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Characterization of germline alterations affecting genes influencing development and prognosis of specific adult cancers
Jelínková, Sandra ; Kleibl, Zdeněk (advisor) ; Hlaváč, Viktor (referee) ; Hájková, Nikola (referee)
In my dissertation, I studied the genetic predisposition of selected types of cancer that have not been systematically studied in the Czech Republic. We used next-generation panel sequencing to identify germline pathogenic variants. Analysis of 1333 patients with ovarian cancer, 527 patients with endometrial cancer, and 334 patients with hepatocellular carcinoma included sequencing using the CZECANCA panel. A specific CZMELAC panel was prepared for the analysis of 264 melanoma patients. We focused on the identification of pathogenic variants in known predisposition genes. We also evaluated candidate genes and phenotypic characteristics in carriers of pathogenic variants. Analysis of high-risk melanoma patients revealed pathogenic variants in melanoma associated genes in 9/264 (3.4%) patients, and an additional 22 (8.3%) patients carried a pathogenic variant in one of the other predisposition genes. The odds of carrying a pathogenic variant were increased in probands with multiple melanomas and in the presence of melanoma in relatives. The incidence of germline pathogenic variants was highest in ovarian cancer, where pathogenic variants were found in 427/1332 (32.0%) patients, with a predominance of mutations in BRCA1, BRCA2, followed by alterations in other ovarian predisposition genes. Breast and...
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Spontaneous Regression of Melanoma in Pigs of the MeLiM Strain
Plánská, Daniela
Melanoma is a skin tumour arising from melanocytes - skin cells bearing pigment melanin. Melanoma belongs among immunogenic tumours, which is probably associated with a relatively high incidence of partial spontaneous regression (SR). Melanoma-bearing Libechov Minipigs (MeLiM) represent a specially bred animal model that is mostly affected by nodular melanomas, which fully regressed in about 2/3 of the affected animals. Our interest was to examine immune response (associated with melanoma cell destruction) and the role of proteins related to the extracellular matrix (reflecting tissue remodeling) during SR of MeLiM melanoma. We performed an extensive time-lapse study of skin melanomas taken from individuals of the MeLiM strain at 3, 4, 6, 8, 10, 12, 20 and 32 weeks (5-10 samples in each age category) in which we immunohistochemically detected the expression of collagen IV, laminin, fibronectin, tenascin C, as well as MMP-2 and we monitored the proportion of basic immune subpopulations in blood and tumour by flow cytometry. The higher expression of collagen IV, laminin and MMP-2 positively correlated with the appearance of melanoma cells. The expression of collagen IV and laminin indicates a possible survival of tumour cells due to the interaction with these proteins, the presence of MMP-2 in these...
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Spontaneous Regression of Melanoma in Pigs of the MeLiM Strain
Plánská, Daniela ; Horák, Vratislav (advisor) ; Smetana, Karel (referee) ; Bartůňková, Jiřina (referee)
Melanoma is a skin tumour arising from melanocytes - skin cells bearing pigment melanin. Melanoma belongs among immunogenic tumours, which is probably associated with a relatively high incidence of partial spontaneous regression (SR). Melanoma-bearing Libechov Minipigs (MeLiM) represent a specially bred animal model that is mostly affected by nodular melanomas, which fully regressed in about 2/3 of the affected animals. Our interest was to examine immune response (associated with melanoma cell destruction) and the role of proteins related to the extracellular matrix (reflecting tissue remodeling) during SR of MeLiM melanoma. We performed an extensive time-lapse study of skin melanomas taken from individuals of the MeLiM strain at 3, 4, 6, 8, 10, 12, 20 and 32 weeks (5-10 samples in each age category) in which we immunohistochemically detected the expression of collagen IV, laminin, fibronectin, tenascin C, as well as MMP-2 and we monitored the proportion of basic immune subpopulations in blood and tumour by flow cytometry. The higher expression of collagen IV, laminin and MMP-2 positively correlated with the appearance of melanoma cells. The expression of collagen IV and laminin indicates a possible survival of tumour cells due to the interaction with these proteins, the presence of MMP-2 in these...
