Abstract

Medical sciences have begun to focus their attention on the use of nanomaterials to improve diagnosis and treatment with the ultimate goal of moving into personalized medicine. The need to develop more efficient drug delivery systems motivated the development of the nanomaterial proposed in this thesis based on lanthanide doped upconverting nanoparticles (UCNPs).
Upconverting nanoparticles have the attractive optical property of higher energy light production from lower energy excitation. Upon near infrared (NIR) excitation, LiYF4:Tm3+/Yb3+ upconverting nanoparticles produce emission in the ultraviolet (UV), visible and NIR. Surface modification of the nanoparticles with a supported lipid bilayer composed of phospholipids and cholesterol provided water dispersiblility and colloidal stability to LiYF4:Tm3+/Yb3+ upconverting nanoparticles, while protecting the emission of the nanoparticle from water-derived quenching. Additionally, the supported lipid bilayer provided a drug-loading feature. Different electron microscopy techniques were used to characterize the supported lipid bilayer on the surface of the LiYF4:Tm3+/Yb3+ upconverting nanoparticles and steady-state spectroscopy was used for the characterization of the emission of the nanoparticles.
The development of a dynamic photo-responsive supported lipid bilayer was achieved with the incorporation of an azobenzene-derivative lipid in the supported lipid bilayer at the surface of LiYF4:Tm3+/Yb3+ upconverting nanoparticles to photo-control the drug release. Azobenzene is a photoswitching molecule that changes from its stable trans-isomer to the cis-isomer under UV irradiation. Upon NIR excitation, UV emission is obtained in situ from LiYF4:Tm3+/Yb3+ upconverting nanoparticles and transferred to the azobenzene-derivative lipid, triggering the photoswitching and disrupting the supported lipid bilayer. The dye Nile red was loaded in the supported lipid bilayer as a model drug and its photo-control release was studied. Upon 30 minutes of NIR irradiation, LiYF4:Tm3+/Yb3+ upconverting nanoparticle transferred 23% of its energy to the azobenzene-derivative lipid, leading to 40% release of the dye encapsulated through the disruption of the photo-responsive supported lipid bilayer. Studies on the Förster resonant energy transfer (FRET) between LiYF4:Tm3+/Yb3+ upconverting nanoparticles and the azobenzene-derivative lipid showed that the energy transfer mechanism is mainly radiative (via reabsorption of light). Measurements on the absolute quantum yields of the individual emissions bands of LiYF4:Tm3+/Yb3+ determined that the quantum yields of the each of the emission bands are significantly different.
Cellular studies of this nanomaterial were conducted using alveolar lung carcinoma cells (A549) where the cellular uptake, cytotoxicity, endocytosis mechanisms and trafficking inside the cells were studied. The cellular uptake was studied as a function of the surface properties of LiYF4:Tm3+/Yb3+, and as a function of incubation time using ICP-MS for the quantification of the nanoparticle internalization. Through inhibitory studies of the cellular endocytosis mechanism it was found that the clathrin-mediated, caveolae-dependent and an energy-independent pathway were the main internalization mechanisms. Additionally, the trafficking and photo-controlled release of the nanomaterial was studied using confocal microscopy taking advantages of the optical properties of LiYF4:Tm3+/Yb3+ upconverting nanoparticles and the fluorescence of the dye Nile red. It was observed that the nanomaterial was trafficked to different organelles including the endoplasmic reticulum, Golgi apparatus, lamellar and lipid bodies. Upon NIR stimulation of the nanomaterial inside the lung cancer A549 cells, a fast release of Nile red, a model hydrophobic drug was produced from the photo-responsive supported lipid bilayer.
The studies herein contribute to the development of photo-responsive nanomaterials using lanthanide upconverting nanoparticles and enrich the understanding of the nanoparticle-cell interactions for the development and design of nanomedicines.