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New biological markers in lymphoproliferative diseases
Špaček, Martin ; Mandys, Václav (advisor) ; Smolej, Lukáš (referee) ; Lysák, Daniel (referee)
New biological markers in lymphoproliferative diseases Summary The prognosis of patients with lymphoid neoplasms is extremely variable. Reliable prognostic markers could allow the identification of patient subsets that may benefit from alternate approaches. Epstein-Barr virus (EBV) is associated with approximately one third of Hodgkin lymphoma (HL) cases. EBV-DNA is often present in the plasma and whole blood of EBV-associated HL patients. However, the significance of EBV-DNA monitoring is debated. In a cohort of 165 adult HL patients EBV-DNA viral load was prospectively monitored. Our results suggest that assessment of plasma EBV-DNA viral load might be of value for estimation of prognosis and follow-up of patients with EBV-positive HL. The role of the TP53 gene's R72P polymorphism in NHL has been analyzed in several studies but it has not been studied in HL. We have evaluated the role of R72P in 340 NHL and 298 HL patients. Our results support the evidence that R72P is not a prognostic factor in Caucasian NHL patients, and they indicate its irrelevance for HL development or prognosis. In a study of 73 patients with chronic lymphocytic leukemia we have assessed IgVH mutational status, presence of mutation in TP53 gene and expression of CD38 and ZAP-70. Overall concordance between ZAP-70 expression and IgVH...
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Implementation of Immunohistochemical and Molecular-Genetic Methods in Differential Diagnosis of Urogenital and Gynecologic Tract Lesions
Ondič, Ondrej ; Hes, Ondřej (advisor) ; Mandys, Václav (referee) ; Vernerová, Zdeňka (referee)
This thesis focuses on gynecopathology. It consists of a collection of seven papers published in pathology journals with impact factor. Introduction section contains selection of examples showing scientific application of molecular genetic methods. Further on the aims of individual research projects are described. The first project comprises histomophologic study of skin endometriosis addressing "mullerian" differentiation. A case report of a rare tumor namely borderline papillary serous tumor of the fimbriated end of the fallopian tube follows with molecular genetic analysis of KRAS, BRAF and p53 gene mutation status. Prospective longitudinal study on high grade squamous dysplasia (HSIL) of the cervix in HPV vaccinated women, so called DAV (dysplasia after vaccination), aims to elucidate pathogenesis of this phenomenon. Two other studies focus on incidence of fumarate hydratase deficient leiomyomas of the uterus and hereditary leiomyomatosis and renal cell carcinoma syndrome (HLRCC). The aim of those studies is to improve our diagnostic capability and increase detection rate of the patients with HLRCC syndrome. Finally a new subtype of HSIL namely bizarre cell dysplasia is described in two separate studies. Conclusion remarks contemplate the role of molecular genetics in surgical pathology.
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Tumors of Urogenital Tract, Molecular Genetic Characteristics and Impact on Biological Behavior
Pivovarčíková, Kristýna ; Hes, Ondřej (advisor) ; Mandys, Václav (referee) ; Vernerová, Zdeňka (referee)
The Ph.D. thesis is a collection of seven commented articles on the topic of molecular-genetic of urogenital tumors. The spectrum of urogenital lesions is very wide, there is lots of new insights in field of urogenital pathology. Therefore, presented Ph.D. thesis is focused only on small part of urogenital neoplasms - renal cell carcinomas characterized by papillary growing pattern. The introduction presents different entities with papillary morphology, there are specified basic morphological, immunohistochemical and molecular-genetic characteristic of these lesions, their differential diagnosis is commented. In the results, there are presented recently published studies dealing with newly described entities of renal neoplasias, heterogenity of molecular genetic studies in individual tumorous groups is established, prognostic significance is commented, typical pitfalls are introduced. In the conclusions, the limitations of the use molecular-genetic methods are discussed and clinical outcome and significance is evaluated.
