PRUSSIAN BLUE NANOIMMUNOTHERAPIES FOR NEUROBLASTOMA
Fisher, John P
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Neuroblastoma is the most common extracranial solid tumor in children, accounting for 15% of cancer-related deaths. Despite improvements in diagnosis and surgical techniques, neuroblastoma remains challenging to treat due to the heterogeneity of the tumor, low neoantigen expression, immunosuppressive tumor environment, and high recurrence rate. We have therefore engineered a nanoimmunotherapy that combines the advantages of nanotechnology and immunotherapy to combat the aforementioned challenges in treating neuroblastoma. Specifically, our ensemble comprises of Prussian blue nanoparticles (PBNPs) biofunctionalized with the immune adjuvant CpG-oligodeoxynucleotide (CpG). We utilize PBNPs for photothermal therapy (PTT), which ablates tumor cells and releases tumor antigens and adjuvants that increase tumor immunogenicity. Additionally, the PBNPs are biofunctionalized with CpG (CpG-PBNPs) to serve as a depot for local delivery of exogenous immune adjuvants that play an important role in breaking tolerance to tumor antigens and improving tumor antigen presentation. We hypothesize that this approach of targeting tumor cells, antigen presenting cells, and T cells, may hold the key in converting a non-responsive “cold” tumor such as neuroblastoma into a responsive “hot” tumor, leading to better treatments. We first describe the synthesis and characterization of CpG-PBNPs using a facile layer-by-layer coating scheme. The resultant nanoparticles exhibit monodisperse size distributions, multiday stability, and are not cytotoxic. The strong, intrinsic absorption of PBNPs in the CpG-PBNPs is leveraged to administer PTT (CpG-PBNP-PTT) that triggers immunogenic tumor cell death releasing tumor antigens, which increases tumor antigenicity. Simultaneously, the CpG coating functions as an exogenous adjuvant that complements the endogenous adjuvants released by the CpG-PBNP-PTT (e.g. ATP, calreticulin, and HMGB1), increasing adjuvanticity. When administered in a murine model of neuroblastoma, CpG-PBNP-PTT results in complete tumor regression in a significantly higher proportion (70%) of treated animals relative to controls. Further, the long-term surviving, CpG-PBNP-PTT-treated animals reject tumor rechallenge suggesting that our nanoimmunotherapy generates immunological memory. When we treat a synchronous model of neuroblastoma, 50% of nanoimmunotherapy-treated mice show complete eradication of both tumors compared to controls, which showed no survival efficacy. Our findings show the importance of simultaneous cytotoxicity, antigenicity, and adjuvanticity in generating robust and persistent antitumor immune responses. The strategies described in this dissertation encompass novel examples of nanoimmunotherapies to be applied in the clinic for the treatment of neuroblastoma.