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Transcriptional changes associated with melanoma resistance to small inhibitor treatment
Dorčáková, Terézia ; Kolář, Michal (advisor) ; Modrák, Martin (referee)
Malignant melanoma is an aggressive skin tumour with increasing incidence and, in advanced stages, limited therapeutic results. The major oncogenic alteration in melanoma is mutational activation of the B-Raf protein, with the predominant mutation V600E, which occurs in 60% of cases and hyperactivates the ERK signaling pathway. This cascade of Raf, MEK and ERK protein kinases is an integral part of an evolu- tionarily conserved signaling network that allows eukaryotic cells to sense a variety of extracellular signals. These protein kinases sequentially activate each other and translate extracellular signals into cellular responses such as proliferation, di↵erentiation, cell cycle changes, apoptosis or cell migration. The central role of the ERK pathway in oncogen- esis has made it one of the targets for therapeutic intervention, and inhibitors targeting mutant Raf, such as vemurafenib or dabrafenib, have been approved for the treatment of melanoma. These molecules specifically target B-Raf(V600E) and have led to a revo- lution in melanoma treatment. Unfortunately, such targeted therapy is complicated by the ability of cancer cells to acquire resistance. The aim of this work is to study at the transcriptional level the emergence of this resistance in patients who respond well to treatment and in...
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Larval cestodes and tumors: characterization of antibody response and cross-reactivity
Vajs, Vojtěch ; Horák, Petr (advisor) ; Panská, Lucie (referee)
In recent years there have been developments in research concerning the effect of certain species of helminths on cancer. For example, a preventative infection with these parasites is capable of slowing or completely preventing the development of certain cancers in experimental models. In our preliminary experiments, the suppressive effect of the infection with Taenia crassiceps and Mesocestoides corti on melanoma cancer cells was discovered. Primary infection with these tapeworms has been able to suppress and completely prevent the growth and metastasis of the B16F10 melanoma cell line. Parasites affect the immune system of their hosts with their presence and products. It was therefore the subject of this work to explore one of the ways how these tapeworms may achieve this effect - cross-reactivity of antibodies with tapeworm and cancer cell antigens. First the antibody response of the mouse organism to the tapeworm infection was examined in C57BL/6J inbred and ICR outbred strains. The growth of immunoglobulin levels was measured with ELISA within the first five weeks post infection, upon which the antigens, with which these immunoglobulins react, were specified with the help of Western blot. After this, these reacting antigens were localised in the tapeworm organism with immunohistochemical...
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Immunohistochemical and histological correlation of melanoma lesions
ČAPKOVÁ, Denisa
The bachelor thesis deals with the problematic of melanoma lesions. This theme is actual because melanoma is the most serious type of skin cancer and its incidence is moving to the younger generation. Because melanomas occur primary due to ultraviolet light exposure, one of the best possible ways to reduce the risk of developing melanoma is to protect yourself from ultraviolet radiation by using the sunscreen, wearing clothing that covers arms and legs and avoid using tanning beds. The prognosis of early melanoma is usually very good, so the detection and treatement is very important. In the theoretical part the current knowledge of melanoma, the characteristic, the occurence, the clasification, the prevention and the current possibilities of treatment were summarized. In the practical part the Hematoxylin eosin staining and the immunohistochemical method was described. There is also comparation of positive HMB-45 marker between each histological groups. The HMB-45 marker is absolutly specific for melanocytic tumors and has great utility fot the pathologist in differentiation of tumors with uncertain origin. From 2012 to 2018 were imunohistologicaly examined at the Departement of Pathology Nemocnice Písek a.s. 61 samples. Out of total 61 samples were 58 HMB-45 positive. The thesis has been focused on the imunohistological profile of each histological group. The imunohistologichemical profile was proven in 92%.
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Intercellular interactions in malignant melanoma
Nedvědová, Tereza ; Dvořánková, Barbora (advisor) ; Brábek, Jan (referee)
Melanomas are one of the most aggressive types of tumours, with increasing incidence, high mortality and high potential to metastasize to a variety of diverse locations. The aim of this thesis was to study the tumour as a complex structure consisting not only of tumour cells but also of tumour stroma. Stromal cells play a major role in cancer biology. This is well documented for example in squamous cell epithelium tumours of the head and neck. Similar mechanisms can be expected to occur in melanomas. In the first experiment, we simulated the conditions in vivo during the metastatic process and studied the influence of non-adhesive environment both with and without the influence of stromal fibroblasts. The presented data demonstrates a change of tumour cells' phenotype leading to increased plasticity of the melanoma cells in these conditions. It also indicates the crucial role of stromal fibroblasts in interactions with melanoma cells. Cancer cell lines show variability in their behaviour, which is in accordance with well-known melanoma heterogeneity in clinical practice. The previous experiments in our laboratory indicate that cancer associated fibroblasts are able to influence the phenotype of a tumour cell line and this effect is based on a tumour type-unspecific mechanism. In the second part of...
