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Glioblastom, glioma tumor gradusa IV, je najčešći maligni tumor mozga kod odraslih i jedan od najsmrtonosnijih tipova tumora. I pored agresivne terapije koja obuhvata hirurško uklanjanje tumora, radio- i hemio-terapiju, prosečno preživljavanje bolesnika sa ovim tipom tumora je oko 15 meseci. Glioblastom (GBM), pored tumorskih ćelija, sadrži i populaciju samo-obnavljajućih tumor-inicirajućih matičnih ćelija (matične ćelije glioblastoma) koje se smatraju odgovornim za nastanak, progresiju, metastaziranje i rezistenciju na terapiju. Geni SOXB1 podgrupe (SOX1, SOX2 i SOX3) kodiraju regulatorne proteine koji imaju značajne uloge u mnogim procesima u toku razvića, kao što su održavanje pluripotentnosti matičnih ćelija i održavanje populacije neuralnih progenitora u pluripotentnom i proliferišućem stanju. Ovi geni imaju i značajne funkcije u procesu karcinogeneze. Ekspresija gena ove podgrupe detektovana je u GBM. Funkcija gena SOX2 je dobro proučena kod ovog tipa tumora; pokazano je da ovaj gen promoviše maligni potencijal ćelija GBM i neophodan je za održavanje tumorogenog potencijala matičnih ćelija glioblastoma. Za razliku od gena SOX2, uloga gena SOX1 i SOX3 u ćelijama GBM još uvek nije dovoljno istražena. Stoga, u okviru ove doktorske disertacije analizirana je uloga ovih gena u ćelijama glioblastoma. Dobijeni rezultati pokazuju da ćelijske linije GBM eksprimiraju gen SOX1. Pored toga, u uslovima utišane ekspresije proteina SOX1 detektovano je smanjenje proliferativnog kapaciteta, vijabilnosti i migratornog potencijala U251 ćelija GBM, kao i povećanje broja ovih ćelija u senescenciji. Nakon dediferencijacije ćelijskih linija GBM detektovano je povećanje ekspresije gena SOX1 u poređenju sa ekspresijom uočenom u njihovim parentalnim ćelijama. Nivo ekspresije gena SOX1 povećan je u kulturama matičnih ćelija glioblastoma u poređenju sa ekspresijom ovog gena u imortalizovanim U87 i U251 ćelijama; pri diferencijaciji ovih kultura uočeno je smanjenje ekspresije gena SOX1. U, one of deadliest cancers. Despite aggressive treatment, including surgical resection, chemotherapy and radiation, the median survival of patients with glioblastoma is 15 months. A growing body of evidence indicates that GBM contains a population of selfrenewing tumor-initiating cells (glioblastoma stem cells - GSCs) that drive tumor initiation, propagation, metastasis and therapy resistance. SOXB1 genes (SOX1, SOX2 and SOX3) encode transcription regulators with important roles in embryonic development and carcinogenesis. Literature date revealed that SOXB1 genes are expressed in GMB tumor samples. The role of one member of this group, SOX2 gene, is well documented in GBM. It was shown that SOX2 gene promotes malignant potential of GBM cells and it is mandatory for maintenance of tumorogenicity of GSCs. Since the function of SOX1 and SOX3 genes in GBM still remains to be established, the aim of this thesis was to analyze the role of these genes in GBM. Obtained results demonstrated that all analyzed GBM cell lines express SOX1. Downregulation of this gene expression decreases proliferation, viability and migration, and induces senescence of U251 cells. Furthermore, dedifferentiation of GBM cells is accompanied by increase of SOX1 level compared to that in parental cells. Expression of this gene was significantly increased in patient-derived GSC cultures compared to that in U87 and U251 cells; downregulation of SOX1 gene expression was seen upon differentiation of GSCs. In addition, knock-down of this gene expression reduced proliferation and viability of GSCs. Results obtained in this thesis reveal SOX3 expression in GBM cell lines and tumor tissue; the expression of this gene was elevated in the most of analyzed GBM samples compared to expression levels detected in non-tumoral brain tissues. A high SOX3 expression was not associated with the overall survival of GBM patients. Ectopic overexpression of this gene increased proliferation, viability, migration and invasi

