3D Bioprinted Blood–Brain–Tumor Barrier Spheroid‐Based In Situ Evaluation of Anticancer Compounds Against Glioblastoma: ROS‐Producing Ruthenium as a Potential Therapeutic Candidate

ABSTRACT The blood–brain–tumor barrier (BBTB) penetration by anticancer drugs and their effects on glioblastoma (GBM) are assessed using 3D bioprinted‐spheroids. Comparative analysis reveals prominent cytoplasmic mixing between pericytes and the human GBM cell line U‐87MG, expressions of tight junction proteins ZO‐1, occludin, and claudin‐5, which act as a barrier to the paracellular transport across BBTB, and the expression of ABC transporter such as P‐glycoprotein (P‐gp) in the vascular endothelial cell layer of BBTB spheroids. Anticancer drug‐induced U‐87 MG cell death is quantified through in situ imaging of spheroids using a cancer cell death probe. Paclitaxel and cisplatin, known P‐gp substrates, do not demonstrate transcellular transport in BBTB spheroids, whereas temozolomide and a ruthenium (Ru) compound induce marked U‐87 MG cell death. Relative BBTB permeability is calculated as a function of IC 50 and treatment concentration. The Ru compound, a partial P‐gp substrate, exhibits greater anticancer efficacy compared with temozolomide, which is not a P‐gp substrate, in BBTB spheroids. The Ru compound attenuates tight junction protein expression (occludin and ZO‐1) in vascular endothelial cells through the released reactive oxygen species (ROS), increases ECM degradation via elevated matrix metalloproteinase‐9 in vascular endothelial and astrocytes, and promotes expression of the vasoconstrictor endothelin‐1.