Thermoplastic microfluidic bioreactors with integrated electrodes to study tumor treating fields on yeast cells

Abstract

Tumor-treating fields (TTFields) are alternating electrical fields of intermediate frequency and low intensity that can slow or inhibit tumor growth by disrupting mitosis division of cancerous cells through cell cycle proteins. In this work, for the first time, an in-house fabricated cyclo-olefin polymer made microfluidic bioreactors are integrated with Cr/Au interdigitated electrodes to test TTFields on yeast cells with fluorescent protein:Nop56 gene. A small gap between electrodes (50 μm) allows small voltages (<150 mV) to be applied on the cells; hence, uninsulated gold electrodes are used in the non-faradaic region without causing any electrochemical reaction at the electrode-medium interface. Electrochemical modeling as well as impedance characterization and analysis of the electrodes are done using four different cell nutrient media. The experiments with yeast cells are done with 150 mV, 150 kHz and 30 mV, 200 kHz sinusoidal signals to generate electrical field magnitudes of 6.58 V/cm and 1.33 V/cm, respectively. In the high electrical field experiment, the cells go through electroporation. In the experiment with the low electrical field magnitude for TTFields, the cells have prolonged mitosis from typical 80–90 min to 200–300 min. Our results confirm the validity of the electrochemical model and the importance of applying a correct magnitude of the electrical field. Compared to the so far reported alternatives with insulated electrodes, the here developed thermoplastic microfluidic bioreactors with uninsulated electrodes provide a new, versatile, and durable platform for in vitro cell studies toward the improvement of anti-cancer therapies including personalized treatment.

A Microfluidic Platform for the Time-Resolved Interrogation of Polarized Retinal Pigment Epithelial Cells

“…Multiple layers of microfluidic elements were laser-cut from preformed acrylic (8560K172; McMaster-Carr, Elmhurst, IL) and silicone (87315K71 and 87315K73; McMaster-Carr) sheets using an infrared laser cutter (Mini 30W; Epilog Laser, Golden, CO). Unlike other low-volume, membrane insert devices in the literature, our approach precluded the need for either molding processes, such as those used to make soft lithography with PDMS 20 or thermoplastics, 21 or the need for three-dimensional printing. 22 A Transwell membrane supporting a polarized monolayer of RPE cells was cut out of a Transwell insert with a biopsy punch to create a 12-mm diameter membrane disk covered with cells.…”

Anti-cancer mechanisms of action of therapeutic alternating electric fields (tumor treating fields [TTFields])

“…Various publications demonstrate that the effect of TTFields on tumor cell growth depends on the frequency, intensity, duration, and field direction for various cell types, with the optimal therapeutic frequency window of 100–300 kHz ( Kirson et al., 2007 , 2009b ; Giladi et al., 2014a ; Voloshin et al., 2016 ; Porat et al., 2017 ; Silginer et al., 2017 ; Jo et al., 2018a , 2019 , 2020 ; Berkelmann et al., 2019 ; Lee et al., 2019 ; Gencturk et al., 2020 ; Bobkova et al., 2021 ; Jeong et al., 2021 ; Linder et al., 2021 ; Mumblat et al., 2021 ; Pfeifer et al., 2021 ; Sabri and Brosseau, 2021 ; Wu et al., 2021 ; Branter et al., 2022 ). Higher frequency and durations of exposure >72 h did not more significantly reduce cell proliferation and did not more significantly improve treatment outcomes ( Kirson et al., 2007 , 2009b ; Giladi et al., 2014a ; Voloshin et al., 2016 ; Porat et al., 2017 ; Silginer et al., 2017 ; Jo et al., 2018a , 2019 ; Berkelmann et al., 2019 ; Lee et al., 2019 ; Gencturk et al., 2020 ). The optimal TTFields frequencies for various cancer cell lines are summarized in Supplementary Table S1 .…”

“…(2017) performed similar experiments in LTCs (LN-18 and LN-229) and glioma initiating cells (ZH-161) but, instead, used 100 kHz TTFields with a reduced exposure duration of 6 h. They did not detect caspase-3 processing upon TTFields exposure, indicating that TTFields-induced cell death occurred in a caspase-independent manner ( Silginer et al., 2017 ). The effect of TTFields on cancer cells has been shown to depend on the cell type as well as the frequency, intensity, duration, number of cycles, and field direction of the TTFields ( Kirson et al., 2007 , 2009b ; Giladi et al., 2014a ; Voloshin et al., 2016 ; Porat et al., 2017 ; Silginer et al., 2017 ; Jo et al., 2018a , 2019 ; Berkelmann et al., 2019 ; Lee et al., 2019 ; Gencturk et al., 2020 ; Pfeifer et al., 2021 ; Wu et al., 2021 ; Ye et al., 2022 ). For instance, Wu et al.…”

Recent Advances in Microscale Electroporation

“…Microscale EP allows for precise control over electrical and delivery conditions exerted on single cells and larger in vitro biological systems. For example, Gencturk et al examined the response of S. cerevisiae cells to electric fields at different voltage magnitudes 352. Cells treated at sub-EP voltages underwent prolonged mitosis and took 200−300 min to divide instead of the normal 80−90 min.…”