Master Degree / Yüksek Lisans Tezleri
Permanent URI for this collectionhttps://hdl.handle.net/11147/3008
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Master Thesis Development of Doxorubicin-Loaded Liposomes Self-Assembled With Polysaccharides for Breast Cancer Therapy(01. Izmir Institute of Technology, 2023) Önol, Ayşenur Başar; Kılıç Özdemir, Sevgi; Polat, HürriyetThis thesis aimed to develop Tariquidar and Doxorubicin-loaded liposomes decorated by Fucoidan coating for breast cancer treatment. Fucoidan is a negatively charged polysaccharide with a special affinity to p-selectins expressed on MDA-MB-231 breast cancer cells and, at the same time, possesses anti-cancer activity. Different liposomes were prepared by extrusion method from the DSPC, Cholesterol, and cationic lipid DSTAP mixtures for coating negatively charged Fucoidan. The most stable liposomes with a size of 200 nm were obtained at a molar ratio of DSPC/Cholesterol/DSTAP:55/30/15, exhibiting a zeta potential above +30 mV. Tariquidar was encapsulated into the liposome bilayer by passive loading, and Doxorubicin into the core of the liposome by active loading. In the final step, liposomes were coated with Fucoidan by electrostatic interaction. Tariquidar loading was determined by UV-Vis spectrophotometry, indicating an optimum TRQ/Lipid molar ratio of 0.012 with encapsulation and loading efficiencies of 50% and 20%, respectively. Fluorescence spectrophotometry determined Doxorubicin loading, showing insignificant encapsulation efficiency change (exhibiting around 70%) by neither Tariquidar content in the bilayer nor DSTAP% in the formulation. An optimum amount of Fucoidan was determined by incubating the liposomes with varying amounts of fucoidan at different dilutions. Size and zeta potential measurements monitored the coating of liposomes with Fucoidan. Our finding showed that zeta potentials of liposomes go from positive to negative with increasing fucoidan, while no trend was observed in the size of liposomes. However, smaller sizes were observed when incubation was performed in diluted solutions.Master Thesis Enhancement of Bioavailability of Vitamin D by Nano-Sized Delivery Systems(01. Izmir Institute of Technology, 2023) Sağlam, Ezgi İrem; Kılıç Özdemir, Sevgi; Bulmuş Zareie, Esma VolgaStudies have indicated that Vitamin D (VitD) may decrease tumor invasiveness and propensity to metastasize. Cholecalciferol (VitD3) is the passive form of VitD3 and converts to active calcitriol through two-step hydroxylation reactions in the body, promoting binding to VitD-receptors (VDR). However, some breast cancer cells, especially MDA-MB-231, have very low levels of VDR. Besides, VitD3 suffers from first pass-effect of the liver which causes deactivation of VitD3. Therefore, new approaches are needed to increase VitD3 level in the cancerous sites. In this study, VitD3 was loaded into liposomes, which were subsequently coated by Fucoidan (FUC) to promote their binding to MDA-MB-231 cancer cells. Fucoidan strongly binds to P-selectins overexpressed in the breast cancer cells, blocking the cancer cells to adhere on the platelets to carry within the body, causing metastasis. Doxorubicin (DOX), being considered as the one of the most effective chemotherapeutic agents against breast cancer, was also loaded into liposomes in a similar manner. By liposomal encapsulations and fucoidan coating, it was aimed to deliver the all-cargo directly to the cancerous site and enhance the bioavailability of both agents at the target site. It was seen that liposomal VitD3 was more effective than free form to inhibit cell proliferation and, therapeutic potential of DOX increased with VitD3.VitD3 loaded FUC coated liposomes at optimized concentrations has a comparable effect with DOX-loaded liposomes with and without FUC coating. Overall, these results suggested that VitD3 and DOX loaded and FUC coated liposomes can be applied as combined therapy in cancer treatment.Master Thesis Determination of Therapeutic Effects of Multifunctional Antibody and Peptide Micelle-Based Nanocarriers on Breast Cancer Cells(01. Izmir Institute of Technology, 2021) Abdulhadi, Nusaibah Abdulsalam Abdulhad; Baran, YusufBreast cancer is the most prevalent type of cancer and a major cause of death among women globally. Currently, many treatments are developed to reduce breast cancer death risks. Targeting therapy represents an advanced and successful approach. It provides targeting specific tumor sites by using specific ligands and modifying physicochemical characterization of nanocarriers to increase drug efficiency. In this study, we aim to determine and compare the therapeutic effects of doxorubicin (DOX)- loaded nanocarrier that was synthesized by using two properties