Evaluation of Arsenic Trioxide Potential for Lung Cancer Treatment: Assessment of Apoptotic Mechanisms and Oxidative Damage.

BACKGROUND: Lung cancer is one of the most lethal and common cancers in the world, causing up to 3 million deaths annually. The chemotherapeutic drugs that have been used in treating lung cancer include cisplatin-pemetrexed, cisplastin-gencitabinoe, carboplatin-paclitaxel and crizotinib. Arsenic trioxide (ATO) has been used in the treatment of acute promyelocytic leukemia. However, its effects on lung cancer are not known. We hypothesize that ATO may also have a bioactivity against lung cancer, and its mechanisms of action may involve apoptosis, DNA damage and changes in stress-related proteins in lung cancer cells. METHODS: To test the above stated hypothesis, lung carcinoma (A549) cells were used as the test model. The effects of ATO were examined by performing 6-diamidine-2 phenylindole (DAPI) nuclear staining for morphological characterization of apoptosis, flow cytometry analysis for early apoptosis, and western blot analysis for stress-related proteins (Hsp70 and cfos) and apoptotic protein expressions. Also, the single cell gel electrophoresis (Comet) assay was used to evaluate the genotoxic effect. RESULTS: ATO-induced apoptosis was evidenced by chromatin condensation and formation of apoptotic bodies as revealed by DAPI nuclear staining. Cell shrinkage and membrane blebbing were observed at 4 and 6 µg/ml of ATO. Data from the western blot analysis revealed a significant dose-dependent increase (p < 0.05) in the Hsp 70, caspase 3 and p53 protein expression, and a significant (p < 0.05) decrease in the cfos, and bcl-2 protein expression at 4 and 6 µg/ml of ATO. There was a slight decrease in cytochrome c protein expression at 4 and 6 µg/ ml of ATO. Comet assay data revealed significant dose-dependent increases in the percentages of DNA damage, Comet tail lengths, and Comet tail moment. CONCLUSION: Taken together our results indicate that ATO is cytotoxic to lung cancer cells and its bioactivity is associated with oxidative damage, changes in cellular morphology, and apoptosis.

Arsenic trioxide modulates DNA synthesis and apoptosis in lung carcinoma cells.

Arsenic trioxide, the trade name Trisenox, is a drug used to treat acute promyleocytic leukemia (APL). Studies have demonstrated that arsenic trioxide slows cancer cells growth. Although arsenic influences numerous signal-transduction pathways, cell-cycle progression, and/or apoptosis, its apoptotic mechanisms are complex and not entirely delineated. The primary objective of this research was to evaluate the effects of arsenic trioxide on DNA synthesis and to determine whether arsenic-induced apoptosis is mediated via caspase activation, p38 mitogen-activated protein kinase (MAPK), and cell cycle arrest. To achieve this goal, lung cancer cells (A549) were exposed to various concentrations (0, 2, 4, 6, 8, and 10 microg/mL) of arsenic trioxide for 48 h. The effect of arsenic trioxide on DNA synthesis was determined by the [3H]thymidine incorporation assay. Apoptosis was determined by the caspase-3 fluorescein isothiocyanate (FITC) assay, p38 MAP kinase activity was determined by an immunoblot assay, and cell-cycle analysis was evaluated by the propidium iodide assay. The [3H]thymidine-incorporation assay revealed a dose-related cytotoxic response at high levels of exposure. Furthermore, arsenic trioxide modulated caspase 3 activity and induced p38 MAP kinase activation in A549 cells. However, cell-cycle studies showed no statistically significant differences in DNA content at subG1 check point between control and arsenic trioxide treated cells.

The effects of arsenic trioxide on DNA synthesis and genotoxicity in human colon cancer cells.

