Browsing Theses and Dissertations School of Pharmacy by Subject "Dendrimers"
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In-vitro Efficacy and Intracellular Mechanism of Riboflavin-Conjugated PEGylated Poly- L-Lysine DendrimerChemotherapeutic drugs have advanced using different drug delivery methods to treat breast cancer specifically. This development has arisen because many classical drugs exhibit physicochemical limitations including solubility, specificity, stability, biodistribution, and therapeutic efficacy. There were numerous adverse effects associated with these limitations because chemotherapeutic drugs enter normal tissues. In order to eliminate off-target side-effect,nanoparticles were developed to target anticancer drugs to a specific carcinogenic area. As one of developing nanomedicines, dendrimers possess ability to be utilized in different administration routes and has potential to stay in the blood circulation longer while showing increased accumulation in tumor cells. Commercially available poly (amidoamine) (PAMAM) dendrimers have the potential to cause toxicity in vivo due to lack of biodegradation at sites of accumulation. Poly-L-Lysine (PLL) dendrimers are an alternative class of dendrimers that possess a biodegradable structure. PEGylated poly-l-lysine (PLL) dendrimers are known to be more favorable due to lessened cytotoxicity manifested by masking of cationic charges and avoiding uptake by Reticulo Endothelial System (RES). Using this biodegradable dendrimer, we sought to examine the effect of PEGylation as well as delivering anti-cancer drug, Doxorubicin (DOX), to a targeted internalization pathway in human breast cancer cells effectively. PEGylated PLL dendrimers also have their limitation, in which some tumor cells are not dependent upon enhanced permeability and retention (EPR) effect. As a result, riboflavin receptor, which is found to be upregulated in the exterior of breast and ovarian cancer cells, was utilized by attaching a riboflavin ligand to PEGylated PLL dendrimers in order to be actively uptaken by breast cancer cells. To target chemotherapeutic drug selectively and efficaciously, riboflavin conjugated PLL dendrimers were assessed in-vitro by investigating cytotoxicity, uptake accumulation, and intracellular colocalization. Further investigation on the endocytosis mechanism and detailed intracellular trafficking in different compartments of the cells were analyzed in order to fully understand the machinery behind delivering chemotherapeutic drugs successfully.
Mechanisms of Dendrimer-Mediated Oral Drug DeliveryOral administration of chemotherapeutics remains a challenge despite the benefits for both the patient and health care system. To overcome the poor solubility and low oral bioavailability of anti-cancer drugs, polymeric delivery systems have been investigated. Dendrimers, a class of highly branched polymers, have proven useful for drug delivery because of their compact, nanoscopic size. Specifically, poly(amidoamine) (PAMAM) dendrimers have been shown to permeate the intestinal epithelium indicating potential as oral drug delivery carriers. While studies in our laboratory have determined the effects of surface modification on dendrimer transport and uptake, a large gap in knowledge exists in the transport and cytotoxicity mechanisms of PAMAM dendrimers. Additionally, alternatives to PAMAM dendrimers such as biodegradable poly-L-lysine (PLL) dendrimers have yet to be investigated for use in oral delivery. In this work we report the mechanisms of tight junction modulation by PAMAM dendrimers. While anionic dendrimers modulated tight junction proteins, cationic dendrimers opened tight junctions through phospholipase C-mediated calcium signaling allowing for paracellular small molecule transport. In comparison, cationic PLL dendrimers also allowed for small molecule transport with similar decreases in transepithelial electrical resistance. Small generation PAMAM and PLL dendrimers (16 and 32 surface amines) activated Caspase-3 and -7 resulting in apoptosis. In contrast, PLL dendrimers showed less long term toxicity compared to PAMAM dendrimers illustrating the benefits of dendrimer biodegradability. We also investigated the mechanisms of PLL dendrimer internalization and subcellular trafficking and the impact conjugation had on these mechanisms. The pH and enzymes present vary within different intracellular vesicles. Knowledge of the environment a drug delivery system will encounter is crucial for proper drug release. While PLL dendrimers were internalized via cholesterol- and dynamin-mediated endocytosis and macropinocytosis, conjugation site impacted uptake and localization. By conjugating a model compound to either the dendrimer core or surface, the uptake and transport properties of the delivery system were modified. Core-conjugated dendrimers had higher uptake and localized to the lysosomes and nucleus while surface conjugation resulted in higher transport and less accumulation in lysosomes. This research provides important knowledge for designing an effective dendrimer-based oral drug delivery system.