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Golden berry-derived 4β-hydroxywithanolide E for selectively killing oral cancer cells by generating ROS, DNA damage,
Chien-Chih Chiu1, Jo-Wen Haung, Fang-Rong Chang
1Department of Biotechnology, Kaohsiung Medical University, Kaohsiung, Taiwan.
Background:
Most chemotherapeutic drugs for killing cancer cells are highly cytotoxic in normal cells, which limits their clinical applications. Therefore, a continuing challenge is identifying a drug that is hypersensitive to cancer cells but has minimal deleterious effects on healthy cells. The aims of this study were to evaluate the potential of 4β-hydroxywithanolide (4βHWE) for selectively killing cancer cells and to elucidate its related mechanisms.
Methodology And Principal Findings:
Changes in survival, oxidative stress, DNA damage, and apoptosis signaling were compared between 4βHWE-treated oral cancer (Ca9-22) and normal fibroblast (HGF-1) cells. At 24 h and 48 h, the numbers of Ca9-22 cells were substantially decreased, but the numbers of HGF-1 cells were only slightly decreased. Additionally, the IC50 values for 4βHWE in the Ca9-22 cells were 3.6 and 1.9 µg/ml at 24 and 48 h, respectively. Time-dependent abnormal increases in ROS and dose-responsive mitochondrial depolarization can be exploited by using 4βHWE in chemotherapies for selectively killing cancer cells. Dose-dependent DNA damage measured by comet-nuclear extract assay and flow cytometry-based γ-H2AX/propidium iodide (PI) analysis showed relatively severer damage in the Ca9-22 cells. At both low and high concentrations, 4βHWE preferably perturbed the cell cycle in Ca9-22 cells by increasing the subG1 population and arrest of G1 or G2/M. Selective induction of apoptosis in Ca9-22 cells was further confirmed by Annexin V/PI assay, by preferential expression of phosphorylated ataxia-telangiectasia- and Rad3-related protein (p-ATR), and by cleavage of caspase 9, caspase 3, and poly ADP-ribose polymerase (PARP).
Conclusions/Significance:
Together, the findings of this study, particularly the improved understanding of the selective killing mechanisms of 4βHWE, can be used to improve efficiency in killing oral cancer cells during chemoprevention and therapy.
Insights
4β-hydroxywithanolide (4βHWE) selectively kills oral cancer cells by inducing DNA damage and apoptosis, sparing normal cells. This natural compound shows promise for targeted cancer therapy and chemoprevention.
Area of Science:
- Pharmacology
- Cancer Biology
- Natural Products Chemistry
Background:
- Chemotherapy faces challenges due to cytotoxicity in normal cells.
- Developing cancer-specific drugs with minimal side effects is crucial.
- 4β-hydroxywithanolide (4βHWE) is investigated for its selective anti-cancer potential.
Purpose of the Study:
- To evaluate 4β-hydroxywithanolide (4βHWE) for selective oral cancer cell killing.
- To elucidate the underlying mechanisms of 4βHWE's action.
- To assess its effects on normal fibroblast cells.
Main Methods:
- Comparison of cell survival, oxidative stress, DNA damage, and apoptosis in oral cancer (Ca9-22) and normal fibroblast (HGF-1) cells treated with 4βHWE.
- Assessment of reactive oxygen species (ROS) levels, mitochondrial depolarization, and DNA damage (comet assay, γ-H2AX/PI).
- Cell cycle perturbation analysis (subG1, G1/G2/M arrest) and apoptosis induction assays (Annexin V/PI, p-ATR, caspase cleavage, PARP cleavage).
Main Results:
- 4βHWE significantly reduced Ca9-22 cell numbers while minimally affecting HGF-1 cells.
- Increased ROS, mitochondrial depolarization, and DNA damage were observed in cancer cells.
- 4βHWE induced cell cycle arrest and apoptosis preferentially in oral cancer cells via ATR and caspase pathways.
Conclusions:
- 4βHWE demonstrates selective cytotoxicity against oral cancer cells.
- Mechanisms involve oxidative stress, DNA damage, cell cycle perturbation, and apoptosis induction.
- Findings support 4βHWE's potential for oral cancer chemoprevention and therapy.
