Investigation of the physicochemical properties and selective anti-cancer efficacy of in-plasma treated PBS using an exclusive liquid-submerged plasma jet

L. Hoebus; P. Shali; N. Caz; J. Van den Bosch; R. Ghobeira; M. Narimisa; R. Morent; E. Wolfs; N. De Geyter

doi:10.5281/zenodo.17435258

Investigation of the physicochemical properties and selective anti-cancer efficacy of in-plasma treated PBS using an exclusive liquid-submerged plasma jet

L. Hoebus; P. Shali; N. Caz; J. Van den Bosch; R. Ghobeira; M. Narimisa; R. Morent; E. Wolfs; N. De Geyter

Publisher: Zenodo
DOI: 10.5281/zenodo.17435258
Repository: zenodo.org
Language: en

Abstract

Despite advances in current treatment options, cancer remains one of the leading causes of death, leading to nearly 10 million deaths in 2020. This highlights the urgent need for novel therapeutic alternatives. Plasma treated liquids (PTLs) have emerged as a promising alternative, primarily due to the generation of reactive oxygen and nitrogen species (RONS), such as H2O2, NO2-, and NO3-, during plasma treatment. In this study, a unique submerged plasma setup was used to produce high RONS concentrations in phosphate-buffered saline (PBS). The effects of plasma parameters, including gas flow rate, voltage, and treatment time, were systematically evaluated for their impact on H2O2 and NO2- concentration. Results showed that increasing treatment time and voltage significantly elevated both H2O2 and NO2- concentrations, whereas gas flow rate had minimal effect on both. Notably, the submerged setup generated exceptionally high H2O2 concentrations (>2000 µM), surpassing those typically reported in literature. Finally, the cytotoxic effects of the plasma treated PBS were assessed on human oral squamous cell carcinoma (OSCC) cells and healthy human keratinocytes (HaCaT) using cell proliferation and viability assays. The results demonstrated selective cytotoxicity, with a clear reduction in OSCC cell proliferation and viability, while sparing HaCaT cells. This selectivity underscores the strong potential of plasma treated PBS as a targeted cancer therapy that minimizes damage to healthy tissue.

Full text

Z Introduction Plasma treated liquids (PTLs) for cancer therapy Materials and methods Investigation of the physicochemical properties and selective anti-cancer efficacy of in-plasma treated PBS using an exclusive liquid-submerged plasma jet L. Hoebus1, P. Shali1, N. Caz2, J. Van den Bosch2, R. Ghobeira1, M. Narimisa1, R. Morent1, E. Wolfs2, N. De Geyter1 1Research Unit Plasma Technology (RUPT), Department of Applied Physics, Faculty of Engineering and Architecture, Ghent University, Ghent, Belgium 2Morphology Research Group, Biomedical Research Institute, Hasselt University, Campus Diepenbeek, Bioville, Diepenbeek, Belgium Optimizing RONS concentration in PBS Conclusion Direct treatment Indirect treatment →Regardless of current treatment options, cancer remains a leading global cause of death, with nearly 10 million deaths in 2020. RONS Plasma treated PBS Phosphate buffered saline solution (PBS) Varying parameters: –Voltage (4.5 kV –6.5 kV) –Flow rate (1 slm –2.5 slm) –Treatment time (1 min –10 min) Spectrophotometric detection of RONS using colorimetric assays pH and conductivity measurements Liquid loss Healthy human keratinocyte HaCaT and UMSCC-14C human oral squamous cell carcinoma (OSCC) cell line Cell proliferation was studied by evaluating cell confluency Cell viability assessment by fluorescence intensity at 530/590 nm Atmospheric pressure plasma jet Treatment time Voltage Flow rate In-liquid plasma treatment Liquid characterization Cell tests Surgery •Complication risks •Not for all cancer types •Not suitable for all patients Radiation •Damage to healthy tissue •Risk of secondary tumours •Various side effects Chemotherapy •Attacks healthy cells •Various side effects like hair loss, fatigue and nausea Direct RONS generation at target location ✓Minimally invasive ✓High local RONS concentration x Limited penetration depth x Safety concerns about direct exposure to plasma electric field and UV radiation RONS generation in liquid ✓Can be used for deeper tissues ✓No direct plasma-patient contact x Reduced reactive species x Diffusion after injection Reactive oxygen and nitrogen species (RONS) can selectively kill cancer cells without damaging healthy tissue Limited effect flow rate on RONS concentration Increasing treatment time →prolonged plasma-liquid interaction →higher concentrations of H202and NO2Increasing voltage →higher electron density in jet →more dissociation reactions → higher concentrations of H202 Cell tests Lowest confluency when exposed to plasma treated PBS →nearly no proliferation after 72h After 72h: •OSCC cell viability is lowest in plasma treated PBS •H2O2is main contributor to cytotoxicity but plasma treated PBS shows strongest effects •Treatment is highly selective Conventional cancer treatment options Optimizing RONS concentration •Very high H2O2concentration due to unique set-up •Treatment time & voltage significantly affect H2O2and NO2production Cell tests •Highly selective •Plasma treated PBS →highest OSCC cytotoxicity →Plasma treated PBS as promising therapeutic strategy for combating OSCC Contact [email protected] www.ugent.be Cell proliferation Cell viability