Cold Physical Plasma in Cancer Therapy: Mechanisms, Signaling, and Immunity

Despite recent advances in therapy, cancer still is a devastating and life-threatening disease, motivating novel research lines in oncology. Cold physical plasma, a partially ionized gas, is a new modality in cancer research. Physical plasma produces various physicochemical factors, primarily reactive oxygen and nitrogen species (ROS/RNS), causing cancer cell death when supplied at supraphysiological concentrations. This review outlines the biomedical consequences of plasma treatment in experimental cancer therapy, including cell death modalities. It also summarizes current knowledge on intracellular signaling pathways triggered by plasma treatment to induce cancer cell death. Besides the inactivation of tumor cells, an equally important aspect is the inflammatory context in which cell death occurs to suppress or promote the responses of immune cells. This is mainly governed by the release of damage-associated molecular patterns (DAMPs) to provoke immunogenic cancer cell death (ICD) that, in turn, activates cells of the innate immune system to promote adaptive antitumor immunity. The pivotal role of the immune system in cancer treatment, in general, is highlighted by many clinical trials and success stories on using checkpoint immunotherapy. Hence, the potential of plasma treatment to induce ICD in tumor cells to promote immunity targeting cancer lesions systemically is also discussed.

Lack of Adverse Effects of Cold Physical Plasma-Treated Blood from Leukemia Patients: A Proof-of-Concept Study

Chronic lymphocytic leukemia (CLL) is the most common blood malignancy with multiple therapeutic challenges. Cold physical plasma has been considered a promising approach in cancer therapy in recent years. In this study, we aimed to evaluate the cytotoxic effect of cold plasma or plasma-treated solutions (PTS) on hematologic parameters in the whole blood of CLL patients. The mean red blood cell count, white blood cell (WBC) count, platelet and hemoglobin counts, and peripheral blood smear images did not significantly differ between treated and untreated samples in either CLL or healthy individuals. However, both direct plasma and indirect PTS treatment increased lipid peroxidation and RNS deposition in the whole blood of CLL patients and in healthy subjects. In addition, the metabolic activity of WBCs was decreased with 120 s of cold plasma or PTS treatment after 24 h and 48 h. However, cold plasma and PTS treatment did not affect the prothrombin time, partial thromboplastin time, nor hemolysis in either CLL patients or in healthy individuals. The present study identifies the components of cold plasma to reach the blood without disturbing the basic parameters important in hematology, confirming the idea that the effect of cold plasma may not be limited to solid tumors and possibly extends to hematological disorders. Further cellular and molecular studies are needed to determine which cells in CLL patients are targeted by cold plasma or PTS.

H2A.X Phosphorylation in Oxidative Stress and Risk Assessment in Plasma Medicine

At serine139-phosphorylated gamma histone H2A.X (γH2A.X) has been established over the decades as sensitive evidence of radiation-induced DNA damage, especially DNA double-strand breaks (DSBs) in radiation biology. Therefore, γH2A.X has been considered a suitable marker for biomedical applications and a general indicator of direct DNA damage with other therapeutic agents, such as cold physical plasma. Medical plasma technology generates a partially ionized gas releasing a plethora of reactive oxygen and nitrogen species (ROS) simultaneously that have been used for therapeutic purposes such as wound healing and cancer treatment. The quantification of γH2A.X as a surrogate parameter of direct DNA damage has often been used to assess genotoxicity in plasma-treated cells, whereas no sustainable mutagenic potential of the medical plasma treatment could be identified despite H2A.X phosphorylation. However, phosphorylated H2A.X occurs during apoptosis, which is associated with exposure to cold plasma and ROS. This review summarizes the current understanding of γH2A.X induction and function in oxidative stress in general and plasma medicine in particular. Due to the progress towards understanding the mechanisms of H2A.X phosphorylation in the absence of DSB and ROS, observations of γH2A.X in medical fields should be carefully interpreted.

