(636c) Glucose Oxidase Delivery Using Polylysine-Grafted-Polyethylene Glycol Nanoparticles for Cancer Therapy

Prostate cancer is the most common type of non-skin cancer among men, affecting a significant portion of the population. Developing new therapeutic strategies for treating prostate cancer will have a significant impact on the quality of life and life expectancy of patients. For the treatment of this type of cancer, different therapeutic agents have been introduced. Among these therapeutic agents, the potential of glucose oxidase (GOX) for prostate cancer treatment has recently received renewed interest. Improving the efficacy of the treatment process and reducing the drawbacks, however, requires a targeted delivery system that retains glucose oxidase within the tumor. In this study, we synthesized and characterized GOX-encapsulating polylysine-grafted-polyethylene glycol (PLL-g-PEG) nanoparticles (NPs) that were labeled with an antibody targeting prostate-specific membrane antigen (PSMA).

Firstly, we synthesized a library of PLL-g-PEG copolymers and GOX nanoparticles with different molecular weights of PLL and different degrees of PEGylation (2%, 10%, and 20%). These copolymers were characterized via NMR to verify the degree of PEGylation. NPs were formed subsequently through electrostatic self-assembly between the copolymer and GOX enzyme and stabilized with a glutaraldehyde cross-linker. Next, the NPs were functionalized with an anti-PSMA antibody. The targeted NPs were characterized based on particle size, zeta potential, stability, and enzyme activity to understand how these properties correlate with the conditions of enzyme encapsulation and particle formation. The uptake of NPs with and without antibody and their cytotoxicity was evaluated on PSMA expressing (LNCaP) and non-PSMA expressing (PC3) cell lines. The results showed that NPs with anti-PSMA antibody had better cytotoxicity against PSMA-expressing prostate cancer cells. The immunogenicity of NPs was determined by quantifying the production of different cytokines (GM-CSF, IFN-γ, IL-1β, IL-2, IL-4, IL-5, IL-10, TNF-α) produced from RAW 264.7 mouse macrophage cells exposed to the particles. Two samples with the best efficacy were chosen for the final evaluation using a xenograft murine model. Subcutaneous prostate tumors composed of LNCaP cells were allowed to develop in a nude mouse, and the mice were then treated with GOX-encapsulating NPs.

In tissue culture, NPs were shown to be resistant to uptake by macrophages and induce high toxicity to PSMA-expressing prostate cancer cells. In the mouse model, intratumoral injection was performed and the location of NPs was tracked using live animal imaging, showing reasonable retention within the tumor. Efficacy was determined by monitoring the size of the tumor where we observed that GOX NPs inhibited tumor growth. Tumor specimens were harvested and tissue sections stained to determine tissue histology including the extent of necrosis and apoptosis.