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Nano and Micro Technologies

Nano and Micro Technologies - Poster Session E

Poster Y9 - Exosome-Coated Gadolinium-Doped Prussian Blue Nanoparticles as a Multi-theranostic Agent for Treatment of Glioblastoma

Saturday, October 26, 2024
10:00 AM - 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  Glioblastoma is one of the most common and aggressive types of brain cancer, with no known cure and a survival of less than 15 months. FDA approved, Prussian Blue nanoparticles (PBNPs) are being explored as a new class of near-infrared (NIR) laser-driven, photothermal ablation/photoacoustic imaging agents. Laser interstitial thermal therapy (LITT) is an emerging, minimally invasive, magnetic resonance imaging (MRI)-guided procedure used for cytoreductive treatment of glioblastoma. Photosensitizing agents such as PBNPs that can enhance thermal ablation efficacy at lower powers are needed to minimize lethal heat exposure to normal brain tissue. The doping of gadolinium (Gd) in the PBNP structure (Gd-doped PBNPs; GdPBNPs), to enhance MRI signals as well as shift their primary light absorbance peak larger to 800 nm, which augments the photoheranostic efficacy of PBNPs. Furthermore, the nanoparticles coated with the exosomes derived from the glioblastoma tumor present a systemic brain-targeting of therapeutics across the blood-brain barrier (BBB). Such agents will be beneficial in precise control of the ablation area, especially near functionally eloquent brain regions. In this study, we develop novel nanohybrids (Exo:GdPB) that can preferentially accumulate in orthotopic glioma and assist in improving the efficacy and safety of LITT procedure of glioma treatment, potentially applied to the clinical LITT system (e.g., Visualase®).

Materials and

Methods:  Synthesis of gadolinium (Gd)-doped Prussian Blue nanoparticles (GdPBNPs) was conducted by initially combining potassium ferricyanide, polyvinylpyrrolidone, citric acid, and 0.1M HCl to create solution A and gadolinium nitrate hexahydrate, polyvinylpyrrolidone, citric acid, and 0.1M HCl to create solution B. Solution B was then slowly dropped into solution A under magnetic stirring at room temperature for 3 hours, then heated to 80oC for 24 hours before washing by centrifugation with deionized water. Gd content was determined via inductively coupled plasma spectroscopy (ICP), and the surface morphology of the particles was characterized via transmission electron microscopy (TEM). Exosome isolation was done through multistep, serial high-speed centrifugation to collect small-sized extracellular vesicles (80 to 150 nm in diameter) using U-251 glioblastoma cell lines. Exosome coating of the GdPBNPs was carried out through the physical extrusion of particle/exosome mixtures through a 200 nm membrane. The coated and uncoated particle sizes were determined through dynamic light scattering measurements and UV-vis spectroscopy. Cell uptake studies were carried out in U-251 cells seeded in a 6 well-plate. Cells were treated with 0.1 mg/mL of DiI-stained exosomes and DiI Exo:GdPB. MTT assays were conducted to determine laser ablation efficacy in U-251 cells seeded in a 96 well-plate. The cells treated with varying concentrations of Exo:GdPB (0 to 0.25 mg/mL) were subjected to an 808 nm laser for 1 min and left overnight before conducting the MTT assay. Statistical analysis was performed using a student’s t-test and considered significant at p< 0.05.

Results, Conclusions, and Discussions: Gadolinium-doped PBNPs (GdPBNPs) that are uniform and monodisperse were successfully synthesized with NIR absorbance. PBNPs (no Gd doping) were 210.4 nm (PDI: 0.053) and had a peak absorption at 750 nm. Synthesized GdPBNPs were 256.4 nm (PDI: 0.028) and measured Gd content of 1.28 µg/mL. The absorbance spectrum showed a broad peak between 800 nm and 900 nm, allowing the particle to be used as a photothermal ablation agent with an 808 nm laser and clinically used 980 nm lasers in the Visualase apparatus. T1 relaxivity of the GdPBNPs were measured against Dotarem, a clinical MR contrast agent. The Gd-doped PBNPs showed T1 values between that of water and Dotarem. TEM images confirmed the cubic morphology of the particles with GdPBNPs having rounder edges compared to Gd-undoped PBNPs. The coating of the particles with exosomes was confirmed with a size shift between the GdPBNPs and Exo:GdPBNPs (253.7 nm to 295.3 nm). Uranyl acetate staining further confirmed the coated uranium stained (exosome) layers, with iron and gadolinium cores from the Exo:GdPB in electron mapping of TEM. Exosomes express surface markers that allow them to be recognized by target glioma cells and help transfer therapeutic agents (nanoparticles) with passive transport. This is specifically done through receptor-mediated phagocytosis. Exosome-mediated delivery was then further validated with cell uptake studies in U-251 glioblastoma cells that showed similar uptake patterns between DiI-stained exosomes and DiI-stained Exo:GdPB as there are similar fluorescent signals in the perinuclear regions after 24 hr incubation. MTT assay results confirm the capability of the GdPBNPs to be used as a photothermal ablation agent with a significant reduction in cell viability at low doses of nanoparticles ( < 55% at 0.03 mg/mL of NPs) after treatment of the cells with Exo:GdPB and 808 nm laser (1.5 W/cm2 for 1 min). It was further confirmed the GdPBNPs showed augmented thermal ablation efficacy in cancer cell killing compared to PBNPs with 808 nm lasers. These results indicate that the nanohybrids (Exo:GdPB) can function as a multi-theranostic agent in glioblastoma treatment with its tumor cell targeting and augmented photoacoustic imaging and photothermal ablation efficacy.

Acknowledgements (Optional):