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.

The Mechanisms of Action of Tumor Treating Fields

“…Using this cell line, it was shown that the administration of frequency-, intensity-, as well as duration- dependent TTFields results in the delocalization of its major tight junction protein claudin-5, as well as the cytoplasmic accessory protein zonula occludens-1 (ZO-1), from the cell membrane boundaries into the cytoplasm ( 82 ). Allowing the cells to recover up to 96h post-TTFields led to the restoration of the original cellular morphology, pointing to the reversibility of the process.…”

An overview of potential novel mechanisms of action underlying Tumor Treating Fields-induced cancer cell death and their clinical implications

“…Moreover, this effect can be modulated with the duration of cell membrane permeability dependent upon the length of TTFields exposure. Increased cancer cell permeability may have clinical implications such as increased uptake of chemotherapeutic agents which would be especially important when considering the potential to open up the blood-brain barrier in the treatment of GBM (Salvador et al 2020).…”

Tumor treating fields for newly diagnosed high‐grade glioma based on the criteria of 2021 WHO CNS5: A retrospective analysis of Chinese patients in a single center

“… 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 The anti‐tumor mechanisms of action of TTFields are also associated with influencing tumor cell migration, increasing the permeability of tumor cell membranes and the blood‐brain barrier, inducing autophagy in tumor cells, increasing DNA damage, and inducing antitumor immunity. 23 , 24 , 25 , 26 , 27 , 28 , 29 , 30 , 31 TTFields have been approved by the FDA for recurrent and newly diagnosed GBM based on the results of clinical studies in EF‐11 and EF‐14. 14 , 32 In addition, TTFields have been approved for the treatment of mesothelioma and are currently being studied in many other tumor types including non‐small cell lung cancer, pancreatic cancer, ovarian cancer and so on.…”

“…TTFields interfere with mitosis in tumor cells by disrupting microtubulin polymerization, septal localization, and cytokinesis, preventing mitotic spindle assembly, and adjusting cellular membrane potential to increase the influx of intracellular Ca 2+ to promote microtubulin depolymerization, which causes cell cycle arrest, and ultimately leads to tumor cell death/apoptosis 15–22 . The anti‐tumor mechanisms of action of TTFields are also associated with influencing tumor cell migration, increasing the permeability of tumor cell membranes and the blood‐brain barrier, inducing autophagy in tumor cells, increasing DNA damage, and inducing antitumor immunity 23–31 . TTFields have been approved by the FDA for recurrent and newly diagnosed GBM based on the results of clinical studies in EF‐11 and EF‐14 14,32 .…”