SCDT-02. BLOOD-BRAIN-BARRIER SPHEROIDS AS AN IN VITRO SCREENING PLATFORM FOR BRAIN-PENETRATING AGENTS

Abstract

The inability of most systemically delivered therapeutics to cross the blood-brain-barrier (BBB) is considered a major barrier to effective brain cancer treatment. In malignant glioma the tumor neovasculature is known to be leaky, however invasive cells remain protected behind an intact BBB, and continue to thrive within the brain, ultimately leading to tumor regrowth and patient death. This leaves us with an urgent unmet need for the development of next-generation therapeutics with improved brain delivery. For the first time, we describe here the utility of 3D multicellular BBB spheroids made of human brain endothelial cells (ECs), pericytes and astrocytes as a screening tool for brain-penetrating agents. We show that the outer surface of the spheroids, composed primarily of ECs and pericytes, form a tight barrier which is permeabilized in the presence of VEGF. The barrier is characterized by the presence of intact tight junctions and efflux-pump activity (i.e., P-glycoprotein). Furthermore, we have used this model to successfully demonstrate the transport of angiopep-2 (a well-known brain delivery vector) and its conjugates (containing cargoes of various sizes such as peptide, protein and affibody), thereby displaying the versatility of this model to screen and study a wide range of therapeutic agents. We demonstrate that this model is superior to the conventional transwell model in maintaining essential BBB characteristics (i.e., tight/adherens junctions and P-glycoprotein expression) and as a drug-screening tool. We have utilized the spheroid model to screen a panel of cell-penetrating peptides (CPPs) to identify several candidates with high brain-penetration potential. We then verified the ability of the top 4 candidate CPPs to cross the BBB in mice. This high-throughput model can lead to better design and analysis of first-in-class glioma therapeutics, and improve prediction of drug penetration in a living model, paving the way for breakthrough discoveries in brain cancer.