Despite recent advancements in cancer treatment, the 5-year survival rate for glioblastoma is only 22%. The greatest challenge in brain cancer treatment is the blood-brain barrier (BBB), a physical barrier that protects the central nervous system (CNS) from circulating solutes in the blood, but in cancer, prevents therapeutics from entering the brain space. While surgery is the gold-standard treatment, this procedure is high-risk. Thus, we are investigating injectable therapies to treat brain tumors by crossing the BBB. To do this, we are developing nanoparticles (NPs) to cross the BBB via receptor-mediated transcytosis (RMT). In RMT, ligands (keys) bind to receptors (locks) expressed by the BBB to gain entrance into the CNS. By attaching the ligand transferrin, we can trick the BBB into granting our NPs access to the brain. Prior to testing particles in vivo, I am developing a cellular model of the BBB to assess the ability of different NP formulations to cross the BBB in vitro. The transwell model comprises two main compartments: the donor and acceptor compartments, separated by a cellular monolayer that only allows transport between compartments via RMT. Here, I developed a co-culture consisting of brain endothelial cells and astrocytes. This co-culture model is more representative of the BBB and provides higher monolayer integrity than single-cell models. In parallel, I am investigating the use of acidification inhibitors to enhance the ability of NPs to cross the BBB. If successful, these inhibitors will be incorporated into a new NP design. Through this project, I am (i) developing a more representative in vitro model of the BBB and (ii) exploring alternative mechanisms to enhance NP transport through the BBB. Ultimately, these two aims will enable us to better direct NP behavior in vivo and cross the BBB.