a core cross-linked and pH sensitivity to increase drug stability and DOX releasing at the tumor site. The effects of DOX-loaded micelles (DM), HER2 targeting peptide (LTVSPWY)-conjugated-DOX-loaded micelles (DMP), and antibody (Herceptin) conjugated-DOX-loaded-micelles (DMA) on HER2 positive SKBR-3 cell line and HER2 negative MCF-10A normal epithelial breast cell line were determined by using cytotoxic, apoptotic, cytostatic, and genotoxic assays. According to the cytotoxic assay, the IC50 value of DM, DMA, and DMP were 0.71-, 0.49-, 0.34-µM, respectively. Additionally, the fluorescence image showed higher DOX uptake by SKBR-3 cells treated with DMP. According to the apoptotic assays, the mitochondrial membrane potential on SKBR-3 cells with treated DMP decreased as well as higher apoptosis and necrosis rate that was regulated by Bcl-2, Pro-Caspase-3, PARP1, Bax, Bak, and Bcl-xL. Besides, the application of DMP caused cell cycle arrest at the G2/M phase. Lastly, DNA damage was observed in response to DMP determined by comet assay. This study provides a novel and effective therapeutic option for breast cancer through using this nanocarrier system with targeting properties.Master Thesis Time Dependent Expression and Localization of Connexin 32: Implication in Epithelial To Mesenchymal Transition of Mammary Epithelial Mcf10a and Triple Negative Breast Cancer Mda Mb 231 Cells(01. Izmir Institute of Technology, 2020) Ünal, Yağmur Ceren; Meşe Özçivici, GülistanBreast cancer is the most frequent and the second leading cause of cancer-related deaths among women worldwide. Epithelial to mesenchymal transition (EMT) is critical driving force in metastasis. Connexins as a basic subunit of gap junctions indicate critical roles in regulation of EMT. In addition to Cx26 and Cx43, Cx32 is associated with breast cancer and elevated levels of Cx32 has been reported in lymph node metastasis compared to primary breast cancer while the role of Cx32 in breast cancer is still elusive. Here we aimed to shed light on the effect of Cx32 on breast cancer. Our study suggested that Cx32 acquired mesenchymal morphology and decreased proliferation in MCF10A cells but not in MDA MB 231 cells. To further elucidate whether Cx32 indicate these changes through EMT, EMT markers were examined and subsequently it was revealed that Cx32 expression was strongly correlated with increased E-cadherin and Vimentin in MCF10A cells while decreased E-cadherin and Snail in MDA MB 231 cells. Importantly majority of Cx32 did not localize to the plasma membrane and indicated dynamic changes in a day dependent manner in both MCF10A and MDA MB 231 cells. Moreover, day dependent expression and localization of Cx32 revealed strong correlation with Zeb2 expression in MCF10A cells. In conclusion, Cx32 indicated differential effects in regulation of EMT between MCF10A and MDA MB 231 cells. It was the first time that the role of Cx32 on EMT was investigated in breast cancer and differential localization of Cx32 was identified.Master Thesis Investigating Oncogenic Role of Sema6d in Breast Cancer Cells(Izmir Institute of Technology, 2019) Günyüz, Zehra Elif; Yalçın Özuysal, ÖzdenBreast cancer, the most commonly diagnosed cancer type and the leading cause of cancer-associated deaths, is the major health issue among women worldwide. In many cancer types, the expression of the semaphorins and their receptors such as plexins and neuropilins are dysregulated. SEMA6D is a member of class-6 family transmembrane semaphorin proteins and acts through Plexin-A1 receptor. It was previously shown that overexpression of SEMA6D in breast cancer cell line MCF-7 leads to a reduction in proliferation and an increase in migration. On the other hand, in the MDA-MB-231 breast cancer cell line, overexpression of SEMA6D had no significant effect on proliferation but enhanced migration. In this study, we aimed to analyze the effects of SEMA6D overexpression in normal breast cell line MCF10A and investigate the invasive behavior and transformation capacity of SEMA6D overexpressing breast cancer cell lines. We demonstrated that overexpression of SEMA6D leads to elevated proliferation, viability and migration in MCF10A cells, whereas it did not trigger their anchorage-independent growth. On the other hand, MDA-MB-231 and MCF7 cells stably expressing SEMA6D showed reduced colony formation in the soft-agar assay. Furthermore, the invasiveness of MDA-MB-231 cells was elevated with SEMA6D overexpression, whereas SEMA6D overexpression did not stimulate the invasiveness of MCF-7 cells through matrigel microenvironment, whereas slightly trigger invasion through bone microenvironment. In conclusion, SEMA6D overexpression has cell-specific effects on breast cancer. The