Additional file 1 of 3-Bromopyruvate-mediated MCT1-dependent metabolic perturbation sensitizes triple negative breast cancer cells to ionizing radiation

Additional file 1: Supplementary methods. Table S1. Charge, mass-to-charge ratio (m/z), retention time and formula of identified metabolites. Figure S1. Full western blots. (A) MCT1 expression in triple negative breast cancer (TNBC) cell lines, as presented in Fig. 1a. (B) MCT1 expression in native BT20 cells and BT20 cells transfected with siRNA against MCT1 (siMCT1), 1 and 2 days post-transfection, as presented in Fig. 1g. (C) Western blots for the assessment of MCT1 and MCT4 expression in the TNBC cell lines, BT20 and MDA-MB-231. Figure S2. RNA expression data for breast cancer cell lines. RNA expression of isoforms MCT1, 2 and 4, encoded by SLC16A1, 7 and 3, respectively were plotted for breast cancer cell lines. The RNA expression data for each isoform were derived from the Cancer Cell Line Encyclopedia (CCLE) database, expressed as log2(Fold Change) and plotted using Prism (GraphPad, CA, USA). Figure S3. Bromide level (78Br and 80Br) normalized to control measured by LC-MS/MS in lysates from BT20 cells and BT20 cells transfected with siRNA targeted against MCT1 (siMCT1-BT20) or scramble siRNA (sc-BT20) (n=6). 3BP uptake is significantly different between BT20 and siMCT1-BT20 at all time points, but no differences evident when comparing BT20 to the scrambled control cells. Error bars represent SD, **** represents P<0.001 as determined by multiple t-tests. Figure S4. Effect of sub-toxic concentration of 3BP on the extracellular acidification rate (ECAR), oxygen consumption rate (OCR) and the OCR/ECAR ratio of TNBC cells. BT20 and MDA-MB-231 cells were treated with 20 μM 3BP for 24 h in DMEM prior to measurement of the OCR and ECAR. Statistical significance was calculated using ‘unmatched’ two-way ANOVA, with the P value corrected for multiple comparisons using the Sidak test. Multiplicity adjusted P value is reported. N=4. Ns P>0.05, *P<0.05. Error bars represent the standard deviation from the mean.