Tumor treating fields effects on the blood-brain barrier in vitro and in vivo.

Abstract

2551 Background: The greatest hurdle, which even potent and effective drugs targeting central nervous system (CNS) tumors and other disorders face, is the blood brain barrier (BBB). The inability to cross the tight regulatory mechanism renders these drugs futile. Of late, administration of tumor treating fields (TTFields) as part of a combined treatment modality for glioblastoma demonstrated increased overall patient survival. Still, the effects of TTFields on the BBB have not yet been investigated. Here, we report the potential of TTFields application to open up the BBB. Methods: Murine brain endothelial cells were treated with 100-300 kHz TTFields for 24-96 h. Cells were also allowed to recover from 24-96 h after treatment. Subsequently, changes in cell morphology, integrity, and permeability were observed via staining of intercellular junction proteins (IJP) as well as transendothelial electrical resistance (TEER)and permeability assays. In vivo, rats were treated with 100 kHz TTFields or heat for 72 h after which they were IV injected with Evan´s Blue (EB)/ TRITC-dextran (TD) which was later quantified from the brain. Rat brain cryosections were also stained for IJPs as well as immunoglobulin G (IgG) to assess vessel structure. Finally, serial dynamic contrast-enhanced (DCE) MRI with gadolinium (Gd) contrast agent was performed pre- and post- TTFields. Results: Upon TTFields application, IJPs such as claudin-5 were delocalized from the cell membrane to the cytoplasm with maximal effects at 100 kHz. In addition, BBB integrity was significantly reduced and permeability for 4 kDa molecules was significantly increased. Cell morphology recovery was first observed at 48 h post-treatment and completely restored to normal after 96 h, indicating a reversibility of the TTFields effect on the BBB. In addition, EB and TD permeated the rat brain post-TTFields treatment. Brain cryosections displayed IJPs delocalization as well as IgG accumulation in the brain parenchyma. Confirming these observations, increased Gd in the brain was shown by DCE-MRI post-TTFields application. A reversion to normal conditions was detected 96 h after end of treatment, which was demonstrated by no difference in contrast enhancement between control and treated rats. Conclusions: TTFields application both in vitro and in vivo points towards its ability to transiently open the BBB. This presents TTFields as a novel aid for drug delivery geared towards treatment of CNS tumors and other related diseases. Hence, it is indicative of the possibility of an enhanced and more effective combinatorial therapeutic strategy.