Rationale: The blood-brain barrier (BBB) is a major impediment to therapeutic intracranial drug delivery for the treatment of neurodegenerative diseases, including Alzheimer's disease (AD). Focused ultrasound applied together with microbubbles (FUS^+MB) is a novel technique to transiently open the BBB and increase drug delivery. Evidence suggests that FUS^+MB is safe, however, the effects of FUS^+MB on human BBB cells, especially in the context of AD, remain sparsely investigated. In addition, there currently are no cell platforms to test for FUS^+MB-mediated drug delivery. Methods: Here we generated BBB cells (induced brain endothelial-like cells (iBECs) and astrocytes (iAstrocytes)) from apolipoprotein E gene allele E4 (APOE4, high sporadic AD risk) and allele E3 (APOE3, lower AD risk) carrying patient-derived induced pluripotent stem cells (iPSCs). We established mono- and co-culture models of human sporadic AD and control BBB cells to investigate the effects of FUS^+MB on BBB cell phenotype and to screen for the delivery of two potentially therapeutic AD antibodies, an Aducanumab-analogue (Aduhelm^TM; anti-amyloid-β) and a novel anti-Tau antibody, RNF5. We then developed a novel hydrogel-based 2.5D BBB model as a step towards a more physiologically relevant FUS^+MB drug delivery platform. Results: When compared to untreated cells, the delivery of Aducanumab-analogue and RNF5 was significantly increased (up to 1.73 fold), across the Transwell-based BBB models following FUS^+MB treatment. Our results also demonstrated the safety of FUS^+MB indicated by minimal changes in iBEC transcriptome as well as little or no changes in iBEC or iAstrocyte viability and inflammatory responses within the first 24 h post FUS^+MB. Furthermore, we demonstrated successful iBEC barrier formation in our novel 2.5D hydrogel-based BBB model with significantly increased delivery (1.4 fold) of Aducanumab-analogue following FUS^+MB. Conclusion: Our results demonstrate a robust and reproducible approach to utilize patient cells for FUS^+MB-mediated drug delivery screening in vitro. With such a cell platform for FUS^+MB research previously not reported, it has the potential to identify novel FUS^+MB-deliverable drugs as well as screen for cell- and patient-specific effects of FUS^+MB, accelerating the use of FUS^+MB as a therapeutic modality in AD.

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