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Diagnostic morphological features of PDGFRA-mutated gastrointestinal
Daum, Ondřej ; Hes, Ondřej (advisor) ; Mandys, Václav (referee) ; Ehrmann, Jiří (referee) ; Zámečník, Josef (referee)
Daum O., Grossmann P., Vanecek T., Sima R., Mukensnabl P., Michal M. (2006): Diagnostic morphological features of PDGFRA-mutated gastrointestinal stromal tumors: Molecular genetic and histological analysis of 60 cases of gastric GISTs. Ann. Diagn. Pathol. In Press Summary In this study, 60 gastrointestinal stromal tumors (GISTs) of the stomach were analyzed to elucidate the possible relation of their morphology to mutation status of KIT and PDGFRA genes. The patients included 27 men and 33 women with a mean age of 63,8 years (range 12 to 92). Only one tumor occurred before the age of 21 years. KIT mutations were detected in 31 cases (51,7%), PDGFRA mutations in 22 cases (36,7%), and seven cases (11,7%) were KIT and PDGFRA wild type. When the mutation status was correlated with histological features of the tumors, epithelioid or mixed epithelioid/spindle cell pattern and mast cell infiltration were found as the most reliable signs of PDGFRA mutation. Neoplastic rhabdoid cells and multinucleated giant cells, also previously reported as features of PDGFRA mutated GISTs, seemed to be less specific but still helpful markers in our study. Finally, tumor infiltrating lymphocytes and myxoid stroma do not seem to be valuable histological signs. Daum O., Klecka J., Ferda J., Treska V., Vanecek T., Sima R., Mukensnabl...
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The role of selected molecules in biologic behaviour of human breast cancer
Gojiš, Ondřej ; Mandys, Václav (advisor) ; Kolář, Zdeněk (referee) ; Kinkor, Zdeněk (referee)
The role of selected molecules in biologic behaviour of human breast cancer Abstract Ondřej Gojiš In human breast cancer the rate of immunohistochemical discordance between primary tumour and secondary deposits were reported to range from 28% - 42% for ER and was 17% for PgR. Depending on the study and techniques utilized, discordance rates of Her2 ranged between 0% - 37%. It has been showed that PAX2 competes with SRC-3 for binding and regulation of HER-2 transcription. Human breast cancers that were PAX2 positive and SRC-3 negative had the lowest recurrence rate, and the relationship between PAX2 and SRC-3, with regard to levels that determine relapses, were found to be inversely dependent. This suggests a transcriptional link between the two subtypes of breast cancer, namely ER positive and HER2 positive tumors. Immunohistochemical profiles were performed for 30 metastatic deposits using antigens for SRC3, Pax2, ER, PgR, Her2, CK5/6, Ki67 and EGFR. SRC-3 was positive in 66.6% of cases and Pax2 in 73.3%. No correlation was observed between the patient's age, cancer grade, histotype, lymphatic node status, and protein expression. Using all seven antigens, the primary tumor's protein expression differed from that of the brain deposits in 13 cases out of 14 (93%). Constant expression of SRC3 and Pax2 was...
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The comparison of morphology, expression, epigenetic changes and mutations of HNF1B in solid tumors and non-neoplastic lesions.
Bártů, Michaela ; Dundr, Pavel (advisor) ; Mandys, Václav (referee) ; Škarda, Jozef (referee)
Introduction HNF1B is a tissue-specific transcription factor, which plays a crucial role in the embryological development of a number of organs, especially kidneys, gastrointestinal system, pancreas and billiary system. While the significance of HNF1B in the development of urinary tract malformations has already been well described, its role in the pathogenesis of solid tumors has not yet been elucidated. Based on the current data it seems that depending on the type of individual tumor HNF1B can either act as an oncogene or a tumor suppressor. However, the precise mechanism of how it exerts its influence is still unclear. Aims: The thesis focuses on expanding the knowledge of the significance of HNF1B changes in selected solid tumors and non-neoplastic lesions. The individual goals include: 1) determining the role which HNF1B plays in the pathogenesis of these lesions, 2) evaluating the significance of HNF1B for differential diagnosis, 3) analysis of the prognostic and predictive meaning of HNF1B, 4) mutation analysis of the HNF1B gene in all the tumor and non-tumor tissues with the aim to identify novel pathogenic mutations, 5) methylation analysis of the HNF1B promoter. Material and methods: Immunohistochemical examination with the antibody against HNF1B was performed on 516 samples of tumor and...
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The Study of Expression and Function of Selected Nuclear Receptors in Caenorhabditis Elegans
Vohánka, Jaroslav ; Kostrouchová, Marta (advisor) ; Nedvídek, Josef (referee) ; Mandys, Václav (referee)
6 Abstract: The genome of Caenorhabditis elegans encodes more than 280 nuclear hormone receptors (NHRs) in contrast to the 48 NHRs in humans and 18 NHRs in Drosophila. The majority of the C. elegans NHRs are categorized as supplementary nuclear receptors (supnrs) that evolved by successive duplications of a single ancestral gene. The evolutionary pressures that lead to the expansion of NHRs in nematodes, as well as the function of the majority of supnrs, are not known. Here, we have studied the expression of seven genes organized in a cluster on chromosome V: nhr-206, nhr-208, nhr-207, nhr-209, nhr-154, nhr-153 and nhr- 136. Reverse transcription-quantitative PCR and analyses using transgenic lines carrying GFP fusion genes with their putative promoters revealed that all seven genes of this cluster are expressed and five have partially overlapping expression patterns including in the pharynx, intestine, certain neurons, the anal sphincter muscle, and male specific cells. Four genes in this cluster are conserved between C. elegans and C. briggsae whereas three genes are present only in C. elegans, the apparent result of a relatively recent expansion. Interestingly, we find that a subset of the conserved and non-conserved genes in this cluster respond transcriptionally to fasting in tissue-specific patterns....
