Asparaginase induces selective dose‐ and time‐dependent cytotoxicity, apoptosis, and reduction of NFκB expression in oral cancer cells

Abstract

Asparaginase is fundamental to the treatment of haematological malignancies. However, little has been studied on the effects that asparaginase could exert on solid tumours. Thus, this study aimed to evaluate the effects of asparaginase on an oral carcinoma cell line. The cytotoxicity of asparaginase in SCC‐9 (tongue squamous cell carcinoma) and HaCaT (human keratinocyte) cell lines was evaluated with MTT cell viability assay. The cells were treated with asparaginase at 0.04, 0.16, 0.63, 1.0, 1.5, 2.5, and 5.0 IU/mL. Dose‐response curves and IC50 values were obtained and the Tumour Selectivity Index (TSI) was calculated. The effect of asparaginase on procaspase‐3 and nuclear factor κB (NFκB) expression was evaluated with western blot because it was reported that the overexpression of NFκB has been shown to contribute to tumour cell survival, proliferation, and migration. Caspase 3/7 staining was performed to identify cell death using flow cytometry. Effective asparaginase concentrations were lower for SCC‐9 cells when compared to HaCaT cells. The cytotoxicity results at 48 and 72 hours were significantly different for SCC‐9 cells. The TSI indicated that asparaginase was selective for the tumour cells. A decrease in procaspase‐3 and NFκB protein levels was observed in SCC‐9 cells. Furthermore, asparaginase resulted in significant apoptosis after 48 and 72 hours. Based on these results, asparaginase was cytotoxic in a dose‐ and time‐dependent manner, induces apoptosis, and reduces NFκB expression in oral cancer cells. These results encourage further studies on the effectiveness of this enzyme as a treatment for solid tumours, especially head and neck cancer.

Understanding the effect of measurement time on drug characterization

“…Our models are also not detailed enough to accurately capture the mechanisms of action of many chemotherapy agents [71]. These limitations are likely the reason there is discrepancy between our findings and experimental measurements [13][14][15][16][17]. However, the models examined here form the basis for many more complex tumor growth models [72][73][74][75], so their behavior needs to be understood in detail.…”

“…Other modeling studies have noted differences in IC 50 values with measurement time [9,12], with one finding a decrease in IC 50 with measurement time [9] and the other finding that IC 50 values increase with measurement time [12]. Experimental studies show more mixed results [9,[13][14][15][16][17], sometimes showing a decrease in IC 50 with measurement time [9,[13][14][15][16][17], sometimes showing an increase in IC 50 with measurement time [14,15,17], but also exhibiting more complex trends [13]. These discrepancies in model predictions indicate that much more work is needed to identify which models most accurately describe tumor growth [38,64] since these growth models are increasingly being used to guide patient treatment decisions [65].…”

“…The current method to find drug effect uses dose-response curves. Unfortunately, the measured IC 50 , as well as measured ε max , depend on the exact day that is chosen to make the measurement; this effect has been observed in both modeling studies [8][9][10][11][12] and in experimental studies [9,[13][14][15][16][17]. Cells are grown in the presence of various concentrations of drug and measurement of the number of cells on a particular day is used to determine the relative drug effect and the dose-response curve.…”