(636a) Leveraging Immortalized Brain Endothelial Cells to Enable High-Throughput, Preclinical Screening of Nanomaterials for Drug Delivery across the Blood Brain Barrier.

With millions of patients suffering from neurological diseases and brain tumors, and current treatment options providing only symptom management or minor improvements in survival, there is a major need to deliver therapeutics across the blood brain barrier (BBB). Nanoparticle drug carriers provide a particularly promising strategy to improve drug accumulation in the brain. However, there is currently no standardized in vitro model of the BBB for high-throughput screening of nanocarrier candidates. Here, we evaluate three high-throughput methods for assessing BBB uptake and transport of nanomaterials: flow cytometry, monolayer association, and Transwell transport. We apply the immortalized human cerebral microcapillary endothelium cell line hCMEC/D3, cultured on standard cell culture plasticware or on Transwell supports, to model the BBB. We assess factors including monolayer development time, Transwell pore size, experiment temperature, nanomaterial stiffness, nanoparticle surface chemistry, changes in endothelial marker gene expression, and availability of transporters on the cell surface, to determine how they affect uptake of nanomaterials in the high-throughput model. Then, we examine brain accumulation of a select group of the nanomaterials in vivo in mice to compare it to the relative predictions made by the three BBB models. In the Transwell model, we demonstrate that there is little difference in particle transport after four or seven days of monolayer development, despite changes in paracellular tight junction permeability. Further, we show that the nanomaterials undergo active, specific transport through the cells, and that core material stiffness and surface chemistry both play roles in determining the amount of nanomaterial taken up by each model. We likewise demonstrate that both of these factors govern accumulation into the brain in vivo. Taken together, our data show that flow cytometry, monolayer association, and Transwell transport each provide advantages and caveats for library screening of nanomaterial uptake across the BBB, and that the choice of model is dependent on the intended application of the nanomaterials undergoing development. This work details the pros and cons of each method and provides a framework for which model(s) are most applicable to screening each candidate set, enabling the rational design and selection of promising nanomaterials to elevate to more complex models of uptake into the brain.

This work is supported by Cancer Research UK grant number C42454/A28596, and we would like to acknowledge use of core facilities, especially the Flow Cytometry and Microscopy cores, in the MIT Koch Institute Swanson Biotechnology Center, which is supported by the Koch Institute Core Grant P30-CA14051 from the NCI.