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Intercellular interactions in skin tumors.
Kučera, Jan ; Smetana, Karel (advisor) ; Masařík, Michal (referee) ; Kovář, Marek (referee)
The dissertation is focused on the study of intercellular interactions in skin tumors. It is based on 5 original publications that cover several topics. We studied the origin of tumor-associated fibroblasts concerning the primary tumor population. We demonstrated using a mouse model that tumor-associated fibroblasts are produced from the host organism and thus did not arise from transformation directly from tumor cells. We also investigated the relationship between tumor-associated fibroblasts and keratinocytes. We have shown that tumor-associated melanoma fibroblasts affect keratinocytes which, under their influence, acquire the features typically observed in migrating cells and cells undergoing epithelial-mesenchymal transition. We studied the interactions between healthy fibroblasts and tumor cells. We have demonstrated that fibroblasts acquired from healthy skin from a patient suffering from melanoma are significantly different from control fibroblasts of healthy donors in the expression profile. Changes in distal fibroblasts support the view of melanoma as a systemic disease. We have further demonstrated that melanoma-associated fibroblasts do not carry a BRAF mutation, in contrast to BRAF positivity of melanoma cells. And therefore, they did not arise from the transition from melanoma. The...
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Morphological and Immunohistochemical Analysis of Melanocytic Lesions
Důra, Miroslav ; Dundr, Pavel (advisor) ; Mandys, Václav (referee) ; Pock, Lumír (referee)
This doctoral thesis deals with morphological and immunohistochemical analysis of melanocytic lesions. Melanoma is the most dangerous malignant skin tumor with its potential occurrence outside the skin. Its early diagnosis is the basic aspect of a favorable disease prognosis. Distant metastases development occurring several years after the initial diagnosis is a typical feature of melanoma. Histopathological differential diagnostics of melanocytic lesions is extremely difficult because of the diversity of melanocytic lesions. Contemporary research is focused on the identification of new surrogate markers based on immunohistochemistry or molecular genetics, which would be enable to estimate the behavior of a certain lesion and clarify the gray zone of melanocytic lesions of uncertain biological behavior. The doctoral thesis is focused on the immunohistochemical analysis of GLUT-1 protein expression in 400 cases of benign and malignant melanocytic lesions. Significant expression of this marker was identified in the group of melanomas, it was not identified in melanocytic nevi. Expression of GLUT-1 corresponded to the worse prognosis of the disease. The doctoral thesis also deals with the issue of TILs. The analysis of TILs (tumor infiltrating lymphocytes) in 213 melanomas compared the prognostic...
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Role of fibroblasts in wound healing and cancer
Mateu Sanz, Rosana ; Smetana, Karel (advisor) ; Masařík, Michal (referee) ; Mokrý, Jaroslav (referee)
Fibroblasts are stromal cells ubiquitously present in the human body. They often appear in a quiescent state and can become activated in response to tissue remodeling signals. Activated fibroblasts acquire biosynthetic, pro-inflammatory and contractile properties, key functions for wound healing. In addition, the presence of permanently activated fibroblasts is one of the hallmarks of cancer. The purpose of this work is to investigate the differences between newborn and adult fibroblasts and keratinocytes in their implication in scarless wound healing, the origin of cancer associated fibroblasts (CAF)s and the influence of fibroblasts in melanoma invasion. Evidence suggests that wounds heal almost without scar in newborns. To understand the mechanisms that contribute to scarless wound healing we focused on the differences between newborn and adult fibroblasts and keratinocytes, which are cells present in human skin and participating in wound healing process. A comparison of the expression profile between newborn and adult fibroblasts showed differentially regulated genes related to the acute phase of the inflammatory response and ECM organization, traits involved in wound healing. We also found that newborn fibroblast showed higher differentiation potential, exhibited markers of pluripotency and...
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