*Article* **Non-Thermal Plasma Couples Oxidative Stress to TRAIL Sensitization through DR5 Upregulation**

**Soon Young Hwang 1,**† **, Ngoc Hoan Nguyen 1,**† **, Tae Jung Kim <sup>2</sup> , Youngsoo Lee <sup>3</sup> , Mi Ae Kang 1,\* and Jong-Soo Lee 1,\***


Received: 8 July 2020; Accepted: 23 July 2020; Published: 26 July 2020

**Abstract:** Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in various tumor cells without affecting most normal cells. Despite being in clinical testing, novel strategies to induce TRAIL-mediated apoptosis are in need to overcome cancer cell unresponsiveness and resistance. Plasma-activated medium (PAM) markedly stimulates reactive oxygen/nitrogen species (ROS/RNS)-dependent apoptosis in cancer cells. We investigate the capability of PAM and TRAIL (PAM/TRAIL) combination therapy to overcome TRAIL resistance and improve the anticancer efficacy of TRAIL. The combinatorial treatment of PAM and TRAIL shows synergistic effects on growth inhibition in TRAIL-resistant cancer cells via augmented apoptosis by two attributes. DR5 (TRAIL-R2) transcription by CHOP is upregulated in a PAM-generated ROS/RNS-dependent manner, and PAM itself upregulates PTEN expression mediated by suppression of miR-425 which is involved in Akt inactivation, leading to increased apoptosis induction. Treatment of cancer cell lines with the antioxidant N-acetylcysteine reduces the extent of membrane dysfunction and the expression of both CHOP-DR5 and miR-425-PTEN axes, attenuating PAM/TRAIL-induced cancer cell apoptosis. These data suggest that PAM/TRAIL treatment is a novel approach to sensitizing cancer cells to TRAIL-induced apoptosis and overcoming TRAIL resistance. PAM is a promising candidate for further investigations as a chemotherapeutic sensitizer in the treatment of cancer.

**Keywords:** plasma-activated medium; TRAIL; DR5; apoptosis; ROS/RNS