Most women die of ovarian metastasis rather than the effects of the primary tumor. However, little is known about the factors that support the survival and secondary outgrowth of exfoliated ovarian cancer cells. In addition to genetic and molecular factors, the unique environment of the peritoneal cavity exposes ovarian cells to biophysical forces, particularly fluid shear stress (FSS). These biomechanical forces, only recently identified as a hallmark of cancer, induce rapid signaling events in attached and aggregated cells, a process termed mechanotransduction. The cellular responses to these forces and their impact on tumor initiation, progression, and metastasis are not understood. In order to delineate these phenomena, dynamic and syngeneic cell models are needed that represent the development of the disease and can be used in relevant engineered testing platforms. Thus, in an interdisciplinary approach, this work bridges molecular and cancer biology, device engineering, fluid mechanics, and biophysics strategies. The results demonstrated that even a low level of continual FSS significantly and differentially affected the viability of epithelial ovarian cancer cells of various stages of progression over time, and enhanced their aggregation, adhesion, and cellular architecture, traits of more aggressive disease. Furthermore, benign cells that survived FSS displayed phenotypic and genotypic changes resembling more aggressive stages of the disease, suggesting an impact of FSS on early stages of tumor development. After identifying a biological affect, we designed an in vitro testing platform for controlled FSS investigations, and we modeled the system fluid mechanics to understand the platform's performance capability. A cylindrical platform divided into annular sections with lid-driven flow was selected to allow continuous experiments sustainable for long durations. Tuning of the lid speed or fluid height resulted in a wide range of FSS magnitudes (0- 20 N/m2) as

Most women die of ovarian metastasis rather than the effects of the primary tumor. However, little is known about the factors that support the survival and secondary outgrowth of exfoliated ovarian cancer cells. In addition to genetic and molecular factors, the unique environment of the peritoneal cavity exposes ovarian cells to biophysical forces, particularly fluid shear stress (FSS). These biomechanical forces, only recently identified as a hallmark of cancer, induce rapid signaling events in attached and aggregated cells, a process termed mechanotransduction. The cellular responses to these forces and their impact on tumor initiation, progression, and metastasis are not understood. In order to delineate these phenomena, dynamic and syngeneic cell models are needed that represent the development of the disease and can be used in relevant engineered testing platforms. Thus, in an interdisciplinary approach, this work bridges molecular and cancer biology, device engineering, fluid mechanics, and biophysics strategies. The results demonstrated that even a low level of continual FSS significantly and differentially affected the viability of epithelial ovarian cancer cells of various stages of progression over time, and enhanced their aggregation, adhesion, and cellular architecture, traits of more aggressive disease. Furthermore, benign cells that survived FSS displayed phenotypic and genotypic changes resembling more aggressive stages of the disease, suggesting an impact of FSS on early stages of tumor development. After identifying a biological affect, we designed an in vitro testing platform for controlled FSS investigations, and we modeled the system fluid mechanics to understand the platform's performance capability. A cylindrical platform divided into annular sections with lid-driven flow was selected to allow continuous experiments sustainable for long durations. Tuning of the lid speed or fluid height resulted in a wide range of FSS magnitudes (0- 20 N/m2) as

A number of anti-angiogenic drugs targeting vascular endothelial growth factor receptors (VEGF-R) have developed and enabled significant advances in cancer therapy including colorectal cancer. However, acquired resistance to the drugs occurs, leading to disease progression, such as invasion and metastasis. How tumors become the resistance and promote their malignancy remains fully uncertain. One of possible mechanisms for the resistance and the progression may be the direct effect of VEGF-R inhibitors on tumor cells expressing VEGF-R. We investigated here the direct effect of a VEGF-R-targeting agent, regorafenib, which is the first small molecule inhibitor of VEGF-Rs for the treatment of patients with colorectal cancer, on phenotype changes in colon cancer HCT116 cells. Treatment of cells with regorafenib for only 2 days activated cell migration and invasion, while vehicle-treated control cells showed less activity. Intriguingly, chronic exposure to regorafenib for 90 days dramatically increased migration and invasion activities and induced a resistance to hypoxia-induced apoptosis. These results suggest that loss of VEGF signaling in cancer cells may induce the acquired resistance to VEGF/VEGF-R targeting therapy by gaining two major malignant phenotypes, apoptosis resistance and activation of migration/invasion.

Chromosomal rearrangements which result in localized increases of genetic material are frequent in breast cancer, and occur consistently in certain genomic regions. The resulting increase in expression of genes contained within these amplifications contributes to the malignant phenotype. Such amplified genes, such as Her2-Neu, provide targets for diagnosis and for the development of inhibitory drugs. The purpose of this study is to use novel genomic technologies to find new genes in breast cancer that are both highly amplified and are suitable targets by virtue of the fact that they are membrane-associated (receptors, membrane antigens, secretory proteins). The aims of the study are (1) To specify intervals of genomic amplification (amplicons) in primary breast cancer cell lines using genomic microarrays; (2) To prepare a database of membrane- associated genes, selected by differential hybridization of RNA prepared from fractionated microsomes; (3) To use the database to select membrane-associated genes that are located within amplicons, and measure their expression in the primary cell line using cDNA arrays, in order to select those that are upregulated. These genes will provide new insights and reagents for diagnosis and treatment of breast cancer.