Master Degree / Yüksek Lisans Tezleri
Permanent URI for this collectionhttps://hdl.handle.net/11147/3008
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Master Thesis Development of Peg-Peptide Conjugate Based Curcumin Delivery Systems(01. Izmir Institute of Technology, 2022) Aydoğan, Gamze; Top, AybenIn this study, a drug delivery system based on Pluronic F127 and a peptide conjugate was proposed. The F127-peptide conjugate was prepared by the reaction between succinimidyl functionalized F127 (SC-F127) and peptide. SC-F127 was synthesized using disuccinimidyl carbonate and DMAP. Folic acid-functionalized F127 (FA-F127) was also prepared to obtain active targeting copolymers. Four peptides containing pH-responsive multiple histidines and endosome disruptive GFWFG domain were synthesized using the Fmoc procedure. H-Gly-2-ClTrt resin and Rink amide MBHA resin were used to synthesize side-chain-protected and deprotected peptides, respectively. 2-chlorotrityl resin failed in synthesizing the high-purity peptides with adjacent histidines in their sequences. Peptide-4 having a sequence of GGH6GFWFG, was prepared with acceptable purity using rink amide MBHA resin and was conjugated to SC-F127. Curcumin was loaded to F127 and F127-peptide using the thin film method with DCM solvent. Almost all curcumin was encapsulated into F127 micelles. However, the entrapment efficiency % of the F127-peptide micelles was ~86% due to the lower solubility of F127-peptide conjugate in DCM. Dynamic light scattering experiments were used to determine the stability and size distribution of the micelles. Number-based size distributions of both micelles indicated that a single peak between 10 and 30 nm was independent of pH. The peak position did not change upon incubating the micelles at 37oC up to a few days. Initially, intensity-based results of both samples indicate bidisperse populations at pH 5.0 and 7.4. Curcumin-loaded F127 micelles aggregated in the three days, as revealed by the formation of the third peak above 1000 nm independent of pH. Curcumin-loaded F127-peptide micelles, on the other hand, retain their stability for up to five days at neutral pH. For this sample, the third peak was observed only at pH 5.0 on days 2 and 5.Master Thesis Preparation of Drug Loaded Albumin Nanoparticles in Water / Ionic Liquids Microemulsion Systems(Izmir Institute of Technology, 2021) Yıldırım, Barış; Akdoğan, YaşarNanoparticles (NPs) have been used in various applications such as biotechnology, nanomedicine, and drug delivery systems. Many nanoparticle drug delivery systems have been promoted for cancer treatment, and numerous materials have been investigated to use as drug delivery agents to enhance the therapeutic efficiency and safety of anticancer drugs. Albumin is a natural biopolymer and the most abundant protein in blood plasma. Due to its versatile binding capacity of widespread therapeutical drugs, albumin becomes an ideal material to obtain nanoparticles. In this study, the ionic liquid (IL) based emulsification methods were investigated. Instead of classical toxic and volatile solvents, using ILs in microemulsions, environment-friendly media were received to synthesize bovine serum albumin (BSA) NPs. In order to obtain BSA NPs, high-speed homogenizer processing was applied by following crosslinker addition. The IL microemulsions are a thermodynamically stable colloidal dispersion containing spherical droplets (W/IL or IL/W) in submicron sizes that act as nanoreactors for NP formation. Chlorambucil (CHL) was used as a model drug to investigate drug loading and releasing kinetics of BSA NPs as a drug delivery candidate. Results showed that chlorambucil loading capacities and release kinetics depended on the synthesized medium such as anion-type of ILs and surfactants. CHL loaded to the BSA NPs synthesized in hydrophilic IL BmimBF4 in relatively higher amounts and released in the same trend. In addition, the cell viability effect of CHL-loaded BSA NPs synthesized in different types of ILs were investigated. The CHL-loaded BSA NPs synthesized in BmimOTf and BmimPF6 reduced the cancer cell viability more than the used same dose of free CHL.