Non-Invasive Physical Plasma Enhances the Membrane Permeability to Low Molecular Weight Compounds and Subsequently Leads to the Loss of Cellular ATP and the Devitalization of Epithelial Cancer Cells

Non-invasive physical plasma (NIPP) achieves biomedical effects primarily through the formation of reactive oxygen and nitrogen species. In clinical use, these species interact with cells of the treated tissue, affecting the cytoplasmic membrane first. The present study investigated the permeability of the cytoplasmic membrane of breast cancer cells with different fluorescent dyes after NIPP treatment and determined the subsequent effects on cell viability. After NIPP treatment and the associated formation of reactive oxygen species, low molecular weight compounds were able to pass through the cytoplasmic membrane in both directions to a higher extent. Consequently, a loss of cellular ATP into the extracellular space was induced. Due to these limitations in cell physiology, apoptosis was induced in the cancer cells and the entire cell population exhibited decreased cell growth. It can be concluded that NIPP treatment disturbs the biochemical functionality of the cytoplasmic membrane of cancer cells, which massively impairs their viability. This observation opens a vast application horizon of NIPP therapy to treat precancerous and malignant diseases beyond breast cancer therapy.

Degradation of glyphosate in water by the application of surface-corona-discharges

Glyphosate (GLP) is one of the most widely applied herbicide, which is found ubiquitously in the environment. The removal of glyphosate from waste water and soil is challenging and can be achieved with chemical or biological methods, which, nevertheless, suffer from different disadvantages. The application of a physical plasma for the removal of GLP in water was examined by the application of surface corona discharges in a wire-to-cylinder setup filled with argon. The plasma was ignited at the liquid surface without any additives. By applying a photometric method, GLP was detected after derivatisation with fluorenyl methoxycarbonyl chloride, whereas phosphate was determined with ammonium molybdate. A GLP degradation rate of 90.8% could be achieved within a treatment time of 30 minutes with an estimated energy efficiency of 0.32 g/kWh.

The Anticancer Efficacy of Plasma-Oxidized Saline (POS) in the Ehrlich Ascites Carcinoma Model In Vitro and In Vivo

Cold physical plasma, a partially ionized gas rich in reactive oxygen species (ROS), is receiving increasing interest as a novel anticancer agent via two modes. The first involves its application to cells and tissues directly, while the second uses physical plasma-derived ROS to oxidize liquids. Saline is a clinically accepted liquid, and here we explored the suitability of plasma-oxidized saline (POS) as anticancer agent technology in vitro and in vivo using the Ehrlich Ascites Carcinoma (EAC) model. EAC mainly grows as a suspension in the peritoneal cavity of mice, making this model ideally suited to test POS as a putative agent against peritoneal carcinomatosis frequently observed with colon, pancreas, and ovarium metastasis. Five POS injections led to a reduction of the tumor burden in vivo as well as in a decline of EAC cell growth and an arrest in metabolic activity ex vivo. The treatment was accompanied by a decreased antioxidant capacity of Ehrlich tumor cells and increased lipid oxidation in the ascites supernatants, while no other side effects were observed. Oxaliplatin and hydrogen peroxide were used as controls and mediated better and worse outcomes, respectively, with the former but not the latter inducing profound changes in the inflammatory milieu among 13 different cytokines investigated in ascites fluid. Modulation of inflammation in the POS group was modest but significant. These results promote POS as a promising candidate for targeting peritoneal carcinomatosis and malignant ascites and suggest EAC to be a suitable and convenient model for analyzing innovative POS approaches and combination therapies.

Plasma-Treated Solutions (PTS) in Cancer Therapy

Cold physical plasma is a partially ionized gas generating various reactive oxygen and nitrogen species (ROS/RNS) simultaneously. ROS/RNS have therapeutic effects when applied to cells and tissues either directly from the plasma or via exposure to solutions that have been treated beforehand using plasma processes. This review addresses the challenges and opportunities of plasma-treated solutions (PTSs) for cancer treatment. These PTSs include plasma-treated cell culture media in experimental research as well as clinically approved solutions such as saline and Ringer’s lactate, which, in principle, already qualify for testing in therapeutic settings. Several types of cancers were found to succumb to the toxic action of PTSs, suggesting a broad mechanism of action based on the tumor-toxic activity of ROS/RNS stored in these solutions. Moreover, it is indicated that the PTS has immuno-stimulatory properties. Two different routes of application are currently envisaged in the clinical setting. One is direct injection into the bulk tumor, and the other is lavage in patients suffering from peritoneal carcinomatosis adjuvant to standard chemotherapy. While many promising results have been achieved so far, several obstacles, such as the standardized generation of large volumes of sterile PTS, remain to be addressed.