High density lipoprotein (HDL), is an important natural nanoparticle that may be modified for biomedical imaging purposes. Here we developed a novel technique to create unique multimodality HDL mimicking nanoparticles by inclusion of gold, iron oxide or quantum dot nanocrystals for computed tomography, magnetic resonance and fluorescence imaging, respectively. By including additional labels in the corona of the particles, they were made multi-functional. The characterization of these nanoparticles, as well as their in vitro and in vivo behavior revealed that they closely mimic native HDL.

The development and application of nanoparticles as in vivo delivery vehicles for therapeutic and/or diagnostic agents has seen a drastic growth over the last decades. Novel imaging techniques allow real-time in vivo study of nanoparticle accumulation kinetics at the level of the cell and targeted tissue. Successful intravenous application of such nanocarriers requires a hydrophilic particle surface coating, of which polyethylene glycol (PEG) has become the most widely studied and applied. In the current study, the effect of nanoparticle PEG surface density on the targeting efficiency of ligand-functionalized nanoemulsions was investigated. We synthesized 100 nm nanoemulsions with a PEG surface density varying from 5 to 50 mol%. Fluorescent and paramagnetic lipids were included to allow their multimodal detection, while RGD peptides were conjugated to the PEG coating to obtain specificity for the αvβ3-integrin. The development of a unique experimental imaging setup allowed us to study, in real time, nanoparticle accumulation kinetics at (sub)-cellular resolution in tumors that were grown in a window chamber model with confocal microscopy imaging, and at the macroscopic tumor level in subcutaneously grown xenografts with magnetic resonance imaging. Accumulation in the tumor occurred more rapidly for the targeted nanoemulsions than for the non-targeted versions, and the PEG surface density had a strong effect on nanoparticle targeting efficiency. Counter intuitively, yet consistent with the PEG density conformation models, the highest specificity and targeting efficiency was observed at a low PEG surface density.

Nanoparticle applications in medicine have seen a tremendous growth in the last decade. In addition to their drug targeting application and their ability to improve bioavailability of drugs, nanoparticles can be designed to allow their detection with a variety of imaging methodologies. In the current study we developed a multimodal nanoparticle platform to enable imaging guided therapy, which was evaluated in a colon cancer mouse model. This “theranostic” platform, is based on oil-in-water nanoemulsions and carries iron oxide nanocrystals for MRI, the fluorescent dye Cy7 for NIRF imaging and the hydrophobic glucocorticoid prednisolone acetate valerate (PAV) for therapeutic purposes. Angiogenesis targeted nanoemulsions functionalized with αvβ3-specific RGD-peptides were evaluated as well. When subcutaneous tumor were palpable the nanoemulsions were administered at a dose of 30 mg FeO/kg and 10 mg PAV/kg. MRI and NIRF imaging showed significant nanoparticle accumulation in the tumors, while tumor growth profiles revealed a potent inhibitory effect in all the PAV-nanoemulsions treated animals as compared to the ones treated with control nanoemulsions, the free drug or saline. In conclusion, this study demonstrated that our nanoemulsions, when loaded with PAV, iron oxide nanocrystals and Cy7, represent a flexible and unique theranostic nanoparticle platform that can be applied for imaging guided therapy of cancer.

Extracellular vesicles (EVs) have been implicated in tumorigenesis. Biomolecules which can block EV binding and uptake into recipient cells may be of therapeutic value as well as enhance understanding of EV biology. Here, we show that heparin interacts with uptake of tumor-derived as well as non-tumor-derived EVs into recipient cells. Incubation of glioma cell-derived EVs with heparin resulted in micron-sized structures observed by transmission electron microscopy, with EVs clearly visible within these structures. Inclusion of heparin greatly diminished transfer of labeled EVs from donor to recipient tumor cells. We also show a direct interaction between heparin and EVs using confocal microscopy. We found that the block in EV uptake was at the level of cell binding and not internalization. Finally, incubation of glioma-derived EVs containing EGFRvIII mRNA with heparin reduced transfer of this message to recipient cells. The effect of heparin on EVs uptake may provide a unique tool to study EV function. It may also foster research of heparin or its derivatives as a therapeutic for disease in which EVs play a role.