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 Development of Ultrasound Triggered Drug Delivery Systems for Cancer Treatment(Izmir Institute of Technology, 2019) Önercan, Cansu; Kılıç Özdemir, SevgiDoxorubicin (DOX) is one of the most commonly used hydrophilic anticancer drug in cancer treatment. However, when it is used in free form, it can attack not only cancer cells but also healthy cells. So as to prevent entering of DOX to the healthy cells, the encapsulation method is employed. Liposomes are suitable for encapsulation of DOX but the most important problems with the use of liposome are hand-foot syndrome and stomatitis. Encapsulation method is not enough because of these reasons, thus delivery of DOX to the desired site by targeted therapy has gained interest in recent years. In this study, DOX was encapsulated into liposomes and the DOX loaded liposomes (LipoDOX) was attached to microbubbles (MBs). MBs as ultrasound contrast agents are widely used in medical imaging. Use of MBs in combination of DOX loaded liposomes facilitates the uptake of the drug because ultrasound cavitation results in opening of transient pores in cell membrane via a process named sonoporation. Herein, MB-LipoDOX complex was engineered to optimize the size of the complex as well as the loaded DOX content. For this purpose, determination of incubation temperature and time for DOX loading into liposome and optimization of liposome formulation for maximum DOX loading were studied. Ratios of Lipid/Cholesterol/PEGylated lipid, PEG chain length and PEG molar ratio in liposome were determined. Also, determination of Strept Avidin (StAv) to Biotin ratio in LipoDOX and the amount of LipoDOX in LipoDOX-MB complex were studied. For characterization, Dynamic Light Scattering (DLS) method, Fluorescence Spectrometry method and Coulter Counter device were used. Lipoosme size was found to be associated with the pore size of polycarbonate membrane (200nm) resulting in liposomes at around 190±5 nm in size . When the PEGylated lipid with PEG chain of 2000 was used in liposome structure, particle size distribution is more monodispersed than the others. The maximum amount of DOX loaded liposomes was obtained at 32% Cholesterol, 5% DSPE-PEG2000, after 90 min. incubation at 65oC incubation. Optimum StAv to Biotin ratio in LipoDOX was determined as 1.0. The optimum molar ratio of Biotinylated lipids in LipoDOX was determined as 0.05% and the optimum molar ratio of Biotinylated lipids in MBs was determined as 8%.Master Thesis Development of Endosome Disruptive Peptide and Peg Conjugate Based Doxorubicin Delivery System(Izmir Institute of Technology, 2019) Özkıyıcı, Selin; Top, AybenIn this study, it was aimed to develop a drug carrier system including a TAT-derived cell penetrating peptide in order to provide fast transport of anticancer drugs from endosomal compartments to nucleus. The drug delivery system, denoted as mPEGpeptide- oxime-DOX, was based on polyethylene glycol, endosome disruptive peptide (G2RQR3QR3G2S), and doxorubicin (DOX) conjugate. Control drug delivery system, lack of the peptide (mPEG-oxime-DOX) was also synthesized to assess the effect of the peptide on the physiochemical and drug release properties of the drug carrier. As the first synthesis step, mPEG-OH was converted to mPEG-aldehyde form using DMSO-acetic anhydride oxidation reaction and aldehyde functionalization was determined by using FTIR and NMR spectroscopy. The peptide and mPEG-peptide were synthesized using solid phase synthesis protocol, and their purities were confirmed using HPLC and MALDI-TOF mass spectroscopy analyses. Prior to DOX conjugation, hydroxyl group of serine residue in the mPEG-peptide system was oxidized to aldehyde. The anticancer drug was attached to the carrier molecules via amine-aldehyde reaction forming an acid cleavable oxime bond. Drug release, size distribution, and stability of the PEG-peptideoxime- DOX system were evaluated and compared with those results of the control drug delivery system. For mPEG-oxime-DOX, a pH programmed DOX release with the respective % DOX release values of ~68 % and ~28 % at pH 5.0 and pH 7.4 was observed. For mPEG-peptide-oxime-DOX, on the other hand, quite low DOX release (~10-15 %) was obtained for both pH values suggesting possible interaction between DOX and the peptide. Mean size value of the mPEG-oxime-DOX was measured as ~24 nm. However, mPEG-peptide-oxime-DOX, had quite lower hydrodynamic diameter values (~3nm and ~6 nm at pH 5.0 and pH 7.4, respectively) possibly due to repulsions between the arginines in the peptide domain. Observation of the morphology and evaluation of the cytotoxicity of these drug delivery systems are underway.Master Thesis Study of Drug Transportation by Esr Spectroscopy(Izmir Institute of Technology, 2018) Tatlıdil, Duygu; Akdoğan, Yaşar; Emrullahoğlu, MustafaThe ability to track drug binding and release makes electron spin resonance (ESR) spectroscopy well suited for drug delivery studies. Using the continuous wave cw ESR technique to extract information about the dynamics of the spin labeled drugs we can simultaneously determine the bound and unbound drugs. In this study, ESR technique was used to detect the binding and release of spinlabeled salicylic acid (SLSA) to and from bovine serum albumin (BSA), and to detect different binding interactions between them. We have labeled salicylic acid with stable nitroxide-based tempo radicals to monitor the BSA bound