Saudi Cultural Missions Theses & Dissertations
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Item Restricted Pharmacokinetics And Tissue Distribution of a Lipid-Based Extended-Release Nano-Formulation Of MO-OH-Nap Tropolone to Enhance Pulmonary Delivery for the Treatment of Pulmonary Metastatic Osteosarcoma(University of Nebraska Medical Center, 2024-08) Aldhafiri, Wafaa Nasser; Murry, DJOsteosarcoma is the most common primary bone tumor in children and adolescents. Metastasis is prevalent in 25% of patients, at diagnosis, with the lungs being the most common metastatic site. Pulmonary metastatic osteosarcoma is a significant therapeutic challenge with a 5-year survival rate of ≤30%. Patients with pulmonary metastatic disease at diagnosis receive the same standard of care as patients with local disease, which hasn’t changed over the past 30 years. The treatment and prevention of pulmonary metastatic osteosarcoma represents a significant critical and unmet need. MO-OH-Nap tropolone is a novel small molecule with cytotoxic activity across multiple human osteosarcoma cell lines, yet poor solubility limits its clinical development. We developed a MO-OH-Nap tropolone liposomal nanoparticle formulation to enhance delivery to pulmonary tissue for the treatment of pulmonary metastatic osteosarcoma. The nanoparticle formulation was prepared using a lipid film hydration method. Dynamic light scattering and ultra-centrifugation were used to determine the average particle size and encapsulation efficiency. The pharmacokinetics and biodistribution of MO-OH-Nap tropolone were determined in CD-1 mice following a single intraperitoneal dose of MO-OH-Nap tropolone aqueous solution or MO-OH-Nap tropolone liposomes nanoparticle formulation (5 mg/kg). Serial plasma and tissue samples were collected over 48 hours. MO-OH-Nap tropolone concentrations were determined utilizing a validated LC-MS/MS method. Phoenix® software was used to determine pharmacokinetic parameters utilizing non-compartmental analysis and a 3-compartment pharmacokinetic model to better characterize the MO-OH-Nap tropolone liposomes nanoparticle plasma and tissue concentrations. The finalized model was used to simulate different dosing regimen responses utilizing Monte Carlo simulations. The liposomes nanoparticle average particle size was 188.3 ± 54 d.nm with 99% encapsulation efficiency. Following single dose administration in mice, MO-OH-Nap tropolone systemic drug exposure was significantly increased (~10 fold) for the nanoparticle formulation (28229 hr*ng/mL) compared to the MO-OH-Nap (2846 hr*ng/mL). The observed clearance was 184.4 mL/hr/kg for the nanoparticle formulation and 1738.6 mL/hr/kg for the drug in aqueous solution. Drug concentrations in lung tissue 24 hours post dose (165.5 ng/g) were significantly increased following nanoparticle formulation administration compared to drug in aqueous solution (1.20 ng/g, student t-test, p-value < 0.001). The developed pharmacokinetic model accurately described the plasma concentration time profile and pulmonary drug accumulation. The model revealed a biphasic absorption pattern with a rapid initial drug release followed by a sustained drug release phase, indicating the controlled release properties of the nanoparticle formulation. Monte Carlo simulation for multiple dosing scenarios identified that the 7.5 mg/kg twice-weekly regimen would significantly improve lung coverage while moderately enhancing plasma levels, aligning better with the goal of maintaining pulmonary concentrations above the IC50 for a 75% of the dosing interval of 28 days. The model's fit was validated through bootstrap analysis, confirming the reliability of the pharmacokinetic parameters. Overall, this pharmacokinetic model provides critical insights into the behavior of MO-OH-Nap tropolone nanoparticle formulation, facilitating the development of effective dosing strategies for enhanced treatment of pulmonary metastatic osteosarcoma. The developed formulation substantially increased systemic exposure and pulmonary delivery of MO-OH-Nap tropolone. Further studies will assess the safety profile of 7.5 mg/kg IP dose twice weekly, as well as evaluate efficacy in mouse models of metastatic OS.17 0Item Restricted Study on Liposomes Integrity and Delivery of siRNA to Lung Cancer(University of Nottingham, 2024-06-12) Aljasser, Abdullah Abdulrahman; Stolnik, Snow; Bosquillon, CynthiaThe thesis focuses on studying liposomes' structural integrity and their interactions with model and plasma cellular membrane in vitro, utilizing fluorescence resonance energy transfer (FRET) as a key analytical tool. The study comprises the fabrication and physicochemical characterization of liposomes, the influence of α-linolenic acid incorporation on their properties and liposomes - cell interactions, and the potential of liposomes as siRNA delivery vehicles for lung cancer treatment. The initial phase of this research involved the fabrication of liposomes from various lipid and optimization of incorporation of DiO and DiI as fluorescent FRET pair probes. The successful embedding of these fluorophores within the liposomal membrane was confirmed by the observed quenching of donor emission and enhancement of acceptor emission. A key focus then moved to the role of an incorporation of an unsaturated fatty acid, α-linolenic acid, into liposomal membrane and the assessment of its potential fusogenic behavior. Through physicochemical characterizations the study evaluated dye incorporation efficiency, liposomal content leakage, and membrane fluidity. The findings revealed that liposomal membrane fluidity increases with the increase of α-linolenic acid content in liposomes. Data further show that on interaction with a membrane model liposome, the FRET ratio for DiO&DiI probes embedded in different liposome formulations decreased, suggesting vesicle membrane mixing / fusion. No significant carboxyfluorescein leakage from liposomes was observed with up