Nanomicelles And Liposomes For Improving Drug Delivery of Small Molecules And siRNA Therapeutic Agents
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PART I: SELF-ASSEMBLING TACROLIMUS NANOMICELLES FOR BACK OF THE EYE DISEASES Vrinda Gote, Candidate for the Doctor of Philosophy Degree University of Missouri-Kansas City, 2021 ABSTRACT Neovascular age-related macular degeneration (AMD) is characterized by increase in reactive oxygen species (ROS) and pro-inflammatory cytokines in retinal pigment epithelium cells. The primary purpose of this study was the development of a clear, tacrolimus nanomicellar formulation (TAC-NMF) for AMD. The optimized formulation had a mean diameter of 15.41 nm, zeta potential of 0.5 mV and an entrapment efficiency of 97.13%. In vitro cytotoxicity studies revealed dose-dependent cytotoxicity of TAC-NMF on human retinal pigment epithelium (D407), monkey retinal choroidal endothelial (RF/6A) cells and human corneal epithelium (CCL 20.2) cells. Cellular uptake and in vitro distribution studies using flow cytometry and confocal microscopy respectively indicated an elevated uptake of TAC-NMF in a time-dependent manner. Biocompatibility assay using macrophage RAW 264.7 cell line resulted low production of inflammatory cytokines IL-6, IL-1β and TNF-α after treatment with TAC-NMF. There was a decrease in ROS in D407 cells pre-treated sodium iodate (ROS inducing agent) after treating with TAC-NMF and tacrolimus drug. Similarly, there was a reduction in the pro-inflammatory cytokines and VEGF-A in D407 cells pretreated with sodium iodate. This can indicate that TAC-NMF could lower pro-inflammatory cytokines and ROS commonly seen in AMD. PART II: NANOFORMULATIONS TO IMPROVE DRUG DELIVERY AND REDUCE MULTI DRUG RESITSANCE IN BREAST CANCER. Vrinda Gote, Candidate for the Doctor of Philosophy Degree University of Missouri-Kansas City, 2021 ABSTRACT Nanoformulations having the ability to specifically target cancer cells along with overcoming multidrug resistance (MDR) can be highly effective in breast cancer targeted therapy. In this study we designed two nanoformulations for targeting breast cancer cells. In the first study, we constructed a hyaluronic acid (HA) decorated mixed nanomicelles encapsulating chemotherapeutic drug paclitaxel (PTX) and P-glycoprotein inhibitor ritonavir (RTV). HA was conjugated to poly (lactide) co-(glycolide) (PLGA) polymer by disulfide bonds, (HA-ss-PLGA). HA is a natural ligand for CD44 receptors overexpressed on breast cancer cells. Disulfide bonds undergo rapid reduction in the presence of glutathione, present in breast cancer cells. The mixed nanomicelles contained HA-ss-PLGA and Vitamin E TPGS amphiphilic polymers. Addition of RTV can inhibit P-gp and CYP3A4 mediated metabolism of PTX, thus increasing the intracellular levels of the chemotherapeutic drug. This can help in reversing MDR and sensitizing the cells towards PTX. In the second study, we designed a targeted liposomal formulation encapsulating siRNA to reduce angiogenesis in breast cancer cells. Lipocalin 2 (Lcn2) is a promising therapeutic target for metastatic breast cancer (MBC) and triple negative breast cancer (TNBC). Lcn2 overexpression in breast cancer lead to its progression by inducing the epithelial to mesenchymal transition and enhancing angiogenesis. Lcn2 concentrations in the urine can act as diagnostic marker for breast cancer detection and disease stage. In this study, we engineered a novel PEGylated liposomal system encapsulating Lcn2 small interfering RNA (Lcn2 siRNA) for selective targeting breast cancer cells. The PEGylated liposomes were targeted with Octreotide (OCT) peptide. OCT is an octapeptide analog of somatostatin (SST) growth hormone, having higher affinity for somatostatin receptors (SSTRs). These receptors are overexpressed on breast cancer cells. Optimized OCT-targeted Lcn2 siRNA encapsulated PEGylated liposomes (OCT-Lcn2-Lipo) was formulated and evaluated for its effectiveness in targeting breast cancer cells. Physiochemical and morphological characteristics were determined for both the nanoformulations. Performance of the nanoformulations were assessed at various stresses including temperature, freeze-thaw, and dilution. In vitro uptake and intracellular distribution studies was assessed in breast cancer cell lines MCF-7 and TNBC cell line MDA-MB-231 and non-neoplastic breast epithelium MCF-12A. Cytotoxicity studies and potency studies were performed in the above-mentioned cell lines.
Table of Contents
Part I: Self-assembling tactolimus nanomicelles for back of the eye diseases -- Chapter 1: Nanomicelles for back of the eye disorders -- Chapter 2: Self-assembling tacrolimus nanomicelles for retinal drug delivery -- Part 2: Nanoformulations to improve drug delivery and reduce multi drug resistance in breast cancer -- Chapter 3: Nanomedicine for metastatic breast cancer -- Chapter 4: Stimuli-sensitive nanomicellar drug delivery for breast cancer to overcome multi drug resistance and achieve hyaluronic acid mediated active targeting -- Chapter 5: Octreotide targeted LCN2 SIRNA pegylated liposomes as a treatment for metastatic breast cancer and triple negative breast cancer -- Chapter 6: Summary and recommendations -- Appendix A. Abbreviations -- Appendix B. Permissions
Ph.D. (Doctor of Philosophy)