PROCESS DESIGN OF NANOGHOST PEGYLATION TO PREVENT AGGREGATION AND ENHANCE TARGETING

We have developed a drug delivery vehicle for cancer therapy derived from cytoplasmic membranes of naturally targeted cells that will be more effective than currently available particulate drug delivery systems. The delivery system is known as a nanoghost (NG); as it is on the nano-scale and incorporates the cytoplasmic membrane of ghost cells. The work presented here focuses on optimizing the device itself, with particular interest on the PEGylation of the device. PEGylation of the NGs was initially examined for its ability to reduce opsonization-dependent uptake and increase circulation time. The specific PEG quantity necessary for the device and the kinetics of the PEGylation reaction were further evaluated in order to determine the optimal PEGylation conditions that prevent particle aggregation, without over-PEGylating, and enhances targeting. Different quantities of PEG-5000 were reacted with fluorescently-labeled NGs for various times, and the resulting fluorescent PEG-NGs were incubated in vitro with A549, PC3, MCF-7, and U87 cancer cells with SMCs as a negative control to observe the effect of PEGylation on targeting. FACS analysis was used to observe the targeting results, and it was demonstrated that reacting 0.5 mg PEG for every 1 mg protein, as determined by Bradford Assay, for 1 h was ideal. Below 0.5 mg PEG, or if the reaction was less than 1 h, the targeting was decreased as a result of particle aggregation. Above 0.5 mg PEG, or for reactions longer than 1 h, the targeting was decreased as a result of masking MSC-surface markers, or over-PEGylation.