The Efficacy and Endocytosis of uPAR-targeted Quantum Dots Öffentlichkeit
Babu, Ranjith (2009)
Abstract
Nanoparticles offer a wide variety of functionalities for both the treatment and imaging of cancer. One method of increasing efficiency is by coupling nanoparticles with ligands that target receptors that are overexpressed on cancer cells. One such receptor is the urokinase plasminogen activator receptor (uPAR). The urokinase plasminogen activator (uPA), which is a serine protease, binds to uPAR through its amino terminal fragment (ATF). This portion of the protein has been sequenced, allowing for its production and purification. This peptide can then be conjugated to nanoparticles, allowing them to target cancer cells that overexpress uPAR. The efficacy of uPAR-targeted nanoparticles in comparison to a free chemotherapeutic agent was investigated using spectroscopy, biomass assays and fluorescence microscopy. It was seen that there was a 3-4 fold increase in the number of targeted nanoparticles within cells compared to their untargeted counterparts. These targeted particles were also more effective in killing cells than the free chemotherapeutic drug. Next, whether uPAR-targeted nanoparticles entered cells via a caveolae-mediated endocytotic mechanism was investigated using caveolin-1 knockout cells. It was seen that there was approximately 50% fewer nanoparticles in caveolin-1 knockout cells compared to the wild-type. The results also showed that uPAR-targeted nanoparticles were not as efficacious in caveolin-1 knockout cells. These results demonstrated that caveolin-1 plays a significant role in uPAR-mediated endocytosis. However, as targeted particles still accumulated within caveolin-1 knockout cells, there are alternate mechanisms by which this can occur.
Table of Contents
LIST OF FIGURES
CHAPTERS
1. INTRODUCTION
1.1 Cancer and Nanomedicine
1.2 Nanoparticles for tumor imaging and drug delivery
1.2.1 Advantages of Nanoparticles
1.2.2 Types of Nanoparticles
1.3 Nanoparticle delivered chemotherapeutic agents
1.4 uPAR as a target for ligand conjugated nanoparticles
1.5 Endocytosis
1.6 Objectives
2. MATERIALS AND METHODS
2.1 Materials
2.2 Cell Cultures
2.3 Production of Mouse ATF Peptides
2.4 Preparation of QD-Dox and QD-mA-Dox
2.5 Preparation of uPAR-targeted QD-ATF
2.6 Cell Treatment
2.7 Crystal Violet Biomass Assay
2.8 Spectroscopy
2.9 Cell Visualization
3. RESULTS
3.1 Internalization of unconjugated and uPAR-targeted quantum dots
3.2 Efficacy of uPAR-targeted quantum dots
3.3 Effect of Caveolin-1 knockout on the internalization of unconjugated and uPAR-targeted quantum dots
3.4 Effect of Caveolin-1 knockout on the efficacy of uPAR-targeted quantum dots
3.5 Quantification of QD and doxorubicin accumulation within cells
4. DISCUSSION
4.1 Increased intracellular accumulation of uPAR-targeted quantum dots
4.2 Effects of quantum dots on cells
4.3 uPAR internalization via caveolin-mediated endocytosis
5. CONCLUSION
REFERENCES
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