and unbound conditions of the drug. Studying with the different concentrations of SLSA-BSA binding showed that the drug-protein stoichiometry increases significantly in the physiological range of BSA concentration. Also, during the release of SLSA from BSA, there is an unchanging balance between the bound and unbound SLSA. In order to study various drug binding interactions, SL-benzoic acid, SL-phenol, SL-benzene, SL-cyclohexane, SL-hexane and SL-methyl were prepared. We showed that the main conjugation in the binding of these drugs to BSA is hydrophobic interaction. In addition, cationic BSA (cBSA) was prepared to investigate the effect of electrostatic interaction on drug binding. The SLSA loading capacity of cBSA is significantly higher than that of BSA, this result indicates the importance of electrostatic interactions for the drug binding. Finally, we examined the competitive binding behaviors of salicylic acid, ibuprofen and aspirin to BSA. Binding sites of SL-salicylic acid and SL-ibuprofen in BSA show 96% of similarities. In addition, our results showed that binding sites of SL-salicylic acid and SL-aspirin in BSA have 73% of similarities.These results demonstrate that cw ESR spectroscopy with the spin labeling technique is an effective technique for the determination of drug-protein interactions and stoichiometric analysis of drug binding.Master Thesis Preparation and Characterization of Polymer Based Composite Nanospheres for Bone Infection Prevention(Izmir Institute of Technology, 2018) Kımna, Ceren; Tıhmınlıoğlu, FundaCurrently, bone tissue engineering applications comprise the development of smart materials that can induce tissue regeneration meanwhile supporting the defect site of the body. Despite of the advancements, inflammation is still a serious problem that threatens human health at the post-implantation period. To overcome potential inflammations, antibiotic therapy is commonly employed in clinical trials. However, antibiotic therapy causes some side effects such as ototoxicity and nephrotoxicity, especially when applied in high doses. Therefore, local drug delivery systems play a vital role in bone disorders due to the elimination of disadvantages introduced by conventional methods. In the presented study, it was aimed to develop chitosan-based composite nanospheres as a controlled drug delivery system against bone infections. Accordingly, chitosan and montmorillonite nanoclay was homogenized with microfluidizer and electrosprayed to obtain spherical nanoparticles. The optimum electrospraying conditions were investigated using response surface methodology. Vancomycin and Gentamicin antibiotics were incorporated in the polymeric matrix to provide controlled release at the defect region to overcome inflammations after implantation. The prepared nanospheres were characterized in terms of morphology, hydrodynamic size distribution, surface charge, drug encapsulation efficiency and release profiles. The dominant drug release mechanism was determined by empirical mathematic models. Drug loaded nanospheres have been successfully produced with a size range of 180-350 nm. High encapsulation efficiency was achieved (80-95%) with a controlled drug release up to 30 days. Fickian diffusion was found as the main mechanism in drug delivery from spherical CS/MMT nanocomposites. The in vitro release medium of nanospheres showed strong antimicrobial activity against gram-positive S. aureus and gram-negative E. coli bacteria. Furthermore, it was found that the nanospheres did not show any cytotoxic effect to 3T3 and SaOS-2 cell lines. These results demonstrated that the prepared nanospheres can be a promising option for bone infection prevention.Master Thesis Development of Drug-Loaded Microbubbles for In-Vitro Applications in Cell Biology(Izmir Institute of Technology, 2017) Coşkun, Sema; Özdemir, Ekrem; Sultan Altun, ZekiyeDoxorubicin (DOX) is one of the drugs for cancer therapy. When DOX is used in solution, it affects not only the cancer cells but also the healthy cells. In order to eliminate possible side effects, DOX was encapsulated within liposomes and applied for the cancer therapy. Because the circulation time for liposomes is longer in the body, they accumulate in capillaries, especially at the finger tips and at the toe of the foot called the hand-and-foot syndrome. Here, we proposed to couple the liposomes containing DOX with the microbubbles as the ultrasound contrast agent and deliver the drug to the area of interest. Therefore, DOX was loaded within the liposomes and characterized for their DOX contents. The DOX containing liposomes were conjugated with microbubbles through the avidin-biotin chemistry. It was found that the loaded- DOX content within the liposomes was Langmuir-type. The loaded DOX content increased at lower DOX concentrations