exact role of SEMA6D in breast cancer development remains undefined and must be further investigated.Master Thesis Increasing Doxorubicin (dox) Release From Liposomes(Izmir Institute of Technology, 2019) Hanoğlu, Berçem Dilan; Özdemir, Ekrem; Altun, Zekiye SultanCancer is the second most common cause of death in the world and its incidence is increasing day by day. Doxorubicin (DOX) is an anthracycline group drug frequently used in many cancer treatments including breast cancer. However, free DOX has many harmful side effects and need to be encapsulated into nanocarrier such as liposomes. Although liposomal DOX has many advantages over its free form, liposomal DOX has undesirable side effects such as hand and foot syndrome. In this thesis, it was aimed to develop a more effective liposomal DOX delivery and release systems. Liposomes were prepared with alkaline solutions containing tris, sodium carbonate, ammonium chloride, and ammonium sulfate. DOX loading into liposomes and the percentage of release from liposomes were examined. A loading efficiency of about 80% was achieved, while the release was found to be below 13% at room temperature. The release of DOX was found to be enhanced from liposomes in the presence of ammonia (NH3), whose content was dependent on pH. Temperature was also found an important parameter and enhances DOX release at higher temperatures than the phase transition temperature of the lipid. A two-component liposomal system was proposed where ammonia (NH3) would be released from one liposome and enhance the DOX release from other liposomes. It was found that temperature, pH, and ammonia (NH3) concentration affected DOX release from liposomes. As a result, DOX was successfully loaded into liposomes and ready to study their effect on breast cancer cells.Master Thesis Cellular Mechanosensing at a Distance(Izmir Institute of Technology, 2019) Can, Ali; Pesen Okvur, Devrim; Özçivici, EnginThe goal of the project is to determine differences in mechanical sensing at a distance between breast cancer cells and normal mammary epithelial cells. To achieve this goal, we aim to: 1. Optimize the device for mechanical sensing at a distance 2. Determine the effect of mechanical sensing at a distance on cell proliferation 3. Determine the effect of mechanical sensing at a distance on cell migration Breast cancer is one of the cancers with the highest incidence and mortality rates in women in Turkey as well as in the world. Tumor microenvironment comprises of cancer and normal cells, extracellular matrix, soluble biological and chemical factors. Biochemical aspects of the interactions of cancer cells with the constituents of the microenvironment are widely studied whereas biophysical studies are at limited numbers. There is increasing evidence that extracellular matrix can change the mechanics and function of cancer and stroma cells. It has been observed that cancer cells show different responses to soft and stiff tissues they are in direct contact with than normal cells. However, it is not known whether the distance at which cancer cells can feel the stiffness of a distant tissue is longer, the same or shorter than that of normal cells. The hypothesis we will test in this project is as follows: The distance at which cancer cells can feel the stiffness of a distant tissue is shorter than that of normal cells.Master Thesis Determination of Therapeutic Effects of Multifunctional Micelle-Based Nanocarriers on Breast Cancer Cells(Izmir Institute of Technology, 2019) Ulu, Gizem Tuğçe; Baran, YusufBreast cancer is the most common and frequent cause of death among women composed to all types of cancer. Current treatment protocols do not provide complete cure or selective drug delivery while targeted therapy can provide an important avenue for successful treatment of breast cancer. In this study, therapeutic effects of drug-conjugated nanocarrier system with enhanced stability and double moiety pH-sensitivity on breast cancer (SKBR-3- HER-2- positive), normal breast epithelial (MCF-10A, HER-2-negative) and chronic myeloid leukemia (K562, HER-2-negative) cells were determined. With this approach, SKBR-3 cells were targeted by single nanocarriers having selectivity with unused peptide ligand (HER-2), stability with cross-linking of core moiety, and cleavage by two sites of pHeffect and drug release properties. After physicochemical characterization of micellebased nanocarriers, cytotoxic, apoptotic and cytostatic effects of doxorubicin conjugated micelles were determined. Doxorubicin conjugated micelles with HER-2 peptide (DOX-HER-2-NCs) had more cytotoxic effects on HER-2 positive cells. Additionally, intracellular amounts of doxorubicin is higher in SKBR-3 cells with applied DOX-HER-2-NCs