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Diagnostic contribution to the evaluation of intervention models in the treatment of type 1 diabetes
Zacharovová, Klára ; Saudek, František (advisor) ; Plecitá, Lydie (referee) ; Mandys, Václav (referee)
During treatment of diabetes mellitus by immunointervention or transplantation, it is necessary to monitor the markers of immune destruction or rejection of surviving insulin producing cells. An aim of this thesis is to improve the possibilities of following autoimmunity and to detect the survival of transplanted pancreatic islet in vivo. Partial aims included vitality testing of isolated islets for transplantation by measurement of respiration activity, observing the process of in vitro labeling of isolated islets with superparamagnetic iron oxide (SPIO) contrast agent for subsequent magnetic resonance imaging (MRI) of islets and observing SPIO particles transport after transplantation. We also studied a new dual paramagnetic contrast agent combined with fluorescein intended for identification of the MRI contrast agent in samples for histology. Further, we assessed autoimmune reaction by evaluation of cytokine response to specific stimulation with auto-antigens. We tried to affect beta-cells destruction by polyclonal anti- thymocyte antibodies in a mouse experimental model. A new method of the islet respiration measurement correlated with other methods of islet quality testing and it was suggested as a diagnostic test before clinical transplantation. Results obtained studying the intercellular...
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Dipeptidyl peptidase IV in orthotopic models of glioma
Hilšer, Marek ; Šedo, Aleksi (advisor) ; Mandys, Václav (referee) ; Šefc, Luděk (referee)
Malignant gliomas belong to a highly aggressive class of tumours. Average patient survival time generally does not exceed 15 months. Despite intensive research, no therapeutic strategies capable of significantly extending the lives of those affected by the disease have been established to date. One potentially viable area of research into possible therapeutic targets in cancer therapy focuses on cell surface proteases. This group of proteins includes dipeptidyl peptidase IV (DPP-IV). Changes to DPP-IV expression have been established in the case of various cancer types including malignant gliomas. Understanding the role of DPP-IV in the biological processes of this malignant disease may thus contribute to the development of new therapeutic modalities. This thesis is therefore dedicated to establishing an orthotopic xenograft model as well as a genetically engineered model (GEM) of the glioma. The effects of DPP-IV on the growth of an experimental glioma were subsequently examined, as was the ratio of catalytic and non- catalytic mechanisms in this process. The GEM model was used for monitoring enzymatic activity and DPP-IV distribution. Non-invasive fluorescence imaging was employed in order to monitor the intraexperimental dynamics of experimental gliomas. The results indicated that DPP-IV...
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Expression of miRNAs in pancreatic adenocarcinoma-relationship to morphological characteristics and disease prognosis.
Szabó, Arpád ; Mandys, Václav (advisor) ; Hermanová, Markéta (referee) ; Daum, Ondřej (referee)
Ductal adenocarcinoma (PDAC) is the most common primary pancreatic neoplasm. It's frequently diagnosed in late inoperable stage and has high resistance to chemotherapy; this situation contributes to its unfavourable prognosis. Therefore, there is a need for biomarkers enabling early detection of PDACs and by this way to improve patient prognosis. MicroRNAs (miRNA), short non-coding RNA molecules involved in post-transcriptional regulation of gene expression, belong to such markers. Contrary to other RNA molecules, miRNAs are stable in biological samples. Their expression is measured by several analytical methods, including real-time quantitative PCR (RT-qPCR). Normalization of methods determining miRNA levels requires adequate endogenous controls. However, variable expression of endogenous controls in tumors may cause bias in determining miRNA levels. The aim of the first study was to investigate the expression of six miRNAs isolated from formalin fixed paraffin embedded (FFPE) samples of PDACs. Four controls were chosen for RT-qPCR result normalization: artificial spike miR-39 from C. elegans, U6 snRNA, miR-16 and snoRNA U91. Expression values of all studied miRNAs in tumors were significantly different depending on selected endogenous controls. Additionally, stability of the controls varied...
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