Low Dose Combined Treatment with Ultraviolet-C and Withaferin a Enhances Selective Killing of Oral Cancer Cells

Sheng-Yao Peng,Jen-Yang Tang,Ting-Hsun Lan,Yen-Yun Wang,Shyng-Shiou F. Yuan,Li-Ching Lin,Hsueh-Wei Chang

doi:10.3390/antiox9111120

Abstract

Withaferin A (WFA), a Withania somnifera-derived triterpenoid, is an anticancer natural product. The anticancer effect of nonionizing radiation such as ultraviolet-C (UVC) as well as the combined treatment of UVC and WFA is rarely investigated. Low dose UVC and/or WFA treatments (12 J/m2 and/or 1 μM) were chosen to evaluate antioral cancer cell line effects by examining cytotoxicity, cell cycle disruption, apoptosis induction, and DNA damage. For two cancer cell lines (Ca9-22 and HSC-3), single treatment (UVC or WFA) showed about 80% viability, while a combined treatment of UVC/WFA showed about 40% viability. In contrast, there was noncytotoxicity to normal oral cell lines (HGF-1). Compared to single treatment and control, low dose UVC/WFA shows high inductions of apoptosis in terms of flow cytometric detections for subG1, annexin V, pancaspase changes as well as Western blotting for detecting cleaved poly (ADP-ribose) polymerase (c-PARP) and caspase 3 (c-Cas 3) and luciferase assay for detecting Cas 3/7 activity. Low dose UVC/WFA also showed high inductions of oxidative stress and DNA damage in terms of flow cytometric detections of reactive oxygen species (ROS), mitochondrial superoxide (MitoSOX) generation, and membrane potential (MitoMP) destruction, γH2AX and 8-oxo-2’deoxyguanosine (8-oxodG) types of DNA damages. For comparison, low dose UVC/WFA show rare inductions of annexin V, Cas 3/7 activity, ROS, MitoSOX, and MitoMP changes to normal oral HGF-1 cells. Therefore, low dose UVC/WFA provides a novel selectively killing mechanism to oral cancer cells, suggesting that WFA is a UVC sensitizer to inhibit the proliferation of oral cancer cells.

Highlights

Radiotherapy and chemotherapy are commonly used for curing oral cancer [1]
Combined treatment with low dose methanolic extracts of Cryptocarya concinna (10 μg/mL; 80.4% viability) and UVC (14 J/m2; 83.2% viability) improve antiproliferation against oral cancer cells compared to single treatment [25]
We demonstrated that a low dose combined treatment of UVC/Withaferin A (WFA) dramatically decreased cell viability (~40%) of two oral cancer cell lines (Ca9-22 and HSC-3) without cytotoxic effects on normal oral cells in a 24-h MTS assay (Figure 1)

Summary

Introduction

Radiotherapy and chemotherapy are commonly used for curing oral cancer [1]. They raise the problems of radio- and chemo-resistance [2]. A novel strategy was developed to reduce chemoresistance by using low dose treatment. In the clinical drug doxorubicin, its low dose treatment improved survival by suppressing the patient-derived chemoresistant leukemia stem cells in an animal model study [3]. Low dose drug treatments have a potential for reducing chemoresistance in oral cancer therapy. Low dose radiation treatments have a potential for reducing radioresistance in cancer therapy. In addition to dose adjustments of radiation, a combined treatment provides an alternative strategy to overcome radioresistance in cancer therapy. DNA-dependent protein kinase (DNA-PK) inhibitors KU57788 and IC87361 [5] and multikinase inhibitors sorafenib and sunitinib [6] displayed radiosensitization to head and neck cancer cells

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