Figure 4.

Clonogenic survival data for HPV-positive SCCHN cell lines and for comparison, a typical TP53 mutant HPV-negative line (UM-SCC-81B). A stated in the methods, clonogenic assays were performed on the indicated SCCHN cell lines exposed to the indicated doses of IR. Cells were either left untreated or pre-treated with 25 mM 2-DG, 20 mM metformin (met), or combined treatment for one hour prior to irradiation. (A) displays clonogenic survival curves for each of the SCCHN cell lines and (B) the survival fractions at 2 Gy for each indicated treatment condition. Whilst 2-DG or metformin alone failed to have any significant effect on radiation response in any of the HPV-positive cell lines, the combination of 2-DG and metformin resulted in a significant reduction in clonogenic survival (in the case of the HPV-positive cell lines p-values represent the difference between the survival curves for the untreated condition and when combined pre-treatment with 2-DG and metformin was administered). In contrast, the addition of 2-DG alone consistently potentiated the effects of IR in UM-SCC-81B cells, while the addition of metformin has no discernible effect on clonogenic survival (the p-value in this case represents the difference between the survival curves for the untreated condition and when pre-treatment with 2-DG alone was administered). The results shown represent the mean values obtained from at least three separate experiments and error bars represent the SEMs. * p < 0.05, ** p < 0.005. (C) depicts representative example images of colony formation at 2 Gy for each treatment condition (a = IR alone; b = IR +2-DG; c = IR + 2-DG + Metformin; d = IR + Metformin) for the indicated cell lines.