Effects of celecoxib, Fenugreek and black seed on lung cancer/ in vitro study

Lung cancer is the leading cause (account for 18%) of cancer death in both men and women world-wide, causing 1.4 million deaths per year. The overall 5-year survival rate for all stages combined is disappointing (15%).
This study was done to evaluate the cytotoxic effects of each of celecoxib, fenugreek and black seed on lung cancer cell line alone and in combination with cisplatin, and their effects on protein 53 (p53) and epidermal growth factor receptor (EGFR) expressions. Large cell lung cancer (QU-DB) cells were cultured in Eagle's minimum essential medium culture media supplemented with 5% fetal bovine serum and antibiotics. The cells were seeded in 96 well plate and the cytotoxic effects of each of cisplatin [25-0.195 μl/ml (or μg/ml)], celecoxib [300-1.1719 μl/ml (=1500-5.86 μg/ml)], fenugreek [300-1.1719 μl/ml (each one μl is extracted from 25 μg of dried seed)] and black seed [300-1.1719 μl/ml (each one μl is extracted from 25 μg of dried seed)] was determined using neutral red uptake (NRU) assay for 24, 48, and 72 hours in comparison with their corresponding control groups. Combined effect of each of the above agents with cisplatin was determined also using NRU assay. Cytotoxicity was further assessed by trypan blue exclusion assay at IC50 of each agent for 48 hours duration. Immunocytochemistry assay was performed also to detect p53 and EGFR expression. Cisplatin induced a directly proportional, dose-dependent and timedependant cytotoxic effect with an IC50 of 8.5 μg/ml and 7.3 μg/ml after 48 hrs and 72 hrs of exposure, respectively. Significant differences (p<0.05) were observed in optic density of cisplatin group from that of the control for all tested concentrations. Celecoxib induced a directly proportional, dose-dependent and timedependant cytotoxic effect with an IC50 of 162 μl/ml (=810 μg/ml) and 88 μl/ml (=440 μg/ml) after 48 hrs and 72 hrs of exposure, respectively. Significant differences (p<0.05) were observed in optic density of celecoxib from that of the control at concentrations of 75 μl/ml (=375 μg/ml) and above. Fenugreek extract also induced a directly proportional, dose-dependent and time-dependant cytotoxic effect in experiments with 48 hrs and 72 hrs of exposure with an IC50 of 88.25 μl/ml and 125 μl/ml, respectively (each one μl is extracted from 25 μg of dried seed). While, it produces a protective effect in 24 hrs exposure experiment. Significant differences (p<0.05) were observed in optic density of fenugreek from that of the control at concentrations of 37.5 μl/ml and above. Black seed extract induced a directly proportional, dose-dependent and time-dependant cytotoxic effect in experiments with 48 hrs and 72 hrs of exposure with an IC50 of 149.5 μl/ml and 130 μl/ml, respectively (each one μl is extracted from 25 μg of dried seed). While, it produces a protective effect in 24 hrs exposure experiment. Significant differences (p<0.05) were observed in optic density of black seed from that of the control at concentrations of 75 μl/ml and above. Celecoxib, fenugreek and black seed produce an antagonistic action when combined with cisplatin, combination index (CI) >1.3. Cisplatin and celecoxib highly significantly (p<0.005) increased EGFR expression at different concentrations. While, fenugreek and black seed highly significantly (p<0.005) reduced EGFR expression at 300 μl/ml. Cisplatin, fenugreek and black seed highly significantly (p<0.005) decreased the expression of p53, while celecoxib highly significantly (p<0.005) increased it. In conclusion, monotherapy of each of celecoxib, fenugreek and black seed have anticancer effects on lung cancer cell line, but an antagonizing effect to cisplatin when combined with it. Also, each of fenugreek and black seed may have a beneficial therapeutic effect in decreasing EGFR expression and decreasing mutant p53 expression. On the other hand, celecoxib may induce its cytotoxic effects by other mechanisms.