Colon cancer is the third leading cause of cancer-related deaths worldwide. Recent studies in our laboratory have demonstrated that arsenic trioxide is cytotoxic in human colon cancer (HT-29), lung (A549) and breast (MCF-7) carcinoma cells. The purpose of the present study is to investigate the effects of arsenic trioxide on DNA synthesis and the possible genotoxic effects on human colon cancer cells. HT-29 cells were cultured according to standard protocol, followed by exposure to various doses (0, 2, 4, 6, 8, 10, and 12 microg/mL) of arsenic trioxide for 24 h. The proliferative response (DNA synthesis) to arsenic trioxide was assessed by [(3)H]thymidine incorporation. The genotoxic effects of arsenic-induced DNA damage in a human colon cancer cell line was evaluated by the alkaline single cell gel electrophoresis. Results indicated that arsenic trioxide affected DNA synthesis in HT-29 cells in a biphasic manner; showing a slight but not significant increase in cell proliferation at lower levels of exposure (2, 4 and 6 microg/mL) followed by a significant inhibition of cell proliferation at higher doses (i.e., 8 and 10 microg/mL). The study also confirmed that arsenic trioxide exposure caused genotoxicity as revealed by the significant increase in DNA damage, comet tail-lengths, and tail moment when compared to non-exposed cells. Results of the [(3)H]thymidine incorporation assay and comet assay revealed that exposure to arsenic trioxide affected DNA synthesis and exhibited genotoxic effects in human colon cancer cells.

Arsenic Trioxide Modulates DNA Synthesis and Apoptosis in Lung Carcinoma Cells.

Arsenic is a heavy metal that is ubiquitous in the environment. The toxicity of arsenic depends upon its chemical form; the organic forms being usually less harmful than inorganic ones. The primary source of human exposure is through drinking water and food. Arsenic acts on cells through a variety of mechanisms, influencing numerous signal transduction pathways and resulting in a vast range of cellular effects that include apoptosis induction, growth inhibition, promotion or inhibition of differentiation, and angiogenesis inhibition. The primary objective of this research is to evaluate the effects of arsenic trioxide on DNA synthesis and to determine whether arsenic induces apoptosis via caspase activation and the activation the mitogen -activated protein kinase (MAPK) in lung carcinoma cells. To achieve this goal, the lung cancer (A549) cells were cultured following standard protocols, and exposed to various doses (0, 2, 4, 6, 8, and 10 mug/ml) of arsenic trioxide for 48 h with LC(50) being 7.8mug/ml. The proliferative response (DNA synthesis) to arsenic trioxide was determined by [(3)H] thymidine incorporation assay. Arsenic trioxide-induced apoptosis was determined by DNA laddering. Caspase -3 activation was assessed by the caspase-3 fluorescein isothiocyanate (FITC) assay. p38 MAP kinase activity was examined by immunoblot analysis using phospho p38 MAPK mab primary antibody in the presence of ATP and transcription factor (ATF-2) as a substrate. [(3)H] thymidine incorporation assay revealed biphasic reaction; showing cell proliferation at a lower level of exposure, and a dose-related cytotoxic response at higher levels of exposure in A549 cell line. Findings from the DNA laddering assay indicated that arsenic trioxide induced apoptosis in the lung carcinoma cells. Our findings revealed that arsenic trioxide modulated caspase 3 activity and induced p38 map kinase activation in lung carcinoma (A549) cells.

Cytotoxic Effect of Arsenic Trioxide in Adenocarcinoma Colorectal Cancer (HT-29) Cells.

Arsenic is a heavy metal that exhibits a high degree of toxicity to various organ systems. In humans, this compound is associated with an increase risk of skin cancer, and may cause cancers of the lung, liver, bladder, kidney, and colon. The mechanism of arsenic-related carcinogenicity remains to be elucidated. Hence, the aim of the present study was to investigate the cytotoxic effects of arsenic trioxide (As(2)O(3)) on adenocarcinoma colorectal cancer (HT-29) cells using the MTT [3-(4,5 dimethylthiazoyl-2-yl)-2,5- diphenyltetrazolium bromide] assay for cell viability. To achieve this objective, HT-29 cells were cultured and exposed to various doses (0, 2, 4, 6, 8, 10, 12, and 14 µg/ml) of arsenic trioxide for 24 h, 48 h, and 72 h respectively, and subsequently assessed for viability following a standard MTT test protocol. Experimental data indicated that arsenic trioxide is cytotoxic to colon cancer cells showing LD(50) values of 9.8, 9.4 and 9.0 µg/ml upon 24, 48 and 72 h of exposure, respectively. There was a dose-dependent response with regard to As(2)O(3) toxicity in HT-29 cells. Although there was a reduction in LD(50) value with increasing exposure time, this decrease was not statistically significant.