as determined by fluorescence imaging. The apoptosis rate was increased on SKBR-3 at 50% cell growth inhibition (IC50) as determined by Annexin-V/Propidium iodide double staining. However, there was not any significant change in loss of mitochondrial membrane potential. Additionally, DOX-HER-2-NCs resulted in cell cycle arrest at G2/M-phase in response to IC50 value. Besides, protein level of Bcl-2 did not change while protein level of Bax and Caspase-3 were increased as determined by Western Blotting. This project provides novel and more effective treatment of breast cancer by using multifunctional properties of nanocarriers.Master Thesis Evaluation of Biophysical Aspedts of Cancer Using Lab-On Chip Devices(Izmir Institute of Technology, 2019) Tahmaz, İsmail; Pesen Okvur, Devrim; Sürmeli, Nur BaşakBreast cancer metastasis is really crucial point from cancer related deaths. As cancer cells from primary tumor are travelling through blood, they hang on to blood vessel and finally they exit from blood vessel into secondary site where is extracellular matrix and/or tissue/organ. This process commonly known as extravasation. Cancer cells sometimes can be highly aggressive when it exposed to hypoxia referred low oxygen amount by activating HIF1α. This transcription factor is activated in malignant cells, normal cells and endothelial cells in blood vessel when oxygen amount decreased to certain levels and it induce several genes expression such as VEGF, LOX, Angiopoietin-like-4 etc. In this study we investigated effect of HIF1α which is hypoxia indicator on breast cancer extravasation by comparing to normal oxygen level. This study represents both anemic hypoxia physiologically and lead to understand underlying mechanism of extravasation into extracellular matrix related to low oxygen circulating through blood. In addition to HIF1α effects, dynamic perfusion mimicking blood flow was applied to determine effects on extravasation. For this purpose, lab-on-a chip device was utilized for real time visualization. In conclusion, although hypoxia is giving permission MDAMB231 to extravasate because of reshaping of vascular geometry, less extravasated cancer cells observed in matrix during hypoxia under both static and flow condition when compared to normoxic and static conditions. Moreover, it was shown that flow triggers extravasation distance in normoxia against static condition and normal breast epithelial cells extravasated away in hypoxia comparing breast cancer cells by means of flow.Master Thesis Mimicking the Tumor Microenvironment in Lab-On Devices(Izmir Institute of Technology, 2019) Bilgen, Müge; Pesen Okvur, Devrim; Sürmeli, Nur BaşakBreast cancer is one of the cancers with the highest incidence and mortality rates in women in the world. The leading cause of death for cancer patients is tumor metastasis. Cancer cells can extravasate the blood vessel, go through the distant organs and form the metastasis. Tumor microenvironment comprises of cancer and normal cells, extracellular matrix, soluble biological and chemical factors. Biochemical aspects of the interactions of cancer cells with the constituents of the microenvironment are widely studied whereas biophysical studies are at limited numbers. There is increasing evidence that extracellular matrix can change the mechanics and function of cancer and stroma cells. It has been observed that cancer cells show different responses to soft and stiff tissues they are in direct contact with than normal cells. New cell culture setups should be developed to better understand the interactions of cancer cells with their microenvironment. To develop a three dimensional (3D) in vitro model will allow the study of stiffness which is one of the mechanical features of extracellular matrix features first, 3D (dimensional) Controlled in vitro Microenvironments (CivMs) that mimic a blood vessel and its neighboring tissue in vivo will be fabricated using UV lithography. Monolayer which was formed by endothelial cells play a role in pathophysiological processes, so it shows a barrier role between both blood and tissues. To form a blood vessel bEnd.3 cell line was used. Collagen which is the most abundant protein in connective tissues were used to mimic extracellular matrix. pH value of collagen was changed and represented two different stiffness value. Here, the in vitro model we define as controlled in vitro microenvironments (CivM) is a lab-on-a-chip (LOC) application. In this microenvironment; MDA-MB-231 cells which are known to be invasive and MCF10A which is normal mammary epithelial cells were used as control. LOC devices were used to investigate cancer cell extravasation which is the prominent step of metastasis and extracellular matrix relation.