and leveled off at higher DOX concentrations. The Langmuir constants can be used in designing DOX loading experiments. The DOX containing liposomes were coupled with the microbubbles and found an optimum of 7.0 for the avidin/biotin mole ratio on the microbubbles. At the optimum avidin/biotin ratio, the conjugated lipo-DOX amount was 3×10-8 μg-DOX/MB. It was concluded that the DOX molecules can be loaded within the liposomes and easily conjugated with the microbubbles and employed in cancer treatments.Master Thesis Peg-Peptide Drug Carrier Systems With Enzymatic Degradation Units(Izmir Institute of Technology, 2016) Yüksel, Nesligül; Top, AybenIn this study, it was aimed to develop drug delivery systems with high drug release rate, capable of overcoming multidrug resistance of cancer cells. The first generation drug delivery system (DDS) denoted as mPEG-AT3-DOX was prepared by methoxy polyethylene glycol (mPEG) and peptide conjugation, and the model anticancer drug, DOX, was attached to the mPEG-peptide carrier system using stable amide linkage. mPEG was used to increase blood circulation time of the DDS. Designed peptide (AT3 = CG3H6R2ALALG3E) controls release of the drug via RRALAL sequence, which is a substrate for a lysosomal enzyme, cathepsin B, overexpressed in most of the tumor cells. pH responsive histidines and reactive amino acids (glutamic acid and cysteine) for drug and mPEG conjugations were also added to the peptide sequence. The peptide synthesized by using Fmoc chemistry was conjugated to mPEG-maleimide via Michael addition reaction. DOX was attached to the carboxylic acid group present in the carrier system (mPEG-AT3) via amide linkage. Mass spectroscopy and HPLC were used to assess the purity of the AT3 and mPEG-AT3. At pH 7.4, mPEG-AT3-DOX exhibited bimodal size (hydrodynamic diameter) distribution at about 15 and 30 nm independent of pH. % DOX release from mPEG-AT3-DOX was observed to be below 10 % at neutral pH and pH 5.0 in the absence of cathepsin B, and increased to 17 ± 2 % in the presence of cathepsin B. Complete degradation of AT3 peptide within three hours in the presence of cathepsin B suggests lower than expected DOX release is due to aggregation tendency of the DDS. Cytotoxicity of the conjugates was evaluated using the lung cancer (A549) and prostate cancer (PC3) cell lines. At the end of 24 hours the absolute IC50 values of free DOX and mPEG-AT3-DOX were obtained as 1.37 ± 0.05 and 1.33 ± 0.11 for the A549 cell line, 1.51 + 0.07 and 1.63 + 0.80 μg equivalent DOX / ml for the PC3 cell line, respectively. Considering, these results, the second generation DDS will be designed to have more pronounced pH sensitive property by increasing the number of histidines in the peptide domain.Master Thesis Use of Fibroin/Hyaluronic Acid Matrices as a Drug Reservoir in Iontophoretic Transdermal Delivery(Izmir Institute of Technology, 2004) Kuduğ, Emre; Batıgün, AyşegülTransdermal drug delivery is gaining importance due to the extensive research in genetics and resulting increase of protein and peptide based drugs in the market. In order to develop materials to be used in iontophoretic transdermal drug delivery systems, various forms of silk fibroin (SF) and blending agents as hyaluronic acid (HA) have been tested for their feasibility as a potential drug reservoir. For this purpose different forms of silk such as raw silk, degummed silk fibroin, insolubilized freezedried fibroin, membranes of fibroin in pure and blended with HA were investigated for their adsorption capacities of timolol maleate, which is used as the model drug. It was found that silk fibroin and derivatives have considerable adsorption capacities for timolol maleate with 0.35 mmol per gram, comparable with commercial membranes. The insolubilization of the membranes was required for drug loading and delivery in aqueous media. Membrane insolubility was achieved by post treatment, manipulation of drying conditions, and blending with different agents. Configurational changes of fibroin protein and interactions between silk fibroin and hyaluronic acid were investigated by Fourier transform infrared spectroscopy, scanning electron microscopy, and X-ray diffraction analyses. Insoluble fibroin glutaraldehyde membranes were produced. The obtained insoluble membranes were investigated for drug delivery performance in a custom-made diffusion cell under passive diffusion and iontophoretic conditions. It was demonstrated that the silk fibroin glutaraldehyde films could be successfully used for controlled drug delivery. It was found that current densities of 1.5 and 3 mA/cm2 were suitable to accomplish controlled delivery of the drug in a pulsatile manner. The results of this study are expected to be useful in controlled transdermal delivery of positively charged drug molecules.