to 30 mol% of α-linolenic acid, indicating their integrity in suspension. These observations establish the foundation for further investigations into the fusion behavior of fabricated liposomes at the cellular level. In the second chapter, the focus was on the impact of α-linolenic acid on liposome - cell interactions. Metabolic activity analysis revealed that liposomes without α-linolenic acid (αLA0) were non-toxic, whereas those with 20 mol% α-linolenic acid (αLA20) displayed dose-dependent cytotoxicity. FRET measurements data indicate that α-linolenic acid incorporation notably increased cell-associated fluorescence, indicating its significant influence on liposome - cell interactions. Both αLA0 and αLA20 liposomes demonstrated a reduction in FRET ratio upon interaction with A549 cells, suggesting membrane mixing or perturbations in liposome membrane integrity. Interestingly, the incorporation of α-linolenic acid resulted in pronounced liposome accumulation at/within A549 cell plasma membrane, as evidenced by localization patterns (Pearson’s correlation coefficient values) in confocal microscopy. These observations highlight the role of α-linolenic acid in enhancing liposomal interaction with cellular plasma membrane. The final chapter explored the feasibility of using liposomes as siRNA delivery vehicles, particularly in lung cancer cells. Due to its impact on liposomes formation in the presence of negatively charged siRNA and cationic lipid DOTAP, α-linolenic acid was excluded from this phase. The encapsulation efficiency of siRNA within liposomes was determined, revealing high encapsulation (> 90%) with liposomes containing 30 mol % DOTAP with N/P ratio of 9. The cytotoxicity assay demonstrated a concentration-dependent effect of DOTAP liposomes on A549 cells. A total lipid concentration greater than 20 mM was found to be toxic, resulting in less than 10% cell viability. In contrast, concentrations below 4 mM were observed to have minimal or no effect on cell viability. Successful cellular uptake of siRNA was demonstrated through flow cytometry and confocal microscopy. The efficiency of liposomal formulations in gene silencing was confirmed by luciferase gene silencing assays on A549-luciferase cells, with different DOTAP liposomes formulations showing significant activity reductions (‘silencing’) compared to control, scrambled siRNA liposomes. These findings highlight the potential of designed DOTAP/siRNA liposomes as a feasible approach to siRNA delivery and targeted gene silencing. To conclude, the thesis aims to provide understanding of the behavior of liposomes incorporating α-linolenic acid, particularly in the context of a potential lung therapy. The study gains understanding on the structural integrity of liposomes upon their interactions with model and cellular membranes, utilizing the FRET technique as a main investigative tool. It points to the potential of incorporating different 'helper’ lipids to influence interactions of liposomes with cells and consequently influence the delivery of cargo loaded into the liposome.14 0Item Restricted Development of drug loaded cationic liposomes for pulmonary delivery(Saudi Digital Library, 2023-10-11) Alharbi, Sayer; Kett, VickyThe lower respiratory tract infections (LRTIs) represent a serious threat to human health, especially with the growing prevalence of antimicrobial resistance (AMR). Consequently, the demand for antibiotics with higher activity and better targeting is overwhelmingly increasing. The cationic liposomes composed of dimethyldioctadecylammoniumbromide (DDAB) and soy phosphatidylcholine (SPC) lipids with d-α-tocophyeryl polyethylene glycol succinate (TPGS) as an adjuvant, could have the potential to be effective drug delivery vehicles to deliver antibiotics to the infected sites via the pulmonary route. However, the current liposomal synthesis methods have some disadvantages such as the usage of organic solvents. So, this study aims to investigate an organic solvent free method (OSF) as a potential alternative for the thin film hydration method (TFH) for the preparation of drug loaded liposomes. In addition, this study aims to investigate the effects of DDAB and TPGS contents on the final characteristics of the proposed cationic liposomes formulation. Towards these aims, the hydrophilic apramycin (APR) and the hydrophobic rifampicin (RIF) were used as model drugs. The OSF method encompasses the manual trituration of the lipid components and mixing it with the aqueous phase. The loaded liposomes were characterised for particle size, surface charge, encapsulation efficiency, powder particle size, and thermal properties. The OSF prepared APR-loaded liposomes showed comparable characteristics to the TFH prepared APR-loaded liposomes. However, the RIF-loaded liposomes prepared by the OSF method showed poor characteristics in comparison with the TFH prepared. Indicating the suitability of the OSF method for the synthesis of APR-loaded liposomes and the unsuitability of the method to be used with RIF. These contradicting outcomes were linked to the differences in the aqueous solubility between the two drugs. DDAB and TPGS exerted no effects on the particle size of the liposomes. DDAB showed a proportional relationship with the liposomal surface charge of both drugs, which was linked to the cationic charge of DDAB. TPGS showed no effects on the encapsulation efficiency of APR, while increasing the TPGS content decreased the encapsulation efficiency of RIF. The decrease in RIF encapsulation was attributed to the reduction of the available space for hydrophobic drugs between the lipid bilayers due to the presence of TPGS. This study proposed a solvent free method for the preparation of APR-loaded liposomes, and it contributed to the knowledge about DDAB and TPGS effects on the liposomal formulation. Furthermore, this study recommended the exploration of other solvent free synthesis methods for the preparation of RIF-loaded liposomes.35 0