Advanced Colloidal Systems for Targeted Chemotherapy

The current project gave a detailed insight of surface modification of different advance colloidal systems along with their in vitro and in vivo targeting capabilities. Three different colloidal systems (nanoparticles, microparticles and liposomes) were evaluated for their efficacies and consistenci...

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Bibliographic Details
Main Author: Ali, Muhammad Yasir
Contributors: Bakowsky, Udo (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2019
Online Access:PDF Full Text
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Summary:The current project gave a detailed insight of surface modification of different advance colloidal systems along with their in vitro and in vivo targeting capabilities. Three different colloidal systems (nanoparticles, microparticles and liposomes) were evaluated for their efficacies and consistencies in results. The introduction contains an overview for the passive and active targeting of chemotherapeutic agents with different colloidal systems. Different methods of preparation and characterization of these colloidal systems were reviewed. This formed the root level for the use of these formulations in the current project. Furthermore, a brief introduction about aptamers and different examples of targeting molecules was also given to elaborate on aptamers’ specific nature. This provided the basis of surface modification of colloidal formulations with aptamer of interest. Sorafenib tosylate (SFB) was selected as a chemotherapeutic agent because it has low solubility and low bioavailability. Its LogP value is 4.54 with biopharmaceutical classification systems class IV. Systemic toxicity due to non-specific drug delivery is also issues with the use of SFB. Another problem associated with the use of this chemotherapeutic agent is the development of drug resistance after consecutive administrations. Therefore, the current study was designed to improve the efficiency of cancer therapy using sorafenib-loaded colloidal systems coupled with anti-ErbB3-aptamer (Apt). There first part of result and discussion included characterization of SFB-loaded PLGA matrix systems i.e. nanoparticles and microparticles. The encapsulation efficiencies revealed the loading of the drug inside these carrier systems. The physicochemical investigation by Fourier transform infrared spectroscopy, elemental analysis and fluorescence analysis elaborated the success of surface modification of these systems with Apt. Furthermore, morphological analysis by atomic force and scanning electron microscopy supported these results and showed an optimal surface roughness profile for cell surface interactions. Cell culture studies showed a positive impact of the combination of SFB and Apt. The presence of SFB and Apt together showed maximum cytotoxicities compared to other formulations. Dose-dependent toxicities were demonstrated using the cell viability assay. Moreover, time-dependent formulation delivery, to the cytoplasm and subsequently to the nuclear membrane, was observed by CLSM visualization. Higher reactive oxygen species production was observed in the presence of both SFB and Apt as compared to blank formulations. However, the aptamer alone did not significantly induce ROS production. Upon treatment of the cells with different concentration of particles, a significant dose-dependent ROS production was noticed. The metastatic inhibition by the particles, especially those with SFB and Apt was evident from the scratch test. The absence of both SFB and Apt resulted in complete healing of wound within 24 h. Ex vivo hemolysis studies demonstrated the hemocompatibility of the PLGA matrices, thus mimicking in vivo safety of these formulations. The presence of SFB as well Apt did not change the hemolytic potential of formulations to much extent. All the formulations were more hemocampatible as compared to pure drug. Moreover, RBC aggregation test showed no profound change in the morphology of RBCs. In vivo assessment by the blood profiles along with serum biochemistry stamped the safety of the formulation. Nevertheless, heart and liver-specific toxicities were evident in the presence of SFB and Apt but the overall body visceral index was normal. Results and discussion also included characterization of SFB-loaded liposomes. The physicochemical investigation of the liposomes using dynamic light scattering and laser Doppler velocimetry revealed nearly monomodel size range from 121 nm to 155 nm suitable for cellular internalization. However, the presence of SFB and Apt influenced the hydrodynamic diameters and zeta potentials of formulations. Furthermore, morphological characteristics were described by atomic force microscopy and showed optimal sizes and surface roughness profile for cell surface interactions. Synergistic dose-dependent cytotoxicities were demonstrated using SFB and Apt in liposomes in 2D cell culture techniques. The evaluation of toxicity was also visualized in 3D cell cultures and revealed a decrease in 3D culture sizes. This effect was also evident in apoptosis assay showing nuclear condensation as a possible mechanism of cell death. The presence of surface-modified liposomes, inside cells was visualized using CLSM. These investigations showed the presence of liposomes inside the cell, especially near the nuclear region (co-localization coefficient; 0.4-0.7). In order to analyze the in vivo safety as well as the transfection potential of surface modified liposomes the chorioallantoic membrane model (CAM) was used. The presence of these formulations in the mesoderm of the CAM was visualized by CLSM. No evidence of clear toxicity was observed on the development of the embryo. Furthermore, the hemocompatibility studies of liposomes also demonstrated the safety of these formulations when compared to pure drug. Therefore, the combination of chemotherapeutic agent and aptamer together with colloidal drug delivery systems will pave the way to a powerful tool in anticancer therapies. Moreover, the presence of aptamer will also solve the problems of side effects of chemotherapeutic agents by specifically delivering the drug to resistant tumors.
Physical Description:132 Pages