapy's success rate or on therapy's efficiency prediction. One of these processes is angiogenesis induced by high production of vascular endothelial growth factor (VEGF). As per immunohistochemical studies VEGF is not expressed in typical colorectum mucous membrane. However it is strongly expressed in case of adenocarcinoma. Another important process of organism's malignancy response is tumour tissue infiltration by CD8+ T lymphocytes (TIL), which are one of the important components of cellular immunity against tumour and which significantly contribute to tumour cells elimination. Potential of CD8+ T lymphocytes is considerable not just in...
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Study of genetic factors modifying the risk of onset and progression of colorectal and pancreatic cancer
Mohelníková Duchoňová, Beatrice ; Souček, Pavel (advisor) ; Skálová, Lenka (referee) ; Macek, Milan (referee)
Introduction: The aim of this study was to evaluate the role of genetic and lifestyle factors in the risk of onset and progression of colorectal and pancreatic cancer. The first part deals with the etiological factors and the importance of polymorphisms in biotransformation enzymes and genetic alterations in the gene CHEK2 in the origin of these malignancies. In the second part, the ABC transporter genes were analyzed as potential prognostic and predictive markers of a treatment's outcome. Materials and methods: The polymorphisms and other genetic alterations were detected using real-time PCR, allelespecific PCR and PCR-RFLP methods in DNA which was extracted from the blood of patients. The frequency of polymorphisms was evaluated and their importance was assessed with regard to the available epidemiological data. Gene expressions were determined by qPCR in paired samples of tumor tissue and adjacent non-tumorous parenchyma. Results: A majority of the observed polymorphisms failed to show a relationship between their presence and the risk of any of these malignancies. CYP2A13 variant allele*7 coding inactive enzyme was found in 7 of 265 controls and in none of 235 pancreatic carcinoma patients. In contrast, GSTP1-codon 105 Val variant allele and GSTT1-null genotype were associated with an elevated...
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Diagnostic and prognostic markers in the era of targeted treatment of CRC.
Veškrňová, Veronika ; Büchler, Tomáš (advisor) ; Mohelníková Duchoňová, Beatrice (referee) ; Valík, Dalibor (referee)
Introduction: Colorectal cancer (CRC) is the most common malignant tumor in both sexes in the Czech Republic. Prognostic factors in CRC can be classified as clinical (stage at the time of diagnosis, histological type of tumor), genetic (RAS, BRAF), immunological (Immunoscore)and biochemical (CEA, CA 19-9, miRNA). MicroRNAs (miRNAs) regulate the expression of oncogenes and tumor suppressors. The regulatory function of miRNAs is influenced by single nucleotide polymorphisms (SNPs) of target miRNA binding sites (miRSNPs). Aims: To evaluate the currently available prognostic factors for CRC patients treated using targeted therapies and assess the role of novel ones, including miRNA. Methods: The thesis includes clinical works focused on targeted treatment of colorectal cancer, original work focused on the role of miRNA in colorectal cancer pathogenesis and especially as a prognostic and predictive marker, work focused on functional polymorphisms of DNA repair genes and a review article summarizing biochemical factors influencing the effect of fluoropyrimidine cytostatics in the treatment of colorectal cancer. Results: We have identified miR-17/92 as a non-invasive biomarker for predicting post-treatment prognosis in patients with a higher risk of relapse, as well as miRSNPs rs8679 polymorphisms as a...
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Analysis of hereditary genetic variants predisposing to the development of familial forms of ovarian cancer.
Lhotová, Klára ; Soukupová, Jana (advisor) ; Mohelníková Duchoňová, Beatrice (referee) ; Weinberger, Vít (referee)
Ovarian cancer (OC) is the deadliest gynecologic malignancy with a substantial proportion of hereditary cases and a frequent association with breast cancer (BC). Genetic testing facilitates preventive management for carriers of mutations in OC-susceptibility genes. However, the prevalence of germline mutations varies among populations and many rarely mutated OC predisposition genes remain to be identified. We analyzed 219 genes in 1333 Czech OC patients and 2278 population-matched controls (PMC) using next-generation sequencing. Altogether, 427/1333 (32%) patients and 58 /2278 (2,5%) PMC carried pathogenic mutations in 18 known/anticipated OC predisposition genes. Mutations in BRCA1, BRCA2, RAD51C, RAD51D, BARD1 and mismatch repair genes conferred a high OC risk (with OR>5). Mutations in BRIP1 and NBN were associated with moderate risk (both OR ≥2 - <5). BRCA1/2 mutations dominated in almost all clinicopathological subgroups including sporadic borderline tumors of ovary (BTO). Analysis of remaining 201 genes revealed somatic mosaics in PPM1D and germline mutations in SHPRH and NAT1 associating with a high/moderate OC risk significantly; however, further studies are warranted to delineate their contribution to OC development in other populations. Results of this study demonstrate the high proportion...
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Pathophysiology of colorectal cancer. Colorectal cancer screening effect and the role of microRNA in pathophysiology of colorectal cancer.
Král, Jan ; Špičák, Julius (advisor) ; Keil, Radan (referee) ; Mohelníková Duchoňová, Beatrice (referee)
Colorectal cancer is a serious malignant disease with an incidence of over 1.8 million new cases per year worldwide. There are about 8 000 patients diagnosed with CRC in the Czech Republic each year, and about half of them present with an advanced disease. Screening program identifies patients in the early stages of CRC resulting in overall better prognosis and survival. There is also a lack of biomarkers of early CRC detection and of response to treatment. The first aim of our project was to conduct a national multicentre prospective observational study to evaluate the impact of CRC screening within the framework of a Czech population screening programme. Between March 2013 and September 2015, a total of 265 patients were enrolled in 12 centres across the Czech Republic. Patients were divided into screening and control groups and compared for pathology status and clinical characteristics. Screening was defined as a primary screening colonoscopy or a colonoscopy after a positive FOBT in an average-risk population. The distribution of CRC stages was significantly favourable in the screening group compared with the control group (stages 0, I and II, 63% versus 43.3%; p <0.001). The presence of distant (M1) and local metastases (N1 and N2) was significantly less prevalent in the screening group (0%,...
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Morphological and Genomic Profiling of Circulating Tumor Cells in Metastatic Colorectal Cancer
Thiele, Jana-Aletta ; Pitule, Pavel (advisor) ; Mohelníková Duchoňová, Beatrice (referee) ; Kasimir-Bauer, Sabine (referee)
Colorectal cancer (CRC) is the third most common cancer worldwide; it is responsible for nearly 10% of all newly diagnosed cancers and is the second most cause of cancer related death in Europe. Biomarkers for therapy guidance, targeted therapy and survival prognosis are still limited. As CRC is a heterogeneous disease, different parts of the tumor might have varying molecular characteristics which may change during therapy or disease progression. Through solid biopsies and screenings, these local or temporal differences are impossible to monitor. To facilitate detection of these possible temporal changes, a regularly and non-invasively accessible biomarker is required for disease monitoring. Circulating tumor cells (CTCs) might represent such a biomarker as they have been shown to be fluid surrogates of the solid tumor. EpCAM positive CTCs have shown to be prognostic in CRC for survival, but their full potential has not yet been evaluated further. By using the High Definition Single Cell Analysis (HD-SCA) workflow, we were able to analyze the entire spectrum of CTCs and categorize them as the regular CTCs (HD-CTC), CTCs with a smaller nuclear area (CTC-Small), CTCs with low expression of epithelial marker cytokeratin (CTC-LowCK) and CTCs undergoing apoptosis and therefore releasing cell free DNA...
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